From b94d105218e570065605bbcdc029be4c5aff6660 Mon Sep 17 00:00:00 2001 From: Maxime U Garcia Date: Wed, 14 Jun 2023 13:15:05 +0200 Subject: [PATCH 001/184] Update public_aws_ecr.config --- conf/public_aws_ecr.config | 5 +++-- 1 file changed, 3 insertions(+), 2 deletions(-) diff --git a/conf/public_aws_ecr.config b/conf/public_aws_ecr.config index f584b3926..fa0da7c7a 100644 --- a/conf/public_aws_ecr.config +++ b/conf/public_aws_ecr.config @@ -8,8 +8,9 @@ ---------------------------------------------------------------------------------------- */ -docker.registry = 'public.ecr.aws' -podman.registry = 'public.ecr.aws' +docker.registry = 'public.ecr.aws' +podman.registry = 'public.ecr.aws' +singularity.registry = 'public.ecr.aws' process { withName: 'CAT_ADDITIONAL_FASTA' { From 2e936fdb40c7af00f0ab8df7587bb839868f26d8 Mon Sep 17 00:00:00 2001 From: Matthias Zepper Date: Wed, 14 Jun 2023 17:01:29 +0200 Subject: [PATCH 002/184] Display a warning when '--star_extra_alignment_args' are used in combination with the STAR-RSEM route. --- CHANGELOG.md | 6 ++++++ lib/WorkflowRnaseq.groovy | 16 ++++++++++++++++ nextflow_schema.json | 2 +- 3 files changed, 23 insertions(+), 1 deletion(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index daae6c447..13d0707a2 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -3,6 +3,12 @@ The format is based on [Keep a Changelog](https://keepachangelog.com/en/1.0.0/) and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0.html). +## v3.13.0dev - [date] + +### Enhancements & fixes + +- [PR #1049](https://github.com/nf-core/rnaseq/pull/1049) - RSEM imposes own arguments to STAR, therefore `--star_extra_alignment_args` are ignored for this route. A warning is displayed when both are used together. + ## [[3.12.0](https://github.com/nf-core/rnaseq/releases/tag/3.12.0)] - 2023-06-02 ### Credits diff --git a/lib/WorkflowRnaseq.groovy b/lib/WorkflowRnaseq.groovy index 08d684e77..ed2f4356a 100755 --- a/lib/WorkflowRnaseq.groovy +++ b/lib/WorkflowRnaseq.groovy @@ -86,6 +86,9 @@ class WorkflowRnaseq { if (params.rsem_index && params.star_index) { rsemStarIndexWarn(log) } + if (params.star_extra_alignment_args) { + rsemStarExtraArgumentsWarn(log) + } } // Warn if --additional_fasta provided with aligner index @@ -394,6 +397,19 @@ class WorkflowRnaseq { "~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~" } + // + // Print a warning if using '--aligner star_rsem' and providing '--star_extra_alignment_args' + // + private static void rsemStarExtraArgumentsWarn(log) { + log.warn "~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~\n" + + " No additional arguments can be passed to STAR when using RSEM.\n" + + " Because RSEM enforces its own parameters for STAR, any extra arguments\n" + + " to STAR will be ignored. Alternatively, choose the STAR+Salmon route.\n\n" + + " This warning has been generated because you have provided both\n" + + " '--aligner star_rsem' and '--extra_star_align_args'.\n\n" + + "~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~" + } + // // Print a warning if using '--additional_fasta' and '--_index' // diff --git a/nextflow_schema.json b/nextflow_schema.json index 4bb08ed2d..e090cd02c 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -417,7 +417,7 @@ }, "extra_star_align_args": { "type": "string", - "description": "Extra arguments to pass to STAR alignment command in addition to defaults defined by the pipeline.", + "description": "Extra arguments to pass to STAR alignment command in addition to defaults defined by the pipeline. Only available for the STAR-Salmon route.", "fa_icon": "fas fa-plus" }, "extra_salmon_quant_args": { From dff30b5c3c3b065c6104539ac27fba345d58ae77 Mon Sep 17 00:00:00 2001 From: Matthias Zepper <6963520+MatthiasZepper@users.noreply.github.com> Date: Wed, 14 Jun 2023 18:20:39 +0200 Subject: [PATCH 003/184] Apply suggestions from code review Co-authored-by: Harshil Patel --- CHANGELOG.md | 2 +- lib/WorkflowRnaseq.groovy | 2 +- 2 files changed, 2 insertions(+), 2 deletions(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index 13d0707a2..90cba0311 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -7,7 +7,7 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 ### Enhancements & fixes -- [PR #1049](https://github.com/nf-core/rnaseq/pull/1049) - RSEM imposes own arguments to STAR, therefore `--star_extra_alignment_args` are ignored for this route. A warning is displayed when both are used together. +- [PR #1049](https://github.com/nf-core/rnaseq/pull/1049) - RSEM imposes own arguments to STAR, therefore `--extra_star_align_args` are ignored for this route. A warning is displayed when both are used together. ## [[3.12.0](https://github.com/nf-core/rnaseq/releases/tag/3.12.0)] - 2023-06-02 diff --git a/lib/WorkflowRnaseq.groovy b/lib/WorkflowRnaseq.groovy index ed2f4356a..0dd946596 100755 --- a/lib/WorkflowRnaseq.groovy +++ b/lib/WorkflowRnaseq.groovy @@ -86,7 +86,7 @@ class WorkflowRnaseq { if (params.rsem_index && params.star_index) { rsemStarIndexWarn(log) } - if (params.star_extra_alignment_args) { + if (params.aligner == 'star_rsem' && params.extra_star_align_args) { rsemStarExtraArgumentsWarn(log) } } From 8261c4e5e0be898f9c0fa9a6556479880c0074b7 Mon Sep 17 00:00:00 2001 From: Adam Talbot <12817534+adamrtalbot@users.noreply.github.com> Date: Tue, 20 Jun 2023 12:00:56 +0000 Subject: [PATCH 004/184] Remove public_aws_ecr profile --- .github/workflows/cloud_tests_full.yml | 2 +- .github/workflows/cloud_tests_small.yml | 2 +- CHANGELOG.md | 4 ++ conf/public_aws_ecr.config | 73 ------------------------- nextflow.config | 7 +-- 5 files changed, 8 insertions(+), 80 deletions(-) delete mode 100644 conf/public_aws_ecr.config diff --git a/.github/workflows/cloud_tests_full.yml b/.github/workflows/cloud_tests_full.yml index 4738d9045..3c4b23106 100644 --- a/.github/workflows/cloud_tests_full.yml +++ b/.github/workflows/cloud_tests_full.yml @@ -30,7 +30,7 @@ jobs: compute_env: ${{ secrets.TOWER_CE_AWS_CPU }} workdir: "${{ secrets.TOWER_BUCKET_AWS }}/work/rnaseq/work-${{ github.sha }}" run_name: "aws_rnaseq_full_${{ matrix.aligner }}" - profiles: test_full_aws,public_aws_ecr + profiles: test_full_aws parameters: | { "hook_url": "${{ secrets.MEGATESTS_ALERTS_SLACK_HOOK_URL }}", diff --git a/.github/workflows/cloud_tests_small.yml b/.github/workflows/cloud_tests_small.yml index 1f430067b..6f2e6d8a9 100644 --- a/.github/workflows/cloud_tests_small.yml +++ b/.github/workflows/cloud_tests_small.yml @@ -25,7 +25,7 @@ jobs: compute_env: ${{ secrets.TOWER_CE_AWS_CPU }} workdir: "${{ secrets.TOWER_BUCKET_AWS }}/work/rnaseq/work-${{ github.sha }}" run_name: "aws_rnaseq_small" - profiles: test,public_aws_ecr + profiles: test parameters: | { "outdir": "${{ secrets.TOWER_BUCKET_AWS }}/rnaseq/results-test-${{ github.sha }}" diff --git a/CHANGELOG.md b/CHANGELOG.md index 90cba0311..de22e651c 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -9,6 +9,10 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 - [PR #1049](https://github.com/nf-core/rnaseq/pull/1049) - RSEM imposes own arguments to STAR, therefore `--extra_star_align_args` are ignored for this route. A warning is displayed when both are used together. +### Removed + +- Remove public_aws_ecr profile. + ## [[3.12.0](https://github.com/nf-core/rnaseq/releases/tag/3.12.0)] - 2023-06-02 ### Credits diff --git a/conf/public_aws_ecr.config b/conf/public_aws_ecr.config deleted file mode 100644 index fa0da7c7a..000000000 --- a/conf/public_aws_ecr.config +++ /dev/null @@ -1,73 +0,0 @@ -/* -~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ - AWS ECR Config -~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ - Config to set public AWS ECR images wherever possible - This improves speed when running on AWS infrastructure. - Use this as an example template when using your own private registry. ----------------------------------------------------------------------------------------- -*/ - -docker.registry = 'public.ecr.aws' -podman.registry = 'public.ecr.aws' -singularity.registry = 'public.ecr.aws' - -process { - withName: 'CAT_ADDITIONAL_FASTA' { - container = 'quay.io/biocontainers/python:3.9--1' - } - withName: 'CAT_FASTQ' { - container = 'quay.io/nf-core/ubuntu:20.04' - } - withName: 'DESEQ2_QC' { - container = 'quay.io/biocontainers/mulled-v2-8849acf39a43cdd6c839a369a74c0adc823e2f91:ab110436faf952a33575c64dd74615a84011450b-0' - } - withName: 'GTF2BED' { - container = 'quay.io/biocontainers/perl:5.26.2' - } - withName: 'GTF_GENE_FILTER' { - container = 'quay.io/biocontainers/python:3.9--1' - } - withName: 'GUNZIP' { - container = 'quay.io/nf-core/ubuntu:20.04' - } - withName: 'HISAT2_ALIGN' { - container = 'quay.io/biocontainers/mulled-v2-a97e90b3b802d1da3d6958e0867610c718cb5eb1:2cdf6bf1e92acbeb9b2834b1c58754167173a410-0' - } - withName: 'MULTIQC_CUSTOM_BIOTYPE' { - container = 'quay.io/biocontainers/python:3.9--1' - } - withName: 'PREPROCESS_TRANSCRIPTS_FASTA_GENCODE' { - container = 'quay.io/nf-core/ubuntu:20.04' - } - withName: 'RSEM_CALCULATEEXPRESSION' { - container = 'quay.io/biocontainers/mulled-v2-cf0123ef83b3c38c13e3b0696a3f285d3f20f15b:64aad4a4e144878400649e71f42105311be7ed87-0' - } - withName: 'RSEM_MERGE_COUNTS' { - container = 'quay.io/nf-core/ubuntu:20.04' - } - withName: 'RSEM_PREPAREREFERENCE' { - container = 'quay.io/biocontainers/mulled-v2-cf0123ef83b3c38c13e3b0696a3f285d3f20f15b:64aad4a4e144878400649e71f42105311be7ed87-0' - } - withName: 'SALMON_TX2GENE' { - container = 'quay.io/biocontainers/python:3.9--1' - } - withName: 'SAMPLESHEET_CHECK' { - container = 'quay.io/biocontainers/python:3.9--1' - } - withName: 'STAR_ALIGN' { - container = 'quay.io/biocontainers/mulled-v2-1fa26d1ce03c295fe2fdcf85831a92fbcbd7e8c2:1df389393721fc66f3fd8778ad938ac711951107-0' - } - withName: 'STAR_ALIGN_IGENOMES' { - container = 'quay.io/biocontainers/mulled-v2-1fa26d1ce03c295fe2fdcf85831a92fbcbd7e8c2:59cdd445419f14abac76b31dd0d71217994cbcc9-0' - } - withName: 'STAR_GENOMEGENERATE' { - container = 'quay.io/biocontainers/mulled-v2-1fa26d1ce03c295fe2fdcf85831a92fbcbd7e8c2:1df389393721fc66f3fd8778ad938ac711951107-0' - } - withName: 'STAR_GENOMEGENERATE_IGENOMES' { - container = 'quay.io/biocontainers/mulled-v2-1fa26d1ce03c295fe2fdcf85831a92fbcbd7e8c2:59cdd445419f14abac76b31dd0d71217994cbcc9-0' - } - withName: 'UNTAR' { - container = 'quay.io/nf-core/ubuntu:20.04' - } -} diff --git a/nextflow.config b/nextflow.config index 09bed2da2..c6dab1e2e 100644 --- a/nextflow.config +++ b/nextflow.config @@ -231,14 +231,11 @@ profiles { executor.cpus = 16 executor.memory = 60.GB } - public_aws_ecr { - includeConfig 'conf/public_aws_ecr.config' - } test { includeConfig 'conf/test.config' } test_cache { includeConfig 'conf/test_cache.config' } test_full { includeConfig 'conf/test_full.config' } - test_full_aws { - includeConfig 'conf/test_full.config' + test_full_aws { + includeConfig 'conf/test_full.config' } test_full_gcp { includeConfig 'conf/test_full.config' From 844893e3b7f626670abf0da435b3354651315faa Mon Sep 17 00:00:00 2001 From: Adam Talbot <12817534+adamrtalbot@users.noreply.github.com> Date: Tue, 20 Jun 2023 12:04:18 +0000 Subject: [PATCH 005/184] CHANGELOG --- CHANGELOG.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index de22e651c..81a0bde99 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -11,7 +11,7 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 ### Removed -- Remove public_aws_ecr profile. +- [PR #1051](https://github.com/nf-core/rnaseq/pull/1051) - Remove public_aws_ecr profile. ## [[3.12.0](https://github.com/nf-core/rnaseq/releases/tag/3.12.0)] - 2023-06-02 From 11577f0fdf7b638c37e7359e8014ea7f348213ab Mon Sep 17 00:00:00 2001 From: nf-core-bot Date: Fri, 30 Jun 2023 16:15:29 +0000 Subject: [PATCH 006/184] Template update for nf-core/tools version 2.9 --- .github/CONTRIBUTING.md | 1 - .github/ISSUE_TEMPLATE/bug_report.yml | 2 +- .github/workflows/awsfulltest.yml | 11 +- .github/workflows/awstest.yml | 10 +- .github/workflows/ci.yml | 2 +- .gitpod.yml | 5 + CHANGELOG.md | 2 +- CITATIONS.md | 6 + README.md | 6 +- assets/methods_description_template.yml | 12 +- assets/multiqc_config.yml | 4 +- assets/nf-core-rnaseq_logo_light.png | Bin 10453 -> 71212 bytes assets/slackreport.json | 2 +- conf/test_full.config | 2 - docs/usage.md | 6 +- lib/NfcoreSchema.groovy | 530 ------------------------ lib/NfcoreTemplate.groovy | 2 +- lib/WorkflowMain.groovy | 37 -- lib/WorkflowRnaseq.groovy | 45 +- main.nf | 16 + nextflow.config | 54 ++- nextflow_schema.json | 36 +- workflows/rnaseq.nf | 25 +- 23 files changed, 177 insertions(+), 639 deletions(-) delete mode 100755 lib/NfcoreSchema.groovy diff --git a/.github/CONTRIBUTING.md b/.github/CONTRIBUTING.md index f40da9db0..fe09aec68 100644 --- a/.github/CONTRIBUTING.md +++ b/.github/CONTRIBUTING.md @@ -116,4 +116,3 @@ To get started: Devcontainer specs: - [DevContainer config](.devcontainer/devcontainer.json) -- [Dockerfile](.devcontainer/Dockerfile) diff --git a/.github/ISSUE_TEMPLATE/bug_report.yml b/.github/ISSUE_TEMPLATE/bug_report.yml index 3ce613833..60b5e3f4e 100644 --- a/.github/ISSUE_TEMPLATE/bug_report.yml +++ b/.github/ISSUE_TEMPLATE/bug_report.yml @@ -42,7 +42,7 @@ body: attributes: label: System information description: | - * Nextflow version _(eg. 22.10.1)_ + * Nextflow version _(eg. 23.04.0)_ * Hardware _(eg. HPC, Desktop, Cloud)_ * Executor _(eg. slurm, local, awsbatch)_ * Container engine: _(e.g. Docker, Singularity, Conda, Podman, Shifter, Charliecloud, or Apptainer)_ diff --git a/.github/workflows/awsfulltest.yml b/.github/workflows/awsfulltest.yml index 2e6baf508..fdf7c0446 100644 --- a/.github/workflows/awsfulltest.yml +++ b/.github/workflows/awsfulltest.yml @@ -14,7 +14,7 @@ jobs: runs-on: ubuntu-latest steps: - name: Launch workflow via tower - uses: seqeralabs/action-tower-launch@v1 + uses: seqeralabs/action-tower-launch@v2 # TODO nf-core: You can customise AWS full pipeline tests as required # Add full size test data (but still relatively small datasets for few samples) # on the `test_full.config` test runs with only one set of parameters @@ -22,13 +22,18 @@ jobs: workspace_id: ${{ secrets.TOWER_WORKSPACE_ID }} access_token: ${{ secrets.TOWER_ACCESS_TOKEN }} compute_env: ${{ secrets.TOWER_COMPUTE_ENV }} + revision: ${{ github.sha }} workdir: s3://${{ secrets.AWS_S3_BUCKET }}/work/rnaseq/work-${{ github.sha }} parameters: | { + "hook_url": "${{ secrets.MEGATESTS_ALERTS_SLACK_HOOK_URL }}", "outdir": "s3://${{ secrets.AWS_S3_BUCKET }}/rnaseq/results-${{ github.sha }}" } - profiles: test_full,aws_tower + profiles: test_full + - uses: actions/upload-artifact@v3 with: name: Tower debug log file - path: tower_action_*.log + path: | + tower_action_*.log + tower_action_*.json diff --git a/.github/workflows/awstest.yml b/.github/workflows/awstest.yml index 9ce9dacb5..ceab65e65 100644 --- a/.github/workflows/awstest.yml +++ b/.github/workflows/awstest.yml @@ -12,18 +12,22 @@ jobs: steps: # Launch workflow using Tower CLI tool action - name: Launch workflow via tower - uses: seqeralabs/action-tower-launch@v1 + uses: seqeralabs/action-tower-launch@v2 with: workspace_id: ${{ secrets.TOWER_WORKSPACE_ID }} access_token: ${{ secrets.TOWER_ACCESS_TOKEN }} compute_env: ${{ secrets.TOWER_COMPUTE_ENV }} + revision: ${{ github.sha }} workdir: s3://${{ secrets.AWS_S3_BUCKET }}/work/rnaseq/work-${{ github.sha }} parameters: | { "outdir": "s3://${{ secrets.AWS_S3_BUCKET }}/rnaseq/results-test-${{ github.sha }}" } - profiles: test,aws_tower + profiles: test + - uses: actions/upload-artifact@v3 with: name: Tower debug log file - path: tower_action_*.log + path: | + tower_action_*.log + tower_action_*.json diff --git a/.github/workflows/ci.yml b/.github/workflows/ci.yml index debe9dcbf..9f7050723 100644 --- a/.github/workflows/ci.yml +++ b/.github/workflows/ci.yml @@ -24,7 +24,7 @@ jobs: strategy: matrix: NXF_VER: - - "22.10.1" + - "23.04.0" - "latest-everything" steps: - name: Check out pipeline code diff --git a/.gitpod.yml b/.gitpod.yml index 85d95ecc8..25488dcc0 100644 --- a/.gitpod.yml +++ b/.gitpod.yml @@ -1,4 +1,9 @@ image: nfcore/gitpod:latest +tasks: + - name: Update Nextflow and setup pre-commit + command: | + pre-commit install --install-hooks + nextflow self-update vscode: extensions: # based on nf-core.nf-core-extensionpack diff --git a/CHANGELOG.md b/CHANGELOG.md index 31a919f72..49da6be25 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -3,7 +3,7 @@ The format is based on [Keep a Changelog](https://keepachangelog.com/en/1.0.0/) and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0.html). -## v3.12.0dev - [date] +## v3.12.0 - [date] Initial release of nf-core/rnaseq, created with the [nf-core](https://nf-co.re/) template. diff --git a/CITATIONS.md b/CITATIONS.md index cee10e582..f7874b710 100644 --- a/CITATIONS.md +++ b/CITATIONS.md @@ -12,7 +12,10 @@ - [FastQC](https://www.bioinformatics.babraham.ac.uk/projects/fastqc/) + > Andrews, S. (2010). FastQC: A Quality Control Tool for High Throughput Sequence Data [Online]. Available online https://www.bioinformatics.babraham.ac.uk/projects/fastqc/. + - [MultiQC](https://pubmed.ncbi.nlm.nih.gov/27312411/) + > Ewels P, Magnusson M, Lundin S, Käller M. MultiQC: summarize analysis results for multiple tools and samples in a single report. Bioinformatics. 2016 Oct 1;32(19):3047-8. doi: 10.1093/bioinformatics/btw354. Epub 2016 Jun 16. PubMed PMID: 27312411; PubMed Central PMCID: PMC5039924. ## Software packaging/containerisation tools @@ -31,5 +34,8 @@ - [Docker](https://dl.acm.org/doi/10.5555/2600239.2600241) + > Merkel, D. (2014). Docker: lightweight linux containers for consistent development and deployment. Linux Journal, 2014(239), 2. doi: 10.5555/2600239.2600241. + - [Singularity](https://pubmed.ncbi.nlm.nih.gov/28494014/) + > Kurtzer GM, Sochat V, Bauer MW. Singularity: Scientific containers for mobility of compute. PLoS One. 2017 May 11;12(5):e0177459. doi: 10.1371/journal.pone.0177459. eCollection 2017. PubMed PMID: 28494014; PubMed Central PMCID: PMC5426675. diff --git a/README.md b/README.md index f86ed8d60..84b18b356 100644 --- a/README.md +++ b/README.md @@ -2,7 +2,7 @@ [![AWS CI](https://img.shields.io/badge/CI%20tests-full%20size-FF9900?labelColor=000000&logo=Amazon%20AWS)](https://nf-co.re/rnaseq/results)[![Cite with Zenodo](http://img.shields.io/badge/DOI-10.5281/zenodo.XXXXXXX-1073c8?labelColor=000000)](https://doi.org/10.5281/zenodo.XXXXXXX) -[![Nextflow](https://img.shields.io/badge/nextflow%20DSL2-%E2%89%A522.10.1-23aa62.svg)](https://www.nextflow.io/) +[![Nextflow](https://img.shields.io/badge/nextflow%20DSL2-%E2%89%A523.04.0-23aa62.svg)](https://www.nextflow.io/) [![run with conda](http://img.shields.io/badge/run%20with-conda-3EB049?labelColor=000000&logo=anaconda)](https://docs.conda.io/en/latest/) [![run with docker](https://img.shields.io/badge/run%20with-docker-0db7ed?labelColor=000000&logo=docker)](https://www.docker.com/) [![run with singularity](https://img.shields.io/badge/run%20with-singularity-1d355c.svg?labelColor=000000)](https://sylabs.io/docs/) @@ -66,11 +66,11 @@ nextflow run nf-core/rnaseq \ > provided by the `-c` Nextflow option can be used to provide any configuration _**except for parameters**_; > see [docs](https://nf-co.re/usage/configuration#custom-configuration-files). -For more details, please refer to the [usage documentation](https://nf-co.re/rnaseq/usage) and the [parameter documentation](https://nf-co.re/rnaseq/parameters). +For more details and further functionality, please refer to the [usage documentation](https://nf-co.re/rnaseq/usage) and the [parameter documentation](https://nf-co.re/rnaseq/parameters). ## Pipeline output -To see the the results of a test run with a full size dataset refer to the [results](https://nf-co.re/rnaseq/results) tab on the nf-core website pipeline page. +To see the results of an example test run with a full size dataset refer to the [results](https://nf-co.re/rnaseq/results) tab on the nf-core website pipeline page. For more details about the output files and reports, please refer to the [output documentation](https://nf-co.re/rnaseq/output). diff --git a/assets/methods_description_template.yml b/assets/methods_description_template.yml index c263da31b..56c961c8e 100644 --- a/assets/methods_description_template.yml +++ b/assets/methods_description_template.yml @@ -3,17 +3,21 @@ description: "Suggested text and references to use when describing pipeline usag section_name: "nf-core/rnaseq Methods Description" section_href: "https://github.com/nf-core/rnaseq" plot_type: "html" -## TODO nf-core: Update the HTML below to your prefered methods description, e.g. add publication citation for this pipeline +## TODO nf-core: Update the HTML below to your preferred methods description, e.g. add publication citation for this pipeline ## You inject any metadata in the Nextflow '${workflow}' object data: |

Methods

-

Data was processed using nf-core/rnaseq v${workflow.manifest.version} ${doi_text} of the nf-core collection of workflows (Ewels et al., 2020).

+

Data was processed using nf-core/rnaseq v${workflow.manifest.version} ${doi_text} of the nf-core collection of workflows (Ewels et al., 2020), utilising reproducible software environments from the Bioconda (Grüning et al., 2018) and Biocontainers (da Veiga Leprevost et al., 2017) projects.

The pipeline was executed with Nextflow v${workflow.nextflow.version} (Di Tommaso et al., 2017) with the following command:

${workflow.commandLine}
+

${tool_citations}

References

    -
  • Di Tommaso, P., Chatzou, M., Floden, E. W., Barja, P. P., Palumbo, E., & Notredame, C. (2017). Nextflow enables reproducible computational workflows. Nature Biotechnology, 35(4), 316-319. https://doi.org/10.1038/nbt.3820
  • -
  • Ewels, P. A., Peltzer, A., Fillinger, S., Patel, H., Alneberg, J., Wilm, A., Garcia, M. U., Di Tommaso, P., & Nahnsen, S. (2020). The nf-core framework for community-curated bioinformatics pipelines. Nature Biotechnology, 38(3), 276-278. https://doi.org/10.1038/s41587-020-0439-x
  • +
  • Di Tommaso, P., Chatzou, M., Floden, E. W., Barja, P. P., Palumbo, E., & Notredame, C. (2017). Nextflow enables reproducible computational workflows. Nature Biotechnology, 35(4), 316-319. doi: 10.1038/nbt.3820
  • +
  • Ewels, P. A., Peltzer, A., Fillinger, S., Patel, H., Alneberg, J., Wilm, A., Garcia, M. U., Di Tommaso, P., & Nahnsen, S. (2020). The nf-core framework for community-curated bioinformatics pipelines. Nature Biotechnology, 38(3), 276-278. doi: 10.1038/s41587-020-0439-x
  • +
  • Grüning, B., Dale, R., Sjödin, A., Chapman, B. A., Rowe, J., Tomkins-Tinch, C. H., Valieris, R., Köster, J., & Bioconda Team. (2018). Bioconda: sustainable and comprehensive software distribution for the life sciences. Nature Methods, 15(7), 475–476. doi: 10.1038/s41592-018-0046-7
  • +
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  • + ${tool_bibliography}
Notes:
diff --git a/assets/multiqc_config.yml b/assets/multiqc_config.yml index 721fdfcb5..59e039077 100644 --- a/assets/multiqc_config.yml +++ b/assets/multiqc_config.yml @@ -1,7 +1,7 @@ report_comment: > - This report has been generated by the nf-core/rnaseq + This report has been generated by the nf-core/rnaseq analysis pipeline. 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"author_name": "sanger-tol/readmapping v${version} - ${runName}", + "author_name": "nf-core/rnaseq v${version} - ${runName}", "author_icon": "https://www.nextflow.io/docs/latest/_static/favicon.ico", "text": "<% if (success) { %>Pipeline completed successfully!<% } else { %>Pipeline completed with errors<% } %>", "fields": [ diff --git a/conf/test_full.config b/conf/test_full.config index 6d70c5c8e..505808b89 100644 --- a/conf/test_full.config +++ b/conf/test_full.config @@ -10,8 +10,6 @@ ---------------------------------------------------------------------------------------- */ -cleanup = true - params { config_profile_name = 'Full test profile' config_profile_description = 'Full test dataset to check pipeline function' diff --git a/docs/usage.md b/docs/usage.md index 1e45b8d78..7191db88e 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -57,7 +57,7 @@ An [example samplesheet](../assets/samplesheet.csv) has been provided with the p The typical command for running the pipeline is as follows: ```bash -nextflow run nf-core/rnaseq --input samplesheet.csv --outdir --genome GRCh37 -profile docker +nextflow run nf-core/rnaseq --input ./samplesheet.csv --outdir ./results --genome GRCh37 -profile docker ``` This will launch the pipeline with the `docker` configuration profile. See below for more information about profiles. @@ -76,7 +76,8 @@ If you wish to repeatedly use the same parameters for multiple runs, rather than Pipeline settings can be provided in a `yaml` or `json` file via `-params-file `. > ⚠️ Do not use `-c ` to specify parameters as this will result in errors. Custom config files specified with `-c` must only be used for [tuning process resource specifications](https://nf-co.re/docs/usage/configuration#tuning-workflow-resources), other infrastructural tweaks (such as output directories), or module arguments (args). -> The above pipeline run specified with a params file in yaml format: + +The above pipeline run specified with a params file in yaml format: ```bash nextflow run nf-core/rnaseq -profile docker -params-file params.yaml @@ -88,7 +89,6 @@ with `params.yaml` containing: input: './samplesheet.csv' outdir: './results/' genome: 'GRCh37' -input: 'data' <...> ``` diff --git a/lib/NfcoreSchema.groovy b/lib/NfcoreSchema.groovy deleted file mode 100755 index 9b34804d6..000000000 --- a/lib/NfcoreSchema.groovy +++ /dev/null @@ -1,530 +0,0 @@ -// -// This file holds several functions used to perform JSON parameter validation, help and summary rendering for the nf-core pipeline template. -// - -import nextflow.Nextflow -import org.everit.json.schema.Schema -import org.everit.json.schema.loader.SchemaLoader -import org.everit.json.schema.ValidationException -import org.json.JSONObject -import org.json.JSONTokener -import org.json.JSONArray -import groovy.json.JsonSlurper -import groovy.json.JsonBuilder - -class NfcoreSchema { - - // - // Resolve Schema path relative to main workflow directory - // - public static String getSchemaPath(workflow, schema_filename='nextflow_schema.json') { - return "${workflow.projectDir}/${schema_filename}" - } - - // - // Function to loop over all parameters defined in schema and check - // whether the given parameters adhere to the specifications - // - /* groovylint-disable-next-line UnusedPrivateMethodParameter */ - public static void validateParameters(workflow, params, log, schema_filename='nextflow_schema.json') { - def has_error = false - //~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~// - // Check for nextflow core params and unexpected params - def json = new File(getSchemaPath(workflow, schema_filename=schema_filename)).text - def Map schemaParams = (Map) new JsonSlurper().parseText(json).get('definitions') - def nf_params = [ - // Options for base `nextflow` command - 'bg', - 'c', - 'C', - 'config', - 'd', - 'D', - 'dockerize', - 'h', - 'log', - 'q', - 'quiet', - 'syslog', - 'v', - - // Options for `nextflow run` command - 'ansi', - 'ansi-log', - 'bg', - 'bucket-dir', - 'c', - 'cache', - 'config', - 'dsl2', - 'dump-channels', - 'dump-hashes', - 'E', - 'entry', - 'latest', - 'lib', - 'main-script', - 'N', - 'name', - 'offline', - 'params-file', - 'pi', - 'plugins', - 'poll-interval', - 'pool-size', - 'profile', - 'ps', - 'qs', - 'queue-size', - 'r', - 'resume', - 'revision', - 'stdin', - 'stub', - 'stub-run', - 'test', - 'w', - 'with-apptainer', - 'with-charliecloud', - 'with-conda', - 'with-dag', - 'with-docker', - 'with-mpi', - 'with-notification', - 'with-podman', - 'with-report', - 'with-singularity', - 'with-timeline', - 'with-tower', - 'with-trace', - 'with-weblog', - 'without-docker', - 'without-podman', - 'work-dir' - ] - def unexpectedParams = [] - - // Collect expected parameters from the schema - def expectedParams = [] - def enums = [:] - for (group in schemaParams) { - for (p in group.value['properties']) { - expectedParams.push(p.key) - if (group.value['properties'][p.key].containsKey('enum')) { - enums[p.key] = group.value['properties'][p.key]['enum'] - } - } - } - - for (specifiedParam in params.keySet()) { - // nextflow params - if (nf_params.contains(specifiedParam)) { - log.error "ERROR: You used a core Nextflow option with two hyphens: '--${specifiedParam}'. Please resubmit with '-${specifiedParam}'" - has_error = true - } - // unexpected params - def params_ignore = params.schema_ignore_params.split(',') + 'schema_ignore_params' - def expectedParamsLowerCase = expectedParams.collect{ it.replace("-", "").toLowerCase() } - def specifiedParamLowerCase = specifiedParam.replace("-", "").toLowerCase() - def isCamelCaseBug = (specifiedParam.contains("-") && !expectedParams.contains(specifiedParam) && expectedParamsLowerCase.contains(specifiedParamLowerCase)) - if (!expectedParams.contains(specifiedParam) && !params_ignore.contains(specifiedParam) && !isCamelCaseBug) { - // Temporarily remove camelCase/camel-case params #1035 - def unexpectedParamsLowerCase = unexpectedParams.collect{ it.replace("-", "").toLowerCase()} - if (!unexpectedParamsLowerCase.contains(specifiedParamLowerCase)){ - unexpectedParams.push(specifiedParam) - } - } - } - - //~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~// - // Validate parameters against the schema - InputStream input_stream = new File(getSchemaPath(workflow, schema_filename=schema_filename)).newInputStream() - JSONObject raw_schema = new JSONObject(new JSONTokener(input_stream)) - - // Remove anything that's in params.schema_ignore_params - raw_schema = removeIgnoredParams(raw_schema, params) - - Schema schema = SchemaLoader.load(raw_schema) - - // Clean the parameters - def cleanedParams = cleanParameters(params) - - // Convert to JSONObject - def jsonParams = new JsonBuilder(cleanedParams) - JSONObject params_json = new JSONObject(jsonParams.toString()) - - // Validate - try { - schema.validate(params_json) - } catch (ValidationException e) { - println '' - log.error 'ERROR: Validation of pipeline parameters failed!' - JSONObject exceptionJSON = e.toJSON() - printExceptions(exceptionJSON, params_json, log, enums) - println '' - has_error = true - } - - // Check for unexpected parameters - if (unexpectedParams.size() > 0) { - Map colors = NfcoreTemplate.logColours(params.monochrome_logs) - println '' - def warn_msg = 'Found unexpected parameters:' - for (unexpectedParam in unexpectedParams) { - warn_msg = warn_msg + "\n* --${unexpectedParam}: ${params[unexpectedParam].toString()}" - } - log.warn warn_msg - log.info "- ${colors.dim}Ignore this warning: params.schema_ignore_params = \"${unexpectedParams.join(',')}\" ${colors.reset}" - println '' - } - - if (has_error) { - Nextflow.error('Exiting!') - } - } - - // - // Beautify parameters for --help - // - public static String paramsHelp(workflow, params, command, schema_filename='nextflow_schema.json') { - Map colors = NfcoreTemplate.logColours(params.monochrome_logs) - Integer num_hidden = 0 - String output = '' - output += 'Typical pipeline command:\n\n' - output += " ${colors.cyan}${command}${colors.reset}\n\n" - Map params_map = paramsLoad(getSchemaPath(workflow, schema_filename=schema_filename)) - Integer max_chars = paramsMaxChars(params_map) + 1 - Integer desc_indent = max_chars + 14 - Integer dec_linewidth = 160 - desc_indent - for (group in params_map.keySet()) { - Integer num_params = 0 - String group_output = colors.underlined + colors.bold + group + colors.reset + '\n' - def group_params = params_map.get(group) // This gets the parameters of that particular group - for (param in group_params.keySet()) { - if (group_params.get(param).hidden && !params.show_hidden_params) { - num_hidden += 1 - continue; - } - def type = '[' + group_params.get(param).type + ']' - def description = group_params.get(param).description - def defaultValue = group_params.get(param).default != null ? " [default: " + group_params.get(param).default.toString() + "]" : '' - def description_default = description + colors.dim + defaultValue + colors.reset - // Wrap long description texts - // Loosely based on https://dzone.com/articles/groovy-plain-text-word-wrap - if (description_default.length() > dec_linewidth){ - List olines = [] - String oline = "" // " " * indent - description_default.split(" ").each() { wrd -> - if ((oline.size() + wrd.size()) <= dec_linewidth) { - oline += wrd + " " - } else { - olines += oline - oline = wrd + " " - } - } - olines += oline - description_default = olines.join("\n" + " " * desc_indent) - } - group_output += " --" + param.padRight(max_chars) + colors.dim + type.padRight(10) + colors.reset + description_default + '\n' - num_params += 1 - } - group_output += '\n' - if (num_params > 0){ - output += group_output - } - } - if (num_hidden > 0){ - output += colors.dim + "!! Hiding $num_hidden params, use --show_hidden_params to show them !!\n" + colors.reset - } - output += NfcoreTemplate.dashedLine(params.monochrome_logs) - return output - } - - // - // Groovy Map summarising parameters/workflow options used by the pipeline - // - public static LinkedHashMap paramsSummaryMap(workflow, params, schema_filename='nextflow_schema.json') { - // Get a selection of core Nextflow workflow options - def Map workflow_summary = [:] - if (workflow.revision) { - workflow_summary['revision'] = workflow.revision - } - workflow_summary['runName'] = workflow.runName - if (workflow.containerEngine) { - workflow_summary['containerEngine'] = workflow.containerEngine - } - if (workflow.container) { - workflow_summary['container'] = workflow.container - } - workflow_summary['launchDir'] = workflow.launchDir - workflow_summary['workDir'] = workflow.workDir - workflow_summary['projectDir'] = workflow.projectDir - workflow_summary['userName'] = workflow.userName - workflow_summary['profile'] = workflow.profile - workflow_summary['configFiles'] = workflow.configFiles.join(', ') - - // Get pipeline parameters defined in JSON Schema - def Map params_summary = [:] - def params_map = paramsLoad(getSchemaPath(workflow, schema_filename=schema_filename)) - for (group in params_map.keySet()) { - def sub_params = new LinkedHashMap() - def group_params = params_map.get(group) // This gets the parameters of that particular group - for (param in group_params.keySet()) { - if (params.containsKey(param)) { - def params_value = params.get(param) - def schema_value = group_params.get(param).default - def param_type = group_params.get(param).type - if (schema_value != null) { - if (param_type == 'string') { - if (schema_value.contains('$projectDir') || schema_value.contains('${projectDir}')) { - def sub_string = schema_value.replace('\$projectDir', '') - sub_string = sub_string.replace('\${projectDir}', '') - if (params_value.contains(sub_string)) { - schema_value = params_value - } - } - if (schema_value.contains('$params.outdir') || schema_value.contains('${params.outdir}')) { - def sub_string = schema_value.replace('\$params.outdir', '') - sub_string = sub_string.replace('\${params.outdir}', '') - if ("${params.outdir}${sub_string}" == params_value) { - schema_value = params_value - } - } - } - } - - // We have a default in the schema, and this isn't it - if (schema_value != null && params_value != schema_value) { - sub_params.put(param, params_value) - } - // No default in the schema, and this isn't empty - else if (schema_value == null && params_value != "" && params_value != null && params_value != false) { - sub_params.put(param, params_value) - } - } - } - params_summary.put(group, sub_params) - } - return [ 'Core Nextflow options' : workflow_summary ] << params_summary - } - - // - // Beautify parameters for summary and return as string - // - public static String paramsSummaryLog(workflow, params) { - Map colors = NfcoreTemplate.logColours(params.monochrome_logs) - String output = '' - def params_map = paramsSummaryMap(workflow, params) - def max_chars = paramsMaxChars(params_map) - for (group in params_map.keySet()) { - def group_params = params_map.get(group) // This gets the parameters of that particular group - if (group_params) { - output += colors.bold + group + colors.reset + '\n' - for (param in group_params.keySet()) { - output += " " + colors.blue + param.padRight(max_chars) + ": " + colors.green + group_params.get(param) + colors.reset + '\n' - } - output += '\n' - } - } - output += "!! Only displaying parameters that differ from the pipeline defaults !!\n" - output += NfcoreTemplate.dashedLine(params.monochrome_logs) - return output - } - - // - // Loop over nested exceptions and print the causingException - // - private static void printExceptions(ex_json, params_json, log, enums, limit=5) { - def causingExceptions = ex_json['causingExceptions'] - if (causingExceptions.length() == 0) { - def m = ex_json['message'] =~ /required key \[([^\]]+)\] not found/ - // Missing required param - if (m.matches()) { - log.error "* Missing required parameter: --${m[0][1]}" - } - // Other base-level error - else if (ex_json['pointerToViolation'] == '#') { - log.error "* ${ex_json['message']}" - } - // Error with specific param - else { - def param = ex_json['pointerToViolation'] - ~/^#\// - def param_val = params_json[param].toString() - if (enums.containsKey(param)) { - def error_msg = "* --${param}: '${param_val}' is not a valid choice (Available choices" - if (enums[param].size() > limit) { - log.error "${error_msg} (${limit} of ${enums[param].size()}): ${enums[param][0..limit-1].join(', ')}, ... )" - } else { - log.error "${error_msg}: ${enums[param].join(', ')})" - } - } else { - log.error "* --${param}: ${ex_json['message']} (${param_val})" - } - } - } - for (ex in causingExceptions) { - printExceptions(ex, params_json, log, enums) - } - } - - // - // Remove an element from a JSONArray - // - private static JSONArray removeElement(json_array, element) { - def list = [] - int len = json_array.length() - for (int i=0;i - if(raw_schema.keySet().contains('definitions')){ - raw_schema.definitions.each { definition -> - for (key in definition.keySet()){ - if (definition[key].get("properties").keySet().contains(ignore_param)){ - // Remove the param to ignore - definition[key].get("properties").remove(ignore_param) - // If the param was required, change this - if (definition[key].has("required")) { - def cleaned_required = removeElement(definition[key].required, ignore_param) - definition[key].put("required", cleaned_required) - } - } - } - } - } - if(raw_schema.keySet().contains('properties') && raw_schema.get('properties').keySet().contains(ignore_param)) { - raw_schema.get("properties").remove(ignore_param) - } - if(raw_schema.keySet().contains('required') && raw_schema.required.contains(ignore_param)) { - def cleaned_required = removeElement(raw_schema.required, ignore_param) - raw_schema.put("required", cleaned_required) - } - } - return raw_schema - } - - // - // Clean and check parameters relative to Nextflow native classes - // - private static Map cleanParameters(params) { - def new_params = params.getClass().newInstance(params) - for (p in params) { - // remove anything evaluating to false - if (!p['value']) { - new_params.remove(p.key) - } - // Cast MemoryUnit to String - if (p['value'].getClass() == nextflow.util.MemoryUnit) { - new_params.replace(p.key, p['value'].toString()) - } - // Cast Duration to String - if (p['value'].getClass() == nextflow.util.Duration) { - new_params.replace(p.key, p['value'].toString().replaceFirst(/d(?!\S)/, "day")) - } - // Cast LinkedHashMap to String - if (p['value'].getClass() == LinkedHashMap) { - new_params.replace(p.key, p['value'].toString()) - } - } - return new_params - } - - // - // This function tries to read a JSON params file - // - private static LinkedHashMap paramsLoad(String json_schema) { - def params_map = new LinkedHashMap() - try { - params_map = paramsRead(json_schema) - } catch (Exception e) { - println "Could not read parameters settings from JSON. $e" - params_map = new LinkedHashMap() - } - return params_map - } - - // - // Method to actually read in JSON file using Groovy. - // Group (as Key), values are all parameters - // - Parameter1 as Key, Description as Value - // - Parameter2 as Key, Description as Value - // .... - // Group - // - - private static LinkedHashMap paramsRead(String json_schema) throws Exception { - def json = new File(json_schema).text - def Map schema_definitions = (Map) new JsonSlurper().parseText(json).get('definitions') - def Map schema_properties = (Map) new JsonSlurper().parseText(json).get('properties') - /* Tree looks like this in nf-core schema - * definitions <- this is what the first get('definitions') gets us - group 1 - title - description - properties - parameter 1 - type - description - parameter 2 - type - description - group 2 - title - description - properties - parameter 1 - type - description - * properties <- parameters can also be ungrouped, outside of definitions - parameter 1 - type - description - */ - - // Grouped params - def params_map = new LinkedHashMap() - schema_definitions.each { key, val -> - def Map group = schema_definitions."$key".properties // Gets the property object of the group - def title = schema_definitions."$key".title - def sub_params = new LinkedHashMap() - group.each { innerkey, value -> - sub_params.put(innerkey, value) - } - params_map.put(title, sub_params) - } - - // Ungrouped params - def ungrouped_params = new LinkedHashMap() - schema_properties.each { innerkey, value -> - ungrouped_params.put(innerkey, value) - } - params_map.put("Other parameters", ungrouped_params) - - return params_map - } - - // - // Get maximum number of characters across all parameter names - // - private static Integer paramsMaxChars(params_map) { - Integer max_chars = 0 - for (group in params_map.keySet()) { - def group_params = params_map.get(group) // This gets the parameters of that particular group - for (param in group_params.keySet()) { - if (param.size() > max_chars) { - max_chars = param.size() - } - } - } - return max_chars - } -} diff --git a/lib/NfcoreTemplate.groovy b/lib/NfcoreTemplate.groovy index 25a0a74a6..408951ae1 100755 --- a/lib/NfcoreTemplate.groovy +++ b/lib/NfcoreTemplate.groovy @@ -128,7 +128,7 @@ class NfcoreTemplate { def email_html = html_template.toString() // Render the sendmail template - def max_multiqc_email_size = params.max_multiqc_email_size as nextflow.util.MemoryUnit + def max_multiqc_email_size = (params.containsKey('max_multiqc_email_size') ? params.max_multiqc_email_size : 0) as nextflow.util.MemoryUnit def smail_fields = [ email: email_address, subject: subject, email_txt: email_txt, email_html: email_html, projectDir: "$projectDir", mqcFile: mqc_report, mqcMaxSize: max_multiqc_email_size.toBytes() ] def sf = new File("$projectDir/assets/sendmail_template.txt") def sendmail_template = engine.createTemplate(sf).make(smail_fields) diff --git a/lib/WorkflowMain.groovy b/lib/WorkflowMain.groovy index ec7707164..3aaabfaa0 100755 --- a/lib/WorkflowMain.groovy +++ b/lib/WorkflowMain.groovy @@ -20,40 +20,11 @@ class WorkflowMain { " https://github.com/${workflow.manifest.name}/blob/master/CITATIONS.md" } - // - // Generate help string - // - public static String help(workflow, params) { - def command = "nextflow run ${workflow.manifest.name} --input samplesheet.csv --genome GRCh37 -profile docker" - def help_string = '' - help_string += NfcoreTemplate.logo(workflow, params.monochrome_logs) - help_string += NfcoreSchema.paramsHelp(workflow, params, command) - help_string += '\n' + citation(workflow) + '\n' - help_string += NfcoreTemplate.dashedLine(params.monochrome_logs) - return help_string - } - - // - // Generate parameter summary log string - // - public static String paramsSummaryLog(workflow, params) { - def summary_log = '' - summary_log += NfcoreTemplate.logo(workflow, params.monochrome_logs) - summary_log += NfcoreSchema.paramsSummaryLog(workflow, params) - summary_log += '\n' + citation(workflow) + '\n' - summary_log += NfcoreTemplate.dashedLine(params.monochrome_logs) - return summary_log - } // // Validate parameters and print summary to screen // public static void initialise(workflow, params, log) { - // Print help to screen if required - if (params.help) { - log.info help(workflow, params) - System.exit(0) - } // Print workflow version and exit on --version if (params.version) { @@ -62,14 +33,6 @@ class WorkflowMain { System.exit(0) } - // Print parameter summary log to screen - log.info paramsSummaryLog(workflow, params) - - // Validate workflow parameters via the JSON schema - if (params.validate_params) { - NfcoreSchema.validateParameters(workflow, params, log) - } - // Check that a -profile or Nextflow config has been provided to run the pipeline NfcoreTemplate.checkConfigProvided(workflow, log) diff --git a/lib/WorkflowRnaseq.groovy b/lib/WorkflowRnaseq.groovy index 69855db20..9f1ed4b11 100755 --- a/lib/WorkflowRnaseq.groovy +++ b/lib/WorkflowRnaseq.groovy @@ -11,6 +11,7 @@ class WorkflowRnaseq { // Check and validate parameters // public static void initialise(params, log) { + genomeExistsError(params, log) @@ -46,15 +47,57 @@ class WorkflowRnaseq { return yaml_file_text } - public static String methodsDescriptionText(run_workflow, mqc_methods_yaml) { + // + // Generate methods description for MultiQC + // + + public static String toolCitationText(params) { + + // TODO Optionally add in-text citation tools to this list. + // Can use ternary operators to dynamically construct based conditions, e.g. params["run_xyz"] ? "Tool (Foo et al. 2023)" : "", + // Uncomment function in methodsDescriptionText to render in MultiQC report + def citation_text = [ + "Tools used in the workflow included:", + "FastQC (Andrews 2010),", + "MultiQC (Ewels et al. 2016)", + "." + ].join(' ').trim() + + return citation_text + } + + public static String toolBibliographyText(params) { + + // TODO Optionally add bibliographic entries to this list. + // Can use ternary operators to dynamically construct based conditions, e.g. params["run_xyz"] ? "

  • Author (2023) Pub name, Journal, DOI
  • " : "", + // Uncomment function in methodsDescriptionText to render in MultiQC report + def reference_text = [ + "
  • Andrews S, (2010) FastQC, URL: https://www.bioinformatics.babraham.ac.uk/projects/fastqc/).
  • ", + "
  • Ewels, P., Magnusson, M., Lundin, S., & Käller, M. (2016). MultiQC: summarize analysis results for multiple tools and samples in a single report. Bioinformatics , 32(19), 3047–3048. doi: /10.1093/bioinformatics/btw354
  • " + ].join(' ').trim() + + return reference_text + } + + public static String methodsDescriptionText(run_workflow, mqc_methods_yaml, params) { // Convert to a named map so can be used as with familar NXF ${workflow} variable syntax in the MultiQC YML file def meta = [:] meta.workflow = run_workflow.toMap() meta["manifest_map"] = run_workflow.manifest.toMap() + // Pipeline DOI meta["doi_text"] = meta.manifest_map.doi ? "(doi: ${meta.manifest_map.doi})" : "" meta["nodoi_text"] = meta.manifest_map.doi ? "": "
  • If available, make sure to update the text to include the Zenodo DOI of version of the pipeline used.
  • " + // Tool references + meta["tool_citations"] = "" + meta["tool_bibliography"] = "" + + // TODO Only uncomment below if logic in toolCitationText/toolBibliographyText has been filled! + //meta["tool_citations"] = toolCitationText(params).replaceAll(", \\.", ".").replaceAll("\\. \\.", ".").replaceAll(", \\.", ".") + //meta["tool_bibliography"] = toolBibliographyText(params) + + def methods_text = mqc_methods_yaml.text def engine = new SimpleTemplateEngine() diff --git a/main.nf b/main.nf index 92b516125..47a5fb558 100644 --- a/main.nf +++ b/main.nf @@ -25,6 +25,22 @@ params.fasta = WorkflowMain.getGenomeAttribute(params, 'fasta') ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ */ +include { validateParameters; paramsHelp } from 'plugin/nf-validation' + +// Print help message if needed +if (params.help) { + def logo = NfcoreTemplate.logo(workflow, params.monochrome_logs) + def citation = '\n' + WorkflowMain.citation(workflow) + '\n' + def String command = "nextflow run ${workflow.manifest.name} --input samplesheet.csv --genome GRCh37 -profile docker" + log.info logo + paramsHelp(command) + citation + NfcoreTemplate.dashedLine(params.monochrome_logs) + System.exit(0) +} + +// Validate input parameters +if (params.validate_params) { + validateParameters() +} + WorkflowMain.initialise(workflow, params, log) /* diff --git a/nextflow.config b/nextflow.config index cc94a96c9..0b6192edd 100644 --- a/nextflow.config +++ b/nextflow.config @@ -12,12 +12,12 @@ params { // TODO nf-core: Specify your pipeline's command line flags // Input options input = null - - // References genome = null igenomes_base = 's3://ngi-igenomes/igenomes' igenomes_ignore = false + + // MultiQC options multiqc_config = null multiqc_title = null @@ -27,7 +27,6 @@ params { // Boilerplate options outdir = null - tracedir = "${params.outdir}/pipeline_info" publish_dir_mode = 'copy' email = null email_on_fail = null @@ -36,19 +35,15 @@ params { hook_url = null help = false version = false - validate_params = true - show_hidden_params = false - schema_ignore_params = 'genomes' - // Config options + config_profile_name = null + config_profile_description = null custom_config_version = 'master' custom_config_base = "https://raw.githubusercontent.com/nf-core/configs/${params.custom_config_version}" - config_profile_description = null config_profile_contact = null config_profile_url = null - config_profile_name = null - + // Max resource options // Defaults only, expecting to be overwritten @@ -56,6 +51,13 @@ params { max_cpus = 16 max_time = '240.h' + // Schema validation default options + validationFailUnrecognisedParams = false + validationLenientMode = false + validationSchemaIgnoreParams = 'genomes' + validationShowHiddenParams = false + validate_params = true + } // Load base.config by default for all pipelines @@ -75,13 +77,11 @@ try { // } catch (Exception e) { // System.err.println("WARNING: Could not load nf-core/config/rnaseq profiles: ${params.custom_config_base}/pipeline/rnaseq.config") // } - - profiles { debug { dumpHashes = true process.beforeScript = 'echo $HOSTNAME' - cleanup = false + cleanup = false } conda { conda.enabled = true @@ -104,7 +104,6 @@ profiles { } docker { docker.enabled = true - docker.registry = 'quay.io' docker.userEmulation = true conda.enabled = false singularity.enabled = false @@ -128,7 +127,6 @@ profiles { } podman { podman.enabled = true - podman.registry = 'quay.io' conda.enabled = false docker.enabled = false singularity.enabled = false @@ -172,6 +170,18 @@ profiles { test_full { includeConfig 'conf/test_full.config' } } +// Set default registry for Apptainer, Docker, Podman and Singularity independent of -profile +// Will not be used unless Apptainer / Docker / Podman / Singularity are enabled +// Set to your registry if you have a mirror of containers +apptainer.registry = 'quay.io' +docker.registry = 'quay.io' +podman.registry = 'quay.io' +singularity.registry = 'quay.io' + +// Nextflow plugins +plugins { + id 'nf-validation' // Validation of pipeline parameters and creation of an input channel from a sample sheet +} // Load igenomes.config if required if (!params.igenomes_ignore) { @@ -179,8 +189,6 @@ if (!params.igenomes_ignore) { } else { params.genomes = [:] } - - // Export these variables to prevent local Python/R libraries from conflicting with those in the container // The JULIA depot path has been adjusted to a fixed path `/usr/local/share/julia` that needs to be used for packages in the container. // See https://apeltzer.github.io/post/03-julia-lang-nextflow/ for details on that. Once we have a common agreement on where to keep Julia packages, this is adjustable. @@ -198,19 +206,19 @@ process.shell = ['/bin/bash', '-euo', 'pipefail'] def trace_timestamp = new java.util.Date().format( 'yyyy-MM-dd_HH-mm-ss') timeline { enabled = true - file = "${params.tracedir}/execution_timeline_${trace_timestamp}.html" + file = "${params.outdir}/pipeline_info/execution_timeline_${trace_timestamp}.html" } report { enabled = true - file = "${params.tracedir}/execution_report_${trace_timestamp}.html" + file = "${params.outdir}/pipeline_info/execution_report_${trace_timestamp}.html" } trace { enabled = true - file = "${params.tracedir}/execution_trace_${trace_timestamp}.txt" + file = "${params.outdir}/pipeline_info/execution_trace_${trace_timestamp}.txt" } dag { enabled = true - file = "${params.tracedir}/pipeline_dag_${trace_timestamp}.html" + file = "${params.outdir}/pipeline_info/pipeline_dag_${trace_timestamp}.html" } manifest { @@ -219,8 +227,8 @@ manifest { homePage = 'https://github.com/nf-core/rnaseq' description = """RNA sequencing analysis pipeline for gene/isoform quantification and extensive quality control.""" mainScript = 'main.nf' - nextflowVersion = '!>=22.10.1' - version = '3.12.0dev' + nextflowVersion = '!>=23.04.0' + version = '3.12.0' doi = '' } diff --git a/nextflow_schema.json b/nextflow_schema.json index c5b27512a..538766597 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -15,9 +15,9 @@ "input": { "type": "string", "format": "file-path", + "exists": true, "mimetype": "text/csv", "pattern": "^\\S+\\.csv$", - "schema": "assets/schema_input.json", "description": "Path to comma-separated file containing information about the samples in the experiment.", "help_text": "You will need to create a design file with information about the samples in your experiment before running the pipeline. Use this parameter to specify its location. It has to be a comma-separated file with 3 columns, and a header row. See [usage docs](https://nf-co.re/rnaseq/usage#samplesheet-input).", "fa_icon": "fas fa-file-csv" @@ -57,6 +57,7 @@ "fasta": { "type": "string", "format": "file-path", + "exists": true, "mimetype": "text/plain", "pattern": "^\\S+\\.fn?a(sta)?(\\.gz)?$", "description": "Path to FASTA genome file.", @@ -157,7 +158,7 @@ "description": "Maximum amount of time that can be requested for any single job.", "default": "240.h", "fa_icon": "far fa-clock", - "pattern": "^(\\d+\\.?\\s*(s|m|h|day)\\s*)+$", + "pattern": "^(\\d+\\.?\\s*(s|m|h|d|day)\\s*)+$", "hidden": true, "help_text": "Use to set an upper-limit for the time requirement for each process. Should be a string in the format integer-unit e.g. `--max_time '2.h'`" } @@ -174,12 +175,14 @@ "type": "boolean", "description": "Display help text.", "fa_icon": "fas fa-question-circle", + "default": false, "hidden": true }, "version": { "type": "boolean", "description": "Display version and exit.", "fa_icon": "fas fa-question-circle", + "default": false, "hidden": true }, "publish_dir_mode": { @@ -203,6 +206,7 @@ "type": "boolean", "description": "Send plain-text email instead of HTML.", "fa_icon": "fas fa-remove-format", + "default": false, "hidden": true }, "max_multiqc_email_size": { @@ -217,6 +221,7 @@ "type": "boolean", "description": "Do not use coloured log outputs.", "fa_icon": "fas fa-palette", + "default": false, "hidden": true }, "hook_url": { @@ -228,6 +233,7 @@ }, "multiqc_config": { "type": "string", + "format": "file-path", "description": "Custom config file to supply to MultiQC.", "fa_icon": "fas fa-cog", "hidden": true @@ -243,13 +249,6 @@ "description": "Custom MultiQC yaml file containing HTML including a methods description.", "fa_icon": "fas fa-cog" }, - "tracedir": { - "type": "string", - "description": "Directory to keep pipeline Nextflow logs and reports.", - "default": "${params.outdir}/pipeline_info", - "fa_icon": "fas fa-cogs", - "hidden": true - }, "validate_params": { "type": "boolean", "description": "Boolean whether to validate parameters against the schema at runtime", @@ -257,12 +256,29 @@ "fa_icon": "fas fa-check-square", "hidden": true }, - "show_hidden_params": { + "validationShowHiddenParams": { "type": "boolean", "fa_icon": "far fa-eye-slash", "description": "Show all params when using `--help`", + "default": false, "hidden": true, "help_text": "By default, parameters set as _hidden_ in the schema are not shown on the command line when a user runs with `--help`. Specifying this option will tell the pipeline to show all parameters." + }, + "validationFailUnrecognisedParams": { + "type": "boolean", + "fa_icon": "far fa-check-circle", + "description": "Validation of parameters fails when an unrecognised parameter is found.", + "default": false, + "hidden": true, + "help_text": "By default, when an unrecognised parameter is found, it returns a warinig." + }, + "validationLenientMode": { + "type": "boolean", + "fa_icon": "far fa-check-circle", + "description": "Validation of parameters in lenient more.", + "default": false, + "hidden": true, + "help_text": "Allows string values that are parseable as numbers or booleans. For further information see [JSONSchema docs](https://github.com/everit-org/json-schema#lenient-mode)." } } } diff --git a/workflows/rnaseq.nf b/workflows/rnaseq.nf index f4210235f..2fedf361c 100644 --- a/workflows/rnaseq.nf +++ b/workflows/rnaseq.nf @@ -1,21 +1,19 @@ /* ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ - VALIDATE INPUTS + PRINT PARAMS SUMMARY ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ */ -def summary_params = NfcoreSchema.paramsSummaryMap(workflow, params) +include { paramsSummaryLog; paramsSummaryMap } from 'plugin/nf-validation' -// Validate input parameters -WorkflowRnaseq.initialise(params, log) +def logo = NfcoreTemplate.logo(workflow, params.monochrome_logs) +def citation = '\n' + WorkflowMain.citation(workflow) + '\n' +def summary_params = paramsSummaryMap(workflow) -// TODO nf-core: Add all file path parameters for the pipeline to the list below -// Check input path parameters to see if they exist -def checkPathParamList = [ params.input, params.multiqc_config, params.fasta ] -for (param in checkPathParamList) { if (param) { file(param, checkIfExists: true) } } +// Print parameter summary log to screen +log.info logo + paramsSummaryLog(workflow) + citation -// Check mandatory parameters -if (params.input) { ch_input = file(params.input) } else { exit 1, 'Input samplesheet not specified!' } +WorkflowRnaseq.initialise(params, log) /* ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ @@ -69,9 +67,12 @@ workflow RNASEQ { // SUBWORKFLOW: Read in samplesheet, validate and stage input files // INPUT_CHECK ( - ch_input + file(params.input) ) ch_versions = ch_versions.mix(INPUT_CHECK.out.versions) + // TODO: OPTIONAL, you can use nf-validation plugin to create an input channel from the samplesheet with Channel.fromSamplesheet("input") + // See the documentation https://nextflow-io.github.io/nf-validation/samplesheets/fromSamplesheet/ + // ! There is currently no tooling to help you write a sample sheet schema // // MODULE: Run FastQC @@ -91,7 +92,7 @@ workflow RNASEQ { workflow_summary = WorkflowRnaseq.paramsSummaryMultiqc(workflow, summary_params) ch_workflow_summary = Channel.value(workflow_summary) - methods_description = WorkflowRnaseq.methodsDescriptionText(workflow, ch_multiqc_custom_methods_description) + methods_description = WorkflowRnaseq.methodsDescriptionText(workflow, ch_multiqc_custom_methods_description, params) ch_methods_description = Channel.value(methods_description) ch_multiqc_files = Channel.empty() From 9a47abe0bf7762f62eaa945c7786f181a2befe63 Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Tue, 4 Jul 2023 12:24:45 +0100 Subject: [PATCH 007/184] Fix nf-core lint --- .nf-core.yml | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/.nf-core.yml b/.nf-core.yml index e6c9d79f0..fdd1b0aea 100644 --- a/.nf-core.yml +++ b/.nf-core.yml @@ -3,5 +3,5 @@ lint: files_unchanged: - assets/email_template.html - assets/email_template.txt - - lib/NfcoreSchema.groovy - lib/NfcoreTemplate.groovy + multiqc_config: false From 7a5c4bb3966852c79a25aea14a1e10bb4e884d5d Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Wed, 5 Jul 2023 09:56:01 +0100 Subject: [PATCH 008/184] Remove versions from multiqc_config.yml --- assets/multiqc_config.yml | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/assets/multiqc_config.yml b/assets/multiqc_config.yml index 3acbb7729..64cbf61f3 100644 --- a/assets/multiqc_config.yml +++ b/assets/multiqc_config.yml @@ -1,7 +1,7 @@ report_comment: > - This report has been generated by the nf-core/rnaseq + This report has been generated by the nf-core/rnaseq analysis pipeline. For information about how to interpret these results, please see the - documentation. + documentation. report_section_order: "nf-core-rnaseq-methods-description": order: -1000 From 37cdf6c9b5d8a160e419e613bc0daa6ac935ad3d Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Wed, 5 Jul 2023 13:47:50 +0100 Subject: [PATCH 009/184] Add file-path and mimetype to all entities in schema --- nextflow_schema.json | 19 ++++++++++--------- 1 file changed, 10 insertions(+), 9 deletions(-) diff --git a/nextflow_schema.json b/nextflow_schema.json index 2381dd366..0400f2805 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -10,7 +10,7 @@ "type": "object", "fa_icon": "fas fa-terminal", "description": "Define where the pipeline should find input data and save output data.", - "required": ["outdir"], + "required": ["input", "outdir"], "properties": { "input": { "type": "string", @@ -120,12 +120,15 @@ "properties": { "bbsplit_fasta_list": { "type": "string", + "format": "file-path", + "mimetype": "text/plain", "fa_icon": "fas fa-list-alt", "description": "Path to comma-separated file containing a list of reference genomes to filter reads against with BBSplit. You have to also explicitly set `--skip_bbsplit false` if you want to use BBSplit.", "help_text": "The file should contain 2 columns: short name and full path to reference genome(s) e.g. \n```\nmm10,/path/to/mm10.fa\necoli,/path/to/ecoli.fa\n```" }, "bbsplit_index": { "type": "string", + "format": "path", "fa_icon": "fas fa-bezier-curve", "description": "Path to directory or tar.gz archive for pre-built BBSplit index.", "help_text": "The BBSplit index will have to be built at least once with this pipeline (see `--save_reference` to save index). It can then be provided via `--bbsplit_index` for future runs." @@ -149,6 +152,8 @@ }, "ribo_database_manifest": { "type": "string", + "format": "file-path", + "mimetype": "text/plain", "default": "${projectDir}/assets/rrna-db-defaults.txt", "fa_icon": "fas fa-database", "description": "Text file containing paths to fasta files (one per line) that will be used to create the database for SortMeRNA.", @@ -401,7 +406,7 @@ }, "min_mapped_reads": { "type": "number", - "default": 5, + "default": 5.0, "fa_icon": "fas fa-percentage", "description": "Minimum percentage of uniquely mapped reads below which samples are removed from further processing.", "help_text": "Some downstream steps in the pipeline will fail if this threshold is too low." @@ -633,14 +638,12 @@ "type": "boolean", "description": "Display help text.", "fa_icon": "fas fa-question-circle", - "default": false, "hidden": true }, "version": { "type": "boolean", "description": "Display version and exit.", "fa_icon": "fas fa-question-circle", - "default": false, "hidden": true }, "publish_dir_mode": { @@ -664,7 +667,6 @@ "type": "boolean", "description": "Send plain-text email instead of HTML.", "fa_icon": "fas fa-remove-format", - "default": false, "hidden": true }, "max_multiqc_email_size": { @@ -678,7 +680,6 @@ "type": "boolean", "description": "Do not use coloured log outputs.", "fa_icon": "fas fa-palette", - "default": false, "hidden": true }, "hook_url": { @@ -691,12 +692,15 @@ "multiqc_config": { "type": "string", "format": "file-path", + "mimetype": "text/plain", "description": "Custom config file to supply to MultiQC.", "fa_icon": "fas fa-cog", "hidden": true }, "multiqc_logo": { "type": "string", + "format": "file-path", + "mimetype": "text/plain", "description": "Custom logo file to supply to MultiQC. File name must also be set in the MultiQC config file", "fa_icon": "fas fa-image", "hidden": true @@ -717,7 +721,6 @@ "type": "boolean", "fa_icon": "far fa-eye-slash", "description": "Show all params when using `--help`", - "default": false, "hidden": true, "help_text": "By default, parameters set as _hidden_ in the schema are not shown on the command line when a user runs with `--help`. Specifying this option will tell the pipeline to show all parameters." }, @@ -725,7 +728,6 @@ "type": "boolean", "fa_icon": "far fa-check-circle", "description": "Validation of parameters fails when an unrecognised parameter is found.", - "default": false, "hidden": true, "help_text": "By default, when an unrecognised parameter is found, it returns a warinig." }, @@ -733,7 +735,6 @@ "type": "boolean", "fa_icon": "far fa-check-circle", "description": "Validation of parameters in lenient more.", - "default": false, "hidden": true, "help_text": "Allows string values that are parseable as numbers or booleans. For further information see [JSONSchema docs](https://github.com/everit-org/json-schema#lenient-mode)." } From d540efb6f12c67fd090b510d0ab7c23697fcb9f8 Mon Sep 17 00:00:00 2001 From: Maxime U Garcia Date: Wed, 5 Jul 2023 17:48:29 +0200 Subject: [PATCH 010/184] Update README.md --- README.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/README.md b/README.md index d48a2912d..31314a457 100644 --- a/README.md +++ b/README.md @@ -107,7 +107,7 @@ These scripts were originally written for use at the [National Genomics Infrastr The pipeline was re-written in Nextflow DSL2 and is primarily maintained by Harshil Patel ([@drpatelh](https://github.com/drpatelh)) from [Seqera Labs, Spain](https://seqera.io/). -The pipeline workflow diagram was designed by Sarah Guinchard ([@G-Sarah](https://github.com/G-Sarah)) and James Fellows Yates ([@jfy133](https://github.com/jfy133)). +The pipeline workflow diagram was initially designed by Sarah Guinchard ([@G-Sarah](https://github.com/G-Sarah)) and James Fellows Yates ([@jfy133](https://github.com/jfy133)), further modifications where made by Harshil Patel ([@drpatelh](https://github.com/drpatelh)) and Maxime Garcia ([@maxulysse](https://github.com/maxulysse)). Many thanks to other who have helped out along the way too, including (but not limited to): From f3ed725c689788767723187e21cc5932570b530c Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Wed, 5 Jul 2023 19:11:18 +0100 Subject: [PATCH 011/184] Delete validation now performed by nf-validation --- lib/WorkflowMain.groovy | 5 ----- lib/WorkflowRnaseq.groovy | 1 - pyproject.toml | 2 +- workflows/rnaseq.nf | 20 +++++--------------- 4 files changed, 6 insertions(+), 22 deletions(-) diff --git a/lib/WorkflowMain.groovy b/lib/WorkflowMain.groovy index dd9b6e9f4..4d94dfe08 100755 --- a/lib/WorkflowMain.groovy +++ b/lib/WorkflowMain.groovy @@ -44,11 +44,6 @@ class WorkflowMain { // Check AWS batch settings NfcoreTemplate.awsBatch(workflow, params) - - // Check input has been provided - if (!params.input) { - Nextflow.error("Please provide an input samplesheet to the pipeline e.g. '--input samplesheet.csv'") - } } // diff --git a/lib/WorkflowRnaseq.groovy b/lib/WorkflowRnaseq.groovy index c85164a9f..fe6010ad8 100755 --- a/lib/WorkflowRnaseq.groovy +++ b/lib/WorkflowRnaseq.groovy @@ -14,7 +14,6 @@ class WorkflowRnaseq { public static void initialise(params, log, valid_params) { genomeExistsError(params, log) - if (!params.fasta) { Nextflow.error("Genome fasta file not specified with e.g. '--fasta genome.fa' or via a detectable config file.") } diff --git a/pyproject.toml b/pyproject.toml index 0d62beb6f..bc01239b3 100644 --- a/pyproject.toml +++ b/pyproject.toml @@ -1,4 +1,4 @@ -# Config file for Python. Mostly used to configure linting of bin/check_samplesheet.py with Black. +# Config file for Python. Mostly used to configure linting of bin/*.py with Black. # Should be kept the same as nf-core/tools to avoid fighting with template synchronisation. [tool.black] line-length = 120 diff --git a/workflows/rnaseq.nf b/workflows/rnaseq.nf index 4a0b96043..3b12b4318 100755 --- a/workflows/rnaseq.nf +++ b/workflows/rnaseq.nf @@ -26,27 +26,17 @@ def valid_params = [ rseqc_modules : ['bam_stat', 'inner_distance', 'infer_experiment', 'junction_annotation', 'junction_saturation', 'read_distribution', 'read_duplication', 'tin'] ] -// Check input path parameters to see if they exist -checkPathParamList = [ - params.input, params.multiqc_config, - params.fasta, params.transcript_fasta, params.additional_fasta, - params.gtf, params.gff, params.gene_bed, - params.ribo_database_manifest, params.splicesites, - params.star_index, params.hisat2_index, params.rsem_index, params.salmon_index -] -for (param in checkPathParamList) { if (param) { file(param, checkIfExists: true) } } - WorkflowRnaseq.initialise(params, log, valid_params) // Check rRNA databases for sortmerna if (params.remove_ribo_rna) { - ch_ribo_db = file(params.ribo_database_manifest, checkIfExists: true) + ch_ribo_db = file(params.ribo_database_manifest) if (ch_ribo_db.isEmpty()) {exit 1, "File provided with --ribo_database_manifest is empty: ${ch_ribo_db.getName()}!"} } // Check if file with list of fastas is provided when running BBSplit if (!params.skip_bbsplit && !params.bbsplit_index && params.bbsplit_fasta_list) { - ch_bbsplit_fasta_list = file(params.bbsplit_fasta_list, checkIfExists: true) + ch_bbsplit_fasta_list = file(params.bbsplit_fasta_list) if (ch_bbsplit_fasta_list.isEmpty()) {exit 1, "File provided with --bbsplit_fasta_list is empty: ${ch_bbsplit_fasta_list.getName()}!"} } @@ -84,9 +74,9 @@ if (params.fasta && params.gtf) { */ ch_multiqc_config = Channel.fromPath("$projectDir/assets/multiqc_config.yml", checkIfExists: true) -ch_multiqc_custom_config = params.multiqc_config ? Channel.fromPath( params.multiqc_config, checkIfExists: true ) : Channel.empty() -ch_multiqc_logo = params.multiqc_logo ? Channel.fromPath( params.multiqc_logo, checkIfExists: true ) : Channel.empty() -ch_multiqc_custom_methods_description = params.multiqc_methods_description ? file(params.multiqc_methods_description, checkIfExists: true) : file("$projectDir/assets/methods_description_template.yml", checkIfExists: true) +ch_multiqc_custom_config = params.multiqc_config ? Channel.fromPath(params.multiqc_config) : Channel.empty() +ch_multiqc_logo = params.multiqc_logo ? Channel.fromPath(params.multiqc_logo) : Channel.empty() +ch_multiqc_custom_methods_description = params.multiqc_methods_description ? file(params.multiqc_methods_description) : file("$projectDir/assets/methods_description_template.yml", checkIfExists: true) // Header files for MultiQC ch_pca_header_multiqc = file("$projectDir/assets/multiqc/deseq2_pca_header.txt", checkIfExists: true) From 9c9d6e536c735f6f40305e61d9054f7e3394b25a Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Wed, 5 Jul 2023 19:38:41 +0100 Subject: [PATCH 012/184] Delete more validation now performed by nf-validation --- lib/WorkflowRnaseq.groovy | 31 +++++++------------------------ nextflow_schema.json | 5 +++-- workflows/rnaseq.nf | 9 +-------- 3 files changed, 11 insertions(+), 34 deletions(-) diff --git a/lib/WorkflowRnaseq.groovy b/lib/WorkflowRnaseq.groovy index fe6010ad8..46d055011 100755 --- a/lib/WorkflowRnaseq.groovy +++ b/lib/WorkflowRnaseq.groovy @@ -11,13 +11,9 @@ class WorkflowRnaseq { // // Check and validate parameters // - public static void initialise(params, log, valid_params) { + public static void initialise(params, log) { genomeExistsError(params, log) - if (!params.fasta) { - Nextflow.error("Genome fasta file not specified with e.g. '--fasta genome.fa' or via a detectable config file.") - } - if (!params.gtf && !params.gff) { Nextflow.error("No GTF or GFF3 annotation specified! The pipeline requires at least one of these files.") } @@ -53,27 +49,13 @@ class WorkflowRnaseq { } } - if (!params.skip_trimming) { - if (!valid_params['trimmers'].contains(params.trimmer)) { - Nextflow.error("Invalid option: '${params.trimmer}'. Valid options for '--trimmer': ${valid_params['trimmers'].join(', ')}.") - } - } - - if (!params.skip_alignment) { - if (!valid_params['aligners'].contains(params.aligner)) { - Nextflow.error("Invalid option: '${params.aligner}'. Valid options for '--aligner': ${valid_params['aligners'].join(', ')}.") - } - } else { + if (params.skip_alignment) { skipAlignmentWarn(log) } if (!params.skip_pseudo_alignment && params.pseudo_aligner) { - if (!valid_params['pseudoaligners'].contains(params.pseudo_aligner)) { - Nextflow.error("Invalid option: '${params.pseudo_aligner}'. Valid options for '--pseudo_aligner': ${valid_params['pseudoaligners'].join(', ')}.") - } else { - if (!(params.salmon_index || params.transcript_fasta || (params.fasta && (params.gtf || params.gff)))) { - Nextflow.error("To use `--pseudo_aligner 'salmon'`, you must provide either --salmon_index or --transcript_fasta or both --fasta and --gtf / --gff.") - } + if (!(params.salmon_index || params.transcript_fasta || (params.fasta && (params.gtf || params.gff)))) { + Nextflow.error("To use `--pseudo_aligner 'salmon'`, you must provide either --salmon_index or --transcript_fasta or both --fasta and --gtf / --gff.") } } @@ -108,9 +90,10 @@ class WorkflowRnaseq { } // Check which RSeQC modules we are running + def valid_rseqc_modules = ['bam_stat', 'inner_distance', 'infer_experiment', 'junction_annotation', 'junction_saturation', 'read_distribution', 'read_duplication', 'tin'] def rseqc_modules = params.rseqc_modules ? params.rseqc_modules.split(',').collect{ it.trim().toLowerCase() } : [] - if ((valid_params['rseqc_modules'] + rseqc_modules).unique().size() != valid_params['rseqc_modules'].size()) { - Nextflow.error("Invalid option: ${params.rseqc_modules}. Valid options for '--rseqc_modules': ${valid_params['rseqc_modules'].join(', ')}") + if ((valid_rseqc_modules + rseqc_modules).unique().size() != valid_rseqc_modules.size()) { + Nextflow.error("Invalid option: ${params.rseqc_modules}. Valid options for '--rseqc_modules': ${valid_rseqc_modules.join(', ')}") } } diff --git a/nextflow_schema.json b/nextflow_schema.json index 0400f2805..d093872b1 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -324,7 +324,8 @@ "hidden": true, "help_text": "Do not load `igenomes.config` when running the pipeline. You may choose this option if you observe clashes between custom parameters and those supplied in `igenomes.config`." } - } + }, + "required": ["fasta"] }, "read_trimming_options": { "title": "Read trimming options", @@ -406,7 +407,7 @@ }, "min_mapped_reads": { "type": "number", - "default": 5.0, + "default": 5, "fa_icon": "fas fa-percentage", "description": "Minimum percentage of uniquely mapped reads below which samples are removed from further processing.", "help_text": "Some downstream steps in the pipeline will fail if this threshold is too low." diff --git a/workflows/rnaseq.nf b/workflows/rnaseq.nf index 3b12b4318..7c90a0f91 100755 --- a/workflows/rnaseq.nf +++ b/workflows/rnaseq.nf @@ -19,14 +19,7 @@ log.info logo + paramsSummaryLog(workflow) + citation ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ */ -def valid_params = [ - aligners : ['star_salmon', 'star_rsem', 'hisat2'], - trimmers : ['trimgalore', 'fastp'], - pseudoaligners : ['salmon'], - rseqc_modules : ['bam_stat', 'inner_distance', 'infer_experiment', 'junction_annotation', 'junction_saturation', 'read_distribution', 'read_duplication', 'tin'] -] - -WorkflowRnaseq.initialise(params, log, valid_params) +WorkflowRnaseq.initialise(params, log) // Check rRNA databases for sortmerna if (params.remove_ribo_rna) { From e886fa598ec4ab43477e8ef861134f4a7db3ab94 Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Thu, 6 Jul 2023 20:15:14 +0100 Subject: [PATCH 013/184] Validate input exclusively using fromSamplesheet from nf-validation --- assets/schema_input.json | 15 ++- bin/check_samplesheet.py | 172 ----------------------------- conf/modules.config | 8 -- lib/WorkflowRnaseq.groovy | 21 ++++ modules/local/samplesheet_check.nf | 31 ------ nextflow_schema.json | 4 + subworkflows/local/input_check.nf | 45 -------- workflows/rnaseq.nf | 56 +++++----- 8 files changed, 66 insertions(+), 286 deletions(-) delete mode 100755 bin/check_samplesheet.py delete mode 100644 modules/local/samplesheet_check.nf delete mode 100644 subworkflows/local/input_check.nf diff --git a/assets/schema_input.json b/assets/schema_input.json index f2eb5ea81..1af6309dd 100644 --- a/assets/schema_input.json +++ b/assets/schema_input.json @@ -10,30 +10,35 @@ "sample": { "type": "string", "pattern": "^\\S+$", - "errorMessage": "Sample name must be provided and cannot contain spaces" + "errorMessage": "Sample name must be provided and cannot contain spaces", + "meta": ["id"] }, "fastq_1": { "type": "string", + "format": "file-path", + "exists": true, "pattern": "^\\S+\\.f(ast)?q\\.gz$", "errorMessage": "FastQ file for reads 1 must be provided, cannot contain spaces and must have extension '.fq.gz' or '.fastq.gz'" }, "fastq_2": { "errorMessage": "FastQ file for reads 2 cannot contain spaces and must have extension '.fq.gz' or '.fastq.gz'", + "type": "string", + "format": "file-path", + "exists": true, "anyOf": [ { - "type": "string", "pattern": "^\\S+\\.f(ast)?q\\.gz$" }, { - "type": "string", "maxLength": 0 } ] }, "strandedness": { "type": "string", - "errorMessage": "Strandedness must be provided and be one of 'forward', 'reverse' or 'unstranded'", - "enum": ["forward", "reverse", "unstranded"] + "errorMessage": "Strandedness must be provided and be one of 'auto', 'forward', 'reverse' or 'unstranded'", + "enum": ["forward", "reverse", "unstranded", "auto"], + "meta": ["strandedness"] } }, "required": ["sample", "fastq_1", "strandedness"] diff --git a/bin/check_samplesheet.py b/bin/check_samplesheet.py deleted file mode 100755 index 05971a5d9..000000000 --- a/bin/check_samplesheet.py +++ /dev/null @@ -1,172 +0,0 @@ -#!/usr/bin/env python3 - -import os -import sys -import errno -import argparse - - -def parse_args(args=None): - Description = "Reformat nf-core/rnaseq samplesheet file and check its contents." - Epilog = "Example usage: python check_samplesheet.py " - - parser = argparse.ArgumentParser(description=Description, epilog=Epilog) - parser.add_argument("FILE_IN", help="Input samplesheet file.") - parser.add_argument("FILE_OUT", help="Output file.") - return parser.parse_args(args) - - -def make_dir(path): - if len(path) > 0: - try: - os.makedirs(path) - except OSError as exception: - if exception.errno != errno.EEXIST: - raise exception - - -def print_error(error, context="Line", context_str=""): - error_str = f"ERROR: Please check samplesheet -> {error}" - if context != "" and context_str != "": - error_str = f"ERROR: Please check samplesheet -> {error}\n{context.strip()}: '{context_str.strip()}'" - print(error_str) - sys.exit(1) - - -def check_samplesheet(file_in, file_out): - """ - This function checks that the samplesheet follows the following structure: - - sample,fastq_1,fastq_2,strandedness - SAMPLE_PE,SAMPLE_PE_RUN1_1.fastq.gz,SAMPLE_PE_RUN1_2.fastq.gz,forward - SAMPLE_PE,SAMPLE_PE_RUN2_1.fastq.gz,SAMPLE_PE_RUN2_2.fastq.gz,forward - SAMPLE_SE,SAMPLE_SE_RUN1_1.fastq.gz,,forward - - For an example see: - https://github.com/nf-core/test-datasets/blob/rnaseq/samplesheet/v3.1/samplesheet_test.csv - """ - - sample_mapping_dict = {} - with open(file_in, "r", encoding="utf-8-sig") as fin: - ## Check header - MIN_COLS = 3 - HEADER = ["sample", "fastq_1", "fastq_2", "strandedness"] - header = [x.strip('"') for x in fin.readline().strip().split(",")] - if header[: len(HEADER)] != HEADER: - print(f"ERROR: Please check samplesheet header -> {','.join(header)} != {','.join(HEADER)}") - sys.exit(1) - - ## Check sample entries - for line in fin: - if line.strip(): - lspl = [x.strip().strip('"') for x in line.strip().split(",")] - - ## Check valid number of columns per row - if len(lspl) < len(HEADER): - print_error( - f"Invalid number of columns (minimum = {len(HEADER)})!", - "Line", - line, - ) - - num_cols = len([x for x in lspl[: len(HEADER)] if x]) - if num_cols < MIN_COLS: - print_error( - f"Invalid number of populated columns (minimum = {MIN_COLS})!", - "Line", - line, - ) - - ## Check sample name entries - sample, fastq_1, fastq_2, strandedness = lspl[: len(HEADER)] - if sample.find(" ") != -1: - print(f"WARNING: Spaces have been replaced by underscores for sample: {sample}") - sample = sample.replace(" ", "_") - if not sample: - print_error("Sample entry has not been specified!", "Line", line) - - ## Check FastQ file extension - for fastq in [fastq_1, fastq_2]: - if fastq: - if fastq.find(" ") != -1: - print_error("FastQ file contains spaces!", "Line", line) - if not fastq.endswith(".fastq.gz") and not fastq.endswith(".fq.gz"): - print_error( - "FastQ file does not have extension '.fastq.gz' or '.fq.gz'!", - "Line", - line, - ) - - ## Check strandedness - strandednesses = ["unstranded", "forward", "reverse", "auto"] - if strandedness: - if strandedness not in strandednesses: - print_error( - f"Strandedness must be one of '{', '.join(strandednesses)}'!", - "Line", - line, - ) - else: - print_error( - f"Strandedness has not been specified! Must be one of {', '.join(strandednesses)}.", - "Line", - line, - ) - - ## Auto-detect paired-end/single-end - sample_info = [] ## [single_end, fastq_1, fastq_2, strandedness] - if sample and fastq_1 and fastq_2: ## Paired-end short reads - sample_info = ["0", fastq_1, fastq_2, strandedness] - elif sample and fastq_1 and not fastq_2: ## Single-end short reads - sample_info = ["1", fastq_1, fastq_2, strandedness] - else: - print_error("Invalid combination of columns provided!", "Line", line) - - ## Create sample mapping dictionary = {sample: [[ single_end, fastq_1, fastq_2, strandedness ]]} - sample_info = sample_info + lspl[len(HEADER) :] - if sample not in sample_mapping_dict: - sample_mapping_dict[sample] = [sample_info] - else: - if sample_info in sample_mapping_dict[sample]: - print_error("Samplesheet contains duplicate rows!", "Line", line) - else: - sample_mapping_dict[sample].append(sample_info) - - ## Write validated samplesheet with appropriate columns - if len(sample_mapping_dict) > 0: - out_dir = os.path.dirname(file_out) - make_dir(out_dir) - with open(file_out, "w") as fout: - fout.write( - ",".join(["sample", "single_end", "fastq_1", "fastq_2", "strandedness"] + header[len(HEADER) :]) + "\n" - ) - for sample in sorted(sample_mapping_dict.keys()): - ## Check that multiple runs of the same sample are of the same datatype i.e. single-end / paired-end - if not all(x[0] == sample_mapping_dict[sample][0][0] for x in sample_mapping_dict[sample]): - print_error( - f"Multiple runs of a sample must be of the same datatype i.e. single-end or paired-end!", - "Sample", - sample, - ) - - ## Check that multiple runs of the same sample are of the same strandedness - if not all(x[3] == sample_mapping_dict[sample][0][3] for x in sample_mapping_dict[sample]): - print_error( - f"Multiple runs of a sample must have the same strandedness!", - "Sample", - sample, - ) - - for idx, val in enumerate(sample_mapping_dict[sample]): - fout.write(",".join([f"{sample}_T{idx+1}"] + val) + "\n") - else: - print_error(f"No entries to process!", "Samplesheet: {file_in}") - - -def main(args=None): - args = parse_args(args) - check_samplesheet(args.FILE_IN, args.FILE_OUT) - - -if __name__ == "__main__": - sys.exit(main()) diff --git a/conf/modules.config b/conf/modules.config index 3c31fb469..eaa35f259 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -23,14 +23,6 @@ process { saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] - withName: 'SAMPLESHEET_CHECK' { - publishDir = [ - path: { "${params.outdir}/pipeline_info" }, - mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } - ] - } - withName: 'CUSTOM_DUMPSOFTWAREVERSIONS' { publishDir = [ path: { "${params.outdir}/pipeline_info" }, diff --git a/lib/WorkflowRnaseq.groovy b/lib/WorkflowRnaseq.groovy index 46d055011..a46bd0bf2 100755 --- a/lib/WorkflowRnaseq.groovy +++ b/lib/WorkflowRnaseq.groovy @@ -97,6 +97,27 @@ class WorkflowRnaseq { } } + // + // Function to validate channels from input samplesheet + // + public static ArrayList validateInput(input) { + def (metas, fastqs) = input[1..2] + + // Check that multiple runs of the same sample are of the same strandedness + def strandedness_ok = metas.collect{ it.strandedness }.unique().size == 1 + if (!strandedness_ok) { + Nextflow.error("Please check input samplesheet -> Multiple runs of a sample must have the same strandedness!: ${metas[0].id}") + } + + // Check that multiple runs of the same sample are of the same datatype i.e. single-end / paired-end + def endedness_ok = metas.collect{ it.single_end }.unique().size == 1 + if (!endedness_ok) { + Nextflow.error("Please check input samplesheet -> Multiple runs of a sample must be of the same datatype i.e. single-end or paired-end: ${metas[0].id}") + } + + return [ metas[0], fastqs ] + } + // // Function to check whether biotype field exists in GTF file // diff --git a/modules/local/samplesheet_check.nf b/modules/local/samplesheet_check.nf deleted file mode 100644 index f8251de88..000000000 --- a/modules/local/samplesheet_check.nf +++ /dev/null @@ -1,31 +0,0 @@ -process SAMPLESHEET_CHECK { - tag "$samplesheet" - label 'process_single' - - conda "conda-forge::python=3.9.5" - container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? - 'https://depot.galaxyproject.org/singularity/python:3.9--1' : - 'biocontainers/python:3.9--1' }" - - input: - path samplesheet - - output: - path '*.csv' , emit: csv - path "versions.yml", emit: versions - - when: - task.ext.when == null || task.ext.when - - script: // This script is bundled with the pipeline, in nf-core/rnaseq/bin/ - """ - check_samplesheet.py \\ - $samplesheet \\ - samplesheet.valid.csv - - cat <<-END_VERSIONS > versions.yml - "${task.process}": - python: \$(python --version | sed 's/Python //g') - END_VERSIONS - """ -} diff --git a/nextflow_schema.json b/nextflow_schema.json index d093872b1..ded0b5169 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -18,6 +18,7 @@ "exists": true, "mimetype": "text/csv", "pattern": "^\\S+\\.csv$", + "schema": "assets/schema_input.json", "description": "Path to comma-separated file containing information about the samples in the experiment.", "help_text": "You will need to create a design file with information about the samples in your experiment before running the pipeline. Use this parameter to specify its location. It has to be a comma-separated file with 4 columns, and a header row. See [usage docs](https://nf-co.re/rnaseq/usage#samplesheet-input).", "fa_icon": "fas fa-file-csv" @@ -708,6 +709,9 @@ }, "multiqc_methods_description": { "type": "string", + "format": "file-path", + "exists": true, + "mimetype": "text/plain", "description": "Custom MultiQC yaml file containing HTML including a methods description.", "fa_icon": "fas fa-cog" }, diff --git a/subworkflows/local/input_check.nf b/subworkflows/local/input_check.nf deleted file mode 100644 index 172d11ea5..000000000 --- a/subworkflows/local/input_check.nf +++ /dev/null @@ -1,45 +0,0 @@ -// -// Check input samplesheet and get read channels -// - -include { SAMPLESHEET_CHECK } from '../../modules/local/samplesheet_check' - -workflow INPUT_CHECK { - take: - samplesheet // file: /path/to/samplesheet.csv - - main: - SAMPLESHEET_CHECK ( samplesheet ) - .csv - .splitCsv ( header:true, sep:',' ) - .map { create_fastq_channel(it) } - .set { reads } - - emit: - reads // channel: [ val(meta), [ reads ] ] - versions = SAMPLESHEET_CHECK.out.versions // channel: [ versions.yml ] -} - -// Function to get list of [ meta, [ fastq_1, fastq_2 ] ] -def create_fastq_channel(LinkedHashMap row) { - // create meta map - def meta = [:] - meta.id = row.sample - meta.single_end = row.single_end.toBoolean() - meta.strandedness = row.strandedness - - // add path(s) of the fastq file(s) to the meta map - def fastq_meta = [] - if (!file(row.fastq_1).exists()) { - exit 1, "ERROR: Please check input samplesheet -> Read 1 FastQ file does not exist!\n${row.fastq_1}" - } - if (meta.single_end) { - fastq_meta = [ meta, [ file(row.fastq_1) ] ] - } else { - if (!file(row.fastq_2).exists()) { - exit 1, "ERROR: Please check input samplesheet -> Read 2 FastQ file does not exist!\n${row.fastq_2}" - } - fastq_meta = [ meta, [ file(row.fastq_1), file(row.fastq_2) ] ] - } - return fastq_meta -} diff --git a/workflows/rnaseq.nf b/workflows/rnaseq.nf index 7c90a0f91..76c9a6a32 100755 --- a/workflows/rnaseq.nf +++ b/workflows/rnaseq.nf @@ -4,7 +4,7 @@ ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ */ -include { paramsSummaryLog; paramsSummaryMap } from 'plugin/nf-validation' +include { paramsSummaryLog; paramsSummaryMap; fromSamplesheet } from 'plugin/nf-validation' def logo = NfcoreTemplate.logo(workflow, params.monochrome_logs) def citation = '\n' + WorkflowMain.citation(workflow) + '\n' @@ -97,7 +97,6 @@ include { UMITOOLS_PREPAREFORRSEM as UMITOOLS_PREPAREFORSALMON } from '../module // // SUBWORKFLOW: Consisting of a mix of local and nf-core/modules // -include { INPUT_CHECK } from '../subworkflows/local/input_check' include { PREPARE_GENOME } from '../subworkflows/local/prepare_genome' include { ALIGN_STAR } from '../subworkflows/local/align_star' include { QUANTIFY_RSEM } from '../subworkflows/local/quantify_rsem' @@ -188,30 +187,37 @@ workflow RNASEQ { } // - // SUBWORKFLOW: Read in samplesheet, validate and stage input files + // Create input channel from input file provided through params.input // - INPUT_CHECK ( - file(params.input) - ) - .reads - .map { - meta, fastq -> - new_id = meta.id - ~/_T\d+/ - [ meta + [id: new_id], fastq ] - } - .groupTuple() - .branch { - meta, fastq -> - single : fastq.size() == 1 - return [ meta, fastq.flatten() ] - multiple: fastq.size() > 1 - return [ meta, fastq.flatten() ] - } - .set { ch_fastq } - ch_versions = ch_versions.mix(INPUT_CHECK.out.versions) - // TODO: OPTIONAL, you can use nf-validation plugin to create an input channel from the samplesheet with Channel.fromSamplesheet("input") - // See the documentation https://nextflow-io.github.io/nf-validation/samplesheets/fromSamplesheet/ - // ! There is currently no tooling to help you write a sample sheet schema + Channel + .fromSamplesheet("input") + .map { + meta, fastq_1, fastq_2 -> + if (!fastq_1.exists()) { + error("Please check input samplesheet -> Read 1 FastQ file does not exist!\n${fastq_1}") + } + if (fastq_2) { + if(!fastq_2.exists()) { + error("Please check input samplesheet -> Read 2 FastQ file does not exist!\n${fastq_2}") + } + return [ meta.id, meta + [ single_end:false ], [ fastq_1, fastq_2 ] ] + } else { + return [ meta.id, meta + [ single_end:true ], [ fastq_1 ] ] + } + } + .unique() + .groupTuple() + .map { + WorkflowRnaseq.validateInput(it) + } + .branch { + meta, fastqs -> + single : fastqs.size() == 1 + return [ meta, fastqs.flatten() ] + multiple: fastqs.size() > 1 + return [ meta, fastqs.flatten() ] + } + .set { ch_fastq } // // MODULE: Concatenate FastQ files from same sample if required From ac94360d1805d3ed1214ca71215fdf7348aca53d Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Thu, 6 Jul 2023 20:21:47 +0100 Subject: [PATCH 014/184] Update CHANGELOG --- CHANGELOG.md | 12 ++++++++++++ 1 file changed, 12 insertions(+) diff --git a/CHANGELOG.md b/CHANGELOG.md index 29aae3151..951921723 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -5,11 +5,23 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 ## v3.13.0dev - [date] +### Credits + +Special thanks to the following for their contributions to the release: + +- [Adam Talbot](https://github.com/adamrtalbot) +- [Júlia Mir Pedrol](https://github.com/mirpedrol) +- [Matthias Zepper](https://github.com/MatthiasZepper) +- [Maxime Garcia](https://github.com/maxulysse) + +Thank you to everyone else that has contributed by reporting bugs, enhancements or in any other way, shape or form. + ### Enhancements & fixes - [PR #1049](https://github.com/nf-core/rnaseq/pull/1049) - Display a warning when `--extra_star_align_args` are used with `--aligner star_rsem` - [PR #1051](https://github.com/nf-core/rnaseq/pull/1051) - Remove `public_aws_ecr` profile - [PR #1054](https://github.com/nf-core/rnaseq/pull/1054) - Template update to nf-core/tools v2.9 +- [PR #1058](https://github.com/nf-core/rnaseq/pull/1058) - Use `nf-validation` plugin for parameter and samplesheet validation ## [[3.12.0](https://github.com/nf-core/rnaseq/releases/tag/3.12.0)] - 2023-06-02 From cee9f54dbe67135341e4808ca477ecb56938b558 Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Thu, 6 Jul 2023 20:22:53 +0100 Subject: [PATCH 015/184] Remove exists until bug is fixed upstream --- nextflow_schema.json | 1 - 1 file changed, 1 deletion(-) diff --git a/nextflow_schema.json b/nextflow_schema.json index ded0b5169..9dd9a2044 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -710,7 +710,6 @@ "multiqc_methods_description": { "type": "string", "format": "file-path", - "exists": true, "mimetype": "text/plain", "description": "Custom MultiQC yaml file containing HTML including a methods description.", "fa_icon": "fas fa-cog" From 3c5fa30c16436fd3e5cbc7655612b25f26e62f49 Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Mon, 10 Jul 2023 23:31:13 +0100 Subject: [PATCH 016/184] Update logic to reflect fixes / enhancements added in nf-validation@0.3.0 --- .nf-core.yml | 1 + nextflow_schema.json | 16 ++++++++++++++++ workflows/rnaseq.nf | 13 +++---------- 3 files changed, 20 insertions(+), 10 deletions(-) diff --git a/.nf-core.yml b/.nf-core.yml index fdd1b0aea..094459361 100644 --- a/.nf-core.yml +++ b/.nf-core.yml @@ -4,4 +4,5 @@ lint: - assets/email_template.html - assets/email_template.txt - lib/NfcoreTemplate.groovy + - pyproject.toml multiqc_config: false diff --git a/nextflow_schema.json b/nextflow_schema.json index 9dd9a2044..e3f16078a 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -122,6 +122,7 @@ "bbsplit_fasta_list": { "type": "string", "format": "file-path", + "exists": true, "mimetype": "text/plain", "fa_icon": "fas fa-list-alt", "description": "Path to comma-separated file containing a list of reference genomes to filter reads against with BBSplit. You have to also explicitly set `--skip_bbsplit false` if you want to use BBSplit.", @@ -130,6 +131,7 @@ "bbsplit_index": { "type": "string", "format": "path", + "exists": true, "fa_icon": "fas fa-bezier-curve", "description": "Path to directory or tar.gz archive for pre-built BBSplit index.", "help_text": "The BBSplit index will have to be built at least once with this pipeline (see `--save_reference` to save index). It can then be provided via `--bbsplit_index` for future runs." @@ -154,6 +156,7 @@ "ribo_database_manifest": { "type": "string", "format": "file-path", + "exists": true, "mimetype": "text/plain", "default": "${projectDir}/assets/rrna-db-defaults.txt", "fa_icon": "fas fa-database", @@ -193,6 +196,7 @@ "gtf": { "type": "string", "format": "file-path", + "exists": true, "mimetype": "text/plain", "pattern": "^\\S+\\.gtf(\\.gz)?$", "description": "Path to GTF annotation file.", @@ -202,6 +206,7 @@ "gff": { "type": "string", "format": "file-path", + "exists": true, "mimetype": "text/plain", "pattern": "^\\S+\\.gff(\\.gz)?$", "fa_icon": "fas fa-code-branch", @@ -211,6 +216,7 @@ "gene_bed": { "type": "string", "format": "file-path", + "exists": true, "mimetype": "text/plain", "pattern": "^\\S+\\.bed(\\.gz)?$", "fa_icon": "fas fa-procedures", @@ -219,6 +225,7 @@ "transcript_fasta": { "type": "string", "format": "file-path", + "exists": true, "mimetype": "text/plain", "pattern": "^\\S+\\.fn?a(sta)?(\\.gz)?$", "fa_icon": "far fa-file-code", @@ -227,6 +234,7 @@ "additional_fasta": { "type": "string", "format": "file-path", + "exists": true, "mimetype": "text/plain", "pattern": "^\\S+\\.fn?a(sta)?(\\.gz)?$", "fa_icon": "far fa-file-code", @@ -236,6 +244,7 @@ "splicesites": { "type": "string", "format": "file-path", + "exists": true, "mimetype": "text/plain", "fa_icon": "fas fa-hand-scissors", "description": "Splice sites file required for HISAT2." @@ -243,24 +252,28 @@ "star_index": { "type": "string", "format": "path", + "exists": true, "fa_icon": "fas fa-bezier-curve", "description": "Path to directory or tar.gz archive for pre-built STAR index." }, "hisat2_index": { "type": "string", "format": "path", + "exists": true, "fa_icon": "fas fa-bezier-curve", "description": "Path to directory or tar.gz archive for pre-built HISAT2 index." }, "rsem_index": { "type": "string", "format": "path", + "exists": true, "fa_icon": "fas fa-bezier-curve", "description": "Path to directory or tar.gz archive for pre-built RSEM index." }, "salmon_index": { "type": "string", "format": "path", + "exists": true, "fa_icon": "fas fa-bezier-curve", "description": "Path to directory or tar.gz archive for pre-built Salmon index." }, @@ -694,6 +707,7 @@ "multiqc_config": { "type": "string", "format": "file-path", + "exists": true, "mimetype": "text/plain", "description": "Custom config file to supply to MultiQC.", "fa_icon": "fas fa-cog", @@ -702,6 +716,7 @@ "multiqc_logo": { "type": "string", "format": "file-path", + "exists": true, "mimetype": "text/plain", "description": "Custom logo file to supply to MultiQC. File name must also be set in the MultiQC config file", "fa_icon": "fas fa-image", @@ -710,6 +725,7 @@ "multiqc_methods_description": { "type": "string", "format": "file-path", + "exists": true, "mimetype": "text/plain", "description": "Custom MultiQC yaml file containing HTML including a methods description.", "fa_icon": "fas fa-cog" diff --git a/workflows/rnaseq.nf b/workflows/rnaseq.nf index 76c9a6a32..72b7619a4 100755 --- a/workflows/rnaseq.nf +++ b/workflows/rnaseq.nf @@ -193,19 +193,12 @@ workflow RNASEQ { .fromSamplesheet("input") .map { meta, fastq_1, fastq_2 -> - if (!fastq_1.exists()) { - error("Please check input samplesheet -> Read 1 FastQ file does not exist!\n${fastq_1}") - } - if (fastq_2) { - if(!fastq_2.exists()) { - error("Please check input samplesheet -> Read 2 FastQ file does not exist!\n${fastq_2}") - } - return [ meta.id, meta + [ single_end:false ], [ fastq_1, fastq_2 ] ] - } else { + if (!fastq_2) { return [ meta.id, meta + [ single_end:true ], [ fastq_1 ] ] + } else { + return [ meta.id, meta + [ single_end:false ], [ fastq_1, fastq_2 ] ] } } - .unique() .groupTuple() .map { WorkflowRnaseq.validateInput(it) From 8d9ccac23cab6c3f14149a4e8659eaea3120c6e3 Mon Sep 17 00:00:00 2001 From: nf-core-bot Date: Mon, 10 Jul 2023 22:34:29 +0000 Subject: [PATCH 017/184] [automated] Fix linting with Prettier --- docs/usage.md | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/docs/usage.md b/docs/usage.md index 856e00c7f..0162472a4 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -91,9 +91,9 @@ The `--umitools_grouping_method` parameter affects [how similar, but non-identic #### Examples: -| UMI type | Source | Pipeline parameters | -| ------------ | --------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------- | -| In read name | [Illumina BCL convert >3.7.5](https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/software_documentation/bcl_convert/bcl-convert-v3-7-5-software-guide-1000000163594-00.pdf) | `--with_umi --skip_umi_extract --umitools_umi_separator ":"` | +| UMI type | Source | Pipeline parameters | +| ------------ | ---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------- | +| In read name | [Illumina BCL convert >3.7.5](https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/software_documentation/bcl_convert/bcl-convert-v3-7-5-software-guide-1000000163594-00.pdf) | `--with_umi --skip_umi_extract --umitools_umi_separator ":"` | | In sequence | [Lexogen QuantSeq® 3’ mRNA-Seq V2 FWD](https://www.lexogen.com/quantseq-3mrna-sequencing) + [UMI Second Strand Synthesis Module](https://faqs.lexogen.com/faq/how-can-i-add-umis-to-my-quantseq-libraries) | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern "^(?P.{6})(?P.{4}).*"` | | In sequence | [Lexogen CORALL® Total RNA-Seq V1](https://www.lexogen.com/corall-total-rna-seq/)
    > _mind [Appendix H](https://www.lexogen.com/wp-content/uploads/2020/04/095UG190V0130_CORALL-Total-RNA-Seq_2020-03-31.pdf) regarding optional trimming_ | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern "^(?P.{12}).*"`
    Optional: `--clip_r2 9 --three_prime_clip_r2 12` | | In sequence | [Takara Bio SMARTer® Stranded Total RNA-Seq Kit v3](https://www.takarabio.com/documents/User%20Manual/SMARTer%20Stranded%20Total%20RNA/SMARTer%20Stranded%20Total%20RNA-Seq%20Kit%20v3%20-%20Pico%20Input%20Mammalian%20User%20Manual-a_114949.pdf) | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern2 "^(?P.{8})(?P.{6}).*"` | From e3d7c26c9800aec8709d4ef81bc49b571c098e2a Mon Sep 17 00:00:00 2001 From: =?UTF-8?q?Steffen=20M=C3=B6ller?= Date: Thu, 13 Jul 2023 17:06:51 +0200 Subject: [PATCH 018/184] README.md: Pointer to downstream workflow 'differential abundance' --- README.md | 2 ++ 1 file changed, 2 insertions(+) diff --git a/README.md b/README.md index 31314a457..7ff5dfaa1 100644 --- a/README.md +++ b/README.md @@ -95,6 +95,8 @@ To see the results of an example test run with a full size dataset refer to the For more details about the output files and reports, please refer to the [output documentation](https://nf-co.re/rnaseq/output). +This pipeline assigns the RNA/cDNA reads to genes in the genome, counts the number of reads per gene and normalizes the data. It does not compare the samples or assign P values to individual genes. For downstream analyses, the output files can be analysed directly in statistics environments like [R](https://www.r-project.org/) or [Julia](https://julialang.org/), or be forwarded to another workflow of Nextflow to determine transcripts with [differential abundance](https://github.com/nf-core/differentialabundance/). + ## Online videos A short talk about the history, current status and functionality on offer in this pipeline was given by Harshil Patel ([@drpatelh](https://github.com/drpatelh)) on [8th February 2022](https://nf-co.re/events/2022/bytesize-32-nf-core-rnaseq) as part of the nf-core/bytesize series. From 6fcf95f99bbf7ddbc152c1a497c1ea87459441b2 Mon Sep 17 00:00:00 2001 From: hmehlan <43400079+hmehlan@users.noreply.github.com> Date: Wed, 19 Jul 2023 11:06:51 +0200 Subject: [PATCH 019/184] fix copy paste typo --- assets/multiqc/deseq2_pca_header.txt | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/assets/multiqc/deseq2_pca_header.txt b/assets/multiqc/deseq2_pca_header.txt index 2599732af..636f9374d 100644 --- a/assets/multiqc/deseq2_pca_header.txt +++ b/assets/multiqc/deseq2_pca_header.txt @@ -2,7 +2,7 @@ #section_name: 'DESeq2 PCA plot' #description: "PCA plot between samples in the experiment. # These values are calculated using DESeq2 -# in the deseq2_qc.r script." +# in the deseq2_qc.r script." #plot_type: 'scatter' #anchor: 'deseq2_pca' #pconfig: From a55caf0580763d9330d0c4ef49c87f7a3d3c3339 Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Wed, 2 Aug 2023 23:12:08 +0200 Subject: [PATCH 020/184] Update README.md --- README.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/README.md b/README.md index 7ff5dfaa1..4ac07704e 100644 --- a/README.md +++ b/README.md @@ -95,7 +95,7 @@ To see the results of an example test run with a full size dataset refer to the For more details about the output files and reports, please refer to the [output documentation](https://nf-co.re/rnaseq/output). -This pipeline assigns the RNA/cDNA reads to genes in the genome, counts the number of reads per gene and normalizes the data. It does not compare the samples or assign P values to individual genes. For downstream analyses, the output files can be analysed directly in statistics environments like [R](https://www.r-project.org/) or [Julia](https://julialang.org/), or be forwarded to another workflow of Nextflow to determine transcripts with [differential abundance](https://github.com/nf-core/differentialabundance/). +This pipeline quantifies RNA-sequenced reads relative to genes/transcripts in the genome and normalizes the resulting data. It does not compare the samples statistically in order to assign significance in the form of FDR or P-values. For downstream analyses, the output files from this pipeline can be analysed directly in statistical environments like [R](https://www.r-project.org/), [Julia](https://julialang.org/) or via the [nf-core/differentialabundance](https://github.com/nf-core/differentialabundance/) pipeline. ## Online videos From 5162e34fd22fd4af60a4516db77d6f2a2572e8e0 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 8 Aug 2023 14:47:24 +0100 Subject: [PATCH 021/184] Upgrade the untar module --- modules.json | 2 +- modules/nf-core/untar/main.nf | 2 +- 2 files changed, 2 insertions(+), 2 deletions(-) diff --git a/modules.json b/modules.json index 030893337..3bfa5aaea 100644 --- a/modules.json +++ b/modules.json @@ -221,7 +221,7 @@ }, "untar": { "branch": "master", - "git_sha": "5c460c5a4736974abde2843294f35307ee2b0e5e", + "git_sha": "d0b4fc03af52a1cc8c6fb4493b921b57352b1dd8", "installed_by": ["modules"] } } diff --git a/modules/nf-core/untar/main.nf b/modules/nf-core/untar/main.nf index 8cd1856c7..61461c391 100644 --- a/modules/nf-core/untar/main.nf +++ b/modules/nf-core/untar/main.nf @@ -2,7 +2,7 @@ process UNTAR { tag "$archive" label 'process_single' - conda "conda-forge::sed=4.7 bioconda::grep=3.4 conda-forge::tar=1.34" + conda "conda-forge::sed=4.7 conda-forge::grep=3.11 conda-forge::tar=1.34" container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? 'https://depot.galaxyproject.org/singularity/ubuntu:20.04' : 'nf-core/ubuntu:20.04' }" From 04887bdc2e8d1aaa7d96c7149cc7fd027311d46c Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 8 Aug 2023 14:52:32 +0100 Subject: [PATCH 022/184] Add to CHANGELOG --- CHANGELOG.md | 1 + 1 file changed, 1 insertion(+) diff --git a/CHANGELOG.md b/CHANGELOG.md index 951921723..1286a2459 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -22,6 +22,7 @@ Thank you to everyone else that has contributed by reporting bugs, enhancements - [PR #1051](https://github.com/nf-core/rnaseq/pull/1051) - Remove `public_aws_ecr` profile - [PR #1054](https://github.com/nf-core/rnaseq/pull/1054) - Template update to nf-core/tools v2.9 - [PR #1058](https://github.com/nf-core/rnaseq/pull/1058) - Use `nf-validation` plugin for parameter and samplesheet validation +- [PR #1068](https://github.com/nf-core/rnaseq/pull/1068) - Update untar ## [[3.12.0](https://github.com/nf-core/rnaseq/releases/tag/3.12.0)] - 2023-06-02 From 22fd5ffc7adeeb4e50dff0278fac0fd23afa3a2d Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Tue, 8 Aug 2023 19:34:02 +0200 Subject: [PATCH 023/184] Update CHANGELOG.md --- CHANGELOG.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index 1286a2459..c589534d3 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -22,7 +22,7 @@ Thank you to everyone else that has contributed by reporting bugs, enhancements - [PR #1051](https://github.com/nf-core/rnaseq/pull/1051) - Remove `public_aws_ecr` profile - [PR #1054](https://github.com/nf-core/rnaseq/pull/1054) - Template update to nf-core/tools v2.9 - [PR #1058](https://github.com/nf-core/rnaseq/pull/1058) - Use `nf-validation` plugin for parameter and samplesheet validation -- [PR #1068](https://github.com/nf-core/rnaseq/pull/1068) - Update untar +- [PR #1068](https://github.com/nf-core/rnaseq/pull/1068) - Update `grep` version for `untar` module ## [[3.12.0](https://github.com/nf-core/rnaseq/releases/tag/3.12.0)] - 2023-06-02 From efe99ab5d5891ab30cc59ec3bdc8a4de198e4890 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Wed, 30 Aug 2023 09:45:25 +0100 Subject: [PATCH 024/184] Update usage.md for igenomes warning --- docs/usage.md | 19 +++++++++++++++++-- 1 file changed, 17 insertions(+), 2 deletions(-) diff --git a/docs/usage.md b/docs/usage.md index 0162472a4..604f82bc6 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -135,9 +135,18 @@ If unique molecular identifiers were used to prepare the library, add the follow Please refer to the [nf-core website](https://nf-co.re/usage/reference_genomes) for general usage docs and guidelines regarding reference genomes. +:::warning +When using the --genome parameter (e.g. --genome GRCh37), you are referring to references from AWS-iGenomes. Please be aware that: + +- The igenomes file usage triggered by this option is outdated with respect to gene annotations. This can be particularly problematic for RNA-seq analysis, which relies on accurate gene annotation. +- Some iGenomes references (e.g., GRCh38) point to annotation files that use gene symbols as the primary identifier. This can cause issues for downstream analysis, such as the nf-core [differential abundance workflow](https://nf-co.re/differentialabundance) workflow. + +We recommend that you provide reference files directly, via `--gtf` and `--fasta`, and that supplied GTF files do not use gene names as `gene_id`. +::: + The minimum reference genome requirements for this pipeline are a FASTA and GTF file, all other files required to run the pipeline can be generated from these files. However, it is more storage and compute friendly if you are able to re-use reference genome files as efficiently as possible. It is recommended to use the `--save_reference` parameter if you are using the pipeline to build new indices (e.g. custom genomes that are unavailable on [AWS iGenomes](https://nf-co.re/usage/reference_genomes#custom-genomes)) so that you can save them somewhere locally. The index building step can be quite a time-consuming process and it permits their reuse for future runs of the pipeline to save disk space. You can then either provide the appropriate reference genome files on the command-line via the appropriate parameters (e.g. `--star_index '/path/to/STAR/index/'`) or via a custom config file. Another option is to run the pipeline once with `--save_reference --skip_alignment --skip_pseudo_alignment` to generate and save all of the required reference files and indices to the results directory. You can then move the reference files in `/genome/` to a more permanent location and use these paths to override the relevant parameters in the pipeline e.g. `--star_index`. -- If `--genome` is provided then the FASTA and GTF files (and existing indices) will be automatically obtained from AWS-iGenomes unless these have already been downloaded locally in the path specified by `--igenomes_base`. +- If `--genome` (discouraged- see warning above) is provided then the FASTA and GTF files (and existing indices) will be automatically obtained from AWS-iGenomes unless these have already been downloaded locally in the path specified by `--igenomes_base`. - If `--gff` is provided as input then this will be converted to a GTF file, or the latter will be used if both are provided. - If `--gene_bed` is not provided then it will be generated from the GTF file. - If `--additional_fasta` is provided then the features in this file (e.g. ERCC spike-ins) will be automatically concatenated onto both the reference FASTA file as well as the GTF annotation before building the appropriate indices. @@ -169,7 +178,13 @@ Please get in touch with us on the #rnaseq channel in the [nf-core Slack workspa The typical command for running the pipeline is as follows: ```bash -nextflow run nf-core/rnaseq --input --outdir --genome GRCh37 -profile docker +nextflow run \ + nf-core/rnaseq \ + --input \ + --outdir \ + --gtf Homo_sapiens.GRCh38.110.gtf.gz \ + --fasta Homo_sapiens.GRCh38.dna_sm.primary_assembly.fa.gz \ + -profile docker ``` This will launch the pipeline with the `docker` configuration profile. See below for more information about profiles. From 20eb224fa54e5db1a87f0642b4c0890e0c759d29 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Wed, 30 Aug 2023 09:49:32 +0100 Subject: [PATCH 025/184] Update CHANGELOG.md --- CHANGELOG.md | 1 + 1 file changed, 1 insertion(+) diff --git a/CHANGELOG.md b/CHANGELOG.md index c589534d3..6df91910c 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -23,6 +23,7 @@ Thank you to everyone else that has contributed by reporting bugs, enhancements - [PR #1054](https://github.com/nf-core/rnaseq/pull/1054) - Template update to nf-core/tools v2.9 - [PR #1058](https://github.com/nf-core/rnaseq/pull/1058) - Use `nf-validation` plugin for parameter and samplesheet validation - [PR #1068](https://github.com/nf-core/rnaseq/pull/1068) - Update `grep` version for `untar` module +- [PR #1073](https://github.com/nf-core/rnaseq/pull/1073) - Update documentation to discourage --genome ## [[3.12.0](https://github.com/nf-core/rnaseq/releases/tag/3.12.0)] - 2023-06-02 From 170af8e9adf218fbab3f6f5a16daf285d9e8680d Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Wed, 30 Aug 2023 09:51:57 +0100 Subject: [PATCH 026/184] Update usage.md to match reference file usage to match other inputs --- docs/usage.md | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/docs/usage.md b/docs/usage.md index 604f82bc6..ae5e00f1a 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -182,8 +182,8 @@ nextflow run \ nf-core/rnaseq \ --input \ --outdir \ - --gtf Homo_sapiens.GRCh38.110.gtf.gz \ - --fasta Homo_sapiens.GRCh38.dna_sm.primary_assembly.fa.gz \ + --gtf \ + --fasta \ -profile docker ``` From 018876cd995dc818e872b4c7b9c18f0b91f01d65 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Mon, 4 Sep 2023 15:05:22 +0100 Subject: [PATCH 027/184] Fix repeated word --- docs/usage.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/docs/usage.md b/docs/usage.md index ae5e00f1a..b8b869421 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -139,7 +139,7 @@ Please refer to the [nf-core website](https://nf-co.re/usage/reference_genomes) When using the --genome parameter (e.g. --genome GRCh37), you are referring to references from AWS-iGenomes. Please be aware that: - The igenomes file usage triggered by this option is outdated with respect to gene annotations. This can be particularly problematic for RNA-seq analysis, which relies on accurate gene annotation. -- Some iGenomes references (e.g., GRCh38) point to annotation files that use gene symbols as the primary identifier. This can cause issues for downstream analysis, such as the nf-core [differential abundance workflow](https://nf-co.re/differentialabundance) workflow. +- Some iGenomes references (e.g., GRCh38) point to annotation files that use gene symbols as the primary identifier. This can cause issues for downstream analysis, such as the nf-core [differential abundance](https://nf-co.re/differentialabundance) workflow. We recommend that you provide reference files directly, via `--gtf` and `--fasta`, and that supplied GTF files do not use gene names as `gene_id`. ::: From 7e09601e7765e45508e4e0ceeaf4d3bc1907a278 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Mon, 4 Sep 2023 17:14:18 +0100 Subject: [PATCH 028/184] Rework reference coverage, include download commands --- docs/usage.md | 53 ++++++++++++++++++++++++++++++++++++--------------- 1 file changed, 38 insertions(+), 15 deletions(-) diff --git a/docs/usage.md b/docs/usage.md index b8b869421..66b5ae865 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -131,38 +131,39 @@ If unique molecular identifiers were used to prepare the library, add the follow --umitools_bc_pattern "^(?P.{6})(?P.{4}).*" ``` -## Reference genome files +## Reference genome options Please refer to the [nf-core website](https://nf-co.re/usage/reference_genomes) for general usage docs and guidelines regarding reference genomes. -:::warning -When using the --genome parameter (e.g. --genome GRCh37), you are referring to references from AWS-iGenomes. Please be aware that: +### Explicit reference file specification (recommended) -- The igenomes file usage triggered by this option is outdated with respect to gene annotations. This can be particularly problematic for RNA-seq analysis, which relies on accurate gene annotation. -- Some iGenomes references (e.g., GRCh38) point to annotation files that use gene symbols as the primary identifier. This can cause issues for downstream analysis, such as the nf-core [differential abundance](https://nf-co.re/differentialabundance) workflow. +The minimum reference genome requirements for this pipeline are a FASTA and GTF file, all other files required to run the pipeline can be generated from these files. For example, the latest reference files for human can be derived from Ensembl like: -We recommend that you provide reference files directly, via `--gtf` and `--fasta`, and that supplied GTF files do not use gene names as `gene_id`. -::: +``` +latest_release=$(curl -s 'http://rest.ensembl.org/info/software?content-type=application/json' | grep -o '"release":[0-9]*' | cut -d: -f2) +wget -L ftp://ftp.ensembl.org/pub/release-${latest_release}/fasta/homo_sapiens/dna/Homo_sapiens.GRCh38.dna_sm.primary_assembly.fa.gz +wget -L ftp://ftp.ensembl.org/pub/release-${latest_release}/gtf/homo_sapiens/Homo_sapiens.GRCh38.${latest_release}.gtf.gz +``` + +These files can then be specified to the workflow with the `--fasta` and `--gtf` parameters. + +> **NB:** Compressed reference files are supported by the pipeline i.e. standard files with the `.gz` extension and indices folders with the `tar.gz` extension. -The minimum reference genome requirements for this pipeline are a FASTA and GTF file, all other files required to run the pipeline can be generated from these files. However, it is more storage and compute friendly if you are able to re-use reference genome files as efficiently as possible. It is recommended to use the `--save_reference` parameter if you are using the pipeline to build new indices (e.g. custom genomes that are unavailable on [AWS iGenomes](https://nf-co.re/usage/reference_genomes#custom-genomes)) so that you can save them somewhere locally. The index building step can be quite a time-consuming process and it permits their reuse for future runs of the pipeline to save disk space. You can then either provide the appropriate reference genome files on the command-line via the appropriate parameters (e.g. `--star_index '/path/to/STAR/index/'`) or via a custom config file. Another option is to run the pipeline once with `--save_reference --skip_alignment --skip_pseudo_alignment` to generate and save all of the required reference files and indices to the results directory. You can then move the reference files in `/genome/` to a more permanent location and use these paths to override the relevant parameters in the pipeline e.g. `--star_index`. +Notes: -- If `--genome` (discouraged- see warning above) is provided then the FASTA and GTF files (and existing indices) will be automatically obtained from AWS-iGenomes unless these have already been downloaded locally in the path specified by `--igenomes_base`. - If `--gff` is provided as input then this will be converted to a GTF file, or the latter will be used if both are provided. - If `--gene_bed` is not provided then it will be generated from the GTF file. - If `--additional_fasta` is provided then the features in this file (e.g. ERCC spike-ins) will be automatically concatenated onto both the reference FASTA file as well as the GTF annotation before building the appropriate indices. +- When using `--aligner star_rsem`, both the STAR and RSEM indices should be present in the path specified by `--rsem_index` (see [#568](https://github.com/nf-core/rnaseq/issues/568)). -When using `--aligner star_rsem`, both the STAR and RSEM indices should be present in the path specified by `--rsem_index` (see [#568](https://github.com/nf-core/rnaseq/issues/568)). - -> **NB:** Compressed reference files are also supported by the pipeline i.e. standard files with the `.gz` extension and indices folders with the `tar.gz` extension. - -As of v3.7 of the pipeline, if you are using a genome downloaded from AWS iGenomes and using `--aligner star_salmon` (default) the version of STAR to use for the alignment will be auto-detected (see [#808](https://github.com/nf-core/rnaseq/issues/808)). +#### Gencode If you are using [GENCODE](https://www.gencodegenes.org/) reference genome files please specify the `--gencode` parameter because the format of these files is slightly different to ENSEMBL genome files: - The `--gtf_group_features_type` parameter will automatically be set to `gene_type` as opposed to `gene_biotype`, respectively. - If you are running Salmon, the `--gencode` flag will also be passed to the index building step to overcome parsing issues resulting from the transcript IDs in GENCODE fasta files being separated by vertical pipes (`|`) instead of spaces (see [this issue](https://github.com/COMBINE-lab/salmon/issues/15)). -## Prokaryotic genome annotations +#### Prokaryotic genome annotations This pipeline uses featureCounts to generate QC metrics based on [biotype](http://www.ensembl.org/info/genome/genebuild/biotypes.html) information available within GFF/GTF genome annotation files. The format of these annotation files can vary significantly depending on the source of the annotation and the type of organism. The default settings in the pipeline are tailored towards Ensembl GTF annotations available for eukaryotic genomes. Prokaryotic genome annotations tend to be distributed in GFF format which are structured differently in terms of the feature naming conventions. There are a number of ways you can tune the behaviour of the pipeline to cater for differences/absence of biotype information: @@ -173,6 +174,28 @@ This pipeline uses featureCounts to generate QC metrics based on [biotype](http: Please get in touch with us on the #rnaseq channel in the [nf-core Slack workspace](https://nf-co.re/join) if you are having problems or need any advice. +#### Indices + +By default, indices are generated dynamically by the workflow for tools such as STAR and Salmon. Since indexing is an expensive process in time and resources you should ensure that it is only done once, by retaining the indices generated from each batch of reference files: + +- the `--save_reference` parameter will save your indices in your results directory +- the `--skip_alignment --skip_pseudo_alignment` will disable other processes if you'd like to do an 'indexing only' workflow run. + +Once you have the indices from a workflow run you should save them somewhere central and reuse them in subsequent runs using custom config files or command line parameters such as `--star_index '/path/to/STAR/index/'`. + +### iGenomes (not recommended) + +If the `--genome` parameter is provided (e.g. `--genome GRCh37`) then the FASTA and GTF files (and existing indices) will be automatically obtained from AWS-iGenomes unless these have already been downloaded locally in the path specified by `--igenomes_base`. + +However this is no longer recommended because: + +- Gene annotations in iGenomes are extremely out of date. This can be particularly problematic for RNA-seq analysis, which relies on accurate gene annotation. +- Some iGenomes references (e.g., GRCh38) point to annotation files that use gene symbols as the primary identifier. This can cause issues for downstream analysis, such as the nf-core [differential abundance](https://nf-co.re/differentialabundance) workflow where a conventional gene identifier distinct from symbol is expected. + +Notes: + +- As of v3.7 of the pipeline, if you are using a genome downloaded from AWS iGenomes and using `--aligner star_salmon` (default) the version of STAR to use for the alignment will be auto-detected (see [#808](https://github.com/nf-core/rnaseq/issues/808)). + ## Running the pipeline The typical command for running the pipeline is as follows: From b69951bcc422b00dcf58295a6984c57a8eeae0b7 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Mon, 4 Sep 2023 17:15:40 +0100 Subject: [PATCH 029/184] Integrate notes --- docs/usage.md | 3 +-- 1 file changed, 1 insertion(+), 2 deletions(-) diff --git a/docs/usage.md b/docs/usage.md index 66b5ae865..b77b21128 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -147,10 +147,9 @@ wget -L ftp://ftp.ensembl.org/pub/release-${latest_release}/gtf/homo_sapiens/Hom These files can then be specified to the workflow with the `--fasta` and `--gtf` parameters. -> **NB:** Compressed reference files are supported by the pipeline i.e. standard files with the `.gz` extension and indices folders with the `tar.gz` extension. - Notes: +- Compressed reference files are supported by the pipeline i.e. standard files with the `.gz` extension and indices folders with the `tar.gz` extension. - If `--gff` is provided as input then this will be converted to a GTF file, or the latter will be used if both are provided. - If `--gene_bed` is not provided then it will be generated from the GTF file. - If `--additional_fasta` is provided then the features in this file (e.g. ERCC spike-ins) will be automatically concatenated onto both the reference FASTA file as well as the GTF annotation before building the appropriate indices. From 75bb8619e8e3a36666cc10b7e02023c3a3c3b4fc Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Mon, 4 Sep 2023 17:18:00 +0100 Subject: [PATCH 030/184] Move index discussion earlier --- docs/usage.md | 18 +++++++++--------- 1 file changed, 9 insertions(+), 9 deletions(-) diff --git a/docs/usage.md b/docs/usage.md index b77b21128..ad0fd3754 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -155,6 +155,15 @@ Notes: - If `--additional_fasta` is provided then the features in this file (e.g. ERCC spike-ins) will be automatically concatenated onto both the reference FASTA file as well as the GTF annotation before building the appropriate indices. - When using `--aligner star_rsem`, both the STAR and RSEM indices should be present in the path specified by `--rsem_index` (see [#568](https://github.com/nf-core/rnaseq/issues/568)). +#### Indices + +By default, indices are generated dynamically by the workflow for tools such as STAR and Salmon. Since indexing is an expensive process in time and resources you should ensure that it is only done once, by retaining the indices generated from each batch of reference files: + +- the `--save_reference` parameter will save your indices in your results directory +- the `--skip_alignment --skip_pseudo_alignment` will disable other processes if you'd like to do an 'indexing only' workflow run. + +Once you have the indices from a workflow run you should save them somewhere central and reuse them in subsequent runs using custom config files or command line parameters such as `--star_index '/path/to/STAR/index/'`. + #### Gencode If you are using [GENCODE](https://www.gencodegenes.org/) reference genome files please specify the `--gencode` parameter because the format of these files is slightly different to ENSEMBL genome files: @@ -173,15 +182,6 @@ This pipeline uses featureCounts to generate QC metrics based on [biotype](http: Please get in touch with us on the #rnaseq channel in the [nf-core Slack workspace](https://nf-co.re/join) if you are having problems or need any advice. -#### Indices - -By default, indices are generated dynamically by the workflow for tools such as STAR and Salmon. Since indexing is an expensive process in time and resources you should ensure that it is only done once, by retaining the indices generated from each batch of reference files: - -- the `--save_reference` parameter will save your indices in your results directory -- the `--skip_alignment --skip_pseudo_alignment` will disable other processes if you'd like to do an 'indexing only' workflow run. - -Once you have the indices from a workflow run you should save them somewhere central and reuse them in subsequent runs using custom config files or command line parameters such as `--star_index '/path/to/STAR/index/'`. - ### iGenomes (not recommended) If the `--genome` parameter is provided (e.g. `--genome GRCh37`) then the FASTA and GTF files (and existing indices) will be automatically obtained from AWS-iGenomes unless these have already been downloaded locally in the path specified by `--igenomes_base`. From 6ee8f6eac515fc60f358518cca9cf97457178b0a Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Mon, 4 Sep 2023 17:19:39 +0100 Subject: [PATCH 031/184] Prettier --- docs/usage.md | 10 +++++----- 1 file changed, 5 insertions(+), 5 deletions(-) diff --git a/docs/usage.md b/docs/usage.md index ad0fd3754..2aedc75e9 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -91,9 +91,9 @@ The `--umitools_grouping_method` parameter affects [how similar, but non-identic #### Examples: -| UMI type | Source | Pipeline parameters | -| ------------ | ---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------- | -| In read name | [Illumina BCL convert >3.7.5](https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/software_documentation/bcl_convert/bcl-convert-v3-7-5-software-guide-1000000163594-00.pdf) | `--with_umi --skip_umi_extract --umitools_umi_separator ":"` | +| UMI type | Source | Pipeline parameters | +| ------------ | --------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------- | +| In read name | [Illumina BCL convert >3.7.5](https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/software_documentation/bcl_convert/bcl-convert-v3-7-5-software-guide-1000000163594-00.pdf) | `--with_umi --skip_umi_extract --umitools_umi_separator ":"` | | In sequence | [Lexogen QuantSeq® 3’ mRNA-Seq V2 FWD](https://www.lexogen.com/quantseq-3mrna-sequencing) + [UMI Second Strand Synthesis Module](https://faqs.lexogen.com/faq/how-can-i-add-umis-to-my-quantseq-libraries) | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern "^(?P.{6})(?P.{4}).*"` | | In sequence | [Lexogen CORALL® Total RNA-Seq V1](https://www.lexogen.com/corall-total-rna-seq/)
    > _mind [Appendix H](https://www.lexogen.com/wp-content/uploads/2020/04/095UG190V0130_CORALL-Total-RNA-Seq_2020-03-31.pdf) regarding optional trimming_ | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern "^(?P.{12}).*"`
    Optional: `--clip_r2 9 --three_prime_clip_r2 12` | | In sequence | [Takara Bio SMARTer® Stranded Total RNA-Seq Kit v3](https://www.takarabio.com/documents/User%20Manual/SMARTer%20Stranded%20Total%20RNA/SMARTer%20Stranded%20Total%20RNA-Seq%20Kit%20v3%20-%20Pico%20Input%20Mammalian%20User%20Manual-a_114949.pdf) | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern2 "^(?P.{8})(?P.{6}).*"` | @@ -145,7 +145,7 @@ wget -L ftp://ftp.ensembl.org/pub/release-${latest_release}/fasta/homo_sapiens/d wget -L ftp://ftp.ensembl.org/pub/release-${latest_release}/gtf/homo_sapiens/Homo_sapiens.GRCh38.${latest_release}.gtf.gz ``` -These files can then be specified to the workflow with the `--fasta` and `--gtf` parameters. +These files can then be specified to the workflow with the `--fasta` and `--gtf` parameters. Notes: @@ -160,7 +160,7 @@ Notes: By default, indices are generated dynamically by the workflow for tools such as STAR and Salmon. Since indexing is an expensive process in time and resources you should ensure that it is only done once, by retaining the indices generated from each batch of reference files: - the `--save_reference` parameter will save your indices in your results directory -- the `--skip_alignment --skip_pseudo_alignment` will disable other processes if you'd like to do an 'indexing only' workflow run. +- the `--skip_alignment --skip_pseudo_alignment` will disable other processes if you'd like to do an 'indexing only' workflow run. Once you have the indices from a workflow run you should save them somewhere central and reuse them in subsequent runs using custom config files or command line parameters such as `--star_index '/path/to/STAR/index/'`. From f706b6f3617b35a3cb03d69abd5ff5cb5f39d63f Mon Sep 17 00:00:00 2001 From: nf-core-bot Date: Mon, 4 Sep 2023 17:02:11 +0000 Subject: [PATCH 032/184] [automated] Fix linting with Prettier --- docs/usage.md | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/docs/usage.md b/docs/usage.md index 2aedc75e9..dd3398f13 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -91,9 +91,9 @@ The `--umitools_grouping_method` parameter affects [how similar, but non-identic #### Examples: -| UMI type | Source | Pipeline parameters | -| ------------ | --------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------- | -| In read name | [Illumina BCL convert >3.7.5](https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/software_documentation/bcl_convert/bcl-convert-v3-7-5-software-guide-1000000163594-00.pdf) | `--with_umi --skip_umi_extract --umitools_umi_separator ":"` | +| UMI type | Source | Pipeline parameters | +| ------------ | ---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------- | +| In read name | [Illumina BCL convert >3.7.5](https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/software_documentation/bcl_convert/bcl-convert-v3-7-5-software-guide-1000000163594-00.pdf) | `--with_umi --skip_umi_extract --umitools_umi_separator ":"` | | In sequence | [Lexogen QuantSeq® 3’ mRNA-Seq V2 FWD](https://www.lexogen.com/quantseq-3mrna-sequencing) + [UMI Second Strand Synthesis Module](https://faqs.lexogen.com/faq/how-can-i-add-umis-to-my-quantseq-libraries) | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern "^(?P.{6})(?P.{4}).*"` | | In sequence | [Lexogen CORALL® Total RNA-Seq V1](https://www.lexogen.com/corall-total-rna-seq/)
    > _mind [Appendix H](https://www.lexogen.com/wp-content/uploads/2020/04/095UG190V0130_CORALL-Total-RNA-Seq_2020-03-31.pdf) regarding optional trimming_ | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern "^(?P.{12}).*"`
    Optional: `--clip_r2 9 --three_prime_clip_r2 12` | | In sequence | [Takara Bio SMARTer® Stranded Total RNA-Seq Kit v3](https://www.takarabio.com/documents/User%20Manual/SMARTer%20Stranded%20Total%20RNA/SMARTer%20Stranded%20Total%20RNA-Seq%20Kit%20v3%20-%20Pico%20Input%20Mammalian%20User%20Manual-a_114949.pdf) | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern2 "^(?P.{8})(?P.{6}).*"` | From 08d492d170e121b99cc16afb7675fd08ab1a763f Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Mon, 4 Sep 2023 18:30:38 +0100 Subject: [PATCH 033/184] Add igenomes_ignore --- docs/usage.md | 2 ++ 1 file changed, 2 insertions(+) diff --git a/docs/usage.md b/docs/usage.md index 2aedc75e9..7afe22e9f 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -206,6 +206,8 @@ nextflow run \ --outdir \ --gtf \ --fasta \ + --igenomes_ignore \ + --genome null \ -profile docker ``` From c4e17a6704c6167663cc239f7f3a456138a1f330 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Mon, 4 Sep 2023 18:40:34 +0100 Subject: [PATCH 034/184] Actually, let's make non-use of iGenomes the default --- docs/usage.md | 2 -- nextflow.config | 4 ++-- nextflow_schema.json | 6 +++--- 3 files changed, 5 insertions(+), 7 deletions(-) diff --git a/docs/usage.md b/docs/usage.md index c72007643..dd3398f13 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -206,8 +206,6 @@ nextflow run \ --outdir \ --gtf \ --fasta \ - --igenomes_ignore \ - --genome null \ -profile docker ``` diff --git a/nextflow.config b/nextflow.config index 692d2df47..017cf11b0 100644 --- a/nextflow.config +++ b/nextflow.config @@ -22,7 +22,7 @@ params { gencode = false save_reference = false igenomes_base = 's3://ngi-igenomes/igenomes' - igenomes_ignore = false + load_igenomes = false // UMI handling with_umi = false @@ -266,7 +266,7 @@ plugins { } // Load igenomes.config if required -if (!params.igenomes_ignore) { +if (params.load_igenomes) { includeConfig 'conf/igenomes.config' } else { params.genomes = [:] diff --git a/nextflow_schema.json b/nextflow_schema.json index e3f16078a..bd2746ce8 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -331,12 +331,12 @@ "fa_icon": "fas fa-cloud-download-alt", "hidden": true }, - "igenomes_ignore": { + "load_igenomes": { "type": "boolean", - "description": "Do not load the iGenomes reference config.", + "description": "Load the iGenomes reference config.", "fa_icon": "fas fa-ban", "hidden": true, - "help_text": "Do not load `igenomes.config` when running the pipeline. You may choose this option if you observe clashes between custom parameters and those supplied in `igenomes.config`." + "help_text": "Load `igenomes.config` when running the pipeline. You may choose this option if you observe clashes between custom parameters and those supplied in `igenomes.config`." } }, "required": ["fasta"] From 5744aa0892f737300d6db197edef4fb47f1c4da4 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Mon, 4 Sep 2023 18:56:25 +0100 Subject: [PATCH 035/184] Fix schema help --- nextflow_schema.json | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/nextflow_schema.json b/nextflow_schema.json index bd2746ce8..02b0a03da 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -181,7 +181,7 @@ "type": "string", "description": "Name of iGenomes reference.", "fa_icon": "fas fa-book", - "help_text": "If using a reference genome configured in the pipeline using iGenomes, use this parameter to give the ID for the reference. This is then used to build the full paths for all required reference genome files e.g. `--genome GRCh38`. \n\nSee the [nf-core website docs](https://nf-co.re/usage/reference_genomes) for more details." + "help_text": "If using a reference genome configured in the pipeline using iGenomes (not recommended), use this parameter to give the ID for the reference. This is then used to build the full paths for all required reference genome files e.g. `--genome GRCh38`. \n\nSee the [nf-core website docs](https://nf-co.re/usage/reference_genomes) for more details." }, "fasta": { "type": "string", @@ -336,7 +336,7 @@ "description": "Load the iGenomes reference config.", "fa_icon": "fas fa-ban", "hidden": true, - "help_text": "Load `igenomes.config` when running the pipeline. You may choose this option if you observe clashes between custom parameters and those supplied in `igenomes.config`." + "help_text": "Load `igenomes.config` when running the pipeline. You may choose this option if you want to use iGenomes reference configurations like `--genome GRCh38` rather than explicit reference file provision." } }, "required": ["fasta"] From 7b063e3c58bdbc42497b0e28e417744eeaa9eff4 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 5 Sep 2023 08:59:19 +0100 Subject: [PATCH 036/184] Restore default igenomes behaviour for testing --- conf/test.config | 1 + 1 file changed, 1 insertion(+) diff --git a/conf/test.config b/conf/test.config index d0430eacb..5b841ea4f 100644 --- a/conf/test.config +++ b/conf/test.config @@ -24,6 +24,7 @@ params { input = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/samplesheet/v3.10/samplesheet_test.csv" // Genome references + load_igenomes = true fasta = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/reference/genome.fasta" gtf = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/reference/genes.gtf.gz" gff = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/reference/genes.gff.gz" From 9d66f0e30dfde99b21a6a9ead48c7322a9c11081 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 5 Sep 2023 09:36:14 +0100 Subject: [PATCH 037/184] Revert "Actually, let's make non-use of iGenomes the default" This reverts commit c4e17a6704c6167663cc239f7f3a456138a1f330. --- docs/usage.md | 2 ++ nextflow.config | 4 ++-- nextflow_schema.json | 6 +++--- 3 files changed, 7 insertions(+), 5 deletions(-) diff --git a/docs/usage.md b/docs/usage.md index dd3398f13..c72007643 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -206,6 +206,8 @@ nextflow run \ --outdir \ --gtf \ --fasta \ + --igenomes_ignore \ + --genome null \ -profile docker ``` diff --git a/nextflow.config b/nextflow.config index 017cf11b0..692d2df47 100644 --- a/nextflow.config +++ b/nextflow.config @@ -22,7 +22,7 @@ params { gencode = false save_reference = false igenomes_base = 's3://ngi-igenomes/igenomes' - load_igenomes = false + igenomes_ignore = false // UMI handling with_umi = false @@ -266,7 +266,7 @@ plugins { } // Load igenomes.config if required -if (params.load_igenomes) { +if (!params.igenomes_ignore) { includeConfig 'conf/igenomes.config' } else { params.genomes = [:] diff --git a/nextflow_schema.json b/nextflow_schema.json index 02b0a03da..8c9247048 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -331,12 +331,12 @@ "fa_icon": "fas fa-cloud-download-alt", "hidden": true }, - "load_igenomes": { + "igenomes_ignore": { "type": "boolean", - "description": "Load the iGenomes reference config.", + "description": "Do not load the iGenomes reference config.", "fa_icon": "fas fa-ban", "hidden": true, - "help_text": "Load `igenomes.config` when running the pipeline. You may choose this option if you want to use iGenomes reference configurations like `--genome GRCh38` rather than explicit reference file provision." + "help_text": "Do not load `igenomes.config` when running the pipeline. You may choose this option if you observe clashes between custom parameters and those supplied in `igenomes.config`." } }, "required": ["fasta"] From 3fdd964c52901f89fdd27d5cf2e8ef90f5279228 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 5 Sep 2023 09:37:46 +0100 Subject: [PATCH 038/184] Revert "Restore default igenomes behaviour for testing" This reverts commit 7b063e3c58bdbc42497b0e28e417744eeaa9eff4. --- conf/test.config | 1 - 1 file changed, 1 deletion(-) diff --git a/conf/test.config b/conf/test.config index 5b841ea4f..d0430eacb 100644 --- a/conf/test.config +++ b/conf/test.config @@ -24,7 +24,6 @@ params { input = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/samplesheet/v3.10/samplesheet_test.csv" // Genome references - load_igenomes = true fasta = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/reference/genome.fasta" gtf = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/reference/genes.gtf.gz" gff = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/reference/genes.gff.gz" From db2b2ac85606afa0f5df4e82e186fe42bd664e7d Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 5 Sep 2023 09:42:54 +0100 Subject: [PATCH 039/184] Add mitigating note on igenomes_ignore --- docs/usage.md | 2 ++ 1 file changed, 2 insertions(+) diff --git a/docs/usage.md b/docs/usage.md index c72007643..c0b2d1ba9 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -211,6 +211,8 @@ nextflow run \ -profile docker ``` +> **NB:** Loading iGenomes configuration remains the default for reasons of consistency with other workflows, but should be disabled when not using iGenomes, applying the recommended usage above. + This will launch the pipeline with the `docker` configuration profile. See below for more information about profiles. Note that the pipeline will create the following files in your working directory: From 85480063f220cd59fa9b4294e4e13b01fa0e6bfd Mon Sep 17 00:00:00 2001 From: nf-core-bot Date: Tue, 26 Sep 2023 08:03:29 +0000 Subject: [PATCH 040/184] Template update for nf-core/tools version 2.10 --- .devcontainer/devcontainer.json | 1 + .github/CONTRIBUTING.md | 4 +- .github/workflows/linting.yml | 2 +- .github/workflows/release-announcments.yml | 68 +++++++++ CHANGELOG.md | 2 +- CITATIONS.md | 2 +- CODE_OF_CONDUCT.md | 133 ++++++++++++++---- README.md | 21 +-- assets/multiqc_config.yml | 4 +- conf/modules.config | 9 ++ docs/output.md | 5 +- docs/usage.md | 16 ++- lib/NfcoreTemplate.groovy | 16 +++ lib/WorkflowRnaseq.groovy | 2 +- main.nf | 3 + modules.json | 6 +- .../custom/dumpsoftwareversions/main.nf | 2 +- modules/nf-core/fastqc/main.nf | 8 +- modules/nf-core/multiqc/main.nf | 2 +- nextflow.config | 9 +- nextflow_schema.json | 15 -- workflows/rnaseq.nf | 1 + 22 files changed, 253 insertions(+), 78 deletions(-) create mode 100644 .github/workflows/release-announcments.yml diff --git a/.devcontainer/devcontainer.json b/.devcontainer/devcontainer.json index ea27a5843..4ecfbfe33 100644 --- a/.devcontainer/devcontainer.json +++ b/.devcontainer/devcontainer.json @@ -2,6 +2,7 @@ "name": "nfcore", "image": "nfcore/gitpod:latest", "remoteUser": "gitpod", + "runArgs": ["--privileged"], // Configure tool-specific properties. "customizations": { diff --git a/.github/CONTRIBUTING.md b/.github/CONTRIBUTING.md index fe09aec68..ec7e537b7 100644 --- a/.github/CONTRIBUTING.md +++ b/.github/CONTRIBUTING.md @@ -9,7 +9,9 @@ Please use the pre-filled template to save time. However, don't be put off by this template - other more general issues and suggestions are welcome! Contributions to the code are even more welcome ;) -> If you need help using or modifying nf-core/rnaseq then the best place to ask is on the nf-core Slack [#rnaseq](https://nfcore.slack.com/channels/rnaseq) channel ([join our Slack here](https://nf-co.re/join/slack)). +:::info +If you need help using or modifying nf-core/rnaseq then the best place to ask is on the nf-core Slack [#rnaseq](https://nfcore.slack.com/channels/rnaseq) channel ([join our Slack here](https://nf-co.re/join/slack)). +::: ## Contribution workflow diff --git a/.github/workflows/linting.yml b/.github/workflows/linting.yml index 888cb4bc3..b8bdd2143 100644 --- a/.github/workflows/linting.yml +++ b/.github/workflows/linting.yml @@ -78,7 +78,7 @@ jobs: - uses: actions/setup-python@v4 with: - python-version: "3.8" + python-version: "3.11" architecture: "x64" - name: Install dependencies diff --git a/.github/workflows/release-announcments.yml b/.github/workflows/release-announcments.yml new file mode 100644 index 000000000..6ad339277 --- /dev/null +++ b/.github/workflows/release-announcments.yml @@ -0,0 +1,68 @@ +name: release-announcements +# Automatic release toot and tweet anouncements +on: + release: + types: [published] + workflow_dispatch: + +jobs: + toot: + runs-on: ubuntu-latest + steps: + - uses: rzr/fediverse-action@master + with: + access-token: ${{ secrets.MASTODON_ACCESS_TOKEN }} + host: "mstdn.science" # custom host if not "mastodon.social" (default) + # GitHub event payload + # https://docs.github.com/en/developers/webhooks-and-events/webhooks/webhook-events-and-payloads#release + message: | + Pipeline release! ${{ github.repository }} v${{ github.event.release.tag_name }} - ${{ github.event.release.name }}! + + Please see the changelog: ${{ github.event.release.html_url }} + + send-tweet: + runs-on: ubuntu-latest + + steps: + - uses: actions/setup-python@v4 + with: + python-version: "3.10" + - name: Install dependencies + run: pip install tweepy==4.14.0 + - name: Send tweet + shell: python + run: | + import os + import tweepy + + client = tweepy.Client( + access_token=os.getenv("TWITTER_ACCESS_TOKEN"), + access_token_secret=os.getenv("TWITTER_ACCESS_TOKEN_SECRET"), + consumer_key=os.getenv("TWITTER_CONSUMER_KEY"), + consumer_secret=os.getenv("TWITTER_CONSUMER_SECRET"), + ) + tweet = os.getenv("TWEET") + client.create_tweet(text=tweet) + env: + TWEET: | + Pipeline release! ${{ github.repository }} v${{ github.event.release.tag_name }} - ${{ github.event.release.name }}! + + Please see the changelog: ${{ github.event.release.html_url }} + TWITTER_CONSUMER_KEY: ${{ secrets.TWITTER_CONSUMER_KEY }} + TWITTER_CONSUMER_SECRET: ${{ secrets.TWITTER_CONSUMER_SECRET }} + TWITTER_ACCESS_TOKEN: ${{ secrets.TWITTER_ACCESS_TOKEN }} + TWITTER_ACCESS_TOKEN_SECRET: ${{ secrets.TWITTER_ACCESS_TOKEN_SECRET }} + + bsky-post: + runs-on: ubuntu-latest + steps: + - uses: zentered/bluesky-post-action@v0.0.2 + with: + post: | + Pipeline release! ${{ github.repository }} v${{ github.event.release.tag_name }} - ${{ github.event.release.name }}! + + Please see the changelog: ${{ github.event.release.html_url }} + env: + BSKY_IDENTIFIER: ${{ secrets.BSKY_IDENTIFIER }} + BSKY_PASSWORD: ${{ secrets.BSKY_PASSWORD }} + # diff --git a/CHANGELOG.md b/CHANGELOG.md index 49da6be25..d06775028 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -3,7 +3,7 @@ The format is based on [Keep a Changelog](https://keepachangelog.com/en/1.0.0/) and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0.html). -## v3.12.0 - [date] +## v3.13.0dev - [date] Initial release of nf-core/rnaseq, created with the [nf-core](https://nf-co.re/) template. diff --git a/CITATIONS.md b/CITATIONS.md index f7874b710..cae7ea444 100644 --- a/CITATIONS.md +++ b/CITATIONS.md @@ -12,7 +12,7 @@ - [FastQC](https://www.bioinformatics.babraham.ac.uk/projects/fastqc/) - > Andrews, S. (2010). FastQC: A Quality Control Tool for High Throughput Sequence Data [Online]. Available online https://www.bioinformatics.babraham.ac.uk/projects/fastqc/. + > Andrews, S. (2010). FastQC: A Quality Control Tool for High Throughput Sequence Data [Online]. - [MultiQC](https://pubmed.ncbi.nlm.nih.gov/27312411/) diff --git a/CODE_OF_CONDUCT.md b/CODE_OF_CONDUCT.md index f4fd052f1..c089ec78c 100644 --- a/CODE_OF_CONDUCT.md +++ b/CODE_OF_CONDUCT.md @@ -1,18 +1,20 @@ -# Code of Conduct at nf-core (v1.0) +# Code of Conduct at nf-core (v1.4) ## Our Pledge -In the interest of fostering an open, collaborative, and welcoming environment, we as contributors and maintainers of nf-core, pledge to making participation in our projects and community a harassment-free experience for everyone, regardless of: +In the interest of fostering an open, collaborative, and welcoming environment, we as contributors and maintainers of nf-core pledge to making participation in our projects and community a harassment-free experience for everyone, regardless of: - Age +- Ability - Body size +- Caste - Familial status - Gender identity and expression - Geographical location - Level of experience - Nationality and national origins - Native language -- Physical and neurological ability +- Neurodiversity - Race or ethnicity - Religion - Sexual identity and orientation @@ -22,80 +24,133 @@ Please note that the list above is alphabetised and is therefore not ranked in a ## Preamble -> Note: This Code of Conduct (CoC) has been drafted by the nf-core Safety Officer and been edited after input from members of the nf-core team and others. "We", in this document, refers to the Safety Officer and members of the nf-core core team, both of whom are deemed to be members of the nf-core community and are therefore required to abide by this Code of Conduct. This document will amended periodically to keep it up-to-date, and in case of any dispute, the most current version will apply. +:::note +This Code of Conduct (CoC) has been drafted by Renuka Kudva, Cris Tuñí, and Michael Heuer, with input from the nf-core Core Team and Susanna Marquez from the nf-core community. "We", in this document, refers to the Safety Officers and members of the nf-core Core Team, both of whom are deemed to be members of the nf-core community and are therefore required to abide by this Code of Conduct. This document will be amended periodically to keep it up-to-date. In case of any dispute, the most current version will apply. +::: -An up-to-date list of members of the nf-core core team can be found [here](https://nf-co.re/about). Our current safety officer is Renuka Kudva. +An up-to-date list of members of the nf-core core team can be found [here](https://nf-co.re/about). + +Our Safety Officers are Saba Nafees, Cris Tuñí, and Michael Heuer. nf-core is a young and growing community that welcomes contributions from anyone with a shared vision for [Open Science Policies](https://www.fosteropenscience.eu/taxonomy/term/8). Open science policies encompass inclusive behaviours and we strive to build and maintain a safe and inclusive environment for all individuals. -We have therefore adopted this code of conduct (CoC), which we require all members of our community and attendees in nf-core events to adhere to in all our workspaces at all times. Workspaces include but are not limited to Slack, meetings on Zoom, Jitsi, YouTube live etc. +We have therefore adopted this CoC, which we require all members of our community and attendees of nf-core events to adhere to in all our workspaces at all times. Workspaces include, but are not limited to, Slack, meetings on Zoom, gather.town, YouTube live etc. -Our CoC will be strictly enforced and the nf-core team reserve the right to exclude participants who do not comply with our guidelines from our workspaces and future nf-core activities. +Our CoC will be strictly enforced and the nf-core team reserves the right to exclude participants who do not comply with our guidelines from our workspaces and future nf-core activities. -We ask all members of our community to help maintain a supportive and productive workspace and to avoid behaviours that can make individuals feel unsafe or unwelcome. Please help us maintain and uphold this CoC. +We ask all members of our community to help maintain supportive and productive workspaces and to avoid behaviours that can make individuals feel unsafe or unwelcome. Please help us maintain and uphold this CoC. -Questions, concerns or ideas on what we can include? Contact safety [at] nf-co [dot] re +Questions, concerns, or ideas on what we can include? Contact members of the Safety Team on Slack or email safety [at] nf-co [dot] re. ## Our Responsibilities -The safety officer is responsible for clarifying the standards of acceptable behavior and are expected to take appropriate and fair corrective action in response to any instances of unacceptable behaviour. +Members of the Safety Team (the Safety Officers) are responsible for clarifying the standards of acceptable behavior and are expected to take appropriate and fair corrective action in response to any instances of unacceptable behaviour. -The safety officer in consultation with the nf-core core team have the right and responsibility to remove, edit, or reject comments, commits, code, wiki edits, issues, and other contributions that are not aligned to this Code of Conduct, or to ban temporarily or permanently any contributor for other behaviors that they deem inappropriate, threatening, offensive, or harmful. +The Safety Team, in consultation with the nf-core core team, have the right and responsibility to remove, edit, or reject comments, commits, code, wiki edits, issues, and other contributions that are not aligned to this CoC, or to ban temporarily or permanently any contributor for other behaviors that they deem inappropriate, threatening, offensive, or harmful. -Members of the core team or the safety officer who violate the CoC will be required to recuse themselves pending investigation. They will not have access to any reports of the violations and be subject to the same actions as others in violation of the CoC. +Members of the core team or the Safety Team who violate the CoC will be required to recuse themselves pending investigation. They will not have access to any reports of the violations and will be subject to the same actions as others in violation of the CoC. -## When are where does this Code of Conduct apply? +## When and where does this Code of Conduct apply? -Participation in the nf-core community is contingent on following these guidelines in all our workspaces and events. This includes but is not limited to the following listed alphabetically and therefore in no order of preference: +Participation in the nf-core community is contingent on following these guidelines in all our workspaces and events, such as hackathons, workshops, bytesize, and collaborative workspaces on gather.town. These guidelines include, but are not limited to, the following (listed alphabetically and therefore in no order of preference): - Communicating with an official project email address. - Communicating with community members within the nf-core Slack channel. - Participating in hackathons organised by nf-core (both online and in-person events). -- Participating in collaborative work on GitHub, Google Suite, community calls, mentorship meetings, email correspondence. -- Participating in workshops, training, and seminar series organised by nf-core (both online and in-person events). This applies to events hosted on web-based platforms such as Zoom, Jitsi, YouTube live etc. +- Participating in collaborative work on GitHub, Google Suite, community calls, mentorship meetings, email correspondence, and on the nf-core gather.town workspace. +- Participating in workshops, training, and seminar series organised by nf-core (both online and in-person events). This applies to events hosted on web-based platforms such as Zoom, gather.town, Jitsi, YouTube live etc. - Representing nf-core on social media. This includes both official and personal accounts. ## nf-core cares 😊 -nf-core's CoC and expectations of respectful behaviours for all participants (including organisers and the nf-core team) include but are not limited to the following (listed in alphabetical order): +nf-core's CoC and expectations of respectful behaviours for all participants (including organisers and the nf-core team) include, but are not limited to, the following (listed in alphabetical order): - Ask for consent before sharing another community member’s personal information (including photographs) on social media. - Be respectful of differing viewpoints and experiences. We are all here to learn from one another and a difference in opinion can present a good learning opportunity. -- Celebrate your accomplishments at events! (Get creative with your use of emojis 🎉 🥳 💯 🙌 !) +- Celebrate your accomplishments! (Get creative with your use of emojis 🎉 🥳 💯 🙌 !) - Demonstrate empathy towards other community members. (We don’t all have the same amount of time to dedicate to nf-core. If tasks are pending, don’t hesitate to gently remind members of your team. If you are leading a task, ask for help if you feel overwhelmed.) - Engage with and enquire after others. (This is especially important given the geographically remote nature of the nf-core community, so let’s do this the best we can) - Focus on what is best for the team and the community. (When in doubt, ask) -- Graciously accept constructive criticism, yet be unafraid to question, deliberate, and learn. +- Accept feedback, yet be unafraid to question, deliberate, and learn. - Introduce yourself to members of the community. (We’ve all been outsiders and we know that talking to strangers can be hard for some, but remember we’re interested in getting to know you and your visions for open science!) -- Show appreciation and **provide clear feedback**. (This is especially important because we don’t see each other in person and it can be harder to interpret subtleties. Also remember that not everyone understands a certain language to the same extent as you do, so **be clear in your communications to be kind.**) +- Show appreciation and **provide clear feedback**. (This is especially important because we don’t see each other in person and it can be harder to interpret subtleties. Also remember that not everyone understands a certain language to the same extent as you do, so **be clear in your communication to be kind.**) - Take breaks when you feel like you need them. -- Using welcoming and inclusive language. (Participants are encouraged to display their chosen pronouns on Zoom or in communication on Slack.) +- Use welcoming and inclusive language. (Participants are encouraged to display their chosen pronouns on Zoom or in communication on Slack) ## nf-core frowns on 😕 -The following behaviours from any participants within the nf-core community (including the organisers) will be considered unacceptable under this code of conduct. Engaging or advocating for any of the following could result in expulsion from nf-core workspaces. +The following behaviours from any participants within the nf-core community (including the organisers) will be considered unacceptable under this CoC. Engaging or advocating for any of the following could result in expulsion from nf-core workspaces: - Deliberate intimidation, stalking or following and sustained disruption of communication among participants of the community. This includes hijacking shared screens through actions such as using the annotate tool in conferencing software such as Zoom. - “Doxing” i.e. posting (or threatening to post) another person’s personal identifying information online. - Spamming or trolling of individuals on social media. -- Use of sexual or discriminatory imagery, comments, or jokes and unwelcome sexual attention. -- Verbal and text comments that reinforce social structures of domination related to gender, gender identity and expression, sexual orientation, ability, physical appearance, body size, race, age, religion or work experience. +- Use of sexual or discriminatory imagery, comments, jokes, or unwelcome sexual attention. +- Verbal and text comments that reinforce social structures of domination related to gender, gender identity and expression, sexual orientation, ability, physical appearance, body size, race, age, religion, or work experience. ### Online Trolling -The majority of nf-core interactions and events are held online. Unfortunately, holding events online comes with the added issue of online trolling. This is unacceptable, reports of such behaviour will be taken very seriously, and perpetrators will be excluded from activities immediately. +The majority of nf-core interactions and events are held online. Unfortunately, holding events online comes with the risk of online trolling. This is unacceptable — reports of such behaviour will be taken very seriously and perpetrators will be excluded from activities immediately. -All community members are required to ask members of the group they are working within for explicit consent prior to taking screenshots of individuals during video calls. +All community members are **required** to ask members of the group they are working with for explicit consent prior to taking screenshots of individuals during video calls. -## Procedures for Reporting CoC violations +## Procedures for reporting CoC violations If someone makes you feel uncomfortable through their behaviours or actions, report it as soon as possible. -You can reach out to members of the [nf-core core team](https://nf-co.re/about) and they will forward your concerns to the safety officer(s). +You can reach out to members of the Safety Team (Saba Nafees, Cris Tuñí, and Michael Heuer) on Slack. Alternatively, contact a member of the nf-core core team [nf-core core team](https://nf-co.re/about), and they will forward your concerns to the Safety Team. + +Issues directly concerning members of the Core Team or the Safety Team will be dealt with by other members of the core team and the safety manager — possible conflicts of interest will be taken into account. nf-core is also in discussions about having an ombudsperson and details will be shared in due course. + +All reports will be handled with the utmost discretion and confidentiality. + +You can also report any CoC violations to safety [at] nf-co [dot] re. In your email report, please do your best to include: + +- Your contact information. +- Identifying information (e.g. names, nicknames, pseudonyms) of the participant who has violated the Code of Conduct. +- The behaviour that was in violation and the circumstances surrounding the incident. +- The approximate time of the behaviour (if different than the time the report was made). +- Other people involved in the incident, if applicable. +- If you believe the incident is ongoing. +- If there is a publicly available record (e.g. mailing list record, a screenshot). +- Any additional information. + +After you file a report, one or more members of our Safety Team will contact you to follow up on your report. + +## Who will read and handle reports + +All reports will be read and handled by the members of the Safety Team at nf-core. + +If members of the Safety Team are deemed to have a conflict of interest with a report, they will be required to recuse themselves as per our Code of Conduct and will not have access to any follow-ups. + +To keep this first report confidential from any of the Safety Team members, please submit your first report by direct messaging on Slack/direct email to any of the nf-core members you are comfortable disclosing the information to, and be explicit about which member(s) you do not consent to sharing the information with. + +## Reviewing reports + +After receiving the report, members of the Safety Team will review the incident report to determine whether immediate action is required, for example, whether there is immediate threat to participants’ safety. + +The Safety Team, in consultation with members of the nf-core core team, will assess the information to determine whether the report constitutes a Code of Conduct violation, for them to decide on a course of action. + +In the case of insufficient information, one or more members of the Safety Team may contact the reporter, the reportee, or any other attendees to obtain more information. -Issues directly concerning members of the core team will be dealt with by other members of the core team and the safety manager, and possible conflicts of interest will be taken into account. nf-core is also in discussions about having an ombudsperson, and details will be shared in due course. +Once additional information is gathered, the Safety Team will collectively review and decide on the best course of action to take, if any. The Safety Team reserves the right to not act on a report. -All reports will be handled with utmost discretion and confidentially. +## Confidentiality + +All reports, and any additional information included, are only shared with the team of safety officers (and possibly members of the core team, in case the safety officer is in violation of the CoC). We will respect confidentiality requests for the purpose of protecting victims of abuse. + +We will not name harassment victims, beyond discussions between the safety officer and members of the nf-core team, without the explicit consent of the individuals involved. + +## Enforcement + +Actions taken by the nf-core’s Safety Team may include, but are not limited to: + +- Asking anyone to stop a behaviour. +- Asking anyone to leave the event and online spaces either temporarily, for the remainder of the event, or permanently. +- Removing access to the gather.town and Slack, either temporarily or permanently. +- Communicating to all participants to reinforce our expectations for conduct and remind what is unacceptable behaviour; this may be public for practical reasons. +- Communicating to all participants that an incident has taken place and how we will act or have acted — this may be for the purpose of letting event participants know we are aware of and dealing with the incident. +- Banning anyone from participating in nf-core-managed spaces, future events, and activities, either temporarily or permanently. +- No action. ## Attribution and Acknowledgements @@ -106,6 +161,22 @@ All reports will be handled with utmost discretion and confidentially. ## Changelog -### v1.0 - March 12th, 2021 +### v1.4 - February 8th, 2022 + +- Included a new member of the Safety Team. Corrected a typographical error in the text. + +### v1.3 - December 10th, 2021 + +- Added a statement that the CoC applies to nf-core gather.town workspaces. Corrected typographical errors in the text. + +### v1.2 - November 12th, 2021 + +- Removed information specific to reporting CoC violations at the Hackathon in October 2021. + +### v1.1 - October 14th, 2021 + +- Updated with names of new Safety Officers and specific information for the hackathon in October 2021. + +### v1.0 - March 15th, 2021 - Complete rewrite from original [Contributor Covenant](http://contributor-covenant.org/) CoC. diff --git a/README.md b/README.md index 84b18b356..78e612f97 100644 --- a/README.md +++ b/README.md @@ -1,6 +1,7 @@ # ![nf-core/rnaseq](docs/images/nf-core-rnaseq_logo_light.png#gh-light-mode-only) ![nf-core/rnaseq](docs/images/nf-core-rnaseq_logo_dark.png#gh-dark-mode-only) -[![AWS CI](https://img.shields.io/badge/CI%20tests-full%20size-FF9900?labelColor=000000&logo=Amazon%20AWS)](https://nf-co.re/rnaseq/results)[![Cite with Zenodo](http://img.shields.io/badge/DOI-10.5281/zenodo.XXXXXXX-1073c8?labelColor=000000)](https://doi.org/10.5281/zenodo.XXXXXXX) +[![GitHub Actions CI Status](https://github.com/nf-core/rnaseq/workflows/nf-core%20CI/badge.svg)](https://github.com/nf-core/rnaseq/actions?query=workflow%3A%22nf-core+CI%22) +[![GitHub Actions Linting Status](https://github.com/nf-core/rnaseq/workflows/nf-core%20linting/badge.svg)](https://github.com/nf-core/rnaseq/actions?query=workflow%3A%22nf-core+linting%22)[![AWS CI](https://img.shields.io/badge/CI%20tests-full%20size-FF9900?labelColor=000000&logo=Amazon%20AWS)](https://nf-co.re/rnaseq/results)[![Cite with Zenodo](http://img.shields.io/badge/DOI-10.5281/zenodo.XXXXXXX-1073c8?labelColor=000000)](https://doi.org/10.5281/zenodo.XXXXXXX) [![Nextflow](https://img.shields.io/badge/nextflow%20DSL2-%E2%89%A523.04.0-23aa62.svg)](https://www.nextflow.io/) [![run with conda](http://img.shields.io/badge/run%20with-conda-3EB049?labelColor=000000&logo=anaconda)](https://docs.conda.io/en/latest/) @@ -29,10 +30,11 @@ ## Usage -> **Note** -> If you are new to Nextflow and nf-core, please refer to [this page](https://nf-co.re/docs/usage/installation) on how -> to set-up Nextflow. Make sure to [test your setup](https://nf-co.re/docs/usage/introduction#how-to-run-a-pipeline) -> with `-profile test` before running the workflow on actual data. +:::note +If you are new to Nextflow and nf-core, please refer to [this page](https://nf-co.re/docs/usage/installation) on how +to set-up Nextflow. Make sure to [test your setup](https://nf-co.re/docs/usage/introduction#how-to-run-a-pipeline) +with `-profile test` before running the workflow on actual data. +::: Y?tY(@ZF}Y%IZw#L{1`h+}TN_GY z%!#6AJiqfGEK~E}hWpd4_Mw!WJW+{P~WF0R&hr}bBLo3AI| z81lV=mJW5%D^Q1Pz-J-E8c?x><{OV}V^`G~in|mh-Sm&D3K`e%T(BEI zk4-|q8dc9)x0eg1m4^y*y0~ zXzTGyO>e^=h9@8C?h{xzjT@kk9txIG!~^g|F>-$$8|s~SI0U{;)d;}*s08CWcorI3 zgoH)I>4^ecY2vT;+K=B2)LsG0TX<}0dX#iy^e%G2$00@1j|ChkyPw~$H5%I`yksYP zbPf4nO^MZcO5wrY1p(PVynOK(TO5<>_IOM$* z2TtohUqx~D89lk-9aI6`zoK^D$seTYVXZkZamk1!tP^b$f$cE_u7VcG3}%tOx;f}w z$kuWcVgn1BWlgf`rcTTzt?M)6S6g-b3R;XD4MmR=3Khi3Vn7FVlolPSPhceF%`8l; zw2xdF76Zk}x!0Ap&r;klOC6RW57VxxSUV2gUh5h$wZ9DuMQ@>Mg1=L9eiTiU@sW+f zUpC=`3%(s$Vg}N} zC7fQBRCPaCv6>_YFH7!a{5Q;P_PZGO*r&Fi{bC_6i70Sp5kUvi@MYO6VYo3%sKLde`ncE$Vr5t`F1b^J&oRj4196K04HyOMJNztGGxSm_>eC|EZpH} zvKQ;3@cc(L4}C$0wUSVTKza)K7^z23I5DRdXWq`Wp66M?iyPA`C$Q*@cW#9vc)LZ& zCjIlIN`)Rq^yinYNTd?s^C-*?U}F@5)*A@KX*#8rG>M}k3~;c5L}O2n2!|mc=6}BY 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Pre-processing4. 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inkscape:snap-others="true" inkscape:snap-nodes="true" @@ -1389,47 +1389,42 @@ y="72.633636" style="font-style:normal;font-variant:normal;font-weight:bold;font-stretch:normal;font-size:19.7556px;font-family:'Maven Pro';-inkscape-font-specification:'Maven Pro Bold';fill:#ffffff;stroke:none;stroke-width:0.264583;stroke-opacity:1">3SalmonKallistoMultiQCMultiQCTSVBIGWIGHTML + sodipodi:nodetypes="cc" + id="path2500-5-22-5-4-7-4-0-3-0-2-4-4-4" + d="M 383.56073,50.279353 V 40.018944" + style="fill:none;stroke:#800080;stroke-width:2.64583;stroke-linecap:butt;stroke-linejoin:miter;stroke-miterlimit:4;stroke-dasharray:none;stroke-opacity:1" + inkscape:connector-curvature="0" /> From 6479792bce5e4fc0c2a8e5e743f75111068084f4 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Fri, 3 Nov 2023 18:18:42 +0000 Subject: [PATCH 092/184] Update strandedness params for kallisto and salmon --- conf/modules.config | 4 ++++ nextflow.config | 1 + nextflow_schema.json | 10 +++++++++- 3 files changed, 14 insertions(+), 1 deletion(-) diff --git a/conf/modules.config b/conf/modules.config index 34dd55587..1e64bdb75 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -1151,6 +1151,10 @@ if (params.pseudo_aligner == 'kallisto') { "--single " + "--fragment-length ${meta.fragment_len ?: params.kallisto_default_fraglen} " + "--sd ${meta.fraglen_se ?: params.kallisto_default_fraglen_se}" : '', + params.kallisto_quant_strandedness ? '--' + params.kallisto_quant_strandedness: + (meta.strandedess == 'forward' ? '--fr-stranded': + (meta.strandedness == 'reverse' ? '--rf-stranded' : '') + ), params.extra_kallisto_quant_args ?: '' ].findAll().join(' ').trim() } diff --git a/nextflow.config b/nextflow.config index f027fee6d..3e72d5d9a 100644 --- a/nextflow.config +++ b/nextflow.config @@ -69,6 +69,7 @@ params { extra_kallisto_quant_args = null kallisto_default_fraglen = 150 kallisto_default_fraglen_se = 75 + kallisto_quant_strandedness = null save_merged_fastq = false save_unaligned = false save_align_intermeds = false diff --git a/nextflow_schema.json b/nextflow_schema.json index 9c5f9a574..662b0737a 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -375,7 +375,8 @@ "type": "string", "fa_icon": "fas fa-fast-forward", "description": " Override Salmon library type inferred based on strandedness defined in meta object.", - "help_text": "See [Salmon docs](https://salmon.readthedocs.io/en/latest/library_type.html)." + "help_text": "See [Salmon docs](https://salmon.readthedocs.io/en/latest/library_type.html).", + "enum": ["A,U,SF,SR,IS,IU,ISF,ISR,OS,OU,OSF,OSR,MS,MU,MSF,MSR"] }, "min_mapped_reads": { "type": "number", @@ -404,6 +405,13 @@ "description": "Extra arguments to pass to Salmon quant command in addition to defaults defined by the pipeline.", "fa_icon": "fas fa-plus" }, + "kallisto_quant_strandedness": { + "type": "string", + "fa_icon": "fas fa-ribbon", + "description": "Override Kallisto library type inferred based on strandedness defined in meta object.", + "enum": ["fr-stranded", "rf-stranded"], + "help_text": "See [Kallisto docs](https://pachterlab.github.io/kallisto/manual)" + }, "extra_kallisto_quant_args": { "type": "string", "description": "Extra arguments to pass to Kallisto quant command in addition to defaults defined by the pipeline.", From 92716b4f4039acf92f8811e0d37861532494d784 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Fri, 3 Nov 2023 18:23:07 +0000 Subject: [PATCH 093/184] [skip ci] appease linter --- conf/modules.config | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/conf/modules.config b/conf/modules.config index 1e64bdb75..0c4dc7dec 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -1151,8 +1151,8 @@ if (params.pseudo_aligner == 'kallisto') { "--single " + "--fragment-length ${meta.fragment_len ?: params.kallisto_default_fraglen} " + "--sd ${meta.fraglen_se ?: params.kallisto_default_fraglen_se}" : '', - params.kallisto_quant_strandedness ? '--' + params.kallisto_quant_strandedness: - (meta.strandedess == 'forward' ? '--fr-stranded': + params.kallisto_quant_strandedness ? '--' + params.kallisto_quant_strandedness: + (meta.strandedess == 'forward' ? '--fr-stranded': (meta.strandedness == 'reverse' ? '--rf-stranded' : '') ), params.extra_kallisto_quant_args ?: '' From cadec095a1e44d52ef526c2bffa48bf793590079 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Fri, 3 Nov 2023 18:50:28 +0000 Subject: [PATCH 094/184] Fix kallisto publishing --- conf/modules.config | 2 +- modules/local/tximport/main.nf | 2 +- 2 files changed, 2 insertions(+), 2 deletions(-) diff --git a/conf/modules.config b/conf/modules.config index 0c4dc7dec..8e13fbe1d 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -1162,7 +1162,7 @@ if (params.pseudo_aligner == 'kallisto') { publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') || filename.endsWith('_meta_info.json') ? null : filename } + saveAs: { filename -> filename.equals('versions.yml') || filename.endsWith('_run_info.json') ? null : filename } ] } } diff --git a/modules/local/tximport/main.nf b/modules/local/tximport/main.nf index 6ee92e4d1..d46dde44e 100644 --- a/modules/local/tximport/main.nf +++ b/modules/local/tximport/main.nf @@ -28,7 +28,7 @@ process TXIMPORT { tximport.r \\ NULL \\ quants \\ - merged \\ + ${quant_type}.merged \\ $quant_type \\ $tx2gene From 908c6f2ee163d5bc61498e31813259e99a6fa344 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Fri, 3 Nov 2023 18:50:59 +0000 Subject: [PATCH 095/184] [skip ci] fix usage.md for Kallisto --- docs/usage.md | 8 +++++--- 1 file changed, 5 insertions(+), 3 deletions(-) diff --git a/docs/usage.md b/docs/usage.md index 9748aa5de..be2de8c65 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -65,17 +65,19 @@ An [example samplesheet](../assets/samplesheet.csv) has been provided with the p By default, the pipeline uses [STAR](https://github.com/alexdobin/STAR) (i.e. `--aligner star_salmon`) to map the raw FastQ reads to the reference genome, project the alignments onto the transcriptome and to perform the downstream BAM-level quantification with [Salmon](https://salmon.readthedocs.io/en/latest/salmon.html). STAR is fast but requires a lot of memory to run, typically around 38GB for the Human GRCh37 reference genome. Since the [RSEM](https://github.com/deweylab/RSEM) (i.e. `--aligner star_rsem`) workflow in the pipeline also uses STAR you should use the [HISAT2](https://ccb.jhu.edu/software/hisat2/index.shtml) aligner (i.e. `--aligner hisat2`) if you have memory limitations. -You also have the option to pseudo-align and quantify your data with [Salmon](https://salmon.readthedocs.io/en/latest/salmon.html) by providing the `--pseudo_aligner salmon` parameter. Salmon will then be run in addition to the standard alignment workflow defined by `--aligner`, mainly because it allows you to obtain QC metrics with respect to the genomic alignments. However, you can provide the `--skip_alignment` parameter if you would like to run Salmon in isolation. By default, the pipeline will use the genome fasta and gtf file to generate the transcripts fasta file, and then to build the Salmon index. You can override these parameters using the `--transcript_fasta` and `--salmon_index` parameters, respectively. The library preparation protocol (library type) used by Salmon quantification is inferred by the pipeline based on the information provided in the samplesheet, however, you can override it using the `--salmon_quant_libtype` parameter. You can find the available options in the [Salmon documentation](https://salmon.readthedocs.io/en/latest/library_type.html). +You also have the option to pseudo-align and quantify your data directly with [Salmon](https://salmon.readthedocs.io/en/latest/salmon.html) or [Kallisto](https://pachterlab.github.io/kallisto/) by specifying `salmon` or `kallisto` to the `--pseudo_aligner` parameter. The selected pseudoaligner will then be run in addition to the standard alignment workflow defined by `--aligner`, mainly because it allows you to obtain QC metrics with respect to the genomic alignments. However, you can provide the `--skip_alignment` parameter if you would like to run Salmon or Kallisto in isolation. By default, the pipeline will use the genome fasta and gtf file to generate the transcripts fasta file, and then to build the Salmon index. You can override these parameters using the `--transcript_fasta` and `--salmon_index` parameters, respectively. + +The library preparation protocol (library type) used by Salmon quantification is inferred by the pipeline based on the information provided in the samplesheet, however, you can override it using the `--salmon_quant_libtype` parameter. You can find the available options in the [Salmon documentation](https://salmon.readthedocs.io/en/latest/library_type.html). Similarly, strandedness is taken from the sample sheet or calculated automatically, and passed to Kallisto on a per-library basis, but you can apply a global override by setting `kallisto_quant_strandednes` see the [Kallisto documentation](https://pachterlab.github.io/kallisto/manual). When running Salmon in mapping-based mode via `--pseudo_aligner salmon` the entire genome of the organism is used by default for the decoy-aware transcriptome when creating the indices (see second bulleted option in [Salmon documentation](https://salmon.readthedocs.io/en/latest/salmon.html#preparing-transcriptome-indices-mapping-based-mode)). -Two additional parameters `--extra_star_align_args` and `--extra_salmon_quant_args` were added in v3.10 of the pipeline that allow you to append any custom parameters to the STAR align and Salmon quant commands, respectively. Note, the `--seqBias` and `--gcBias` are not provided to Salmon quant by default so you can provide these via `--extra_salmon_quant_args '--seqBias --gcBias'` if required. +Two additional parameters `--extra_star_align_args` and `--extra_salmon_quant_args` were added in v3.10 of the pipeline that allow you to append any custom parameters to the STAR align and Salmon quant commands, respectively. Note, the `--seqBias` and `--gcBias` are not provided to Salmon quant by default so you can provide these via `--extra_salmon_quant_args '--seqBias --gcBias'` if required. You can now also supply additional arguments to Kallisto via `extra_kallisto_quant_args`. > **NB:** You can use `--skip_alignment --skip_pseudo_alignment` if you only want to run the pre-processing QC steps in the pipeline like FastQ, trimming etc. This will skip alignment, pseudo-alignment and any post-alignment processing steps. ## Quantification options -The current options align with STAR and quantify using either Salmon (`--aligner star_salmon`) / RSEM (`--aligner star_rsem`). You also have the option to pseudo-align and quantify your data with Salmon by providing the `--pseudo_aligner salmon` parameter. +The current options align with STAR and quantify using either Salmon (`--aligner star_salmon`) / RSEM (`--aligner star_rsem`). You also have the option to pseudo-align and quantify your data with Salmon or Kallisto by providing the `--pseudo_aligner salmon` parameter. Since v3.0 of the pipeline, featureCounts is no longer used to perform gene/transcript quantification, however it is still used to generate QC metrics based on [biotype](http://www.ensembl.org/info/genome/genebuild/biotypes.html) information available within GFF/GTF genome annotation files. This decision was made primarily because of the limitations of featureCounts to appropriately quantify gene expression data. Please see [Zhao et al., 2015](https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0141910#pone-0141910-t001) and [Soneson et al., 2015](https://f1000research.com/articles/4-1521/v1). From 0fc54076d5ee02b72f98f4d9b3af312394168930 Mon Sep 17 00:00:00 2001 From: nf-core-bot Date: Fri, 3 Nov 2023 22:57:59 +0000 Subject: [PATCH 096/184] [automated] Fix linting with Prettier --- docs/usage.md | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/docs/usage.md b/docs/usage.md index be2de8c65..bd776e66e 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -65,9 +65,9 @@ An [example samplesheet](../assets/samplesheet.csv) has been provided with the p By default, the pipeline uses [STAR](https://github.com/alexdobin/STAR) (i.e. `--aligner star_salmon`) to map the raw FastQ reads to the reference genome, project the alignments onto the transcriptome and to perform the downstream BAM-level quantification with [Salmon](https://salmon.readthedocs.io/en/latest/salmon.html). STAR is fast but requires a lot of memory to run, typically around 38GB for the Human GRCh37 reference genome. Since the [RSEM](https://github.com/deweylab/RSEM) (i.e. `--aligner star_rsem`) workflow in the pipeline also uses STAR you should use the [HISAT2](https://ccb.jhu.edu/software/hisat2/index.shtml) aligner (i.e. `--aligner hisat2`) if you have memory limitations. -You also have the option to pseudo-align and quantify your data directly with [Salmon](https://salmon.readthedocs.io/en/latest/salmon.html) or [Kallisto](https://pachterlab.github.io/kallisto/) by specifying `salmon` or `kallisto` to the `--pseudo_aligner` parameter. The selected pseudoaligner will then be run in addition to the standard alignment workflow defined by `--aligner`, mainly because it allows you to obtain QC metrics with respect to the genomic alignments. However, you can provide the `--skip_alignment` parameter if you would like to run Salmon or Kallisto in isolation. By default, the pipeline will use the genome fasta and gtf file to generate the transcripts fasta file, and then to build the Salmon index. You can override these parameters using the `--transcript_fasta` and `--salmon_index` parameters, respectively. +You also have the option to pseudo-align and quantify your data directly with [Salmon](https://salmon.readthedocs.io/en/latest/salmon.html) or [Kallisto](https://pachterlab.github.io/kallisto/) by specifying `salmon` or `kallisto` to the `--pseudo_aligner` parameter. The selected pseudoaligner will then be run in addition to the standard alignment workflow defined by `--aligner`, mainly because it allows you to obtain QC metrics with respect to the genomic alignments. However, you can provide the `--skip_alignment` parameter if you would like to run Salmon or Kallisto in isolation. By default, the pipeline will use the genome fasta and gtf file to generate the transcripts fasta file, and then to build the Salmon index. You can override these parameters using the `--transcript_fasta` and `--salmon_index` parameters, respectively. -The library preparation protocol (library type) used by Salmon quantification is inferred by the pipeline based on the information provided in the samplesheet, however, you can override it using the `--salmon_quant_libtype` parameter. You can find the available options in the [Salmon documentation](https://salmon.readthedocs.io/en/latest/library_type.html). Similarly, strandedness is taken from the sample sheet or calculated automatically, and passed to Kallisto on a per-library basis, but you can apply a global override by setting `kallisto_quant_strandednes` see the [Kallisto documentation](https://pachterlab.github.io/kallisto/manual). +The library preparation protocol (library type) used by Salmon quantification is inferred by the pipeline based on the information provided in the samplesheet, however, you can override it using the `--salmon_quant_libtype` parameter. You can find the available options in the [Salmon documentation](https://salmon.readthedocs.io/en/latest/library_type.html). Similarly, strandedness is taken from the sample sheet or calculated automatically, and passed to Kallisto on a per-library basis, but you can apply a global override by setting `kallisto_quant_strandednes` see the [Kallisto documentation](https://pachterlab.github.io/kallisto/manual). When running Salmon in mapping-based mode via `--pseudo_aligner salmon` the entire genome of the organism is used by default for the decoy-aware transcriptome when creating the indices (see second bulleted option in [Salmon documentation](https://salmon.readthedocs.io/en/latest/salmon.html#preparing-transcriptome-indices-mapping-based-mode)). From e9ed01dafd57c3c0162e786ffed9b06c3c58570d Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Fri, 3 Nov 2023 22:59:32 +0000 Subject: [PATCH 097/184] Update fraglen defaults --- nextflow.config | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/nextflow.config b/nextflow.config index 3e72d5d9a..09e49b36a 100644 --- a/nextflow.config +++ b/nextflow.config @@ -67,8 +67,8 @@ params { extra_star_align_args = null extra_salmon_quant_args = null extra_kallisto_quant_args = null - kallisto_default_fraglen = 150 - kallisto_default_fraglen_se = 75 + kallisto_default_fraglen = 200 + kallisto_default_fraglen_se = 200 kallisto_quant_strandedness = null save_merged_fastq = false save_unaligned = false From 1467c81cbc242c4acc37dfecc82a0d80946427da Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Mon, 6 Nov 2023 11:09:53 +0000 Subject: [PATCH 098/184] MultiQC needs STDOUT from Kallisto --- subworkflows/local/quantify_pseudo/main.nf | 4 ++++ workflows/rnaseq.nf | 2 +- 2 files changed, 5 insertions(+), 1 deletion(-) diff --git a/subworkflows/local/quantify_pseudo/main.nf b/subworkflows/local/quantify_pseudo/main.nf index 473002bae..cb3fb7399 100644 --- a/subworkflows/local/quantify_pseudo/main.nf +++ b/subworkflows/local/quantify_pseudo/main.nf @@ -30,13 +30,16 @@ workflow QUANTIFY_PSEUDO { // Quantify and merge counts across samples // + // Note: MultiQC needs Salmon outputs, but Kallisto logs if (pseudo_aligner == 'salmon') { SALMON_QUANT ( reads, index, gtf, transcript_fasta, alignment_mode, lib_type ) ch_pseudo_results = SALMON_QUANT.out.results + ch_pseudo_multiqc = ch_pseudo_results ch_versions = ch_versions.mix(SALMON_QUANT.out.versions.first()) }else { KALLISTO_QUANT ( reads, index, gtf, []) ch_pseudo_results = KALLISTO_QUANT.out.results + ch_pseudo_multiqc = KALLISTO_QUANT.out.log ch_versions = ch_versions.mix(KALLISTO_QUANT.out.versions.first()) } @@ -73,6 +76,7 @@ workflow QUANTIFY_PSEUDO { emit: results = ch_pseudo_results // channel: [ val(meta), results_dir ] + multiqc = ch_pseudo_multiqc // channel: [ val(meta), files_for_multiqc ] tpm_gene = TXIMPORT.out.tpm_gene // channel: [ val(meta), counts ] counts_gene = TXIMPORT.out.counts_gene // channel: [ val(meta), counts ] diff --git a/workflows/rnaseq.nf b/workflows/rnaseq.nf index dc26b96a1..6e9d81c9d 100755 --- a/workflows/rnaseq.nf +++ b/workflows/rnaseq.nf @@ -813,7 +813,7 @@ workflow RNASEQ { false, params.salmon_quant_libtype ?: '' ) - ch_pseudo_multiqc = QUANTIFY_PSEUDO.out.results + ch_pseudo_multiqc = QUANTIFY_PSEUDO.out.multiqc ch_versions = ch_versions.mix(QUANTIFY_PSEUDO.out.versions) ch_counts_gene_length_scaled = QUANTIFY_PSEUDO.out.counts_gene_length_scaled From 7d048863a3a4e1e0c73b843bda81d0549533ea7f Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Mon, 6 Nov 2023 12:28:41 +0000 Subject: [PATCH 099/184] Fix Kallisto MultiQC --- assets/multiqc_config.yml | 1 + conf/modules.config | 2 +- modules/local/multiqc/main.nf | 2 +- 3 files changed, 3 insertions(+), 2 deletions(-) diff --git a/assets/multiqc_config.yml b/assets/multiqc_config.yml index 4af748689..cbc8db099 100644 --- a/assets/multiqc_config.yml +++ b/assets/multiqc_config.yml @@ -23,6 +23,7 @@ run_modules: - hisat2 - rsem - salmon + - kallisto - samtools - picard - preseq diff --git a/conf/modules.config b/conf/modules.config index 8e13fbe1d..c0924e879 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -1205,7 +1205,7 @@ if (!params.skip_pseudo_alignment) { "--count_col 3", params.deseq2_vst ? '--vst TRUE' : '' ].join(' ').trim() - ext.args2 = 'salmon' + ext.args2 = params.pseudo_aligner publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}/deseq2_qc" }, mode: params.publish_dir_mode, diff --git a/modules/local/multiqc/main.nf b/modules/local/multiqc/main.nf index 06fbb9b29..e5fcf88f2 100644 --- a/modules/local/multiqc/main.nf +++ b/modules/local/multiqc/main.nf @@ -23,7 +23,7 @@ process MULTIQC { path ('star/*') path ('hisat2/*') path ('rsem/*') - path ('salmon/*') + path ('pseudoalignment/*') path ('samtools/stats/*') path ('samtools/flagstat/*') path ('samtools/idxstats/*') From 8420e997f3ac3ef833b890c7d912d92be9507824 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Mon, 6 Nov 2023 12:34:40 +0000 Subject: [PATCH 100/184] Update docs --- README.md | 2 +- docs/output.md | 58 ++++++++++++++++++++++++++++++++++---------------- 2 files changed, 41 insertions(+), 19 deletions(-) diff --git a/README.md b/README.md index 9fe5c0ab8..b56768f3a 100644 --- a/README.md +++ b/README.md @@ -39,7 +39,7 @@ 3. [`dupRadar`](https://bioconductor.org/packages/release/bioc/html/dupRadar.html) 4. [`Preseq`](http://smithlabresearch.org/software/preseq/) 5. [`DESeq2`](https://bioconductor.org/packages/release/bioc/html/DESeq2.html) -15. Pseudo-alignment and quantification ([`Salmon`](https://combine-lab.github.io/salmon/); _optional_) +15. Pseudo-alignment and quantification ([`Salmon`](https://combine-lab.github.io/salmon/) or ['Kallisto'](https://pachterlab.github.io/kallisto/); _optional_) 16. Present QC for raw read, alignment, gene biotype, sample similarity, and strand-specificity checks ([`MultiQC`](http://multiqc.info/), [`R`](https://www.r-project.org/)) > **Note** diff --git a/docs/output.md b/docs/output.md index af9721f00..000b0c445 100644 --- a/docs/output.md +++ b/docs/output.md @@ -43,6 +43,8 @@ The pipeline is built using [Nextflow](https://www.nextflow.io/) and processes d - [MultiQC](#multiqc) - Present QC for raw reads, alignment, read counting and sample similiarity - [Pseudo-alignment and quantification](#pseudo-alignment-and-quantification) - [Salmon](#salmon) - Wicked fast gene and isoform quantification relative to the transcriptome + - [Kallisto](#kallisto) - Near-optimal probabilistic RNA-seq quantification +Wicked fast gene and isoform quantification relative to the transcriptome - [Workflow reporting and genomes](#workflow-reporting-and-genomes) - [Reference genome files](#reference-genome-files) - Saving reference genome indices/files - [Pipeline information](#pipeline-information) - Report metrics generated during the workflow execution @@ -180,7 +182,7 @@ When `--remove_ribo_rna` is specified, the pipeline uses [SortMeRNA](https://git ## Alignment and quantification -### STAR and Salmon +### STAR, Salmon and Kallisto

    Output files @@ -199,12 +201,12 @@ When `--remove_ribo_rna` is specified, the pipeline uses [SortMeRNA](https://git [STAR](https://github.com/alexdobin/STAR) is a read aligner designed for splice aware mapping typical of RNA sequencing data. STAR stands for *S*pliced *T*ranscripts *A*lignment to a *R*eference, and has been shown to have high accuracy and outperforms other aligners by more than a factor of 50 in mapping speed, but it is memory intensive. Using `--aligner star_salmon` is the default alignment and quantification option. -[Salmon](https://salmon.readthedocs.io/en/latest/salmon.html) from [Ocean Genomics](https://oceangenomics.com/) is a tool for wicked-fast transcript quantification from RNA-seq data. It requires a set of target transcripts (either from a reference or de-novo assembly) in order to perform quantification. All you need to run Salmon is a FASTA file containing your reference transcripts and a set of FASTA/FASTQ/BAM file(s) containing your reads. The transcriptome-level BAM files generated by STAR are provided to Salmon for downstream quantification. You can of course also provide FASTQ files directly as input to Salmon in order to pseudo-align and quantify your data by providing the `--pseudo_aligner salmon` parameter. The results generated by the pipeline are exactly the same whether you provide BAM or FASTQ input so please see the [Salmon](#salmon) results section for more details. - The STAR section of the MultiQC report shows a bar plot with alignment rates: good samples should have most reads as _Uniquely mapped_ and few _Unmapped_ reads. ![MultiQC - STAR alignment scores plot](images/mqc_star.png) +[Salmon](https://salmon.readthedocs.io/en/latest/salmon.html) from [Ocean Genomics](https://oceangenomics.com/) and [Kallisto](https://pachterlab.github.io/kallisto/), from the Pachter Lab, are provided as options for pseudo-alignment. Both allow quantification of reads against an index generated from a reference set of target transcripts. By default, the transcriptome-level BAM files generated by STAR are provided to Salmon for downstream quantification, and Kallisto is not an option here (it does not allow BAM file input). But you can provide FASTQ files directly as input to either Salmon or Kallisto in order to pseudo-align and quantify your data by providing the `--pseudo_aligner (salmon or kallisto)` parameter. See the [Salmon](#salmon) and (Kallisto)[#kallisto] results sections for more details. + ### STAR via RSEM
    @@ -670,23 +672,32 @@ Results generated by MultiQC collate pipeline QC from supported tools i.e. FastQ ## Pseudo-alignment and quantification -### Salmon +### Pseudo-alignment + +The principal output files are the same between Salmon and Kallsto: + +
    +Output files + +- `/` + - `.merged.gene_counts.tsv`: Matrix of gene-level raw counts across all samples. + - `.gene_tpm.tsv`: Matrix of gene-level TPM values across all samples. + - `.gene_counts.rds`: RDS object that can be loaded in R that contains a [SummarizedExperiment](https://bioconductor.org/packages/release/bioc/html/SummarizedExperiment.html) container with the TPM (`abundance`), estimated counts (`counts`) and transcript length (`length`) in the assays slot for genes. + - `.merged.gene_counts_scaled.tsv`: Matrix of gene-level library size-scaled counts across all samples. + - `.merged.gene_counts_scaled.rds`: RDS object that can be loaded in R that contains a [SummarizedExperiment](https://bioconductor.org/packages/release/bioc/html/SummarizedExperiment.html) container with the TPM (`abundance`), estimated library size-scaled counts (`counts`) and transcript length (`length`) in the assays slot for genes. + - `.merged.gene_counts_length_scaled.tsv`: Matrix of gene-level length-scaled counts across all samples. + - `.merged.gene_counts_length_scaled.rds`: RDS object that can be loaded in R that contains a [SummarizedExperiment](https://bioconductor.org/packages/release/bioc/html/SummarizedExperiment.html) container with the TPM (`abundance`), estimated length-scaled counts (`counts`) and transcript length (`length`) in the assays slot for genes. + - `.merged.transcript_counts.tsv`: Matrix of isoform-level raw counts across all samples. + - `.merged.transcript_tpm.tsv`: Matrix of isoform-level TPM values across all samples. + - `.merged.transcript_counts.rds`: RDS object that can be loaded in R that contains a [SummarizedExperiment](https://bioconductor.org/packages/release/bioc/html/SummarizedExperiment.html) container with the TPM (`abundance`), estimated isoform-level raw counts (`counts`) and transcript length (`length`) in the assays slot for transcripts. + - `tx2gene.tsv`: Tab-delimited file containing gene to transcripts ids mappings. +
    + +An additional subset of files are distinct to each tool, for Salmon:
    Output files -- `salmon/` - - `salmon.merged.gene_counts.tsv`: Matrix of gene-level raw counts across all samples. - - `salmon.merged.gene_tpm.tsv`: Matrix of gene-level TPM values across all samples. - - `salmon.merged.gene_counts.rds`: RDS object that can be loaded in R that contains a [SummarizedExperiment](https://bioconductor.org/packages/release/bioc/html/SummarizedExperiment.html) container with the TPM (`abundance`), estimated counts (`counts`) and transcript length (`length`) in the assays slot for genes. - - `salmon.merged.gene_counts_scaled.tsv`: Matrix of gene-level library size-scaled counts across all samples. - - `salmon.merged.gene_counts_scaled.rds`: RDS object that can be loaded in R that contains a [SummarizedExperiment](https://bioconductor.org/packages/release/bioc/html/SummarizedExperiment.html) container with the TPM (`abundance`), estimated library size-scaled counts (`counts`) and transcript length (`length`) in the assays slot for genes. - - `salmon.merged.gene_counts_length_scaled.tsv`: Matrix of gene-level length-scaled counts across all samples. - - `salmon.merged.gene_counts_length_scaled.rds`: RDS object that can be loaded in R that contains a [SummarizedExperiment](https://bioconductor.org/packages/release/bioc/html/SummarizedExperiment.html) container with the TPM (`abundance`), estimated length-scaled counts (`counts`) and transcript length (`length`) in the assays slot for genes. - - `salmon.merged.transcript_counts.tsv`: Matrix of isoform-level raw counts across all samples. - - `salmon.merged.transcript_tpm.tsv`: Matrix of isoform-level TPM values across all samples. - - `salmon.merged.transcript_counts.rds`: RDS object that can be loaded in R that contains a [SummarizedExperiment](https://bioconductor.org/packages/release/bioc/html/SummarizedExperiment.html) container with the TPM (`abundance`), estimated isoform-level raw counts (`counts`) and transcript length (`length`) in the assays slot for transcripts. - - `salmon_tx2gene.tsv`: Tab-delimited file containing gene to transcripts ids mappings. - `salmon//` - `aux_info/`: Auxiliary info e.g. versions and number of mapped reads. - `cmd_info.json`: Information about the Salmon quantification command, version and options. @@ -695,14 +706,25 @@ Results generated by MultiQC collate pipeline QC from supported tools i.e. FastQ - `logs/`: Contains the file `salmon_quant.log` giving a record of Salmon's quantification. - `quant.genes.sf`: Salmon _gene_-level quantification of the sample, including feature length, effective length, TPM, and number of reads. - `quant.sf`: Salmon _transcript_-level quantification of the sample, including feature length, effective length, TPM, and number of reads. +
    + +... and Kallisto: + +
    +Output files +- `kallisto//` + - `abundance.h5`: a HDF5 binary file containing run info, abundance esimates, bootstrap estimates, and transcript length information length. This file can be read in by sleuth. + - `abundance.tsv`: a plaintext file of the abundance estimates. It does not contains bootstrap estimates. + - `run_info.json`: a json file containing information about the run. +- `.log.txt`: standard output from the Kallisto process per sample.
    -As described in the [STAR and Salmon](#star-and-salmon) section, you can choose to pseudo-align and quantify your data with [Salmon](https://salmon.readthedocs.io/en/latest/salmon.html) by providing the `--pseudo_aligner salmon` parameter. By default, Salmon is run in addition to the standard alignment workflow defined by `--aligner`, mainly because it allows you to obtain QC metrics with respect to the genomic alignments. However, you can provide the `--skip_alignment` parameter if you would like to run Salmon in isolation. If Salmon is run in isolation, the outputs mentioned above will be found in a folder named `salmon`. If Salmon is run alongside STAR, the folder will be named `star_salmon`. +As described in the [STAR and Salmon](#star-and-salmon) section, you can choose to pseudo-align and quantify your data with [Salmon](https://salmon.readthedocs.io/en/latest/salmon.html) or [Kallisto](https://pachterlab.github.io/kallisto/) by providing the `--pseudo_aligner` parameter. By default, Salmon is run in addition to the standard alignment workflow defined by `--aligner`, mainly because it allows you to obtain QC metrics with respect to the genomic alignments. However, you can provide the `--skip_alignment` parameter if you would like to run Salmon or Kallisto in isolation. If Salmon or Kallisto are run in isolation, the outputs mentioned above will be found in a folder named `salmon` or `kallisto`. If Salmon is run alongside STAR, the folder will be named `star_salmon`. Transcripts with large inferential uncertainty won't be assigned the exact number of reads reproducibly, every time Salmon is run. Read more about this on the [nf-core/rnaseq](https://github.com/nf-core/rnaseq/issues/585) and [salmon](https://github.com/COMBINE-lab/salmon/issues/613) Github repos. -The [tximport](https://bioconductor.org/packages/release/bioc/html/tximport.html) package is used in this pipeline to summarise the results generated by Salmon into matrices for use with downstream differential analysis packages. We use tximport with different options to summarize count and TPM quantifications at the gene- and transcript-level. Please see [#499](https://github.com/nf-core/rnaseq/issues/499) for discussion and links regarding which counts are suitable for different types of analysis. +The [tximport](https://bioconductor.org/packages/release/bioc/html/tximport.html) package is used in this pipeline to summarise the results generated by Salmon or Kallisto into matrices for use with downstream differential analysis packages. We use tximport with different options to summarize count and TPM quantifications at the gene- and transcript-level. Please see [#499](https://github.com/nf-core/rnaseq/issues/499) for discussion and links regarding which counts are suitable for different types of analysis. According to the `txtimport` documentation you can do one of the following: From 46866ae764035849e5d64bc5fbdfd533d61510e0 Mon Sep 17 00:00:00 2001 From: nf-core-bot Date: Mon, 6 Nov 2023 12:35:50 +0000 Subject: [PATCH 101/184] [automated] Fix linting with Prettier --- docs/output.md | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/docs/output.md b/docs/output.md index 000b0c445..e6c6af75c 100644 --- a/docs/output.md +++ b/docs/output.md @@ -44,7 +44,7 @@ The pipeline is built using [Nextflow](https://www.nextflow.io/) and processes d - [Pseudo-alignment and quantification](#pseudo-alignment-and-quantification) - [Salmon](#salmon) - Wicked fast gene and isoform quantification relative to the transcriptome - [Kallisto](#kallisto) - Near-optimal probabilistic RNA-seq quantification -Wicked fast gene and isoform quantification relative to the transcriptome + Wicked fast gene and isoform quantification relative to the transcriptome - [Workflow reporting and genomes](#workflow-reporting-and-genomes) - [Reference genome files](#reference-genome-files) - Saving reference genome indices/files - [Pipeline information](#pipeline-information) - Report metrics generated during the workflow execution @@ -691,7 +691,7 @@ The principal output files are the same between Salmon and Kallsto: - `.merged.transcript_tpm.tsv`: Matrix of isoform-level TPM values across all samples. - `.merged.transcript_counts.rds`: RDS object that can be loaded in R that contains a [SummarizedExperiment](https://bioconductor.org/packages/release/bioc/html/SummarizedExperiment.html) container with the TPM (`abundance`), estimated isoform-level raw counts (`counts`) and transcript length (`length`) in the assays slot for transcripts. - `tx2gene.tsv`: Tab-delimited file containing gene to transcripts ids mappings. -
    +
    An additional subset of files are distinct to each tool, for Salmon: @@ -706,7 +706,7 @@ An additional subset of files are distinct to each tool, for Salmon: - `logs/`: Contains the file `salmon_quant.log` giving a record of Salmon's quantification. - `quant.genes.sf`: Salmon _gene_-level quantification of the sample, including feature length, effective length, TPM, and number of reads. - `quant.sf`: Salmon _transcript_-level quantification of the sample, including feature length, effective length, TPM, and number of reads. - + ... and Kallisto: From b39e7e838e122cc8dac9e343c64bbff378358ba0 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Mon, 6 Nov 2023 12:48:19 +0000 Subject: [PATCH 102/184] Correct kallisto out path --- conf/modules.config | 10 ++++++++++ docs/output.md | 1 + 2 files changed, 11 insertions(+) diff --git a/conf/modules.config b/conf/modules.config index c0924e879..15163a1af 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -88,6 +88,16 @@ process { ] } + withName: 'KALLISTO_INDEX' { + ext.args = params.gencode ? '--gencode' : '' + publishDir = [ + path: { "${params.outdir}/genome/index" }, + mode: params.publish_dir_mode, + saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, + enabled: params.save_reference + ] + } + withName: 'RSEM_PREPAREREFERENCE_GENOME' { ext.args = '--star' publishDir = [ diff --git a/docs/output.md b/docs/output.md index 000b0c445..8c2e2fed1 100644 --- a/docs/output.md +++ b/docs/output.md @@ -750,6 +750,7 @@ According to the `txtimport` documentation you can do one of the following: - `hisat2/`: Directory containing HISAT2 indices. - `rsem/`: Directory containing STAR and RSEM indices. - `salmon/`: Directory containing Salmon indices. + - `salmon/`: Directory containing Kallisto indices. From 58bbc278ad3fd39bc0b28d63caec07ac8d83b162 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 7 Nov 2023 09:40:05 +0000 Subject: [PATCH 103/184] Add prefix usage to local modules to facilitate batched runs --- conf/modules.config | 6 ++++++ modules/local/deseq2_qc/main.nf | 2 ++ modules/local/multiqc/main.nf | 2 ++ modules/local/tximport/main.nf | 3 ++- 4 files changed, 12 insertions(+), 1 deletion(-) diff --git a/conf/modules.config b/conf/modules.config index 15163a1af..5e7392409 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -618,6 +618,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { } withName: '.*:QUANTIFY_STAR_SALMON:TXIMPORT' { + ext.prefix = { "${quant_type}.merged" } publishDir = [ path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, @@ -759,6 +760,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { if (!params.skip_qc & !params.skip_deseq2_qc) { process { withName: 'DESEQ2_QC_STAR_SALMON' { + ext.prefix = { "deseq2" } ext.args = [ "--id_col 1", "--sample_suffix ''", @@ -823,6 +825,7 @@ if (!params.skip_alignment && params.aligner == 'star_rsem') { if (!params.skip_qc & !params.skip_deseq2_qc) { process { withName: 'DESEQ2_QC_RSEM' { + ext.prefix = { "deseq2" } ext.args = [ "--id_col 1", "--sample_suffix ''", @@ -1123,6 +1126,7 @@ if (!params.skip_multiqc) { process { withName: 'MULTIQC' { ext.args = params.multiqc_title ? "--title \"$params.multiqc_title\"" : '' + ext.prefix = { "$multiqc_report" } publishDir = [ path: { [ "${params.outdir}/multiqc", @@ -1189,6 +1193,7 @@ if (!params.skip_pseudo_alignment) { } withName: '.*:QUANTIFY_PSEUDO:TXIMPORT' { + ext.prefix = { "${quant_type}.merged" } publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}" }, mode: params.publish_dir_mode, @@ -1208,6 +1213,7 @@ if (!params.skip_pseudo_alignment) { if (!params.skip_qc & !params.skip_deseq2_qc) { process { withName: 'DESEQ2_QC_PSEUDO' { + ext.prefix = { "deseq2" } ext.args = [ "--id_col 1", "--sample_suffix ''", diff --git a/modules/local/deseq2_qc/main.nf b/modules/local/deseq2_qc/main.nf index 7b3fac57b..466194934 100644 --- a/modules/local/deseq2_qc/main.nf +++ b/modules/local/deseq2_qc/main.nf @@ -32,11 +32,13 @@ process DESEQ2_QC { def args2 = task.ext.args2 ?: '' def label_lower = args2.toLowerCase() def label_upper = args2.toUpperCase() + prefix = task.ext.prefix ?: "deseq2" """ deseq2_qc.r \\ --count_file $counts \\ --outdir ./ \\ --cores $task.cpus \\ + --outprefix $prefix \\ $args if [ -f "R_sessionInfo.log" ]; then diff --git a/modules/local/multiqc/main.nf b/modules/local/multiqc/main.nf index e5fcf88f2..7ecc28f87 100644 --- a/modules/local/multiqc/main.nf +++ b/modules/local/multiqc/main.nf @@ -57,8 +57,10 @@ process MULTIQC { script: def args = task.ext.args ?: '' def custom_config = params.multiqc_config ? "--config $multiqc_custom_config" : '' + prefix = task.ext.prefix ?: "multiqc_report" """ multiqc \\ + -n ${prefix}.html \\ -f \\ $args \\ $custom_config \\ diff --git a/modules/local/tximport/main.nf b/modules/local/tximport/main.nf index d46dde44e..56d826a7d 100644 --- a/modules/local/tximport/main.nf +++ b/modules/local/tximport/main.nf @@ -24,11 +24,12 @@ process TXIMPORT { task.ext.when == null || task.ext.when script: // This script is bundled with the pipeline, in nf-core/rnaseq/bin/ + prefix = task.ext.prefix ?: "${quant_type}.merged" """ tximport.r \\ NULL \\ quants \\ - ${quant_type}.merged \\ + $prefix \\ $quant_type \\ $tx2gene From ab4c662ae4d44aa5abcc8ddea8c7b8ce73ada02c Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 7 Nov 2023 15:08:29 +0000 Subject: [PATCH 104/184] [skip ci] Address easy review feedback --- CHANGELOG.md | 2 ++ bin/summarizedexperiment.r | 2 +- bin/tximport.r | 2 +- conf/modules.config | 20 ++++++++--------- docs/output.md | 26 +++++++++++----------- docs/usage.md | 4 ++-- nextflow.config | 4 ++-- nextflow_schema.json | 4 ++-- subworkflows/local/quantify_pseudo/main.nf | 2 +- workflows/rnaseq.nf | 18 +++++++-------- 10 files changed, 43 insertions(+), 41 deletions(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index c6f3aac74..2512ccd26 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -29,6 +29,8 @@ Thank you to everyone else that has contributed by reporting bugs, enhancements - [PR #1088](https://github.com/nf-core/rnaseq/pull/1088) - Updates contributing and code of conduct documents with nf-core template 2.10 - [PR #1091](https://github.com/nf-core/rnaseq/pull/1091) - Reorganise parameters in schema for better usability - [PR #1106](https://github.com/nf-core/rnaseq/pull/1106) - Kallisto quantification +- [PR #1106](https://github.com/nf-core/rnaseq/pull/1106) - MultiQC [version bump](https://github.com/nf-core/rnaseq/pull/1106/commits/aebad067a10a45510a2b421da852cb436ae65fd8) +- [#1050](https://github.com/nf-core/rnaseq/issues/1050) - Provide custom prefix/suffix for summary files to avoid overwriting ### Software dependencies diff --git a/bin/summarizedexperiment.r b/bin/summarizedexperiment.r index 73f6d9f38..410663ae6 100755 --- a/bin/summarizedexperiment.r +++ b/bin/summarizedexperiment.r @@ -6,7 +6,7 @@ library(SummarizedExperiment) args <- commandArgs(trailingOnly = TRUE) if (length(args) < 3) { - stop("Usage: salmon_se.r ", call. = FALSE) + stop("Usage: summarizedexperiment.r ", call. = FALSE) } coldata <- args[1] diff --git a/bin/tximport.r b/bin/tximport.r index 21f5fa6e4..e1bad7511 100755 --- a/bin/tximport.r +++ b/bin/tximport.r @@ -5,7 +5,7 @@ library(tximport) args = commandArgs(trailingOnly=TRUE) if (length(args) < 4) { - stop("Usage: salmon_tximport.r ", call.=FALSE) + stop("Usage: tximport.r ", call.=FALSE) } coldata = args[1] diff --git a/conf/modules.config b/conf/modules.config index 5e7392409..024daeb18 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -760,7 +760,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { if (!params.skip_qc & !params.skip_deseq2_qc) { process { withName: 'DESEQ2_QC_STAR_SALMON' { - ext.prefix = { "deseq2" } + ext.prefix = "deseq2" ext.args = [ "--id_col 1", "--sample_suffix ''", @@ -825,7 +825,7 @@ if (!params.skip_alignment && params.aligner == 'star_rsem') { if (!params.skip_qc & !params.skip_deseq2_qc) { process { withName: 'DESEQ2_QC_RSEM' { - ext.prefix = { "deseq2" } + ext.prefix = "deseq2" ext.args = [ "--id_col 1", "--sample_suffix ''", @@ -1126,7 +1126,7 @@ if (!params.skip_multiqc) { process { withName: 'MULTIQC' { ext.args = params.multiqc_title ? "--title \"$params.multiqc_title\"" : '' - ext.prefix = { "$multiqc_report" } + ext.prefix = "multiqc_report" publishDir = [ path: { [ "${params.outdir}/multiqc", @@ -1145,7 +1145,7 @@ if (!params.skip_multiqc) { if (params.pseudo_aligner == 'salmon') { process { - withName: '.*:QUANTIFY_PSEUDO:SALMON_QUANT' { + withName: '.*:QUANTIFY_PSEUDO_ALIGNMENT:SALMON_QUANT' { ext.args = params.extra_salmon_quant_args ?: '' publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}" }, @@ -1158,13 +1158,13 @@ if (params.pseudo_aligner == 'salmon') { if (params.pseudo_aligner == 'kallisto') { process { - withName: '.*:QUANTIFY_PSEUDO:KALLISTO_QUANT' { + withName: '.*:QUANTIFY_PSEUDO_ALIGNMENT:KALLISTO_QUANT' { ext.args = { [ meta.single_end ? "--single " + - "--fragment-length ${meta.fragment_len ?: params.kallisto_default_fraglen} " + - "--sd ${meta.fraglen_se ?: params.kallisto_default_fraglen_se}" : '', + "--fragment-length ${meta.fragment_len ?: params.kallisto_quant_fraglen} " + + "--sd ${meta.fraglen_se ?: params.kallisto_quant_fraglen_sd}" : '', params.kallisto_quant_strandedness ? '--' + params.kallisto_quant_strandedness: (meta.strandedess == 'forward' ? '--fr-stranded': (meta.strandedness == 'reverse' ? '--rf-stranded' : '') @@ -1184,7 +1184,7 @@ if (params.pseudo_aligner == 'kallisto') { if (!params.skip_pseudo_alignment) { process { - withName: '.*:QUANTIFY_PSEUDO:TX2GENE' { + withName: '.*:QUANTIFY_PSEUDO_ALIGNMENT:TX2GENE' { publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}" }, mode: params.publish_dir_mode, @@ -1192,7 +1192,7 @@ if (!params.skip_pseudo_alignment) { ] } - withName: '.*:QUANTIFY_PSEUDO:TXIMPORT' { + withName: '.*:QUANTIFY_PSEUDO_ALIGNMENT:TXIMPORT' { ext.prefix = { "${quant_type}.merged" } publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}" }, @@ -1201,7 +1201,7 @@ if (!params.skip_pseudo_alignment) { ] } - withName: '.*:QUANTIFY_PSEUDO:SE_.*' { + withName: '.*:QUANTIFY_PSEUDO_ALIGNMENT:SE_.*' { publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}" }, mode: params.publish_dir_mode, diff --git a/docs/output.md b/docs/output.md index 5cfdac4fa..b5cec204a 100644 --- a/docs/output.md +++ b/docs/output.md @@ -679,17 +679,17 @@ The principal output files are the same between Salmon and Kallsto:
    Output files -- `/` - - `.merged.gene_counts.tsv`: Matrix of gene-level raw counts across all samples. - - `.gene_tpm.tsv`: Matrix of gene-level TPM values across all samples. - - `.gene_counts.rds`: RDS object that can be loaded in R that contains a [SummarizedExperiment](https://bioconductor.org/packages/release/bioc/html/SummarizedExperiment.html) container with the TPM (`abundance`), estimated counts (`counts`) and transcript length (`length`) in the assays slot for genes. - - `.merged.gene_counts_scaled.tsv`: Matrix of gene-level library size-scaled counts across all samples. - - `.merged.gene_counts_scaled.rds`: RDS object that can be loaded in R that contains a [SummarizedExperiment](https://bioconductor.org/packages/release/bioc/html/SummarizedExperiment.html) container with the TPM (`abundance`), estimated library size-scaled counts (`counts`) and transcript length (`length`) in the assays slot for genes. - - `.merged.gene_counts_length_scaled.tsv`: Matrix of gene-level length-scaled counts across all samples. - - `.merged.gene_counts_length_scaled.rds`: RDS object that can be loaded in R that contains a [SummarizedExperiment](https://bioconductor.org/packages/release/bioc/html/SummarizedExperiment.html) container with the TPM (`abundance`), estimated length-scaled counts (`counts`) and transcript length (`length`) in the assays slot for genes. - - `.merged.transcript_counts.tsv`: Matrix of isoform-level raw counts across all samples. - - `.merged.transcript_tpm.tsv`: Matrix of isoform-level TPM values across all samples. - - `.merged.transcript_counts.rds`: RDS object that can be loaded in R that contains a [SummarizedExperiment](https://bioconductor.org/packages/release/bioc/html/SummarizedExperiment.html) container with the TPM (`abundance`), estimated isoform-level raw counts (`counts`) and transcript length (`length`) in the assays slot for transcripts. +- `/` + - `.merged.gene_counts.tsv`: Matrix of gene-level raw counts across all samples. + - `.gene_tpm.tsv`: Matrix of gene-level TPM values across all samples. + - `.gene_counts.rds`: RDS object that can be loaded in R that contains a [SummarizedExperiment](https://bioconductor.org/packages/release/bioc/html/SummarizedExperiment.html) container with the TPM (`abundance`), estimated counts (`counts`) and transcript length (`length`) in the assays slot for genes. + - `.merged.gene_counts_scaled.tsv`: Matrix of gene-level library size-scaled counts across all samples. + - `.merged.gene_counts_scaled.rds`: RDS object that can be loaded in R that contains a [SummarizedExperiment](https://bioconductor.org/packages/release/bioc/html/SummarizedExperiment.html) container with the TPM (`abundance`), estimated library size-scaled counts (`counts`) and transcript length (`length`) in the assays slot for genes. + - `.merged.gene_counts_length_scaled.tsv`: Matrix of gene-level length-scaled counts across all samples. + - `.merged.gene_counts_length_scaled.rds`: RDS object that can be loaded in R that contains a [SummarizedExperiment](https://bioconductor.org/packages/release/bioc/html/SummarizedExperiment.html) container with the TPM (`abundance`), estimated length-scaled counts (`counts`) and transcript length (`length`) in the assays slot for genes. + - `.merged.transcript_counts.tsv`: Matrix of isoform-level raw counts across all samples. + - `.merged.transcript_tpm.tsv`: Matrix of isoform-level TPM values across all samples. + - `.merged.transcript_counts.rds`: RDS object that can be loaded in R that contains a [SummarizedExperiment](https://bioconductor.org/packages/release/bioc/html/SummarizedExperiment.html) container with the TPM (`abundance`), estimated isoform-level raw counts (`counts`) and transcript length (`length`) in the assays slot for transcripts. - `tx2gene.tsv`: Tab-delimited file containing gene to transcripts ids mappings.
    @@ -714,7 +714,7 @@ An additional subset of files are distinct to each tool, for Salmon: Output files - `kallisto//` - - `abundance.h5`: a HDF5 binary file containing run info, abundance esimates, bootstrap estimates, and transcript length information length. This file can be read in by sleuth. + - `abundance.h5`: a HDF5 binary file containing run info, abundance esimates, bootstrap estimates, and transcript length information length. This file can be read in by [sleuth](https://pachterlab.github.io/sleuth/about). - `abundance.tsv`: a plaintext file of the abundance estimates. It does not contains bootstrap estimates. - `run_info.json`: a json file containing information about the run. - `.log.txt`: standard output from the Kallisto process per sample. @@ -750,7 +750,7 @@ According to the `txtimport` documentation you can do one of the following: - `hisat2/`: Directory containing HISAT2 indices. - `rsem/`: Directory containing STAR and RSEM indices. - `salmon/`: Directory containing Salmon indices. - - `salmon/`: Directory containing Kallisto indices. + - `kallisto/`: Directory containing Kallisto indices. diff --git a/docs/usage.md b/docs/usage.md index bd776e66e..c543ec033 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -71,13 +71,13 @@ The library preparation protocol (library type) used by Salmon quantification is When running Salmon in mapping-based mode via `--pseudo_aligner salmon` the entire genome of the organism is used by default for the decoy-aware transcriptome when creating the indices (see second bulleted option in [Salmon documentation](https://salmon.readthedocs.io/en/latest/salmon.html#preparing-transcriptome-indices-mapping-based-mode)). -Two additional parameters `--extra_star_align_args` and `--extra_salmon_quant_args` were added in v3.10 of the pipeline that allow you to append any custom parameters to the STAR align and Salmon quant commands, respectively. Note, the `--seqBias` and `--gcBias` are not provided to Salmon quant by default so you can provide these via `--extra_salmon_quant_args '--seqBias --gcBias'` if required. You can now also supply additional arguments to Kallisto via `extra_kallisto_quant_args`. +Two additional parameters `--extra_star_align_args` and `--extra_salmon_quant_args` were added in v3.10 of the pipeline that allow you to append any custom parameters to the STAR align and Salmon quant commands, respectively. Note, the `--seqBias` and `--gcBias` are not provided to Salmon quant by default so you can provide these via `--extra_salmon_quant_args '--seqBias --gcBias'` if required. You can now also supply additional arguments to Kallisto via `--extra_kallisto_quant_args`. > **NB:** You can use `--skip_alignment --skip_pseudo_alignment` if you only want to run the pre-processing QC steps in the pipeline like FastQ, trimming etc. This will skip alignment, pseudo-alignment and any post-alignment processing steps. ## Quantification options -The current options align with STAR and quantify using either Salmon (`--aligner star_salmon`) / RSEM (`--aligner star_rsem`). You also have the option to pseudo-align and quantify your data with Salmon or Kallisto by providing the `--pseudo_aligner salmon` parameter. +The current options align with STAR and quantify using either Salmon (`--aligner star_salmon`) / RSEM (`--aligner star_rsem`). You also have the option to pseudo-align and quantify your data with Salmon or Kallisto by providing the `--pseudo_aligner salmon` or `--pseudo_aligner kallisto` parameter, respectively. Since v3.0 of the pipeline, featureCounts is no longer used to perform gene/transcript quantification, however it is still used to generate QC metrics based on [biotype](http://www.ensembl.org/info/genome/genebuild/biotypes.html) information available within GFF/GTF genome annotation files. This decision was made primarily because of the limitations of featureCounts to appropriately quantify gene expression data. Please see [Zhao et al., 2015](https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0141910#pone-0141910-t001) and [Soneson et al., 2015](https://f1000research.com/articles/4-1521/v1). diff --git a/nextflow.config b/nextflow.config index 09e49b36a..bc38b6124 100644 --- a/nextflow.config +++ b/nextflow.config @@ -67,8 +67,8 @@ params { extra_star_align_args = null extra_salmon_quant_args = null extra_kallisto_quant_args = null - kallisto_default_fraglen = 200 - kallisto_default_fraglen_se = 200 + kallisto_quant_fraglen = 200 + kallisto_quant_fraglen_sd = 200 kallisto_quant_strandedness = null save_merged_fastq = false save_unaligned = false diff --git a/nextflow_schema.json b/nextflow_schema.json index 662b0737a..55ff1d843 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -417,13 +417,13 @@ "description": "Extra arguments to pass to Kallisto quant command in addition to defaults defined by the pipeline.", "fa_icon": "fas fa-plus" }, - "kallisto_default_fraglen": { + "kallisto_quant_fraglen": { "type": "integer", "description": "In single-end mode Kallisto requires an estimated fragment length. Specify a default value for that here. TODO: use existing RSeQC results to do this dynamically.", "default": 150, "fa_icon": "fas fa-ruler-horizontal" }, - "kallisto_default_fraglen_se": { + "kallisto_quant_fraglen_sd": { "type": "integer", "description": "In single-end mode Kallisto requires an estimated standard error for fragment length. Specify a default value for that here. TODO: use existing RSeQC results to do this dynamically.", "default": 75, diff --git a/subworkflows/local/quantify_pseudo/main.nf b/subworkflows/local/quantify_pseudo/main.nf index cb3fb7399..1b712c76b 100644 --- a/subworkflows/local/quantify_pseudo/main.nf +++ b/subworkflows/local/quantify_pseudo/main.nf @@ -12,7 +12,7 @@ include { SUMMARIZEDEXPERIMENT as SE_GENE_LENGTH_SCALED } from '../../../modules include { SUMMARIZEDEXPERIMENT as SE_GENE_SCALED } from '../../../modules/local/summarizedexperiment' include { SUMMARIZEDEXPERIMENT as SE_TRANSCRIPT } from '../../../modules/local/summarizedexperiment' -workflow QUANTIFY_PSEUDO { +workflow QUANTIFY_PSEUDO_ALIGNMENT { take: reads // channel: [ val(meta), [ reads ] ] index // channel: /path/to//index/ diff --git a/workflows/rnaseq.nf b/workflows/rnaseq.nf index 6e9d81c9d..3632e1a84 100755 --- a/workflows/rnaseq.nf +++ b/workflows/rnaseq.nf @@ -97,11 +97,11 @@ include { UMITOOLS_PREPAREFORRSEM as UMITOOLS_PREPAREFORSALMON } from '../module // // SUBWORKFLOW: Consisting of a mix of local and nf-core/modules // -include { PREPARE_GENOME } from '../subworkflows/local/prepare_genome' -include { ALIGN_STAR } from '../subworkflows/local/align_star' -include { QUANTIFY_RSEM } from '../subworkflows/local/quantify_rsem' -include { QUANTIFY_PSEUDO as QUANTIFY_STAR_SALMON } from '../subworkflows/local/quantify_pseudo' -include { QUANTIFY_PSEUDO } from '../subworkflows/local/quantify_pseudo' +include { PREPARE_GENOME } from '../subworkflows/local/prepare_genome' +include { ALIGN_STAR } from '../subworkflows/local/align_star' +include { QUANTIFY_RSEM } from '../subworkflows/local/quantify_rsem' +include { QUANTIFY_PSEUDO_ALIGNMENT as QUANTIFY_STAR_SALMON } from '../subworkflows/local/quantify_pseudo' +include { QUANTIFY_PSEUDO_ALIGNMENT } from '../subworkflows/local/quantify_pseudo' /* ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ @@ -804,7 +804,7 @@ workflow RNASEQ { ch_pseudo_index = PREPARE_GENOME.out.kallisto_index } - QUANTIFY_PSEUDO ( + QUANTIFY_PSEUDO_ALIGNMENT ( ch_filtered_reads, ch_pseudo_index, ch_dummy_file, @@ -813,9 +813,9 @@ workflow RNASEQ { false, params.salmon_quant_libtype ?: '' ) - ch_pseudo_multiqc = QUANTIFY_PSEUDO.out.multiqc - ch_versions = ch_versions.mix(QUANTIFY_PSEUDO.out.versions) - ch_counts_gene_length_scaled = QUANTIFY_PSEUDO.out.counts_gene_length_scaled + ch_pseudo_multiqc = QUANTIFY_PSEUDO_ALIGNMENT.out.multiqc + ch_versions = ch_versions.mix(QUANTIFY_PSEUDO_ALIGNMENT.out.versions) + ch_counts_gene_length_scaled = QUANTIFY_PSEUDO_ALIGNMENT.out.counts_gene_length_scaled if (!params.skip_qc & !params.skip_deseq2_qc) { DESEQ2_QC_PSEUDO ( From 688d19b74fd88e5ea34b7140870a8497412fe72a Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 7 Nov 2023 18:37:33 +0000 Subject: [PATCH 105/184] Add special GTF handling for stringtie --- modules/local/gtf_for_stringtie/main.nf | 30 +++++++++++++++++++++++ subworkflows/local/prepare_genome/main.nf | 13 ++++++++++ workflows/rnaseq.nf | 2 +- 3 files changed, 44 insertions(+), 1 deletion(-) create mode 100644 modules/local/gtf_for_stringtie/main.nf diff --git a/modules/local/gtf_for_stringtie/main.nf b/modules/local/gtf_for_stringtie/main.nf new file mode 100644 index 000000000..b3b1db1a8 --- /dev/null +++ b/modules/local/gtf_for_stringtie/main.nf @@ -0,0 +1,30 @@ +process GTF_FOR_STRINGTIE { + tag "$gtf" + label 'process_low' + + conda "conda-forge::gawk=5.1.0" + container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? + 'https://depot.galaxyproject.org/singularity/gawk:5.1.0' : + 'biocontainers/gawk:5.1.0' }" + + input: + path gtf + + output: + path '*_for_stringtie.gtf' , emit: gtf + path "versions.yml" , emit: versions + + when: + task.ext.when == null || task.ext.when + + """ + # Remove gene entries from a GTF, if they don't contain a transcript_id + # attribute, which is common in Ensembl GTFs + awk '$3 == "\gene" { print; next } /transcript_id/ && \$0 ~ /transcript_id "[^"]+"/' $gtf > ${gtf}_for_stringtie.gtf + + cat <<-END_VERSIONS > versions.yml + "${task.process}": + bash: \$(echo \$(bash --version | grep -Eo 'version [[:alnum:].]+' | sed 's/version //')) + END_VERSIONS + """ +} diff --git a/subworkflows/local/prepare_genome/main.nf b/subworkflows/local/prepare_genome/main.nf index eceb2c1ce..8381fc545 100644 --- a/subworkflows/local/prepare_genome/main.nf +++ b/subworkflows/local/prepare_genome/main.nf @@ -30,6 +30,7 @@ include { GTF2BED } from '../../../modules/local/gt include { CAT_ADDITIONAL_FASTA } from '../../../modules/local/cat_additional_fasta' include { GTF_GENE_FILTER } from '../../../modules/local/gtf_gene_filter' include { STAR_GENOMEGENERATE_IGENOMES } from '../../../modules/local/star_genomegenerate_igenomes' +include { GTF_FOR_STRINGTIE } from '../../../modules/local/gtf_for_stringtie' workflow PREPARE_GENOME { take: @@ -117,6 +118,17 @@ workflow PREPARE_GENOME { ch_versions = ch_versions.mix(GTF2BED.out.versions) } + // + // Prepare a GTF for StringTie + // + + ch_gtf_for_stringtie = Channel.empty() + if (!params.skip_alignment && !params.skip_stringtie) { + GTF_FOR_STRINGTIE( ch_gtf ) + ch_gtf_for_stringtie = GTF_FOR_STRINGTIE.out.gtf + ch_versions = ch_versions.mix(GTF_FOR_STRINGTIE.out.versions) + } + // // Uncompress transcript fasta file / create if required // @@ -263,6 +275,7 @@ workflow PREPARE_GENOME { gtf = ch_gtf // channel: path(genome.gtf) fai = ch_fai // channel: path(genome.fai) gene_bed = ch_gene_bed // channel: path(gene.bed) + gtf_for_stringtie= ch_gtf_for_stringtie // channel: path(gtf) transcript_fasta = ch_transcript_fasta // channel: path(transcript.fasta) chrom_sizes = ch_chrom_sizes // channel: path(genome.sizes) splicesites = ch_splicesites // channel: path(genome.splicesites.txt) diff --git a/workflows/rnaseq.nf b/workflows/rnaseq.nf index 955a727c3..4a13cb3ee 100755 --- a/workflows/rnaseq.nf +++ b/workflows/rnaseq.nf @@ -648,7 +648,7 @@ workflow RNASEQ { if (!params.skip_alignment && !params.skip_stringtie) { STRINGTIE_STRINGTIE ( ch_genome_bam, - PREPARE_GENOME.out.gtf + PREPARE_GENOME.out.gtf_for_stringtie ) ch_versions = ch_versions.mix(STRINGTIE_STRINGTIE.out.versions.first()) } From c6d354df5f66cb8a48bd06dae0d8ed0ffba806dc Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 7 Nov 2023 18:39:30 +0000 Subject: [PATCH 106/184] Update CHANGELOG --- CHANGELOG.md | 1 + 1 file changed, 1 insertion(+) diff --git a/CHANGELOG.md b/CHANGELOG.md index a235dd00f..e64510937 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -28,6 +28,7 @@ Thank you to everyone else that has contributed by reporting bugs, enhancements - [PR #1083](https://github.com/nf-core/rnaseq/pull/1083) - Move local modules and subworkflows to subfolders - [PR #1088](https://github.com/nf-core/rnaseq/pull/1088) - Updates contributing and code of conduct documents with nf-core template 2.10 - [PR #1091](https://github.com/nf-core/rnaseq/pull/1091) - Reorganise parameters in schema for better usability +- [PR #1107](https://github.com/nf-core/rnaseq/pull/1107) - Prepare a GTF for stringtie ### Software dependencies From 82b67e16202685ee7907536773fa78b825cd5e3f Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 7 Nov 2023 18:40:00 +0000 Subject: [PATCH 107/184] appease eclint --- modules/local/gtf_for_stringtie/main.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/modules/local/gtf_for_stringtie/main.nf b/modules/local/gtf_for_stringtie/main.nf index b3b1db1a8..dd25b632d 100644 --- a/modules/local/gtf_for_stringtie/main.nf +++ b/modules/local/gtf_for_stringtie/main.nf @@ -16,7 +16,7 @@ process GTF_FOR_STRINGTIE { when: task.ext.when == null || task.ext.when - + """ # Remove gene entries from a GTF, if they don't contain a transcript_id # attribute, which is common in Ensembl GTFs From 5dbbe0c96c97ee14db3bc1b2f480f2b974bad102 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 7 Nov 2023 18:41:04 +0000 Subject: [PATCH 108/184] Comment dollar --- modules/local/gtf_for_stringtie/main.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/modules/local/gtf_for_stringtie/main.nf b/modules/local/gtf_for_stringtie/main.nf index dd25b632d..2dadbc24f 100644 --- a/modules/local/gtf_for_stringtie/main.nf +++ b/modules/local/gtf_for_stringtie/main.nf @@ -20,7 +20,7 @@ process GTF_FOR_STRINGTIE { """ # Remove gene entries from a GTF, if they don't contain a transcript_id # attribute, which is common in Ensembl GTFs - awk '$3 == "\gene" { print; next } /transcript_id/ && \$0 ~ /transcript_id "[^"]+"/' $gtf > ${gtf}_for_stringtie.gtf + awk '\$3 == "gene" { print; next } /transcript_id/ && \$0 ~ /transcript_id "[^"]+"/' $gtf > ${gtf}_for_stringtie.gtf cat <<-END_VERSIONS > versions.yml "${task.process}": From 124c5701d472f0b29af6298b46d664073a9ac76f Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Fri, 3 Nov 2023 10:31:10 +0000 Subject: [PATCH 109/184] Fix multiqc to avoid imp module error --- modules/local/multiqc/main.nf | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/modules/local/multiqc/main.nf b/modules/local/multiqc/main.nf index 44565a9f7..06fbb9b29 100644 --- a/modules/local/multiqc/main.nf +++ b/modules/local/multiqc/main.nf @@ -1,10 +1,10 @@ process MULTIQC { label 'process_medium' - conda "bioconda::multiqc=1.15" + conda "bioconda::multiqc=1.17" container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? - 'https://depot.galaxyproject.org/singularity/multiqc:1.15--pyhdfd78af_0' : - 'biocontainers/multiqc:1.15--pyhdfd78af_0' }" + 'https://depot.galaxyproject.org/singularity/multiqc:1.17--pyhdfd78af_0' : + 'biocontainers/multiqc:1.17--pyhdfd78af_0' }" input: path multiqc_config From 57d4bde66d329271ca02fa3e8c6ba9792ff3585d Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 9 Nov 2023 12:58:22 +0000 Subject: [PATCH 110/184] Actually, unify GTF filtering --- bin/filter_gtf_for_genes_in_genome.py | 56 +++++++++++++++++------ conf/test.config | 2 +- modules/local/gtf_for_stringtie/main.nf | 30 ------------ modules/local/gtf_gene_filter/main.nf | 5 +- subworkflows/local/prepare_genome/main.nf | 50 +++++++++----------- workflows/rnaseq.nf | 4 +- 6 files changed, 71 insertions(+), 76 deletions(-) delete mode 100644 modules/local/gtf_for_stringtie/main.nf diff --git a/bin/filter_gtf_for_genes_in_genome.py b/bin/filter_gtf_for_genes_in_genome.py index 9f876eaa0..cb2002a02 100755 --- a/bin/filter_gtf_for_genes_in_genome.py +++ b/bin/filter_gtf_for_genes_in_genome.py @@ -9,18 +9,25 @@ logger = logging.getLogger(__file__) logger.setLevel(logging.INFO) - -def is_header(line): +def is_header(line: str) -> bool: + """Returns True if the given line is a header line in a FASTA file.""" return line[0] == ">" - -def extract_fasta_seq_names(fasta_name): - """ +def extract_fasta_seq_names(fasta_name: str) -> set: + """Extracts the sequence names from a FASTA file. + modified from Brent Pedersen Correct Way To Parse A Fasta File In Python given a fasta file. yield tuples of header, sequence from https://www.biostars.org/p/710/ + + Args: + fasta_name: The path to the FASTA file. + + Returns: + A set of the sequence names in the FASTA file. """ + # first open the file outside fh = open(fasta_name) @@ -35,8 +42,15 @@ def extract_fasta_seq_names(fasta_name): headerStr = line[1:].strip().split()[0] yield headerStr +def extract_genes_in_genome(fasta: str, gtf_in: str, prefix: str) -> None: + """Extracts the genes in the genome from a GTF file. -def extract_genes_in_genome(fasta, gtf_in, gtf_out): + Args: + fasta: The path to the FASTA file. + gtf_in: The path to the input GTF file. + prefix: Prefix for output GTF + """ + gtf_out = prefix + '_in_genome.gtf' seq_names_in_genome = set(extract_fasta_seq_names(fasta)) logger.info("Extracted chromosome sequence names from : %s" % fasta) logger.info("All chromosome names: " + ", ".join(sorted(x for x in seq_names_in_genome))) @@ -60,19 +74,35 @@ def extract_genes_in_genome(fasta, gtf_in, gtf_out): logger.info("Wrote matching lines to %s" % gtf_out) +def remove_features_without_transcript_id(gtf_in, prefix): + """ + Removes gene rows with absent or empty transcript_id attributes from a GTF file. + + Args: + gtf_in: Path to the input GTF file. + prefix: Path to the output GTF file. + """ + gtf_out = prefix + '_with_transcript_ids.gtf' + + with open(gtf_in, "r") as f_in, open(gtf_out, "w") as f_out: + for line in f_in: + transcript_id = line.split("\t")[8].split(" transcript_id ")[1].split(";")[0].replace('"', '') + if transcript_id and transcript_id.isalnum(): + f_out.write(line) if __name__ == "__main__": - parser = argparse.ArgumentParser(description="""Filter GTF only for features in the genome""") + parser = argparse.ArgumentParser(description="""Filter GTF for various reasons""") parser.add_argument("--gtf", type=str, help="GTF file") parser.add_argument("--fasta", type=str, help="Genome fasta file") parser.add_argument( - "-o", - "--output", - dest="output", - default="genes_in_genome.gtf", + "-p", + "--prefix", + dest="prefix", + default="genes", type=str, - help="GTF features on fasta genome sequences", + help="Prefix for output GTF files", ) args = parser.parse_args() - extract_genes_in_genome(args.fasta, args.gtf, args.output) + extract_genes_in_genome(args.fasta, args.gtf, args.prefix) + remove_features_without_transcript_id(args.gtf, args.prefix) diff --git a/conf/test.config b/conf/test.config index d0430eacb..f78e4e567 100644 --- a/conf/test.config +++ b/conf/test.config @@ -21,7 +21,7 @@ params { // Input data - input = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/samplesheet/v3.10/samplesheet_test.csv" + input = "/Users/jonathan.manning/projects/rnaseq/samplesheet_test.csv" // Genome references fasta = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/reference/genome.fasta" diff --git a/modules/local/gtf_for_stringtie/main.nf b/modules/local/gtf_for_stringtie/main.nf deleted file mode 100644 index 2dadbc24f..000000000 --- a/modules/local/gtf_for_stringtie/main.nf +++ /dev/null @@ -1,30 +0,0 @@ -process GTF_FOR_STRINGTIE { - tag "$gtf" - label 'process_low' - - conda "conda-forge::gawk=5.1.0" - container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? - 'https://depot.galaxyproject.org/singularity/gawk:5.1.0' : - 'biocontainers/gawk:5.1.0' }" - - input: - path gtf - - output: - path '*_for_stringtie.gtf' , emit: gtf - path "versions.yml" , emit: versions - - when: - task.ext.when == null || task.ext.when - - """ - # Remove gene entries from a GTF, if they don't contain a transcript_id - # attribute, which is common in Ensembl GTFs - awk '\$3 == "gene" { print; next } /transcript_id/ && \$0 ~ /transcript_id "[^"]+"/' $gtf > ${gtf}_for_stringtie.gtf - - cat <<-END_VERSIONS > versions.yml - "${task.process}": - bash: \$(echo \$(bash --version | grep -Eo 'version [[:alnum:].]+' | sed 's/version //')) - END_VERSIONS - """ -} diff --git a/modules/local/gtf_gene_filter/main.nf b/modules/local/gtf_gene_filter/main.nf index cd8e16adb..b4e0578d7 100644 --- a/modules/local/gtf_gene_filter/main.nf +++ b/modules/local/gtf_gene_filter/main.nf @@ -11,8 +11,9 @@ process GTF_GENE_FILTER { path gtf output: - path "*.gtf" , emit: gtf - path "versions.yml", emit: versions + path "*_in_genome.gtf" , emit: genome_gtf + path "*_with_transcript_ids.gtf" , emit: transcript_id_gtf + path "versions.yml" , emit: versions when: task.ext.when == null || task.ext.when diff --git a/subworkflows/local/prepare_genome/main.nf b/subworkflows/local/prepare_genome/main.nf index 8381fc545..cec93e86b 100644 --- a/subworkflows/local/prepare_genome/main.nf +++ b/subworkflows/local/prepare_genome/main.nf @@ -30,7 +30,6 @@ include { GTF2BED } from '../../../modules/local/gt include { CAT_ADDITIONAL_FASTA } from '../../../modules/local/cat_additional_fasta' include { GTF_GENE_FILTER } from '../../../modules/local/gtf_gene_filter' include { STAR_GENOMEGENERATE_IGENOMES } from '../../../modules/local/star_genomegenerate_igenomes' -include { GTF_FOR_STRINGTIE } from '../../../modules/local/gtf_for_stringtie' workflow PREPARE_GENOME { take: @@ -87,6 +86,13 @@ workflow PREPARE_GENOME { ch_versions = ch_versions.mix(GFFREAD.out.versions) } + // + // Apply filtering we may need for GTFs + // + GTF_GENE_FILTER ( ch_fasta, ch_gtf ) + ch_gtf_with_transcript_ids = GTF_GENE_FILTER.out.transcript_id_gtf + ch_gtf_genome = GTF_GENE_FILTER.out.genome_gtf + // // Uncompress additional fasta file and concatenate with reference fasta and gtf files // @@ -118,17 +124,6 @@ workflow PREPARE_GENOME { ch_versions = ch_versions.mix(GTF2BED.out.versions) } - // - // Prepare a GTF for StringTie - // - - ch_gtf_for_stringtie = Channel.empty() - if (!params.skip_alignment && !params.skip_stringtie) { - GTF_FOR_STRINGTIE( ch_gtf ) - ch_gtf_for_stringtie = GTF_FOR_STRINGTIE.out.gtf - ch_versions = ch_versions.mix(GTF_FOR_STRINGTIE.out.versions) - } - // // Uncompress transcript fasta file / create if required // @@ -145,8 +140,7 @@ workflow PREPARE_GENOME { ch_versions = ch_versions.mix(PREPROCESS_TRANSCRIPTS_FASTA_GENCODE.out.versions) } } else { - ch_filter_gtf = GTF_GENE_FILTER ( ch_fasta, ch_gtf ).gtf - ch_transcript_fasta = MAKE_TRANSCRIPTS_FASTA ( ch_fasta, ch_filter_gtf ).transcript_fasta + ch_transcript_fasta = MAKE_TRANSCRIPTS_FASTA ( ch_fasta, ch_gtf_genome ).transcript_fasta ch_versions = ch_versions.mix(GTF_GENE_FILTER.out.versions) ch_versions = ch_versions.mix(MAKE_TRANSCRIPTS_FASTA.out.versions) } @@ -271,19 +265,19 @@ workflow PREPARE_GENOME { } emit: - fasta = ch_fasta // channel: path(genome.fasta) - gtf = ch_gtf // channel: path(genome.gtf) - fai = ch_fai // channel: path(genome.fai) - gene_bed = ch_gene_bed // channel: path(gene.bed) - gtf_for_stringtie= ch_gtf_for_stringtie // channel: path(gtf) - transcript_fasta = ch_transcript_fasta // channel: path(transcript.fasta) - chrom_sizes = ch_chrom_sizes // channel: path(genome.sizes) - splicesites = ch_splicesites // channel: path(genome.splicesites.txt) - bbsplit_index = ch_bbsplit_index // channel: path(bbsplit/index/) - star_index = ch_star_index // channel: path(star/index/) - rsem_index = ch_rsem_index // channel: path(rsem/index/) - hisat2_index = ch_hisat2_index // channel: path(hisat2/index/) - salmon_index = ch_salmon_index // channel: path(salmon/index/) + fasta = ch_fasta // channel: path(genome.fasta) + gtf = ch_gtf // channel: path(genome.gtf) + fai = ch_fai // channel: path(genome.fai) + gene_bed = ch_gene_bed // channel: path(gene.bed) + gtf_with_transcript_ids = ch_gtf_with_transcript_ids // channel: path(gtf) + transcript_fasta = ch_transcript_fasta // channel: path(transcript.fasta) + chrom_sizes = ch_chrom_sizes // channel: path(genome.sizes) + splicesites = ch_splicesites // channel: path(genome.splicesites.txt) + bbsplit_index = ch_bbsplit_index // channel: path(bbsplit/index/) + star_index = ch_star_index // channel: path(star/index/) + rsem_index = ch_rsem_index // channel: path(rsem/index/) + hisat2_index = ch_hisat2_index // channel: path(hisat2/index/) + salmon_index = ch_salmon_index // channel: path(salmon/index/) - versions = ch_versions.ifEmpty(null) // channel: [ versions.yml ] + versions = ch_versions.ifEmpty(null) // channel: [ versions.yml ] } diff --git a/workflows/rnaseq.nf b/workflows/rnaseq.nf index 4a13cb3ee..09de86260 100755 --- a/workflows/rnaseq.nf +++ b/workflows/rnaseq.nf @@ -648,7 +648,7 @@ workflow RNASEQ { if (!params.skip_alignment && !params.skip_stringtie) { STRINGTIE_STRINGTIE ( ch_genome_bam, - PREPARE_GENOME.out.gtf_for_stringtie + PREPARE_GENOME.out.gtf_with_transcript_ids ) ch_versions = ch_versions.mix(STRINGTIE_STRINGTIE.out.versions.first()) } @@ -798,7 +798,7 @@ workflow RNASEQ { ch_filtered_reads, PREPARE_GENOME.out.salmon_index, ch_dummy_file, - PREPARE_GENOME.out.gtf, + PREPARE_GENOME.out.gtf_with_transcript_ids, false, params.salmon_quant_libtype ?: '' ) From 7908ca3e01c6faa0ceaeba1a4773c55147dc3fd7 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 9 Nov 2023 13:02:23 +0000 Subject: [PATCH 111/184] Reverse test profile change --- conf/test.config | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/conf/test.config b/conf/test.config index f78e4e567..d0430eacb 100644 --- a/conf/test.config +++ b/conf/test.config @@ -21,7 +21,7 @@ params { // Input data - input = "/Users/jonathan.manning/projects/rnaseq/samplesheet_test.csv" + input = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/samplesheet/v3.10/samplesheet_test.csv" // Genome references fasta = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/reference/genome.fasta" From f791e8af6a9d35acd06636bb15e29a2b5b1dca14 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 9 Nov 2023 13:58:31 +0000 Subject: [PATCH 112/184] misc fixes --- ...filter_gtf_for_genes_in_genome.py => filter_gtf.py} | 0 modules/local/gtf_gene_filter/main.nf | 6 +++--- subworkflows/local/prepare_genome/main.nf | 10 +++++----- 3 files changed, 8 insertions(+), 8 deletions(-) rename bin/{filter_gtf_for_genes_in_genome.py => filter_gtf.py} (100%) diff --git a/bin/filter_gtf_for_genes_in_genome.py b/bin/filter_gtf.py similarity index 100% rename from bin/filter_gtf_for_genes_in_genome.py rename to bin/filter_gtf.py diff --git a/modules/local/gtf_gene_filter/main.nf b/modules/local/gtf_gene_filter/main.nf index b4e0578d7..c12cf4116 100644 --- a/modules/local/gtf_gene_filter/main.nf +++ b/modules/local/gtf_gene_filter/main.nf @@ -1,4 +1,4 @@ -process GTF_GENE_FILTER { +process GTF_FILTER { tag "$fasta" conda "conda-forge::python=3.9.5" @@ -20,10 +20,10 @@ process GTF_GENE_FILTER { script: // filter_gtf_for_genes_in_genome.py is bundled with the pipeline, in nf-core/rnaseq/bin/ """ - filter_gtf_for_genes_in_genome.py \\ + filter_gtf.py \\ --gtf $gtf \\ --fasta $fasta \\ - -o ${fasta.baseName}_genes.gtf + --prefix ${fasta.baseName} cat <<-END_VERSIONS > versions.yml "${task.process}": diff --git a/subworkflows/local/prepare_genome/main.nf b/subworkflows/local/prepare_genome/main.nf index cec93e86b..906590402 100644 --- a/subworkflows/local/prepare_genome/main.nf +++ b/subworkflows/local/prepare_genome/main.nf @@ -28,7 +28,7 @@ include { RSEM_PREPAREREFERENCE as MAKE_TRANSCRIPTS_FASTA } from '../../.. include { PREPROCESS_TRANSCRIPTS_FASTA_GENCODE } from '../../../modules/local/preprocess_transcripts_fasta_gencode' include { GTF2BED } from '../../../modules/local/gtf2bed' include { CAT_ADDITIONAL_FASTA } from '../../../modules/local/cat_additional_fasta' -include { GTF_GENE_FILTER } from '../../../modules/local/gtf_gene_filter' +include { GTF_FILTER } from '../../../modules/local/gtf_filter' include { STAR_GENOMEGENERATE_IGENOMES } from '../../../modules/local/star_genomegenerate_igenomes' workflow PREPARE_GENOME { @@ -89,9 +89,9 @@ workflow PREPARE_GENOME { // // Apply filtering we may need for GTFs // - GTF_GENE_FILTER ( ch_fasta, ch_gtf ) - ch_gtf_with_transcript_ids = GTF_GENE_FILTER.out.transcript_id_gtf - ch_gtf_genome = GTF_GENE_FILTER.out.genome_gtf + GTF_FILTER ( ch_fasta, ch_gtf ) + ch_gtf_with_transcript_ids = GTF_FILTER.out.transcript_id_gtf + ch_gtf_genome = GTF_FILTER.out.genome_gtf // // Uncompress additional fasta file and concatenate with reference fasta and gtf files @@ -141,7 +141,7 @@ workflow PREPARE_GENOME { } } else { ch_transcript_fasta = MAKE_TRANSCRIPTS_FASTA ( ch_fasta, ch_gtf_genome ).transcript_fasta - ch_versions = ch_versions.mix(GTF_GENE_FILTER.out.versions) + ch_versions = ch_versions.mix(GTF_FILTER.out.versions) ch_versions = ch_versions.mix(MAKE_TRANSCRIPTS_FASTA.out.versions) } From 19a72f6f168f004e5c6cd15fd5367a86ecfb0344 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 9 Nov 2023 14:00:34 +0000 Subject: [PATCH 113/184] Blackify --- bin/filter_gtf.py | 17 +++++++++++------ 1 file changed, 11 insertions(+), 6 deletions(-) diff --git a/bin/filter_gtf.py b/bin/filter_gtf.py index cb2002a02..055bc5197 100755 --- a/bin/filter_gtf.py +++ b/bin/filter_gtf.py @@ -9,13 +9,15 @@ logger = logging.getLogger(__file__) logger.setLevel(logging.INFO) + def is_header(line: str) -> bool: """Returns True if the given line is a header line in a FASTA file.""" return line[0] == ">" + def extract_fasta_seq_names(fasta_name: str) -> set: """Extracts the sequence names from a FASTA file. - + modified from Brent Pedersen Correct Way To Parse A Fasta File In Python given a fasta file. yield tuples of header, sequence @@ -42,6 +44,7 @@ def extract_fasta_seq_names(fasta_name: str) -> set: headerStr = line[1:].strip().split()[0] yield headerStr + def extract_genes_in_genome(fasta: str, gtf_in: str, prefix: str) -> None: """Extracts the genes in the genome from a GTF file. @@ -50,7 +53,7 @@ def extract_genes_in_genome(fasta: str, gtf_in: str, prefix: str) -> None: gtf_in: The path to the input GTF file. prefix: Prefix for output GTF """ - gtf_out = prefix + '_in_genome.gtf' + gtf_out = prefix + "_in_genome.gtf" seq_names_in_genome = set(extract_fasta_seq_names(fasta)) logger.info("Extracted chromosome sequence names from : %s" % fasta) logger.info("All chromosome names: " + ", ".join(sorted(x for x in seq_names_in_genome))) @@ -74,6 +77,7 @@ def extract_genes_in_genome(fasta: str, gtf_in: str, prefix: str) -> None: logger.info("Wrote matching lines to %s" % gtf_out) + def remove_features_without_transcript_id(gtf_in, prefix): """ Removes gene rows with absent or empty transcript_id attributes from a GTF file. @@ -82,13 +86,14 @@ def remove_features_without_transcript_id(gtf_in, prefix): gtf_in: Path to the input GTF file. prefix: Path to the output GTF file. """ - gtf_out = prefix + '_with_transcript_ids.gtf' + gtf_out = prefix + "_with_transcript_ids.gtf" with open(gtf_in, "r") as f_in, open(gtf_out, "w") as f_out: for line in f_in: - transcript_id = line.split("\t")[8].split(" transcript_id ")[1].split(";")[0].replace('"', '') - if transcript_id and transcript_id.isalnum(): - f_out.write(line) + transcript_id = line.split("\t")[8].split(" transcript_id ")[1].split(";")[0].replace('"', "") + if transcript_id and transcript_id.isalnum(): + f_out.write(line) + if __name__ == "__main__": parser = argparse.ArgumentParser(description="""Filter GTF for various reasons""") From 2cfea9d97fe8cce5f0a7eb618f81be2e350021c5 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 9 Nov 2023 14:03:59 +0000 Subject: [PATCH 114/184] move module to right path --- modules/local/{gtf_gene_filter => gtf_filter}/main.nf | 0 1 file changed, 0 insertions(+), 0 deletions(-) rename modules/local/{gtf_gene_filter => gtf_filter}/main.nf (100%) diff --git a/modules/local/gtf_gene_filter/main.nf b/modules/local/gtf_filter/main.nf similarity index 100% rename from modules/local/gtf_gene_filter/main.nf rename to modules/local/gtf_filter/main.nf From 9b4e1406391c53c9e1fd16e611b84f4aaf79d891 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 9 Nov 2023 14:05:25 +0000 Subject: [PATCH 115/184] update changelog --- CHANGELOG.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index e64510937..a850a5353 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -28,7 +28,7 @@ Thank you to everyone else that has contributed by reporting bugs, enhancements - [PR #1083](https://github.com/nf-core/rnaseq/pull/1083) - Move local modules and subworkflows to subfolders - [PR #1088](https://github.com/nf-core/rnaseq/pull/1088) - Updates contributing and code of conduct documents with nf-core template 2.10 - [PR #1091](https://github.com/nf-core/rnaseq/pull/1091) - Reorganise parameters in schema for better usability -- [PR #1107](https://github.com/nf-core/rnaseq/pull/1107) - Prepare a GTF for stringtie +- [PR #1107](https://github.com/nf-core/rnaseq/pull/1107) - Expand GTF filtering to remove rows with empty transcript ID when required ### Software dependencies From ec6f80e76694b44056a312ff6baa87531d0d426e Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 9 Nov 2023 14:26:18 +0000 Subject: [PATCH 116/184] Correct selector --- conf/modules.config | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/conf/modules.config b/conf/modules.config index eaa35f259..abaf220f9 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -116,7 +116,7 @@ process { ] } - withName: 'GTF_GENE_FILTER' { + withName: 'GTF_FILTER' { publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, From 11f979a605b82f2ac971b1e7fff9f883b787d587 Mon Sep 17 00:00:00 2001 From: maxulysse Date: Thu, 9 Nov 2023 15:30:17 +0100 Subject: [PATCH 117/184] enabled -> saveAs --- conf/modules.config | 108 +++++++++++++++----------------------------- 1 file changed, 37 insertions(+), 71 deletions(-) diff --git a/conf/modules.config b/conf/modules.config index eaa35f259..a2b3ca76a 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -42,7 +42,7 @@ process { path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - enabled: params.save_reference + saveAs: { params.save_reference ? it : null } ] } @@ -55,7 +55,7 @@ process { path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - enabled: params.save_reference + saveAs: { params.save_reference ? it : null } ] } @@ -65,7 +65,7 @@ process { path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - enabled: params.save_reference + saveAs: { params.save_reference ? it : null } ] } @@ -74,7 +74,7 @@ process { path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - enabled: params.save_reference + saveAs: { params.save_reference ? it : null } ] } @@ -84,7 +84,7 @@ process { path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - enabled: params.save_reference + saveAs: { params.save_reference ? it : null } ] } @@ -94,7 +94,7 @@ process { path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - enabled: params.save_reference + saveAs: { params.save_reference ? it : null } ] } @@ -103,7 +103,7 @@ process { path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - enabled: params.save_reference + saveAs: { params.save_reference ? it : null } ] } @@ -112,7 +112,7 @@ process { path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - enabled: params.save_reference + saveAs: { params.save_reference ? it : null } ] } @@ -121,7 +121,7 @@ process { path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - enabled: params.save_reference + saveAs: { params.save_reference ? it : null } ] } @@ -130,7 +130,7 @@ process { path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - enabled: params.save_reference + saveAs: { params.save_reference ? it : null } ] } @@ -139,7 +139,7 @@ process { path: { "${params.outdir}/fastq" }, mode: params.publish_dir_mode, saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - enabled: params.save_merged_fastq + saveAs: { params.save_merged_fastq ? it : null } ] } } @@ -151,8 +151,7 @@ if (!params.skip_bbsplit && params.bbsplit_fasta_list) { publishDir = [ path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - enabled: params.save_reference + saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } ] } } @@ -236,7 +235,7 @@ if (!params.skip_trimming) { path: { "${params.outdir}/${params.trimmer}" }, mode: params.publish_dir_mode, pattern: "*.fq.gz", - enabled: params.save_trimmed + saveAs: { params.save_trimmed ? it : null } ], [ path: { "${params.outdir}/${params.trimmer}" }, @@ -267,7 +266,7 @@ if (!params.skip_trimming) { path: { "${params.outdir}/${params.trimmer}" }, mode: params.publish_dir_mode, pattern: "*.fastq.gz", - enabled: params.save_trimmed + saveAs: { params.save_trimmed ? it : null } ] ] } @@ -294,7 +293,7 @@ if (params.with_umi && !params.skip_umi_extract) { path: { "${params.outdir}/umitools" }, mode: params.publish_dir_mode, pattern: "*.fastq.gz", - enabled: params.save_umi_intermeds + saveAs: { params.save_umi_intermeds ? it : null } ] ] } @@ -319,7 +318,7 @@ if (!params.skip_bbsplit) { path: { "${params.outdir}/bbsplit" }, mode: params.publish_dir_mode, pattern: '*.fastq.gz', - enabled: params.save_bbsplit_reads + saveAs: { params.save_bbsplit_reads ? it : null } ] ] } @@ -340,7 +339,7 @@ if (params.remove_ribo_rna) { path: { "${params.outdir}/sortmerna" }, mode: params.publish_dir_mode, pattern: "*.fastq.gz", - enabled: params.save_non_ribo_reads + saveAs: { params.save_non_ribo_reads ? it : null } ] ] } @@ -368,11 +367,9 @@ if (!params.skip_alignment) { path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, pattern: "*.bam", - enabled: ( ['star_salmon','hisat2'].contains(params.aligner) && - ( params.save_align_intermeds || - ( !params.with_umi && params.skip_markduplicates ) - ) - ) || params.save_align_intermeds || params.skip_markduplicates + saveAs: { ( ['star_salmon','hisat2'].contains(params.aligner) && + ( params.save_align_intermeds || ( !params.with_umi && params.skip_markduplicates ) ) + ) || params.save_align_intermeds || params.skip_markduplicates ? it : null } ] } @@ -382,11 +379,9 @@ if (!params.skip_alignment) { path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, pattern: "*.{bai,csi}", - enabled: ( ['star_salmon','hisat2'].contains(params.aligner) && - ( params.save_align_intermeds || - ( !params.with_umi && params.skip_markduplicates ) - ) - ) || params.save_align_intermeds || params.skip_markduplicates + saveAs: { ( ['star_salmon','hisat2'].contains(params.aligner) && + ( params.save_align_intermeds || ( !params.with_umi && params.skip_markduplicates ) ) + ) || params.save_align_intermeds || params.skip_markduplicates ? it : null } ] } } @@ -450,11 +445,7 @@ if (!params.skip_alignment) { path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, pattern: '*.bam', - enabled: ( - params.save_align_intermeds || - params.with_umi || - params.save_umi_intermeds - ) + saveAs: { params.save_align_intermeds || params.with_umi || params.save_umi_intermeds } ] ] } @@ -466,11 +457,7 @@ if (!params.skip_alignment) { path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, pattern: '*.{bai,csi}', - enabled: ( - params.save_align_intermeds || - params.with_umi || - params.save_umi_intermeds - ) + saveAs: { params.save_align_intermeds || params.with_umi || params.save_umi_intermeds } ] } @@ -579,13 +566,13 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, pattern: '*.bam', - enabled: params.save_align_intermeds + saveAs: { params.save_align_intermeds ? it : null } ], [ path: { "${params.outdir}/${params.aligner}/unmapped" }, mode: params.publish_dir_mode, pattern: '*.fastq.gz', - enabled: params.save_unaligned + saveAs: { params.save_unaligned ? it : null } ] ] } @@ -633,10 +620,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, pattern: '*.bam', - enabled: ( - params.save_align_intermeds || - params.save_umi_intermeds - ) + saveAs: { params.save_align_intermeds || params.save_umi_intermeds ? it : null } ] } @@ -652,10 +636,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, pattern: '*.bam', - enabled: ( - params.save_align_intermeds || - params.save_umi_intermeds - ) + saveAs: { params.save_align_intermeds || params.save_umi_intermeds ? it : null } ] ] } @@ -666,10 +647,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, pattern: '*.bam', - enabled: ( - params.save_align_intermeds || - params.save_umi_intermeds - ) + saveAs: { params.save_align_intermeds || params.save_umi_intermeds ? it : null } ] } @@ -678,10 +656,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, pattern: '*.bai', - enabled: ( - params.save_align_intermeds || - params.save_umi_intermeds - ) + saveAs: { params.save_align_intermeds || params.save_umi_intermeds ? it : null } ] } @@ -691,10 +666,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { path: { "${params.outdir}/${params.aligner}/samtools_stats" }, mode: params.publish_dir_mode, pattern: '*.{stats,flagstat,idxstats}', - enabled: ( - params.save_align_intermeds || - params.save_umi_intermeds - ) + saveAs: { params.save_align_intermeds || params.save_umi_intermeds ? it : null } ] } @@ -715,10 +687,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, pattern: '*.bam', - enabled: ( - params.save_align_intermeds || - params.save_umi_intermeds - ) + saveAs: { params.save_align_intermeds || params.save_umi_intermeds ? it : null } ] ] } @@ -728,10 +697,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, pattern: '*.bai', - enabled: ( - params.save_align_intermeds || - params.save_umi_intermeds - ) + saveAs: { params.save_align_intermeds || params.save_umi_intermeds ? it : null } ] } @@ -791,7 +757,7 @@ if (!params.skip_alignment && params.aligner == 'star_rsem') { path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, pattern: "*.bam", - enabled: params.save_align_intermeds + saveAs: { params.save_align_intermeds ? it : null } ], [ path: { "${params.outdir}/${params.aligner}/log" }, @@ -849,13 +815,13 @@ if (!params.skip_alignment && params.aligner == 'hisat2') { path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, pattern: '*.bam', - enabled: params.save_align_intermeds + saveAs: { params.save_align_intermeds ? it : null } ], [ path: { "${params.outdir}/${params.aligner}/unmapped" }, mode: params.publish_dir_mode, pattern: '*.fastq.gz', - enabled: params.save_unaligned + saveAs: { params.save_unaligned ? it : null } ] ] } From 7eaad5221fb032a04934c3612d7755278023a004 Mon Sep 17 00:00:00 2001 From: maxulysse Date: Thu, 9 Nov 2023 15:39:58 +0100 Subject: [PATCH 118/184] add closure to ext directive with params --- conf/modules.config | 42 +++++++++++++++++++++--------------------- 1 file changed, 21 insertions(+), 21 deletions(-) diff --git a/conf/modules.config b/conf/modules.config index a2b3ca76a..17a3d71af 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -250,7 +250,7 @@ if (!params.skip_trimming) { if (params.trimmer == 'fastp') { process { withName: '.*:FASTQ_FASTQC_UMITOOLS_FASTP:FASTP' { - ext.args = params.extra_fastp_args ?: '' + ext.args = { params.extra_fastp_args ?: '' } publishDir = [ [ path: { "${params.outdir}/${params.trimmer}" }, @@ -277,12 +277,12 @@ if (!params.skip_trimming) { if (params.with_umi && !params.skip_umi_extract) { process { withName: 'UMITOOLS_EXTRACT' { - ext.args = [ + ext.args = { [ params.umitools_extract_method ? "--extract-method=${params.umitools_extract_method}" : '', params.umitools_bc_pattern ? "--bc-pattern='${params.umitools_bc_pattern}'" : '', params.umitools_bc_pattern2 ? "--bc-pattern2='${params.umitools_bc_pattern2}'" : '', params.umitools_umi_separator ? "--umi-separator='${params.umitools_umi_separator}'" : '' - ].join(' ').trim() + ].join(' ').trim() } publishDir = [ [ path: { "${params.outdir}/umitools" }, @@ -374,7 +374,7 @@ if (!params.skip_alignment) { } withName: 'NFCORE_RNASEQ:RNASEQ:.*:BAM_SORT_STATS_SAMTOOLS:SAMTOOLS_INDEX' { - ext.args = params.bam_csi_index ? '-c' : '' + ext.args = { params.bam_csi_index ? '-c' : '' } publishDir = [ path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, @@ -406,7 +406,7 @@ if (!params.skip_alignment) { } withName: '.*:BAM_MARKDUPLICATES_PICARD:SAMTOOLS_INDEX' { - ext.args = params.bam_csi_index ? '-c' : '' + ext.args = { params.bam_csi_index ? '-c' : '' } ext.prefix = { "${meta.id}.markdup.sorted" } publishDir = [ path: { "${params.outdir}/${params.aligner}" }, @@ -451,7 +451,7 @@ if (!params.skip_alignment) { } withName: '.*:BAM_DEDUP_STATS_SAMTOOLS_UMITOOLS_GENOME:SAMTOOLS_INDEX' { - ext.args = params.bam_csi_index ? '-c' : '' + ext.args = { params.bam_csi_index ? '-c' : '' } ext.prefix = { "${meta.id}.umi_dedup.sorted" } publishDir = [ path: { "${params.outdir}/${params.aligner}" }, @@ -521,10 +521,10 @@ if (!params.skip_alignment) { if (!params.skip_stringtie) { process { withName: 'STRINGTIE_STRINGTIE' { - ext.args = [ + ext.args = { [ '-v', params.stringtie_ignore_gtf ? '' : '-e' - ].join(' ').trim() + ].join(' ').trim() } publishDir = [ path: { "${params.outdir}/${params.aligner}/stringtie" }, mode: params.publish_dir_mode, @@ -542,7 +542,7 @@ if (!params.skip_alignment) { if (!params.skip_alignment && params.aligner == 'star_salmon') { process { withName: '.*:ALIGN_STAR:STAR_ALIGN|.*:ALIGN_STAR:STAR_ALIGN_IGENOMES' { - ext.args = [ + ext.args = { [ '--quantMode TranscriptomeSAM', '--twopassMode Basic', '--outSAMtype BAM Unsorted', @@ -555,7 +555,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { '--outSAMstrandField intronMotif', params.save_unaligned ? '--outReadsUnmapped Fastx' : '', params.extra_star_align_args ? params.extra_star_align_args.split("\\s(?=--)") : '' - ].flatten().unique(false).join(' ').trim() + ].flatten().unique(false).join(' ').trim() } publishDir = [ [ path: { "${params.outdir}/${params.aligner}/log" }, @@ -578,7 +578,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { } withName: '.*:QUANTIFY_STAR_SALMON:SALMON_QUANT' { - ext.args = params.extra_salmon_quant_args ?: '' + ext.args = { params.extra_salmon_quant_args ?: '' } publishDir = [ path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, @@ -715,13 +715,13 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { if (!params.skip_qc & !params.skip_deseq2_qc) { process { withName: 'DESEQ2_QC_STAR_SALMON' { - ext.args = [ + ext.args = { [ "--id_col 1", "--sample_suffix ''", "--outprefix deseq2", "--count_col 3", params.deseq2_vst ? '--vst TRUE' : '' - ].join(' ').trim() + ].join(' ').trim() } ext.args2 = 'star_salmon' publishDir = [ path: { "${params.outdir}/${params.aligner}/deseq2_qc" }, @@ -779,13 +779,13 @@ if (!params.skip_alignment && params.aligner == 'star_rsem') { if (!params.skip_qc & !params.skip_deseq2_qc) { process { withName: 'DESEQ2_QC_RSEM' { - ext.args = [ + ext.args = { [ "--id_col 1", "--sample_suffix ''", "--outprefix deseq2", "--count_col 3", params.deseq2_vst ? '--vst TRUE' : '' - ].join(' ').trim() + ].join(' ').trim() } ext.args2 = 'star_rsem' publishDir = [ path: { "${params.outdir}/${params.aligner}/deseq2_qc" }, @@ -902,11 +902,11 @@ if (!params.skip_alignment && !params.skip_qc) { if (!params.skip_biotype_qc && params.featurecounts_group_type) { process { withName: 'SUBREAD_FEATURECOUNTS' { - ext.args = [ + ext.args = { [ '-B -C', params.gencode ? "-g gene_type" : "-g $params.featurecounts_group_type", "-t $params.featurecounts_feature_type" - ].join(' ').trim() + ].join(' ').trim() } publishDir = [ path: { "${params.outdir}/${params.aligner}/featurecounts" }, mode: params.publish_dir_mode, @@ -1078,7 +1078,7 @@ if (!params.skip_alignment && !params.skip_qc) { if (!params.skip_multiqc) { process { withName: 'MULTIQC' { - ext.args = params.multiqc_title ? "--title \"$params.multiqc_title\"" : '' + ext.args = { params.multiqc_title ? "--title \"$params.multiqc_title\"" : '' } publishDir = [ path: { [ "${params.outdir}/multiqc", @@ -1098,7 +1098,7 @@ if (!params.skip_multiqc) { if (!params.skip_pseudo_alignment && params.pseudo_aligner == 'salmon') { process { withName: '.*:QUANTIFY_SALMON:SALMON_QUANT' { - ext.args = params.extra_salmon_quant_args ?: '' + ext.args = { params.extra_salmon_quant_args ?: '' } publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}" }, mode: params.publish_dir_mode, @@ -1134,13 +1134,13 @@ if (!params.skip_pseudo_alignment && params.pseudo_aligner == 'salmon') { if (!params.skip_qc & !params.skip_deseq2_qc) { process { withName: 'DESEQ2_QC_SALMON' { - ext.args = [ + ext.args = { [ "--id_col 1", "--sample_suffix ''", "--outprefix deseq2", "--count_col 3", params.deseq2_vst ? '--vst TRUE' : '' - ].join(' ').trim() + ].join(' ').trim() } ext.args2 = 'salmon' publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}/deseq2_qc" }, From 71efea24cbfe960d12dde6cdf5fe19b1182f9b52 Mon Sep 17 00:00:00 2001 From: maxulysse Date: Thu, 9 Nov 2023 15:44:49 +0100 Subject: [PATCH 119/184] remove double saveAs directives --- conf/modules.config | 33 +++++++++++---------------------- 1 file changed, 11 insertions(+), 22 deletions(-) diff --git a/conf/modules.config b/conf/modules.config index 17a3d71af..30495dd1a 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -41,8 +41,7 @@ process { publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } ] } @@ -54,8 +53,7 @@ process { publishDir = [ path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } ] } @@ -64,8 +62,7 @@ process { publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } ] } @@ -73,8 +70,7 @@ process { publishDir = [ path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } ] } @@ -83,8 +79,7 @@ process { publishDir = [ path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } ] } @@ -93,8 +88,7 @@ process { publishDir = [ path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } ] } @@ -102,8 +96,7 @@ process { publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } ] } @@ -111,8 +104,7 @@ process { publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } ] } @@ -120,8 +112,7 @@ process { publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } ] } @@ -129,8 +120,7 @@ process { publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } ] } @@ -138,8 +128,7 @@ process { publishDir = [ path: { "${params.outdir}/fastq" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - saveAs: { params.save_merged_fastq ? it : null } + saveAs: { filename -> params.save_merged_fastq ? filename.equals('versions.yml') ? null : filename : null } ] } } From 25324ba729fd692da432368f43b7eeafc39d27b4 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 9 Nov 2023 14:45:48 +0000 Subject: [PATCH 120/184] Make gtf filtering dependent on gtf --- subworkflows/local/prepare_genome/main.nf | 46 ++++++++++++----------- 1 file changed, 24 insertions(+), 22 deletions(-) diff --git a/subworkflows/local/prepare_genome/main.nf b/subworkflows/local/prepare_genome/main.nf index 906590402..d82f65b07 100644 --- a/subworkflows/local/prepare_genome/main.nf +++ b/subworkflows/local/prepare_genome/main.nf @@ -68,30 +68,32 @@ workflow PREPARE_GENOME { // // Uncompress GTF annotation file or create from GFF3 if required // - if (gtf) { - if (gtf.endsWith('.gz')) { - ch_gtf = GUNZIP_GTF ( [ [:], gtf ] ).gunzip.map { it[1] } - ch_versions = ch_versions.mix(GUNZIP_GTF.out.versions) - } else { - ch_gtf = Channel.value(file(gtf)) - } - } else if (gff) { - if (gff.endsWith('.gz')) { - ch_gff = GUNZIP_GFF ( [ [:], gff ] ).gunzip.map { it[1] } - ch_versions = ch_versions.mix(GUNZIP_GFF.out.versions) - } else { - ch_gff = Channel.value(file(gff)) + if (gtf || gff) { + if (gtf) { + if (gtf.endsWith('.gz')) { + ch_gtf = GUNZIP_GTF ( [ [:], gtf ] ).gunzip.map { it[1] } + ch_versions = ch_versions.mix(GUNZIP_GTF.out.versions) + } else { + ch_gtf = Channel.value(file(gtf)) + } + } else if (gff) { + if (gff.endsWith('.gz')) { + ch_gff = GUNZIP_GFF ( [ [:], gff ] ).gunzip.map { it[1] } + ch_versions = ch_versions.mix(GUNZIP_GFF.out.versions) + } else { + ch_gff = Channel.value(file(gff)) + } + ch_gtf = GFFREAD ( ch_gff ).gtf + ch_versions = ch_versions.mix(GFFREAD.out.versions) } - ch_gtf = GFFREAD ( ch_gff ).gtf - ch_versions = ch_versions.mix(GFFREAD.out.versions) - } - // - // Apply filtering we may need for GTFs - // - GTF_FILTER ( ch_fasta, ch_gtf ) - ch_gtf_with_transcript_ids = GTF_FILTER.out.transcript_id_gtf - ch_gtf_genome = GTF_FILTER.out.genome_gtf + // + // Apply filtering we may need for GTFs + // + GTF_FILTER ( ch_fasta, ch_gtf ) + ch_gtf_with_transcript_ids = GTF_FILTER.out.transcript_id_gtf + ch_gtf_genome = GTF_FILTER.out.genome_gtf + } // // Uncompress additional fasta file and concatenate with reference fasta and gtf files From 04f8bebd523cfe6b917b7093122d87479b6548a3 Mon Sep 17 00:00:00 2001 From: maxulysse Date: Thu, 9 Nov 2023 15:46:15 +0100 Subject: [PATCH 121/184] no need for anything in publishdir with enabled set to false --- conf/modules.config | 9 --------- 1 file changed, 9 deletions(-) diff --git a/conf/modules.config b/conf/modules.config index 30495dd1a..70b08e5c6 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -155,9 +155,6 @@ process { ext.args = '--record-count 1000000 --seed 1' ext.prefix = { "${meta.id}.subsampled" } publishDir = [ - path: { "${params.outdir}/sample_fastq/fastq" }, - mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, enabled: false ] } @@ -165,9 +162,6 @@ process { withName: '.*:FASTQ_SUBSAMPLE_FQ_SALMON:SALMON_QUANT' { ext.args = '--skipQuant' publishDir = [ - path: { "${params.outdir}/sample_fastq/salmon" }, - mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') || filename.endsWith('_meta_info.json') ? null : filename }, enabled: false ] } @@ -466,7 +460,6 @@ if (!params.skip_alignment) { withName: 'BEDTOOLS_GENOMECOV' { ext.args = '-split -du' publishDir = [ - path: { "${params.outdir}/bedtools/${meta.id}" }, enabled: false ] } @@ -474,7 +467,6 @@ if (!params.skip_alignment) { withName: '.*:BEDGRAPH_BEDCLIP_BEDGRAPHTOBIGWIG_FORWARD:UCSC_BEDCLIP' { ext.prefix = { "${meta.id}.clip.forward" } publishDir = [ - path: { "${params.outdir}/${params.aligner}" }, enabled: false ] } @@ -491,7 +483,6 @@ if (!params.skip_alignment) { withName: '.*:BEDGRAPH_BEDCLIP_BEDGRAPHTOBIGWIG_REVERSE:UCSC_BEDCLIP' { ext.prefix = { "${meta.id}.clip.reverse" } publishDir = [ - path: { "${params.outdir}/${params.aligner}" }, enabled: false ] } From 47e219ab4f6e69d1f81fe7637428f2bf3d5a9e28 Mon Sep 17 00:00:00 2001 From: maxulysse Date: Thu, 9 Nov 2023 16:05:49 +0100 Subject: [PATCH 122/184] better usage of pattern --- conf/modules.config | 80 ++++++++++++++++++++++++++------------------- 1 file changed, 46 insertions(+), 34 deletions(-) diff --git a/conf/modules.config b/conf/modules.config index 70b08e5c6..68beaa68a 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -41,7 +41,8 @@ process { publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, - saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } + pattern: "{!versions.yml}", + saveAs: { params.save_reference ? it : null } ] } @@ -53,7 +54,8 @@ process { publishDir = [ path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, - saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } + pattern: "{!versions.yml}", + saveAs: { params.save_reference ? it : null } ] } @@ -62,7 +64,8 @@ process { publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, - saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } + pattern: "{!versions.yml}", + saveAs: { params.save_reference ? it : null } ] } @@ -70,7 +73,8 @@ process { publishDir = [ path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, - saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } + pattern: "{!versions.yml}", + saveAs: { params.save_reference ? it : null } ] } @@ -79,7 +83,8 @@ process { publishDir = [ path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, - saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } + pattern: "{!versions.yml}", + saveAs: { params.save_reference ? it : null } ] } @@ -88,7 +93,8 @@ process { publishDir = [ path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, - saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } + pattern: "{!versions.yml}", + saveAs: { params.save_reference ? it : null } ] } @@ -96,7 +102,8 @@ process { publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, - saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } + pattern: "{!versions.yml}", + saveAs: { params.save_reference ? it : null } ] } @@ -104,7 +111,8 @@ process { publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, - saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } + pattern: "{!versions.yml}", + saveAs: { params.save_reference ? it : null } ] } @@ -112,7 +120,8 @@ process { publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, - saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } + pattern: "{!versions.yml}", + saveAs: { params.save_reference ? it : null } ] } @@ -120,7 +129,8 @@ process { publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, - saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } + pattern: "{!versions.yml}", + saveAs: { params.save_reference ? it : null } ] } @@ -128,7 +138,8 @@ process { publishDir = [ path: { "${params.outdir}/fastq" }, mode: params.publish_dir_mode, - saveAs: { filename -> params.save_merged_fastq ? filename.equals('versions.yml') ? null : filename : null } + pattern: "{!versions.yml}", + saveAs: { params.save_merged_fastq ? it : null } ] } } @@ -140,7 +151,8 @@ if (!params.skip_bbsplit && params.bbsplit_fasta_list) { publishDir = [ path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, - saveAs: { filename -> params.save_reference ? filename.equals('versions.yml') ? null : filename : null } + pattern: "{!versions.yml}", + saveAs: { params.save_reference ? it : null } ] } } @@ -191,7 +203,7 @@ if (!(params.skip_fastqc || params.skip_qc)) { publishDir = [ path: { "${params.outdir}/${params.trimmer}/fastqc" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ] } } @@ -476,7 +488,7 @@ if (!params.skip_alignment) { publishDir = [ path: { "${params.outdir}/${params.aligner}/bigwig" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ] } @@ -492,7 +504,7 @@ if (!params.skip_alignment) { publishDir = [ path: { "${params.outdir}/${params.aligner}/bigwig" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ] } } @@ -508,7 +520,7 @@ if (!params.skip_alignment) { publishDir = [ path: { "${params.outdir}/${params.aligner}/stringtie" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ] } } @@ -562,7 +574,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { publishDir = [ path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') || filename.endsWith('_meta_info.json') ? null : filename } + pattern: "{!versions.yml,!*_meta_info.json}" ] } @@ -570,7 +582,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { publishDir = [ path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ] } @@ -578,7 +590,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { publishDir = [ path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ] } @@ -586,7 +598,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { publishDir = [ path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ] } } @@ -751,7 +763,7 @@ if (!params.skip_alignment && params.aligner == 'star_rsem') { publishDir = [ path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ] } } @@ -839,7 +851,7 @@ if (!params.skip_alignment && !params.skip_qc) { publishDir = [ path: { "${params.outdir}/${params.aligner}/qualimap" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ] } } @@ -890,7 +902,7 @@ if (!params.skip_alignment && !params.skip_qc) { publishDir = [ path: { "${params.outdir}/${params.aligner}/featurecounts" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ] } @@ -898,7 +910,7 @@ if (!params.skip_alignment && !params.skip_qc) { publishDir = [ path: { "${params.outdir}/${params.aligner}/featurecounts" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ] } } @@ -910,7 +922,7 @@ if (!params.skip_alignment && !params.skip_qc) { publishDir = [ path: { "${params.outdir}/${params.aligner}/rseqc/bam_stat" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ] } } @@ -922,7 +934,7 @@ if (!params.skip_alignment && !params.skip_qc) { publishDir = [ path: { "${params.outdir}/${params.aligner}/rseqc/infer_experiment" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ] } } @@ -1011,7 +1023,7 @@ if (!params.skip_alignment && !params.skip_qc) { publishDir = [ path: { "${params.outdir}/${params.aligner}/rseqc/read_distribution" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ] } } @@ -1025,7 +1037,7 @@ if (!params.skip_alignment && !params.skip_qc) { path: { "${params.outdir}/${params.aligner}/rseqc/inner_distance/txt" }, mode: params.publish_dir_mode, pattern: '*.txt', - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ], [ path: { "${params.outdir}/${params.aligner}/rseqc/inner_distance/pdf" }, @@ -1048,7 +1060,7 @@ if (!params.skip_alignment && !params.skip_qc) { publishDir = [ path: { "${params.outdir}/${params.aligner}/rseqc/tin" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ] } } @@ -1065,7 +1077,7 @@ if (!params.skip_multiqc) { params.skip_alignment? '' : "/${params.aligner}" ].join('') }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ] } } @@ -1082,7 +1094,7 @@ if (!params.skip_pseudo_alignment && params.pseudo_aligner == 'salmon') { publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') || filename.endsWith('_meta_info.json') ? null : filename } + pattern: "{!versions.yml,!*_meta_info.json}" ] } @@ -1090,7 +1102,7 @@ if (!params.skip_pseudo_alignment && params.pseudo_aligner == 'salmon') { publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ] } @@ -1098,7 +1110,7 @@ if (!params.skip_pseudo_alignment && params.pseudo_aligner == 'salmon') { publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ] } @@ -1106,7 +1118,7 @@ if (!params.skip_pseudo_alignment && params.pseudo_aligner == 'salmon') { publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename } + pattern: "{!versions.yml}" ] } } From 26e9c1c28884e1aeea23039f1674e101bf2e71d4 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Fri, 3 Nov 2023 10:31:10 +0000 Subject: [PATCH 123/184] Fix multiqc to avoid imp module error --- modules/local/multiqc/main.nf | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/modules/local/multiqc/main.nf b/modules/local/multiqc/main.nf index 44565a9f7..06fbb9b29 100644 --- a/modules/local/multiqc/main.nf +++ b/modules/local/multiqc/main.nf @@ -1,10 +1,10 @@ process MULTIQC { label 'process_medium' - conda "bioconda::multiqc=1.15" + conda "bioconda::multiqc=1.17" container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? - 'https://depot.galaxyproject.org/singularity/multiqc:1.15--pyhdfd78af_0' : - 'biocontainers/multiqc:1.15--pyhdfd78af_0' }" + 'https://depot.galaxyproject.org/singularity/multiqc:1.17--pyhdfd78af_0' : + 'biocontainers/multiqc:1.17--pyhdfd78af_0' }" input: path multiqc_config From 251e368543cb823cca23c0635fb46655bb5ca9c5 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 9 Nov 2023 15:35:29 +0000 Subject: [PATCH 124/184] Fix transcript_id pattern --- bin/filter_gtf.py | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/bin/filter_gtf.py b/bin/filter_gtf.py index 055bc5197..13b7360ff 100755 --- a/bin/filter_gtf.py +++ b/bin/filter_gtf.py @@ -3,6 +3,7 @@ import logging from itertools import groupby import argparse +import re # Create a logger logging.basicConfig(format="%(name)s - %(asctime)s %(levelname)s: %(message)s") @@ -90,11 +91,10 @@ def remove_features_without_transcript_id(gtf_in, prefix): with open(gtf_in, "r") as f_in, open(gtf_out, "w") as f_out: for line in f_in: - transcript_id = line.split("\t")[8].split(" transcript_id ")[1].split(";")[0].replace('"', "") - if transcript_id and transcript_id.isalnum(): + transcript_id_match = re.search(r'transcript_id "([^"]+)"', line) + if transcript_id_match: f_out.write(line) - if __name__ == "__main__": parser = argparse.ArgumentParser(description="""Filter GTF for various reasons""") parser.add_argument("--gtf", type=str, help="GTF file") From 6a3ea7ee8725b4c1c8dbaf6af31b385f73b733c1 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 9 Nov 2023 15:47:00 +0000 Subject: [PATCH 125/184] blackify --- bin/filter_gtf.py | 1 + 1 file changed, 1 insertion(+) diff --git a/bin/filter_gtf.py b/bin/filter_gtf.py index 13b7360ff..3d4aa1fc9 100755 --- a/bin/filter_gtf.py +++ b/bin/filter_gtf.py @@ -95,6 +95,7 @@ def remove_features_without_transcript_id(gtf_in, prefix): if transcript_id_match: f_out.write(line) + if __name__ == "__main__": parser = argparse.ArgumentParser(description="""Filter GTF for various reasons""") parser.add_argument("--gtf", type=str, help="GTF file") From bb47808c550e3c0324200c1b341141fc2195a35d Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 9 Nov 2023 17:11:24 +0000 Subject: [PATCH 126/184] emit chromosome-filtered gtf from genome preparation --- subworkflows/local/prepare_genome/main.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/subworkflows/local/prepare_genome/main.nf b/subworkflows/local/prepare_genome/main.nf index d82f65b07..337d65615 100644 --- a/subworkflows/local/prepare_genome/main.nf +++ b/subworkflows/local/prepare_genome/main.nf @@ -268,7 +268,7 @@ workflow PREPARE_GENOME { emit: fasta = ch_fasta // channel: path(genome.fasta) - gtf = ch_gtf // channel: path(genome.gtf) + gtf = ch_gtf_genome // channel: path(genome.gtf) fai = ch_fai // channel: path(genome.fai) gene_bed = ch_gene_bed // channel: path(gene.bed) gtf_with_transcript_ids = ch_gtf_with_transcript_ids // channel: path(gtf) From 7c8b39994d5ce6af4fef189b3c3462bd2cf65c35 Mon Sep 17 00:00:00 2001 From: maxulysse Date: Thu, 9 Nov 2023 18:28:43 +0100 Subject: [PATCH 127/184] cannot exclude with glob patterns --- conf/modules.config | 80 +++++++++++++++++++-------------------------- 1 file changed, 34 insertions(+), 46 deletions(-) diff --git a/conf/modules.config b/conf/modules.config index 68beaa68a..d1578b7d9 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -41,8 +41,7 @@ process { publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}", - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> filename.equals('versions.yml') ? null : params.save_reference ? filename : null } ] } @@ -54,8 +53,7 @@ process { publishDir = [ path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}", - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> filename.equals('versions.yml') ? null : params.save_reference ? filename : null } ] } @@ -64,8 +62,7 @@ process { publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}", - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> filename.equals('versions.yml') ? null : params.save_reference ? filename : null } ] } @@ -73,8 +70,7 @@ process { publishDir = [ path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}", - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> filename.equals('versions.yml') ? null : params.save_reference ? filename : null } ] } @@ -83,8 +79,7 @@ process { publishDir = [ path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}", - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> filename.equals('versions.yml') ? null : params.save_reference ? filename : null } ] } @@ -93,8 +88,7 @@ process { publishDir = [ path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}", - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> filename.equals('versions.yml') ? null : params.save_reference ? filename : null } ] } @@ -102,8 +96,7 @@ process { publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}", - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> filename.equals('versions.yml') ? null : params.save_reference ? filename : null } ] } @@ -111,8 +104,7 @@ process { publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}", - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> filename.equals('versions.yml') ? null : params.save_reference ? filename : null } ] } @@ -120,8 +112,7 @@ process { publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}", - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> filename.equals('versions.yml') ? null : params.save_reference ? filename : null } ] } @@ -129,8 +120,7 @@ process { publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}", - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> filename.equals('versions.yml') ? null : params.save_reference ? filename : null } ] } @@ -138,8 +128,7 @@ process { publishDir = [ path: { "${params.outdir}/fastq" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}", - saveAs: { params.save_merged_fastq ? it : null } + saveAs: { filename -> filename.equals('versions.yml') ? null : params.save_merged_fastq ? filename : null } ] } } @@ -151,8 +140,7 @@ if (!params.skip_bbsplit && params.bbsplit_fasta_list) { publishDir = [ path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}", - saveAs: { params.save_reference ? it : null } + saveAs: { filename -> filename.equals('versions.yml') ? null : params.save_reference ? filename : null } ] } } @@ -203,7 +191,7 @@ if (!(params.skip_fastqc || params.skip_qc)) { publishDir = [ path: { "${params.outdir}/${params.trimmer}/fastqc" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } } @@ -488,7 +476,7 @@ if (!params.skip_alignment) { publishDir = [ path: { "${params.outdir}/${params.aligner}/bigwig" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } @@ -504,7 +492,7 @@ if (!params.skip_alignment) { publishDir = [ path: { "${params.outdir}/${params.aligner}/bigwig" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } } @@ -520,7 +508,7 @@ if (!params.skip_alignment) { publishDir = [ path: { "${params.outdir}/${params.aligner}/stringtie" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } } @@ -574,7 +562,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { publishDir = [ path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml,!*_meta_info.json}" + saveAs: { filename -> filename.equals('versions.yml') || filename.endsWith('_meta_info.json') ? null : filename } ] } @@ -582,7 +570,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { publishDir = [ path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } @@ -590,7 +578,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { publishDir = [ path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } @@ -598,7 +586,7 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { publishDir = [ path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } } @@ -763,7 +751,7 @@ if (!params.skip_alignment && params.aligner == 'star_rsem') { publishDir = [ path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } } @@ -851,7 +839,7 @@ if (!params.skip_alignment && !params.skip_qc) { publishDir = [ path: { "${params.outdir}/${params.aligner}/qualimap" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } } @@ -902,7 +890,7 @@ if (!params.skip_alignment && !params.skip_qc) { publishDir = [ path: { "${params.outdir}/${params.aligner}/featurecounts" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } @@ -910,7 +898,7 @@ if (!params.skip_alignment && !params.skip_qc) { publishDir = [ path: { "${params.outdir}/${params.aligner}/featurecounts" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } } @@ -922,7 +910,7 @@ if (!params.skip_alignment && !params.skip_qc) { publishDir = [ path: { "${params.outdir}/${params.aligner}/rseqc/bam_stat" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } } @@ -934,7 +922,7 @@ if (!params.skip_alignment && !params.skip_qc) { publishDir = [ path: { "${params.outdir}/${params.aligner}/rseqc/infer_experiment" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } } @@ -1023,7 +1011,7 @@ if (!params.skip_alignment && !params.skip_qc) { publishDir = [ path: { "${params.outdir}/${params.aligner}/rseqc/read_distribution" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } } @@ -1037,7 +1025,7 @@ if (!params.skip_alignment && !params.skip_qc) { path: { "${params.outdir}/${params.aligner}/rseqc/inner_distance/txt" }, mode: params.publish_dir_mode, pattern: '*.txt', - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ], [ path: { "${params.outdir}/${params.aligner}/rseqc/inner_distance/pdf" }, @@ -1060,7 +1048,7 @@ if (!params.skip_alignment && !params.skip_qc) { publishDir = [ path: { "${params.outdir}/${params.aligner}/rseqc/tin" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } } @@ -1077,7 +1065,7 @@ if (!params.skip_multiqc) { params.skip_alignment? '' : "/${params.aligner}" ].join('') }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } } @@ -1094,7 +1082,7 @@ if (!params.skip_pseudo_alignment && params.pseudo_aligner == 'salmon') { publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml,!*_meta_info.json}" + saveAs: { filename -> filename.equals('versions.yml') || filename.endsWith('_meta_info.json') ? null : filename } ] } @@ -1102,7 +1090,7 @@ if (!params.skip_pseudo_alignment && params.pseudo_aligner == 'salmon') { publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } @@ -1110,7 +1098,7 @@ if (!params.skip_pseudo_alignment && params.pseudo_aligner == 'salmon') { publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } @@ -1118,7 +1106,7 @@ if (!params.skip_pseudo_alignment && params.pseudo_aligner == 'salmon') { publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}" }, mode: params.publish_dir_mode, - pattern: "{!versions.yml}" + saveAs: { filename -> filename.equals('versions.yml') ? null : filename } ] } } From 047f7494a256b6b28aa6f98cfcd4dac18bb2ba93 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 9 Nov 2023 17:31:16 +0000 Subject: [PATCH 128/184] [skip ci] fix changelog --- CHANGELOG.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index e64510937..b38303f49 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -28,7 +28,7 @@ Thank you to everyone else that has contributed by reporting bugs, enhancements - [PR #1083](https://github.com/nf-core/rnaseq/pull/1083) - Move local modules and subworkflows to subfolders - [PR #1088](https://github.com/nf-core/rnaseq/pull/1088) - Updates contributing and code of conduct documents with nf-core template 2.10 - [PR #1091](https://github.com/nf-core/rnaseq/pull/1091) - Reorganise parameters in schema for better usability -- [PR #1107](https://github.com/nf-core/rnaseq/pull/1107) - Prepare a GTF for stringtie +- [PR #1107](https://github.com/nf-core/rnaseq/pull/1107) - Expand GTF filtering to remove rows with empty transcript ID when required, fix STAR GTF usage ### Software dependencies From c612dd043a74048225e01b3b5061135922d0c2bf Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 9 Nov 2023 17:45:16 +0000 Subject: [PATCH 129/184] [skip ci] Update config per latest Co-authored-by: Maxime U Garcia --- conf/modules.config | 7 +++---- 1 file changed, 3 insertions(+), 4 deletions(-) diff --git a/conf/modules.config b/conf/modules.config index 024daeb18..65d1243bb 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -89,12 +89,11 @@ process { } withName: 'KALLISTO_INDEX' { - ext.args = params.gencode ? '--gencode' : '' + ext.args = { params.gencode ? '--gencode' : '' } publishDir = [ path: { "${params.outdir}/genome/index" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') ? null : filename }, - enabled: params.save_reference + saveAs: { filename -> filename.equals('versions.yml') ? null : params.save_reference ? filename : null } ] } @@ -1221,7 +1220,7 @@ if (!params.skip_pseudo_alignment) { "--count_col 3", params.deseq2_vst ? '--vst TRUE' : '' ].join(' ').trim() - ext.args2 = params.pseudo_aligner + ext.args2 = { params.pseudo_aligner } publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}/deseq2_qc" }, mode: params.publish_dir_mode, From 4675be06b66cd75ad2f787896939c7909c0d1272 Mon Sep 17 00:00:00 2001 From: maxulysse Date: Thu, 9 Nov 2023 18:52:38 +0100 Subject: [PATCH 130/184] fix statement --- conf/modules.config | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/conf/modules.config b/conf/modules.config index d1578b7d9..d3c22f00c 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -428,7 +428,7 @@ if (!params.skip_alignment) { path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, pattern: '*.bam', - saveAs: { params.save_align_intermeds || params.with_umi || params.save_umi_intermeds } + saveAs: { params.save_align_intermeds || params.with_umi || params.save_umi_intermeds ? it : null } ] ] } @@ -440,7 +440,7 @@ if (!params.skip_alignment) { path: { "${params.outdir}/${params.aligner}" }, mode: params.publish_dir_mode, pattern: '*.{bai,csi}', - saveAs: { params.save_align_intermeds || params.with_umi || params.save_umi_intermeds } + saveAs: { params.save_align_intermeds || params.with_umi || params.save_umi_intermeds ? it : null } ] } From 4a81e64fa59b8bee3af0df1970b1849569527eca Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 9 Nov 2023 17:53:17 +0000 Subject: [PATCH 131/184] [skip ci] fix font sizes in metro map file icons --- docs/images/nf-core-rnaseq_metro_map_grey.png | Bin 308202 -> 307103 bytes docs/images/nf-core-rnaseq_metro_map_grey.svg | 44 +++++++++--------- 2 files changed, 22 insertions(+), 22 deletions(-) diff --git a/docs/images/nf-core-rnaseq_metro_map_grey.png b/docs/images/nf-core-rnaseq_metro_map_grey.png index e8e3d2e0d6d8b106bb8f6000f46f2c44f9f3f614..f2b5e4e878715c0720357cac59169594213829ea 100644 GIT binary patch literal 307103 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47.5386 v -20.9983 c 0,-1.1338 0.91983,-2.0521 2.05208,-2.0521 h 18.12807 v -10.2809 h -67.71875 z m 67.71894,-95.336 h -67.71894 v 19.7705 h 67.71875 v -19.7705 z" id="path10553-46-0-7" />GTFGTFFASTAFASTAFASTQFASTQTSVTSVTSVTSVBAMBAMBAIBAIBIGWIGBIGWIGHTMLHTML Date: Thu, 9 Nov 2023 17:59:39 +0000 Subject: [PATCH 132/184] thank myself to poke ci --- CHANGELOG.md | 1 + modules.json | 2 +- modules/nf-core/kallisto/quant/main.nf | 18 ++++++++++++++++++ modules/nf-core/kallisto/quant/meta.yml | 7 +++++++ .../nf-core/kallisto/quant/tests/main.nf.test | 4 ++++ .../kallisto/quant/tests/single_end.config | 3 --- 6 files changed, 31 insertions(+), 4 deletions(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index 2512ccd26..9242b1cff 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -13,6 +13,7 @@ Special thanks to the following for their contributions to the release: - [Júlia Mir Pedrol](https://github.com/mirpedrol) - [Matthias Zepper](https://github.com/MatthiasZepper) - [Maxime Garcia](https://github.com/maxulysse) +- [Jonathan Manning](https://github.com/pinin4fjords) Thank you to everyone else that has contributed by reporting bugs, enhancements or in any other way, shape or form. diff --git a/modules.json b/modules.json index 431d7dfda..3bd2a9132 100644 --- a/modules.json +++ b/modules.json @@ -72,7 +72,7 @@ }, "kallisto/quant": { "branch": "master", - "git_sha": "bdbbd9a849f9dd2b6078b303d36bf26e6adb8a9e", + "git_sha": "2680fb696b197fceb2113790b19364a8da35eab0", "installed_by": ["modules"] }, "picard/markduplicates": { diff --git a/modules/nf-core/kallisto/quant/main.nf b/modules/nf-core/kallisto/quant/main.nf index 06579746e..49ec2cee2 100644 --- a/modules/nf-core/kallisto/quant/main.nf +++ b/modules/nf-core/kallisto/quant/main.nf @@ -12,6 +12,8 @@ process KALLISTO_QUANT { tuple val(meta2), path(index) path gtf path chromosomes + val fragment_length + val fragment_length_sd output: tuple val(meta), path("${prefix}") , emit: results @@ -27,12 +29,28 @@ process KALLISTO_QUANT { prefix = task.ext.prefix ?: "${meta.id}" def gtf_input = gtf ? "--gtf ${gtf}" : '' def chromosomes_input = chromosomes ? "--chromosomes ${chromosomes}" : '' + + def single_end_params = '' + if (meta.single_end) { + if (!(fragment_length =~ /^\d+$/)) { + error "fragment_length must be set and numeric for single-end data" + } + if (!(fragment_length_sd =~ /^\d+$/)) { + error "fragment_length_sd must be set and numeric for single-end data" + } + single_end_params = "--single --fragment-length=${fragment_length} --sd=${fragment_length_sd}" + } + + def strandedness = (meta.strandedness == 'forward') ? '--fr-stranded' : + (meta.strandedness == 'reverse') ? '--rf-stranded' : '' """ kallisto quant \\ --threads ${task.cpus} \\ --index ${index} \\ ${gtf_input} \\ ${chromosomes_input} \\ + ${single_end_params} \\ + ${strandedness} \\ ${args} \\ -o $prefix \\ ${reads} 2> >(tee -a ${prefix}.log.txt >&2) diff --git a/modules/nf-core/kallisto/quant/meta.yml b/modules/nf-core/kallisto/quant/meta.yml index d133847e8..2dce7b639 100644 --- a/modules/nf-core/kallisto/quant/meta.yml +++ b/modules/nf-core/kallisto/quant/meta.yml @@ -36,6 +36,13 @@ input: type: file description: Optional tab separated file with chromosome names and lengths. pattern: "*.tsv" + - fragment_length: + type: integer + description: For single-end mode only, the estimated average fragment length. + - fragment_length_sd: + type: integer + description: For single-end mode only, the estimated standard deviation of the fragment length. + output: - meta: type: map diff --git a/modules/nf-core/kallisto/quant/tests/main.nf.test b/modules/nf-core/kallisto/quant/tests/main.nf.test index 4c5eac84b..f716e5e6d 100644 --- a/modules/nf-core/kallisto/quant/tests/main.nf.test +++ b/modules/nf-core/kallisto/quant/tests/main.nf.test @@ -39,6 +39,8 @@ nextflow_process { input[1] = KALLISTO_INDEX.out.index input[2] = [] input[3] = [] + input[4] = 150 + input[5] = 75 """ } } @@ -73,6 +75,8 @@ nextflow_process { input[1] = KALLISTO_INDEX.out.index input[2] = [] input[3] = [] + input[4] = [] + input[5] = [] """ } } diff --git a/modules/nf-core/kallisto/quant/tests/single_end.config b/modules/nf-core/kallisto/quant/tests/single_end.config index 5aa978aba..7022246bc 100644 --- a/modules/nf-core/kallisto/quant/tests/single_end.config +++ b/modules/nf-core/kallisto/quant/tests/single_end.config @@ -2,7 +2,4 @@ process { publishDir = { "${params.outdir}/${task.process.tokenize(':')[-1].tokenize('_')[0].toLowerCase()}" } - withName: 'KALLISTO_QUANT' { - ext.args='--single --fragment-length 150 --sd 75' - } } From a8010136cb064dd43269f8af8afada30db4ebff0 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 9 Nov 2023 18:04:29 +0000 Subject: [PATCH 133/184] Thank myself to poke CI --- CHANGELOG.md | 1 + 1 file changed, 1 insertion(+) diff --git a/CHANGELOG.md b/CHANGELOG.md index b38303f49..66afef72f 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -13,6 +13,7 @@ Special thanks to the following for their contributions to the release: - [Júlia Mir Pedrol](https://github.com/mirpedrol) - [Matthias Zepper](https://github.com/MatthiasZepper) - [Maxime Garcia](https://github.com/maxulysse) +- [Jonathan Manning](https://github.com/pinin4fjords) Thank you to everyone else that has contributed by reporting bugs, enhancements or in any other way, shape or form. From d75d5076618d15f05a98580c50ead9aa1d7534fb Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 9 Nov 2023 18:13:04 +0000 Subject: [PATCH 134/184] Reflect updated kallisto module --- conf/modules.config | 14 +------------- subworkflows/local/quantify_pseudo/main.nf | 2 +- 2 files changed, 2 insertions(+), 14 deletions(-) diff --git a/conf/modules.config b/conf/modules.config index 65d1243bb..c2a375e8c 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -1158,19 +1158,7 @@ if (params.pseudo_aligner == 'salmon') { if (params.pseudo_aligner == 'kallisto') { process { withName: '.*:QUANTIFY_PSEUDO_ALIGNMENT:KALLISTO_QUANT' { - ext.args = { - [ - meta.single_end ? - "--single " + - "--fragment-length ${meta.fragment_len ?: params.kallisto_quant_fraglen} " + - "--sd ${meta.fraglen_se ?: params.kallisto_quant_fraglen_sd}" : '', - params.kallisto_quant_strandedness ? '--' + params.kallisto_quant_strandedness: - (meta.strandedess == 'forward' ? '--fr-stranded': - (meta.strandedness == 'reverse' ? '--rf-stranded' : '') - ), - params.extra_kallisto_quant_args ?: '' - ].findAll().join(' ').trim() - } + ext.args = params.extra_kallisto_quant_args ?: '' publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}" }, diff --git a/subworkflows/local/quantify_pseudo/main.nf b/subworkflows/local/quantify_pseudo/main.nf index 1b712c76b..4688bcca4 100644 --- a/subworkflows/local/quantify_pseudo/main.nf +++ b/subworkflows/local/quantify_pseudo/main.nf @@ -37,7 +37,7 @@ workflow QUANTIFY_PSEUDO_ALIGNMENT { ch_pseudo_multiqc = ch_pseudo_results ch_versions = ch_versions.mix(SALMON_QUANT.out.versions.first()) }else { - KALLISTO_QUANT ( reads, index, gtf, []) + KALLISTO_QUANT ( reads, index, gtf, [], params.kallisto_quant_fraglen, params.kallisto_quant_fraglen_sd) ch_pseudo_results = KALLISTO_QUANT.out.results ch_pseudo_multiqc = KALLISTO_QUANT.out.log ch_versions = ch_versions.mix(KALLISTO_QUANT.out.versions.first()) From 16430c2f27c2f596930136181e492de1a51b80c4 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Fri, 10 Nov 2023 09:40:57 +0000 Subject: [PATCH 135/184] filter_gtf improvements --- bin/filter_gtf.py | 64 ++++++++++++++++++++++++----------------------- 1 file changed, 33 insertions(+), 31 deletions(-) diff --git a/bin/filter_gtf.py b/bin/filter_gtf.py index 3d4aa1fc9..0f87dc615 100755 --- a/bin/filter_gtf.py +++ b/bin/filter_gtf.py @@ -45,49 +45,51 @@ def extract_fasta_seq_names(fasta_name: str) -> set: headerStr = line[1:].strip().split()[0] yield headerStr - -def extract_genes_in_genome(fasta: str, gtf_in: str, prefix: str) -> None: +def extract_genes_in_genome(fasta: str, gtf_in: str, gtf_out: str) -> None: """Extracts the genes in the genome from a GTF file. Args: fasta: The path to the FASTA file. gtf_in: The path to the input GTF file. - prefix: Prefix for output GTF - """ - gtf_out = prefix + "_in_genome.gtf" - seq_names_in_genome = set(extract_fasta_seq_names(fasta)) - logger.info("Extracted chromosome sequence names from : %s" % fasta) - logger.info("All chromosome names: " + ", ".join(sorted(x for x in seq_names_in_genome))) - seq_names_in_gtf = set([]) - - n_total_lines = 0 - n_lines_in_genome = 0 - with open(gtf_out, "w") as f: - with open(gtf_in) as g: - for line in g.readlines(): - n_total_lines += 1 - seq_name_gtf = line.split("\t")[0] - seq_names_in_gtf.add(seq_name_gtf) - if seq_name_gtf in seq_names_in_genome: - n_lines_in_genome += 1 - f.write(line) - logger.info( - "Extracted %d / %d lines from %s matching sequences in %s" % (n_lines_in_genome, n_total_lines, gtf_in, fasta) - ) - logger.info("All sequence IDs from GTF: " + ", ".join(sorted(x for x in seq_name_gtf))) + gtf_out: The path to the output GTF file. - logger.info("Wrote matching lines to %s" % gtf_out) + Raises: + ValueError: If no overlap is found or if the GTF file is not tab delimited. + """ + def is_tab_delimited(file): + with open(file, 'r') as f: + return '\t' in f.readline() + if not is_tab_delimited(gtf_in): + raise ValueError("The GTF file is not tab delimited.") -def remove_features_without_transcript_id(gtf_in, prefix): + seq_names_in_genome = set(extract_fasta_seq_names(fasta)) + logger.info(f"Extracted chromosome sequence names from {fasta}") + logger.info("All chromosome names: " + ", ".join(sorted(seq_names_in_genome))) + + with open(gtf_in) as gtf, open(gtf_out, "w") as out: + seq_names_in_gtf = {line.split("\t")[0] for line in gtf if line.strip()} + overlap = seq_names_in_genome & seq_names_in_gtf + if not overlap: + raise ValueError("No overlapping scaffolds found.") + + gtf.seek(0) # Reset file pointer to the start of the file + for line in gtf: + if line.split("\t")[0] in overlap: + out.write(line) + + logger.info(f"Extracted {len(overlap)} matching sequences from {gtf_in} into {gtf_out}") + logger.info("All sequence IDs from GTF: " + ", ".join(sorted(seq_names_in_gtf))) + logger.info(f"Wrote matching lines to {gtf_out}") + +def remove_features_without_transcript_id(gtf_in: str, gtf_out: str) -> None: """ Removes gene rows with absent or empty transcript_id attributes from a GTF file. Args: gtf_in: Path to the input GTF file. - prefix: Path to the output GTF file. + gtf_out: The path to the output GTF file. """ - gtf_out = prefix + "_with_transcript_ids.gtf" with open(gtf_in, "r") as f_in, open(gtf_out, "w") as f_out: for line in f_in: @@ -110,5 +112,5 @@ def remove_features_without_transcript_id(gtf_in, prefix): ) args = parser.parse_args() - extract_genes_in_genome(args.fasta, args.gtf, args.prefix) - remove_features_without_transcript_id(args.gtf, args.prefix) + extract_genes_in_genome(args.fasta, args.gtf, args.prefix + "_in_genome.gtf" ) + remove_features_without_transcript_id(args.prefix + "_in_genome.gtf", args.prefix + "_with_transcript_ids.gtf") From 65ac5ef9f4a3333f8ab788598dab299d63c39c25 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Fri, 10 Nov 2023 09:46:56 +0000 Subject: [PATCH 136/184] Blackify --- bin/filter_gtf.py | 13 ++++++++----- 1 file changed, 8 insertions(+), 5 deletions(-) diff --git a/bin/filter_gtf.py b/bin/filter_gtf.py index 0f87dc615..cb7a85dd1 100755 --- a/bin/filter_gtf.py +++ b/bin/filter_gtf.py @@ -45,6 +45,7 @@ def extract_fasta_seq_names(fasta_name: str) -> set: headerStr = line[1:].strip().split()[0] yield headerStr + def extract_genes_in_genome(fasta: str, gtf_in: str, gtf_out: str) -> None: """Extracts the genes in the genome from a GTF file. @@ -56,9 +57,10 @@ def extract_genes_in_genome(fasta: str, gtf_in: str, gtf_out: str) -> None: Raises: ValueError: If no overlap is found or if the GTF file is not tab delimited. """ + def is_tab_delimited(file): - with open(file, 'r') as f: - return '\t' in f.readline() + with open(file, "r") as f: + return "\t" in f.readline() if not is_tab_delimited(gtf_in): raise ValueError("The GTF file is not tab delimited.") @@ -66,7 +68,7 @@ def is_tab_delimited(file): seq_names_in_genome = set(extract_fasta_seq_names(fasta)) logger.info(f"Extracted chromosome sequence names from {fasta}") logger.info("All chromosome names: " + ", ".join(sorted(seq_names_in_genome))) - + with open(gtf_in) as gtf, open(gtf_out, "w") as out: seq_names_in_gtf = {line.split("\t")[0] for line in gtf if line.strip()} overlap = seq_names_in_genome & seq_names_in_gtf @@ -82,6 +84,7 @@ def is_tab_delimited(file): logger.info("All sequence IDs from GTF: " + ", ".join(sorted(seq_names_in_gtf))) logger.info(f"Wrote matching lines to {gtf_out}") + def remove_features_without_transcript_id(gtf_in: str, gtf_out: str) -> None: """ Removes gene rows with absent or empty transcript_id attributes from a GTF file. @@ -112,5 +115,5 @@ def remove_features_without_transcript_id(gtf_in: str, gtf_out: str) -> None: ) args = parser.parse_args() - extract_genes_in_genome(args.fasta, args.gtf, args.prefix + "_in_genome.gtf" ) - remove_features_without_transcript_id(args.prefix + "_in_genome.gtf", args.prefix + "_with_transcript_ids.gtf") + extract_genes_in_genome(args.fasta, args.gtf, args.prefix + "_in_genome.gtf") + remove_features_without_transcript_id(args.prefix + "_in_genome.gtf", args.prefix + "_with_transcript_ids.gtf") From b0cf29533a0f506e4e2b889dfd5b36e3028ace79 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Fri, 10 Nov 2023 09:48:01 +0000 Subject: [PATCH 137/184] [skip ci] test file not supposed to be there --- test.sh | 6 ------ 1 file changed, 6 deletions(-) delete mode 100755 test.sh diff --git a/test.sh b/test.sh deleted file mode 100755 index 8222248a9..000000000 --- a/test.sh +++ /dev/null @@ -1,6 +0,0 @@ -#!/usr/bin/env bash - -nextflow run . \ - --outdir output \ - --pseudo_aligner kallisto \ - -resume -profile conda,test From c4823eaae012c9f67681ca98f622e53da87a33d0 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Fri, 10 Nov 2023 10:01:38 +0000 Subject: [PATCH 138/184] [skip ci] Add resolved issues to changelog --- CHANGELOG.md | 3 +++ 1 file changed, 3 insertions(+) diff --git a/CHANGELOG.md b/CHANGELOG.md index 66afef72f..e370c60cb 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -30,6 +30,9 @@ Thank you to everyone else that has contributed by reporting bugs, enhancements - [PR #1088](https://github.com/nf-core/rnaseq/pull/1088) - Updates contributing and code of conduct documents with nf-core template 2.10 - [PR #1091](https://github.com/nf-core/rnaseq/pull/1091) - Reorganise parameters in schema for better usability - [PR #1107](https://github.com/nf-core/rnaseq/pull/1107) - Expand GTF filtering to remove rows with empty transcript ID when required, fix STAR GTF usage +- [#1082](https://github.com/nf-core/rnaseq/issues/1082) - More informative error message for filter_gtf_for_genes_in_genome.py +- [#1102](https://github.com/nf-core/rnaseq/issues/1102) - gene entries with empty transcript_id fields +- [#1074](https://github.com/nf-core/rnaseq/issues/1074) - Enable quantification using StringTie AND a custom Ensembl genome ### Software dependencies From ecb3a1480136af13a4d3a64af7a0156ef619aa83 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Fri, 10 Nov 2023 10:08:45 +0000 Subject: [PATCH 139/184] reinstate publish modes --- conf/modules.config | 2 ++ 1 file changed, 2 insertions(+) diff --git a/conf/modules.config b/conf/modules.config index 0c6bc646d..aabe7bfe7 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -155,6 +155,7 @@ process { ext.args = '--record-count 1000000 --seed 1' ext.prefix = { "${meta.id}.subsampled" } publishDir = [ + mode: params.publish_dir_mode, enabled: false ] } @@ -162,6 +163,7 @@ process { withName: '.*:FASTQ_SUBSAMPLE_FQ_SALMON:SALMON_QUANT' { ext.args = '--skipQuant' publishDir = [ + mode: params.publish_dir_mode, enabled: false ] } From 4f556ee9ab3722012ba096bb42c05957bb6dc465 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Mon, 13 Nov 2023 15:31:14 +0000 Subject: [PATCH 140/184] [skip ci] Add Kallisto outputs to Tower yml --- tower.yml | 9 +++++++++ 1 file changed, 9 insertions(+) diff --git a/tower.yml b/tower.yml index b6983530a..818ef13bf 100644 --- a/tower.yml +++ b/tower.yml @@ -21,6 +21,15 @@ reports: display: "All samples Salmon merged transcript raw counts" "**/salmon/salmon.merged.transcript_tpm.tsv": display: "All samples Salmon merged transcript TPM counts" + "**/kallisto/**/deseq2.plots.pdf": + display: "All samples Kallisto DESeq2 QC PDF plots" + "**/kallisto/kallisto.merged.gene_counts.tsv": + display: "All samples Kallisto merged gene raw counts" + "**/kallisto/kallisto.merged.gene_tpm.tsv": + display: "All samples Kallisto merged gene TPM counts" + "**/kallisto/kallisto.merged.transcript_counts.tsv": + display: "All samples Kallisto merged transcript raw counts" + "**/kallisto/kallisto.merged.transcript_tpm.tsv": "**/star_rsem/**/deseq2.plots.pdf": display: "All samples STAR RSEM DESeq2 QC PDF plots" "**/star_rsem/rsem.merged.gene_counts.tsv": From 1c23a4a0991e7f71faba1c64cde39956594f255f Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 14 Nov 2023 09:53:52 +0000 Subject: [PATCH 141/184] [skip ci] Reduce logging in GTF filtering --- bin/filter_gtf.py | 5 ++--- 1 file changed, 2 insertions(+), 3 deletions(-) diff --git a/bin/filter_gtf.py b/bin/filter_gtf.py index cb7a85dd1..10cb646b9 100755 --- a/bin/filter_gtf.py +++ b/bin/filter_gtf.py @@ -10,7 +10,6 @@ logger = logging.getLogger(__file__) logger.setLevel(logging.INFO) - def is_header(line: str) -> bool: """Returns True if the given line is a header line in a FASTA file.""" return line[0] == ">" @@ -67,7 +66,7 @@ def is_tab_delimited(file): seq_names_in_genome = set(extract_fasta_seq_names(fasta)) logger.info(f"Extracted chromosome sequence names from {fasta}") - logger.info("All chromosome names: " + ", ".join(sorted(seq_names_in_genome))) + logger.debug("All chromosome names: " + ", ".join(sorted(seq_names_in_genome))) with open(gtf_in) as gtf, open(gtf_out, "w") as out: seq_names_in_gtf = {line.split("\t")[0] for line in gtf if line.strip()} @@ -81,7 +80,7 @@ def is_tab_delimited(file): out.write(line) logger.info(f"Extracted {len(overlap)} matching sequences from {gtf_in} into {gtf_out}") - logger.info("All sequence IDs from GTF: " + ", ".join(sorted(seq_names_in_gtf))) + logger.debug("All sequence IDs from GTF: " + ", ".join(sorted(seq_names_in_gtf))) logger.info(f"Wrote matching lines to {gtf_out}") From 0a70d58289bf4031355844b51b1e6f336ae0135f Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 14 Nov 2023 09:56:39 +0000 Subject: [PATCH 142/184] [skip ci] Apply suggestions from code review Co-authored-by: Matthias Zepper <6963520+MatthiasZepper@users.noreply.github.com> --- bin/filter_gtf.py | 37 +++++++++++++++++------------------ modules/local/multiqc/main.nf | 4 ++-- 2 files changed, 20 insertions(+), 21 deletions(-) diff --git a/bin/filter_gtf.py b/bin/filter_gtf.py index 10cb646b9..cafc14c39 100755 --- a/bin/filter_gtf.py +++ b/bin/filter_gtf.py @@ -29,20 +29,12 @@ def extract_fasta_seq_names(fasta_name: str) -> set: Returns: A set of the sequence names in the FASTA file. """ - - # first open the file outside - fh = open(fasta_name) - - # ditch the boolean (x[0]) and just keep the header or sequence since - # we know they alternate. - faiter = (x[1] for x in groupby(fh, is_header)) - - for i, header in enumerate(faiter): - line = next(header) - if is_header(line): - # drop the ">" - headerStr = line[1:].strip().split()[0] - yield headerStr + seqnames = set() + with open(fasta_name) as fasta: + for line in fasta: + if line[0] == ">": + seqnames.add(line[1:].split(None, 1)[0]) + return seqnames def extract_genes_in_genome(fasta: str, gtf_in: str, gtf_out: str) -> None: @@ -57,14 +49,21 @@ def extract_genes_in_genome(fasta: str, gtf_in: str, gtf_out: str) -> None: ValueError: If no overlap is found or if the GTF file is not tab delimited. """ - def is_tab_delimited(file): + def tab_delimited(file) -> float: + + import statistics. # put to the top + with open(file, "r") as f: - return "\t" in f.readline() + data = f.read(1024) + lines = data.split("\n") + # most lines should have 9 tab-separated columns + return statistics.median([line.count("\t") for line in lines]) - if not is_tab_delimited(gtf_in): - raise ValueError("The GTF file is not tab delimited.") + num_sep = tab_delimited(gtf_in) + if num_sep != 8: + raise ValueError("No valid tab-delimited GTF file.") - seq_names_in_genome = set(extract_fasta_seq_names(fasta)) + seq_names_in_genome = extract_fasta_seq_names(fasta) logger.info(f"Extracted chromosome sequence names from {fasta}") logger.debug("All chromosome names: " + ", ".join(sorted(seq_names_in_genome))) diff --git a/modules/local/multiqc/main.nf b/modules/local/multiqc/main.nf index 06fbb9b29..6ab3a38d0 100644 --- a/modules/local/multiqc/main.nf +++ b/modules/local/multiqc/main.nf @@ -3,8 +3,8 @@ process MULTIQC { conda "bioconda::multiqc=1.17" container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? - 'https://depot.galaxyproject.org/singularity/multiqc:1.17--pyhdfd78af_0' : - 'biocontainers/multiqc:1.17--pyhdfd78af_0' }" + 'https://depot.galaxyproject.org/singularity/multiqc:1.17--pyhdfd78af_1' : + 'biocontainers/multiqc:1.17--pyhdfd78af_1' }" input: path multiqc_config From 996abcc5c6ed9a23079acefc91c957035c072dea Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 14 Nov 2023 10:04:02 +0000 Subject: [PATCH 143/184] [skip ci] post-suggestion integration work --- bin/filter_gtf.py | 51 +++++++++++++++++++---------------------------- 1 file changed, 20 insertions(+), 31 deletions(-) diff --git a/bin/filter_gtf.py b/bin/filter_gtf.py index cafc14c39..11cfbdc30 100755 --- a/bin/filter_gtf.py +++ b/bin/filter_gtf.py @@ -37,13 +37,15 @@ def extract_fasta_seq_names(fasta_name: str) -> set: return seqnames -def extract_genes_in_genome(fasta: str, gtf_in: str, gtf_out: str) -> None: +def filter_gtf(fasta: str, gtf_in: str, gtf_in_genome_out: str, gtf_transcript_out: str) -> None: """Extracts the genes in the genome from a GTF file. Args: fasta: The path to the FASTA file. gtf_in: The path to the input GTF file. - gtf_out: The path to the output GTF file. + gtf_in_genome_out: The path to the output GTF file with scaffolds filtered. + gtf_transcript_out: The path to the output GTF file with scaffolds + filtered and rows without transcript ID removed. Raises: ValueError: If no overlap is found or if the GTF file is not tab delimited. @@ -67,36 +69,24 @@ def tab_delimited(file) -> float: logger.info(f"Extracted chromosome sequence names from {fasta}") logger.debug("All chromosome names: " + ", ".join(sorted(seq_names_in_genome))) - with open(gtf_in) as gtf, open(gtf_out, "w") as out: - seq_names_in_gtf = {line.split("\t")[0] for line in gtf if line.strip()} - overlap = seq_names_in_genome & seq_names_in_gtf - if not overlap: - raise ValueError("No overlapping scaffolds found.") - - gtf.seek(0) # Reset file pointer to the start of the file + with open(gtf_in) as gtf, open(gtf_in_genome_out, "w") as out, open( + gtf_transcript_out, "w" + ) as out2: + line_count_all = 0 + line_count_transcript = 0 for line in gtf: - if line.split("\t")[0] in overlap: + if ( + line.split("\t")[0] in seq_names_in_genome + and line.count("\t") == num_sep + ): out.write(line) + line_count_all += 1 + if re.search(r'transcript_id "([^"]+)"', line): + out2.write(line) + line_count_transcript += 1 + if line_count_all == 0: + raise ValueError("No overlapping scaffolds found.") - logger.info(f"Extracted {len(overlap)} matching sequences from {gtf_in} into {gtf_out}") - logger.debug("All sequence IDs from GTF: " + ", ".join(sorted(seq_names_in_gtf))) - logger.info(f"Wrote matching lines to {gtf_out}") - - -def remove_features_without_transcript_id(gtf_in: str, gtf_out: str) -> None: - """ - Removes gene rows with absent or empty transcript_id attributes from a GTF file. - - Args: - gtf_in: Path to the input GTF file. - gtf_out: The path to the output GTF file. - """ - - with open(gtf_in, "r") as f_in, open(gtf_out, "w") as f_out: - for line in f_in: - transcript_id_match = re.search(r'transcript_id "([^"]+)"', line) - if transcript_id_match: - f_out.write(line) if __name__ == "__main__": @@ -113,5 +103,4 @@ def remove_features_without_transcript_id(gtf_in: str, gtf_out: str) -> None: ) args = parser.parse_args() - extract_genes_in_genome(args.fasta, args.gtf, args.prefix + "_in_genome.gtf") - remove_features_without_transcript_id(args.prefix + "_in_genome.gtf", args.prefix + "_with_transcript_ids.gtf") + filter_gtf(args.fasta, args.gtf, args.prefix + "_in_genome.gtf", args.prefix + "_with_transcript_ids.gtf") From 01e05f74a96d86d9ea4fcc8c8ee9655420ada095 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 14 Nov 2023 10:11:43 +0000 Subject: [PATCH 144/184] [skip ci] more post-review tidy-up to GTF filtering script --- bin/filter_gtf.py | 117 +++++++++++++--------------------------------- 1 file changed, 33 insertions(+), 84 deletions(-) mode change 100755 => 100644 bin/filter_gtf.py diff --git a/bin/filter_gtf.py b/bin/filter_gtf.py old mode 100755 new mode 100644 index 11cfbdc30..8a6836e4e --- a/bin/filter_gtf.py +++ b/bin/filter_gtf.py @@ -1,106 +1,55 @@ #!/usr/bin/env python -from __future__ import print_function import logging -from itertools import groupby import argparse import re +import statistics +from typing import Set # Create a logger logging.basicConfig(format="%(name)s - %(asctime)s %(levelname)s: %(message)s") -logger = logging.getLogger(__file__) +logger = logging.getLogger("fasta_gtf_filter") logger.setLevel(logging.INFO) -def is_header(line: str) -> bool: - """Returns True if the given line is a header line in a FASTA file.""" - return line[0] == ">" - - -def extract_fasta_seq_names(fasta_name: str) -> set: - """Extracts the sequence names from a FASTA file. - - modified from Brent Pedersen - Correct Way To Parse A Fasta File In Python - given a fasta file. yield tuples of header, sequence - from https://www.biostars.org/p/710/ - - Args: - fasta_name: The path to the FASTA file. - - Returns: - A set of the sequence names in the FASTA file. - """ - seqnames = set() +def extract_fasta_seq_names(fasta_name: str) -> Set[str]: + """Extracts the sequence names from a FASTA file.""" with open(fasta_name) as fasta: - for line in fasta: - if line[0] == ">": - seqnames.add(line[1:].split(None, 1)[0]) - return seqnames + return {line[1:].split(None, 1)[0] for line in fasta if line.startswith(">")} +def tab_delimited(file: str) -> float: + """Check if file is tab-delimited and return median number of tabs.""" + with open(file, "r") as f: + data = f.read(1024) + return statistics.median(line.count("\t") for line in data.split("\n")) def filter_gtf(fasta: str, gtf_in: str, gtf_in_genome_out: str, gtf_transcript_out: str) -> None: - """Extracts the genes in the genome from a GTF file. - - Args: - fasta: The path to the FASTA file. - gtf_in: The path to the input GTF file. - gtf_in_genome_out: The path to the output GTF file with scaffolds filtered. - gtf_transcript_out: The path to the output GTF file with scaffolds - filtered and rows without transcript ID removed. - - Raises: - ValueError: If no overlap is found or if the GTF file is not tab delimited. - """ - - def tab_delimited(file) -> float: - - import statistics. # put to the top - - with open(file, "r") as f: - data = f.read(1024) - lines = data.split("\n") - # most lines should have 9 tab-separated columns - return statistics.median([line.count("\t") for line in lines]) - - num_sep = tab_delimited(gtf_in) - if num_sep != 8: - raise ValueError("No valid tab-delimited GTF file.") + """Filter GTF file based on FASTA sequence names.""" + if tab_delimited(gtf_in) != 8: + raise ValueError("Invalid GTF file: Expected 8 tab-separated columns.") seq_names_in_genome = extract_fasta_seq_names(fasta) logger.info(f"Extracted chromosome sequence names from {fasta}") - logger.debug("All chromosome names: " + ", ".join(sorted(seq_names_in_genome))) - - with open(gtf_in) as gtf, open(gtf_in_genome_out, "w") as out, open( - gtf_transcript_out, "w" - ) as out2: - line_count_all = 0 - line_count_transcript = 0 - for line in gtf: - if ( - line.split("\t")[0] in seq_names_in_genome - and line.count("\t") == num_sep - ): - out.write(line) - line_count_all += 1 - if re.search(r'transcript_id "([^"]+)"', line): - out2.write(line) - line_count_transcript += 1 - if line_count_all == 0: - raise ValueError("No overlapping scaffolds found.") - + try: + with open(gtf_in) as gtf, open(gtf_in_genome_out, "w") as out, open(gtf_transcript_out, "w") as out2: + line_count_all, line_count_transcript = 0, 0 + for line in gtf: + if line.split("\t")[0] in seq_names_in_genome: + out.write(line) + line_count_all += 1 + if re.search(r'transcript_id "([^"]+)"', line): + out2.write(line) + line_count_transcript += 1 + if line_count_all == 0: + raise ValueError("No overlapping scaffolds found.") + except IOError as e: + logger.error(f"File operation failed: {e}") if __name__ == "__main__": - parser = argparse.ArgumentParser(description="""Filter GTF for various reasons""") - parser.add_argument("--gtf", type=str, help="GTF file") - parser.add_argument("--fasta", type=str, help="Genome fasta file") - parser.add_argument( - "-p", - "--prefix", - dest="prefix", - default="genes", - type=str, - help="Prefix for output GTF files", - ) + parser = argparse.ArgumentParser(description="Filters a GTF file based on sequence names in a FASTA file.") + parser.add_argument("--gtf", type=str, required=True, help="GTF file") + parser.add_argument("--fasta", type=str, required=True, help="Genome fasta file") + parser.add_argument("--prefix", dest="prefix", default="genes", type=str, help="Prefix for output GTF files") args = parser.parse_args() filter_gtf(args.fasta, args.gtf, args.prefix + "_in_genome.gtf", args.prefix + "_with_transcript_ids.gtf") + From 556af914e30289533b0d851d2b49b9ccaf2c90fb Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 14 Nov 2023 10:52:39 +0000 Subject: [PATCH 145/184] [skip ci] reinstate logging --- bin/filter_gtf.py | 11 ++++++++++- 1 file changed, 10 insertions(+), 1 deletion(-) mode change 100644 => 100755 bin/filter_gtf.py diff --git a/bin/filter_gtf.py b/bin/filter_gtf.py old mode 100644 new mode 100755 index 8a6836e4e..18bbcb2d4 --- a/bin/filter_gtf.py +++ b/bin/filter_gtf.py @@ -29,11 +29,15 @@ def filter_gtf(fasta: str, gtf_in: str, gtf_in_genome_out: str, gtf_transcript_o seq_names_in_genome = extract_fasta_seq_names(fasta) logger.info(f"Extracted chromosome sequence names from {fasta}") + seq_names_in_gtf = set() try: with open(gtf_in) as gtf, open(gtf_in_genome_out, "w") as out, open(gtf_transcript_out, "w") as out2: line_count_all, line_count_transcript = 0, 0 for line in gtf: - if line.split("\t")[0] in seq_names_in_genome: + seq_name = line.split("\t")[0] + seq_names_in_gtf.add(seq_name) # Add sequence name to the set + + if seq_name in seq_names_in_genome: out.write(line) line_count_all += 1 if re.search(r'transcript_id "([^"]+)"', line): @@ -43,6 +47,11 @@ def filter_gtf(fasta: str, gtf_in: str, gtf_in_genome_out: str, gtf_transcript_o raise ValueError("No overlapping scaffolds found.") except IOError as e: logger.error(f"File operation failed: {e}") + return + + logger.debug("All sequence IDs from GTF: " + ", ".join(sorted(seq_names_in_gtf))) + logger.info(f"Extracted {line_count_all} matching sequences from {gtf_in} into {gtf_in_genome_out}") + logger.info(f"Wrote {line_count_transcript} lines with transcript IDs to {gtf_transcript_out}") if __name__ == "__main__": parser = argparse.ArgumentParser(description="Filters a GTF file based on sequence names in a FASTA file.") From ad998c4be6b73c97b4b2280132cac514c9e15215 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 14 Nov 2023 10:59:27 +0000 Subject: [PATCH 146/184] [skip ci] reinstate logging --- bin/filter_gtf.py | 1 + 1 file changed, 1 insertion(+) diff --git a/bin/filter_gtf.py b/bin/filter_gtf.py index 18bbcb2d4..f815a8d52 100755 --- a/bin/filter_gtf.py +++ b/bin/filter_gtf.py @@ -28,6 +28,7 @@ def filter_gtf(fasta: str, gtf_in: str, gtf_in_genome_out: str, gtf_transcript_o seq_names_in_genome = extract_fasta_seq_names(fasta) logger.info(f"Extracted chromosome sequence names from {fasta}") + logger.debug("All sequence IDs from FASTA: " + ", ".join(sorted(seq_names_in_genome))) seq_names_in_gtf = set() try: From 80a95430aa2ebe617dc3a49b91a2eba4f8b48498 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 14 Nov 2023 11:09:20 +0000 Subject: [PATCH 147/184] [skip ci] Fix salmon subworkflow call for gtf --- workflows/rnaseq.nf | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/workflows/rnaseq.nf b/workflows/rnaseq.nf index 09de86260..0912c661c 100755 --- a/workflows/rnaseq.nf +++ b/workflows/rnaseq.nf @@ -473,7 +473,7 @@ workflow RNASEQ { ch_transcriptome_bam, ch_dummy_file, PREPARE_GENOME.out.transcript_fasta, - PREPARE_GENOME.out.gtf, + PREPARE_GENOME.out.gtf_with_transcript_ids, true, params.salmon_quant_libtype ?: '' ) From 519e42adb361239232d36aac91daaec12a08bf87 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 14 Nov 2023 11:18:47 +0000 Subject: [PATCH 148/184] [skip ci] also raise value for no transcript ids --- bin/filter_gtf.py | 3 +++ 1 file changed, 3 insertions(+) diff --git a/bin/filter_gtf.py b/bin/filter_gtf.py index f815a8d52..316813fa0 100755 --- a/bin/filter_gtf.py +++ b/bin/filter_gtf.py @@ -46,6 +46,9 @@ def filter_gtf(fasta: str, gtf_in: str, gtf_in_genome_out: str, gtf_transcript_o line_count_transcript += 1 if line_count_all == 0: raise ValueError("No overlapping scaffolds found.") + if line_count_transcript == 0: + raise ValueError("No transcript_id values found in the GTF file.") + except IOError as e: logger.error(f"File operation failed: {e}") return From 5e1378e23cafb7bd79b9fbb09ee68cf75f6eb4de Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 14 Nov 2023 12:16:24 +0000 Subject: [PATCH 149/184] [skip ci] @drpatelh feedback: Try to avoid filtering process where not required --- bin/filter_gtf.py | 15 ++++++++---- conf/modules.config | 1 + nextflow.config | 2 ++ nextflow_schema.json | 11 +++++++++ subworkflows/local/prepare_genome/main.nf | 30 ++++++++++++++++++++--- 5 files changed, 51 insertions(+), 8 deletions(-) diff --git a/bin/filter_gtf.py b/bin/filter_gtf.py index 316813fa0..345d4a68e 100755 --- a/bin/filter_gtf.py +++ b/bin/filter_gtf.py @@ -21,7 +21,7 @@ def tab_delimited(file: str) -> float: data = f.read(1024) return statistics.median(line.count("\t") for line in data.split("\n")) -def filter_gtf(fasta: str, gtf_in: str, gtf_in_genome_out: str, gtf_transcript_out: str) -> None: +def filter_gtf(fasta: str, gtf_in: str, gtf_in_genome_out: str, gtf_transcript_out: str, skip_transcript_id_check: bool) -> None: """Filter GTF file based on FASTA sequence names.""" if tab_delimited(gtf_in) != 8: raise ValueError("Invalid GTF file: Expected 8 tab-separated columns.") @@ -41,12 +41,16 @@ def filter_gtf(fasta: str, gtf_in: str, gtf_in_genome_out: str, gtf_transcript_o if seq_name in seq_names_in_genome: out.write(line) line_count_all += 1 - if re.search(r'transcript_id "([^"]+)"', line): - out2.write(line) + + if not skip_transcript_id_check and re.search(r'transcript_id "([^"]+)"', line): + with open(gtf_transcript_out, "a") as out2: + out2.write(line) line_count_transcript += 1 + if line_count_all == 0: raise ValueError("No overlapping scaffolds found.") - if line_count_transcript == 0: + + if not skip_transcript_id_check and line_count_transcript == 0: raise ValueError("No transcript_id values found in the GTF file.") except IOError as e: @@ -62,7 +66,8 @@ def filter_gtf(fasta: str, gtf_in: str, gtf_in_genome_out: str, gtf_transcript_o parser.add_argument("--gtf", type=str, required=True, help="GTF file") parser.add_argument("--fasta", type=str, required=True, help="Genome fasta file") parser.add_argument("--prefix", dest="prefix", default="genes", type=str, help="Prefix for output GTF files") + parser.add_argument("--skip_transcript_id_check", action='store_true', help="Skip checking for transcript IDs in the GTF file") args = parser.parse_args() - filter_gtf(args.fasta, args.gtf, args.prefix + "_in_genome.gtf", args.prefix + "_with_transcript_ids.gtf") + filter_gtf(args.fasta, args.gtf, args.prefix + "_in_genome.gtf", args.prefix + "_with_transcript_ids.gtf", args.skip_transcript_id_check) diff --git a/conf/modules.config b/conf/modules.config index aabe7bfe7..683ccb1db 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -109,6 +109,7 @@ process { } withName: 'GTF_FILTER' { + ext.args = { params.skip_gtf_transcript_filter ?: '--skip_transcript_id_check' } publishDir = [ path: { "${params.outdir}/genome" }, mode: params.publish_dir_mode, diff --git a/nextflow.config b/nextflow.config index f786d1daa..ce4188962 100644 --- a/nextflow.config +++ b/nextflow.config @@ -17,6 +17,8 @@ params { splicesites = null gtf_extra_attributes = 'gene_name' gtf_group_features = 'gene_id' + skip_gtf_filter = false + skip_gtf_transcript_filter = false featurecounts_feature_type = 'exon' featurecounts_group_type = 'gene_biotype' gencode = false diff --git a/nextflow_schema.json b/nextflow_schema.json index a22baecef..412e3b398 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -85,6 +85,17 @@ "description": "Path to GFF3 annotation file.", "help_text": "This parameter must be specified if `--genome` or `--gtf` are not specified." }, + "skip_gtf_filter": { + "type": "boolean", + "fa_icon": "fas fa-forward", + "description": "Skip filtering of GTF for valid scaffolds and/ or transcript IDs.", + "help_text": "If you're confident on the validity of the GTF with respect to the genome file, or wish to disregard failures thriggered by the filtering module, activate this option." + }, + "skip_gtf_transcript_filter": { + "type": "boolean", + "fa_icon": "fas fa-forward", + "description": "Skip just the transcript_id check of GTF filtering." + }, "gene_bed": { "type": "string", "format": "file-path", diff --git a/subworkflows/local/prepare_genome/main.nf b/subworkflows/local/prepare_genome/main.nf index 337d65615..e743c1dab 100644 --- a/subworkflows/local/prepare_genome/main.nf +++ b/subworkflows/local/prepare_genome/main.nf @@ -90,9 +90,33 @@ workflow PREPARE_GENOME { // // Apply filtering we may need for GTFs // - GTF_FILTER ( ch_fasta, ch_gtf ) - ch_gtf_with_transcript_ids = GTF_FILTER.out.transcript_id_gtf - ch_gtf_genome = GTF_FILTER.out.genome_gtf + + filtering_useful = + ( + // Condition 1: Alignment is required and aligner is set to 'star_salmon' + !params.skip_alignment && params.aligner == 'star_salmon' + ) || + ( + // Condition 2: Pseudo-alignment is required and pseudo-aligner is set to 'salmon' + !params.skip_pseudo_alignment && params.pseudo_aligner == 'salmon' + ) || + ( + // Condition 3: Neither alignment nor stringtie are to be skipped + !params.skip_alignment && !params.skip_stringtie + ) || + ( + // Condition 4: Transcript FASTA file is not provided + !transcript_fasta + ) + + if (params.skip_gtf_filter || ! filtering_useful){ + ch_gtf_with_transcript_ids = ch_gtf + ch_gtf_genome = ch_gtf + } else { + GTF_FILTER ( ch_fasta, ch_gtf ) + ch_gtf_with_transcript_ids = GTF_FILTER.out.transcript_id_gtf + ch_gtf_genome = GTF_FILTER.out.genome_gtf + } } // From 9ab9f7c45b74a27390e51f34b767fc8e037335ec Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Tue, 14 Nov 2023 12:51:37 +0000 Subject: [PATCH 150/184] Post-review changes --- CHANGELOG.md | 8 +++--- README.md | 2 +- bin/summarizedexperiment.r | 2 +- conf/modules.config | 14 +++------ docs/output.md | 16 +++++------ docs/usage.md | 8 +++--- modules/nf-core/kallisto/quant/main.nf | 6 +++- nextflow_schema.json | 29 +++++++++++++++---- subworkflows/local/quantify_pseudo/main.nf | 33 +++++++++++----------- tower.yml | 1 + workflows/rnaseq.nf | 18 +++++++----- 11 files changed, 79 insertions(+), 58 deletions(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index 9242b1cff..5d13bc92c 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -10,10 +10,10 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 Special thanks to the following for their contributions to the release: - [Adam Talbot](https://github.com/adamrtalbot) +- [Jonathan Manning](https://github.com/pinin4fjords) - [Júlia Mir Pedrol](https://github.com/mirpedrol) - [Matthias Zepper](https://github.com/MatthiasZepper) - [Maxime Garcia](https://github.com/maxulysse) -- [Jonathan Manning](https://github.com/pinin4fjords) Thank you to everyone else that has contributed by reporting bugs, enhancements or in any other way, shape or form. @@ -38,7 +38,7 @@ Thank you to everyone else that has contributed by reporting bugs, enhancements | Dependency | Old version | New version | | ----------------------- | ----------- | ----------- | | `fastqc` | 0.11.9 | 0.12.1 | -| `multiqc` | 1.14 | 1.15 | +| `multiqc` | 1.14 | 1.17 | | `ucsc-bedgraphtobigwig` | 377 | 445 | > **NB:** Dependency has been **updated** if both old and new version information is present. @@ -65,7 +65,7 @@ Thank you to everyone else that has contributed by reporting bugs, enhancements ### Enhancements & fixes - [[#1011](https://github.com/nf-core/rnaseq/issues/1011)] - FastQ files from UMI-tools not being passed to fastp -- [[#1018](https://github.com/nf-core/rnaseq/issues/1018)] - Ability to skip both alignment and pseudo-alignment to only run pre-processing QC steps. +- [[#1018](https://github.com/nf-core/rnaseq/issues/1018)] - Ability to skip both alignment and pseudoalignment to only run pre-processing QC steps. - [PR #1016](https://github.com/nf-core/rnaseq/pull/1016) - Updated pipeline template to [nf-core/tools 2.8](https://github.com/nf-core/tools/releases/tag/2.8) - [PR #1025](https://github.com/nf-core/fetchngs/pull/1025) - Add `public_aws_ecr.config` to source mulled containers when using `public.ecr.aws` Docker Biocontainer registry - [PR #1038](https://github.com/nf-core/rnaseq/pull/1038) - Updated error log for count values when supplying `--additional_fasta` @@ -813,7 +813,7 @@ Major novel changes include: - Added options to skip several steps - Skip trimming using `--skipTrimming` - Skip BiotypeQC using `--skipBiotypeQC` - - Skip Alignment using `--skipAlignment` to only use pseudo-alignment using Salmon + - Skip Alignment using `--skipAlignment` to only use pseudoalignment using Salmon ### Documentation updates diff --git a/README.md b/README.md index b56768f3a..b45db8c7a 100644 --- a/README.md +++ b/README.md @@ -39,7 +39,7 @@ 3. [`dupRadar`](https://bioconductor.org/packages/release/bioc/html/dupRadar.html) 4. [`Preseq`](http://smithlabresearch.org/software/preseq/) 5. [`DESeq2`](https://bioconductor.org/packages/release/bioc/html/DESeq2.html) -15. Pseudo-alignment and quantification ([`Salmon`](https://combine-lab.github.io/salmon/) or ['Kallisto'](https://pachterlab.github.io/kallisto/); _optional_) +15. Pseudoalignment and quantification ([`Salmon`](https://combine-lab.github.io/salmon/) or ['Kallisto'](https://pachterlab.github.io/kallisto/); _optional_) 16. Present QC for raw read, alignment, gene biotype, sample similarity, and strand-specificity checks ([`MultiQC`](http://multiqc.info/), [`R`](https://www.r-project.org/)) > **Note** diff --git a/bin/summarizedexperiment.r b/bin/summarizedexperiment.r index 410663ae6..d55101ecf 100755 --- a/bin/summarizedexperiment.r +++ b/bin/summarizedexperiment.r @@ -2,7 +2,7 @@ library(SummarizedExperiment) -## Create SummarizedExperiment (se) object from Salmon counts +## Create SummarizedExperiment (se) object from counts args <- commandArgs(trailingOnly = TRUE) if (length(args) < 3) { diff --git a/conf/modules.config b/conf/modules.config index 19e336715..6061e3c76 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -705,11 +705,9 @@ if (!params.skip_alignment && params.aligner == 'star_salmon') { if (!params.skip_qc & !params.skip_deseq2_qc) { process { withName: 'DESEQ2_QC_STAR_SALMON' { - ext.prefix = "deseq2" ext.args = { [ "--id_col 1", "--sample_suffix ''", - "--outprefix deseq2", "--count_col 3", params.deseq2_vst ? '--vst TRUE' : '' ].join(' ').trim() } @@ -770,11 +768,9 @@ if (!params.skip_alignment && params.aligner == 'star_rsem') { if (!params.skip_qc & !params.skip_deseq2_qc) { process { withName: 'DESEQ2_QC_RSEM' { - ext.prefix = "deseq2" ext.args = { [ "--id_col 1", "--sample_suffix ''", - "--outprefix deseq2", "--count_col 3", params.deseq2_vst ? '--vst TRUE' : '' ].join(' ').trim() } @@ -1085,10 +1081,10 @@ if (!params.skip_multiqc) { } // -// Salmon/ Kallisto pseudo-alignment options +// Salmon/ Kallisto pseudoalignment options // -if (params.pseudo_aligner == 'salmon') { +if (!params.skip_pseudo_alignment && params.pseudo_aligner == 'salmon') { process { withName: '.*:QUANTIFY_PSEUDO_ALIGNMENT:SALMON_QUANT' { @@ -1102,15 +1098,14 @@ if (params.pseudo_aligner == 'salmon') { } } -if (params.pseudo_aligner == 'kallisto') { +if (!params.skip_pseudo_alignment && params.pseudo_aligner == 'kallisto') { process { withName: '.*:QUANTIFY_PSEUDO_ALIGNMENT:KALLISTO_QUANT' { ext.args = params.extra_kallisto_quant_args ?: '' - publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') || filename.endsWith('_run_info.json') ? null : filename } + saveAs: { filename -> filename.equals('versions.yml') || filename.endsWith('.run_info.json') || filename.endsWith('.log.txt') ? null : filename } ] } } @@ -1150,7 +1145,6 @@ if (!params.skip_pseudo_alignment) { ext.args = { [ "--id_col 1", "--sample_suffix ''", - "--outprefix deseq2", "--count_col 3", params.deseq2_vst ? '--vst TRUE' : '' ].join(' ').trim() } diff --git a/docs/output.md b/docs/output.md index b5cec204a..0cb60b829 100644 --- a/docs/output.md +++ b/docs/output.md @@ -41,9 +41,9 @@ The pipeline is built using [Nextflow](https://www.nextflow.io/) and processes d - [featureCounts](#featurecounts) - Read counting relative to gene biotype - [DESeq2](#deseq2) - PCA plot and sample pairwise distance heatmap and dendrogram - [MultiQC](#multiqc) - Present QC for raw reads, alignment, read counting and sample similiarity -- [Pseudo-alignment and quantification](#pseudo-alignment-and-quantification) - - [Salmon](#salmon) - Wicked fast gene and isoform quantification relative to the transcriptome - - [Kallisto](#kallisto) - Near-optimal probabilistic RNA-seq quantification +- [Pseudoalignment and quantification](#pseudoalignment-and-quantification) + - [Salmon](#pseudoalignment) - Wicked fast gene and isoform quantification relative to the transcriptome + - [Kallisto](#pseudoalignment) - Near-optimal probabilistic RNA-seq quantification Wicked fast gene and isoform quantification relative to the transcriptome - [Workflow reporting and genomes](#workflow-reporting-and-genomes) - [Reference genome files](#reference-genome-files) - Saving reference genome indices/files @@ -205,7 +205,7 @@ The STAR section of the MultiQC report shows a bar plot with alignment rates: go ![MultiQC - STAR alignment scores plot](images/mqc_star.png) -[Salmon](https://salmon.readthedocs.io/en/latest/salmon.html) from [Ocean Genomics](https://oceangenomics.com/) and [Kallisto](https://pachterlab.github.io/kallisto/), from the Pachter Lab, are provided as options for pseudo-alignment. Both allow quantification of reads against an index generated from a reference set of target transcripts. By default, the transcriptome-level BAM files generated by STAR are provided to Salmon for downstream quantification, and Kallisto is not an option here (it does not allow BAM file input). But you can provide FASTQ files directly as input to either Salmon or Kallisto in order to pseudo-align and quantify your data by providing the `--pseudo_aligner (salmon or kallisto)` parameter. See the [Salmon](#salmon) and (Kallisto)[#kallisto] results sections for more details. +[Salmon](https://salmon.readthedocs.io/en/latest/salmon.html) from [Ocean Genomics](https://oceangenomics.com/) and [Kallisto](https://pachterlab.github.io/kallisto/), from the Pachter Lab, are provided as options for pseudoalignment. Both allow quantification of reads against an index generated from a reference set of target transcripts. By default, the transcriptome-level BAM files generated by STAR are provided to Salmon for downstream quantification, and Kallisto is not an option here (it does not allow BAM file input). But you can provide FASTQ files directly as input to either Salmon or Kallisto in order to pseudoalign and quantify your data by providing the `--pseudo_aligner salmon` or `--pseudo_aligner kallisto` parameter. See the [Salmon](#pseudoalignment) and [Kallisto](#pseudoalignment) results sections for more details. ### STAR via RSEM @@ -670,9 +670,9 @@ The plot on the left hand side shows the standard PC plot - notice the variable Results generated by MultiQC collate pipeline QC from supported tools i.e. FastQC, Cutadapt, SortMeRNA, STAR, RSEM, HISAT2, Salmon, SAMtools, Picard, RSeQC, Qualimap, Preseq and featureCounts. Additionally, various custom content has been added to the report to assess the output of dupRadar, DESeq2 and featureCounts biotypes, and to highlight samples failing a mimimum mapping threshold or those that failed to match the strand-specificity provided in the input samplesheet. The pipeline has special steps which also allow the software versions to be reported in the MultiQC output for future traceability. For more information about how to use MultiQC reports, see . -## Pseudo-alignment and quantification +## Pseudoalignment and quantification -### Pseudo-alignment +### Pseudoalignment The principal output files are the same between Salmon and Kallsto: @@ -717,10 +717,10 @@ An additional subset of files are distinct to each tool, for Salmon: - `abundance.h5`: a HDF5 binary file containing run info, abundance esimates, bootstrap estimates, and transcript length information length. This file can be read in by [sleuth](https://pachterlab.github.io/sleuth/about). - `abundance.tsv`: a plaintext file of the abundance estimates. It does not contains bootstrap estimates. - `run_info.json`: a json file containing information about the run. -- `.log.txt`: standard output from the Kallisto process per sample. + - `kallisto_quant.log`: standard output from the Kallisto process per sample. -As described in the [STAR and Salmon](#star-and-salmon) section, you can choose to pseudo-align and quantify your data with [Salmon](https://salmon.readthedocs.io/en/latest/salmon.html) or [Kallisto](https://pachterlab.github.io/kallisto/) by providing the `--pseudo_aligner` parameter. By default, Salmon is run in addition to the standard alignment workflow defined by `--aligner`, mainly because it allows you to obtain QC metrics with respect to the genomic alignments. However, you can provide the `--skip_alignment` parameter if you would like to run Salmon or Kallisto in isolation. If Salmon or Kallisto are run in isolation, the outputs mentioned above will be found in a folder named `salmon` or `kallisto`. If Salmon is run alongside STAR, the folder will be named `star_salmon`. +As described in the [STAR and Salmon](#star-and-salmon) section, you can choose to pseudoalign and quantify your data with [Salmon](https://salmon.readthedocs.io/en/latest/salmon.html) or [Kallisto](https://pachterlab.github.io/kallisto/) by providing the `--pseudo_aligner` parameter. By default, Salmon is run in addition to the standard alignment workflow defined by `--aligner`, mainly because it allows you to obtain QC metrics with respect to the genomic alignments. However, you can provide the `--skip_alignment` parameter if you would like to run Salmon or Kallisto in isolation. If Salmon or Kallisto are run in isolation, the outputs mentioned above will be found in a folder named `salmon` or `kallisto`. If Salmon is run alongside STAR, the folder will be named `star_salmon`. Transcripts with large inferential uncertainty won't be assigned the exact number of reads reproducibly, every time Salmon is run. Read more about this on the [nf-core/rnaseq](https://github.com/nf-core/rnaseq/issues/585) and [salmon](https://github.com/COMBINE-lab/salmon/issues/613) Github repos. diff --git a/docs/usage.md b/docs/usage.md index c543ec033..9c5cd902c 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -65,19 +65,19 @@ An [example samplesheet](../assets/samplesheet.csv) has been provided with the p By default, the pipeline uses [STAR](https://github.com/alexdobin/STAR) (i.e. `--aligner star_salmon`) to map the raw FastQ reads to the reference genome, project the alignments onto the transcriptome and to perform the downstream BAM-level quantification with [Salmon](https://salmon.readthedocs.io/en/latest/salmon.html). STAR is fast but requires a lot of memory to run, typically around 38GB for the Human GRCh37 reference genome. Since the [RSEM](https://github.com/deweylab/RSEM) (i.e. `--aligner star_rsem`) workflow in the pipeline also uses STAR you should use the [HISAT2](https://ccb.jhu.edu/software/hisat2/index.shtml) aligner (i.e. `--aligner hisat2`) if you have memory limitations. -You also have the option to pseudo-align and quantify your data directly with [Salmon](https://salmon.readthedocs.io/en/latest/salmon.html) or [Kallisto](https://pachterlab.github.io/kallisto/) by specifying `salmon` or `kallisto` to the `--pseudo_aligner` parameter. The selected pseudoaligner will then be run in addition to the standard alignment workflow defined by `--aligner`, mainly because it allows you to obtain QC metrics with respect to the genomic alignments. However, you can provide the `--skip_alignment` parameter if you would like to run Salmon or Kallisto in isolation. By default, the pipeline will use the genome fasta and gtf file to generate the transcripts fasta file, and then to build the Salmon index. You can override these parameters using the `--transcript_fasta` and `--salmon_index` parameters, respectively. +You also have the option to pseudoalign and quantify your data directly with [Salmon](https://salmon.readthedocs.io/en/latest/salmon.html) or [Kallisto](https://pachterlab.github.io/kallisto/) by specifying `salmon` or `kallisto` to the `--pseudo_aligner` parameter. The selected pseudoaligner will then be run in addition to the standard alignment workflow defined by `--aligner`, mainly because it allows you to obtain QC metrics with respect to the genomic alignments. However, you can provide the `--skip_alignment` parameter if you would like to run Salmon or Kallisto in isolation. By default, the pipeline will use the genome fasta and gtf file to generate the transcripts fasta file, and then to build the Salmon index. You can override these parameters using the `--transcript_fasta` and `--salmon_index` parameters, respectively. -The library preparation protocol (library type) used by Salmon quantification is inferred by the pipeline based on the information provided in the samplesheet, however, you can override it using the `--salmon_quant_libtype` parameter. You can find the available options in the [Salmon documentation](https://salmon.readthedocs.io/en/latest/library_type.html). Similarly, strandedness is taken from the sample sheet or calculated automatically, and passed to Kallisto on a per-library basis, but you can apply a global override by setting `kallisto_quant_strandednes` see the [Kallisto documentation](https://pachterlab.github.io/kallisto/manual). +The library preparation protocol (library type) used by Salmon quantification is inferred by the pipeline based on the information provided in the samplesheet, however, you can override it using the `--salmon_quant_libtype` parameter. You can find the available options in the [Salmon documentation](https://salmon.readthedocs.io/en/latest/library_type.html). Similarly, strandedness is taken from the sample sheet or calculated automatically, and passed to Kallisto on a per-library basis, but you can apply a global override by setting `--kallisto_quant_strandedness` see the [Kallisto documentation](https://pachterlab.github.io/kallisto/manual). When running Salmon in mapping-based mode via `--pseudo_aligner salmon` the entire genome of the organism is used by default for the decoy-aware transcriptome when creating the indices (see second bulleted option in [Salmon documentation](https://salmon.readthedocs.io/en/latest/salmon.html#preparing-transcriptome-indices-mapping-based-mode)). Two additional parameters `--extra_star_align_args` and `--extra_salmon_quant_args` were added in v3.10 of the pipeline that allow you to append any custom parameters to the STAR align and Salmon quant commands, respectively. Note, the `--seqBias` and `--gcBias` are not provided to Salmon quant by default so you can provide these via `--extra_salmon_quant_args '--seqBias --gcBias'` if required. You can now also supply additional arguments to Kallisto via `--extra_kallisto_quant_args`. -> **NB:** You can use `--skip_alignment --skip_pseudo_alignment` if you only want to run the pre-processing QC steps in the pipeline like FastQ, trimming etc. This will skip alignment, pseudo-alignment and any post-alignment processing steps. +> **NB:** You can use `--skip_alignment --skip_pseudo_alignment` if you only want to run the pre-processing QC steps in the pipeline like FastQ, trimming etc. This will skip alignment, pseudoalignment and any post-alignment processing steps. ## Quantification options -The current options align with STAR and quantify using either Salmon (`--aligner star_salmon`) / RSEM (`--aligner star_rsem`). You also have the option to pseudo-align and quantify your data with Salmon or Kallisto by providing the `--pseudo_aligner salmon` or `--pseudo_aligner kallisto` parameter, respectively. +The current options align with STAR and quantify using either Salmon (`--aligner star_salmon`) / RSEM (`--aligner star_rsem`). You also have the option to pseudoalign and quantify your data with Salmon or Kallisto by providing the `--pseudo_aligner salmon` or `--pseudo_aligner kallisto` parameter, respectively. Since v3.0 of the pipeline, featureCounts is no longer used to perform gene/transcript quantification, however it is still used to generate QC metrics based on [biotype](http://www.ensembl.org/info/genome/genebuild/biotypes.html) information available within GFF/GTF genome annotation files. This decision was made primarily because of the limitations of featureCounts to appropriately quantify gene expression data. Please see [Zhao et al., 2015](https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0141910#pone-0141910-t001) and [Soneson et al., 2015](https://f1000research.com/articles/4-1521/v1). diff --git a/modules/nf-core/kallisto/quant/main.nf b/modules/nf-core/kallisto/quant/main.nf index 49ec2cee2..983887b98 100644 --- a/modules/nf-core/kallisto/quant/main.nf +++ b/modules/nf-core/kallisto/quant/main.nf @@ -56,7 +56,11 @@ process KALLISTO_QUANT { ${reads} 2> >(tee -a ${prefix}.log.txt >&2) if [ -f $prefix/run_info.json ]; then - cp $prefix/run_info.json "${prefix}_run_info.json" + cp $prefix/run_info.json "${prefix}.run_info.json" + fi + + if [ -f ${prefix}.log.txt ]; then + cp ${prefix}.log.txt ${prefix}/kallisto_quant.log fi cat <<-END_VERSIONS > versions.yml diff --git a/nextflow_schema.json b/nextflow_schema.json index 55ff1d843..47171d860 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -376,11 +376,28 @@ "fa_icon": "fas fa-fast-forward", "description": " Override Salmon library type inferred based on strandedness defined in meta object.", "help_text": "See [Salmon docs](https://salmon.readthedocs.io/en/latest/library_type.html).", - "enum": ["A,U,SF,SR,IS,IU,ISF,ISR,OS,OU,OSF,OSR,MS,MU,MSF,MSR"] + "enum": [ + "A", + "U", + "SF", + "SR", + "IS", + "IU", + "ISF", + "ISR", + "OS", + "OU", + "OSF", + "OSR", + "MS", + "MU", + "MSF", + "MSR" + ] }, "min_mapped_reads": { "type": "number", - "default": 5, + "default": 5.0, "fa_icon": "fas fa-percentage", "description": "Minimum percentage of uniquely mapped reads below which samples are removed from further processing.", "help_text": "Some downstream steps in the pipeline will fail if this threshold is too low." @@ -420,13 +437,13 @@ "kallisto_quant_fraglen": { "type": "integer", "description": "In single-end mode Kallisto requires an estimated fragment length. Specify a default value for that here. TODO: use existing RSeQC results to do this dynamically.", - "default": 150, + "default": 200, "fa_icon": "fas fa-ruler-horizontal" }, "kallisto_quant_fraglen_sd": { "type": "integer", - "description": "In single-end mode Kallisto requires an estimated standard error for fragment length. Specify a default value for that here. TODO: use existing RSeQC results to do this dynamically.", - "default": 75, + "description": "In single-end mode, Kallisto requires an estimated standard error for fragment length. Specify a default value for that here. TODO: use existing RSeQC results to do this dynamically.", + "default": 200, "fa_icon": "fas fa-sort-amount-up-alt" } } @@ -535,7 +552,7 @@ "skip_pseudo_alignment": { "type": "boolean", "fa_icon": "fas fa-fast-forward", - "description": "Skip all of the pseudo-alignment-based processes within the pipeline." + "description": "Skip all of the pseudoalignment-based processes within the pipeline." }, "skip_markduplicates": { "type": "boolean", diff --git a/subworkflows/local/quantify_pseudo/main.nf b/subworkflows/local/quantify_pseudo/main.nf index 4688bcca4..886b45e7b 100644 --- a/subworkflows/local/quantify_pseudo/main.nf +++ b/subworkflows/local/quantify_pseudo/main.nf @@ -1,11 +1,11 @@ // -// Pseudo-alignment and quantification with Salmon or Kallisto +// Pseudoalignment and quantification with Salmon or Kallisto // -include { SALMON_QUANT } from '../../../modules/nf-core/salmon/quant' -include { KALLISTO_QUANT } from '../../../modules/nf-core/kallisto/quant' -include { TX2GENE } from '../../../modules/local/tx2gene' -include { TXIMPORT } from '../../../modules/local/tximport' +include { SALMON_QUANT } from '../../../modules/nf-core/salmon/quant' +include { KALLISTO_QUANT } from '../../../modules/nf-core/kallisto/quant' +include { TX2GENE } from '../../../modules/local/tx2gene' +include { TXIMPORT } from '../../../modules/local/tximport' include { SUMMARIZEDEXPERIMENT as SE_GENE } from '../../../modules/local/summarizedexperiment' include { SUMMARIZEDEXPERIMENT as SE_GENE_LENGTH_SCALED } from '../../../modules/local/summarizedexperiment' @@ -14,13 +14,15 @@ include { SUMMARIZEDEXPERIMENT as SE_TRANSCRIPT } from '../../../modules workflow QUANTIFY_PSEUDO_ALIGNMENT { take: - reads // channel: [ val(meta), [ reads ] ] - index // channel: /path/to//index/ - transcript_fasta // channel: /path/to/transcript.fasta - gtf // channel: /path/to/genome.gtf - pseudo_aligner // val: kallisto or salmon - alignment_mode // bool: Run Salmon in alignment mode - lib_type // val: String to override salmon library type + reads // channel: [ val(meta), [ reads ] ] + index // channel: /path/to//index/ + transcript_fasta // channel: /path/to/transcript.fasta + gtf // channel: /path/to/genome.gtf + pseudo_aligner // val: kallisto or salmon + alignment_mode // bool: Run Salmon in alignment mode + lib_type // val: String to override Salmon library type + kallisto_quant_fraglen // val: Estimated fragment length required by Kallisto in single-end mode + kallisto_quant_fraglen_sd // val: Estimated standard error for fragment length required by Kallisto in single-end mode main: @@ -29,15 +31,14 @@ workflow QUANTIFY_PSEUDO_ALIGNMENT { // // Quantify and merge counts across samples // - - // Note: MultiQC needs Salmon outputs, but Kallisto logs + // NOTE: MultiQC needs Salmon outputs, but Kallisto logs if (pseudo_aligner == 'salmon') { SALMON_QUANT ( reads, index, gtf, transcript_fasta, alignment_mode, lib_type ) ch_pseudo_results = SALMON_QUANT.out.results ch_pseudo_multiqc = ch_pseudo_results ch_versions = ch_versions.mix(SALMON_QUANT.out.versions.first()) - }else { - KALLISTO_QUANT ( reads, index, gtf, [], params.kallisto_quant_fraglen, params.kallisto_quant_fraglen_sd) + } else { + KALLISTO_QUANT ( reads, index, gtf, [], kallisto_quant_fraglen, kallisto_quant_fraglen_sd) ch_pseudo_results = KALLISTO_QUANT.out.results ch_pseudo_multiqc = KALLISTO_QUANT.out.log ch_versions = ch_versions.mix(KALLISTO_QUANT.out.versions.first()) diff --git a/tower.yml b/tower.yml index 818ef13bf..21f821a53 100644 --- a/tower.yml +++ b/tower.yml @@ -30,6 +30,7 @@ reports: "**/kallisto/kallisto.merged.transcript_counts.tsv": display: "All samples Kallisto merged transcript raw counts" "**/kallisto/kallisto.merged.transcript_tpm.tsv": + display: "All samples Kallisto merged transcript TPM counts" "**/star_rsem/**/deseq2.plots.pdf": display: "All samples STAR RSEM DESeq2 QC PDF plots" "**/star_rsem/rsem.merged.gene_counts.tsv": diff --git a/workflows/rnaseq.nf b/workflows/rnaseq.nf index 3632e1a84..66ee061f5 100755 --- a/workflows/rnaseq.nf +++ b/workflows/rnaseq.nf @@ -236,7 +236,7 @@ workflow RNASEQ { .set { ch_strand_fastq } // - // SUBWORKFLOW: Sub-sample FastQ files and pseudo-align with Salmon to auto-infer strandedness + // SUBWORKFLOW: Sub-sample FastQ files and pseudoalign with Salmon to auto-infer strandedness // // Return empty channel if ch_strand_fastq.auto_strand is empty so salmon index isn't created PREPARE_GENOME.out.fasta @@ -477,7 +477,9 @@ workflow RNASEQ { PREPARE_GENOME.out.gtf, 'salmon', true, - params.salmon_quant_libtype ?: '' + params.salmon_quant_libtype ?: '', + params.kallisto_quant_fraglen, + params.kallisto_quant_fraglen_sd ) ch_versions = ch_versions.mix(QUANTIFY_STAR_SALMON.out.versions) @@ -790,7 +792,7 @@ workflow RNASEQ { } // - // SUBWORKFLOW: Pseudo-alignment and quantification with Salmon + // SUBWORKFLOW: Pseudoalignment and quantification with Salmon // ch_pseudo_multiqc = Channel.empty() ch_pseudoaligner_pca_multiqc = Channel.empty() @@ -798,7 +800,7 @@ workflow RNASEQ { if (!params.skip_pseudo_alignment) { - if (params.pseudo_aligner == 'salmon'){ + if (params.pseudo_aligner == 'salmon') { ch_pseudo_index = PREPARE_GENOME.out.salmon_index } else { ch_pseudo_index = PREPARE_GENOME.out.kallisto_index @@ -811,11 +813,13 @@ workflow RNASEQ { PREPARE_GENOME.out.gtf, params.pseudo_aligner, false, - params.salmon_quant_libtype ?: '' + params.salmon_quant_libtype ?: '', + params.kallisto_quant_fraglen, + params.kallisto_quant_fraglen_sd ) - ch_pseudo_multiqc = QUANTIFY_PSEUDO_ALIGNMENT.out.multiqc - ch_versions = ch_versions.mix(QUANTIFY_PSEUDO_ALIGNMENT.out.versions) + ch_pseudo_multiqc = QUANTIFY_PSEUDO_ALIGNMENT.out.multiqc ch_counts_gene_length_scaled = QUANTIFY_PSEUDO_ALIGNMENT.out.counts_gene_length_scaled + ch_versions = ch_versions.mix(QUANTIFY_PSEUDO_ALIGNMENT.out.versions) if (!params.skip_qc & !params.skip_deseq2_qc) { DESEQ2_QC_PSEUDO ( From 0c9c0aaa2708536bc2c2e8f900afaee78e09390e Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Tue, 14 Nov 2023 13:03:57 +0000 Subject: [PATCH 151/184] Fix Kallisto sample name rendering in MultiQC --- assets/multiqc_config.yml | 1 + docs/output.md | 2 +- docs/usage.md | 6 +++--- 3 files changed, 5 insertions(+), 4 deletions(-) diff --git a/assets/multiqc_config.yml b/assets/multiqc_config.yml index cbc8db099..100536f4e 100644 --- a/assets/multiqc_config.yml +++ b/assets/multiqc_config.yml @@ -67,6 +67,7 @@ extra_fn_clean_exts: - ".umi_dedup" - "_val" - ".markdup" + - "_primary" # Customise the module search patterns to speed up execution time # - Skip module sub-tools that we are not interested in diff --git a/docs/output.md b/docs/output.md index 0cb60b829..f5d272c6b 100644 --- a/docs/output.md +++ b/docs/output.md @@ -718,7 +718,7 @@ An additional subset of files are distinct to each tool, for Salmon: - `abundance.tsv`: a plaintext file of the abundance estimates. It does not contains bootstrap estimates. - `run_info.json`: a json file containing information about the run. - `kallisto_quant.log`: standard output from the Kallisto process per sample. - + As described in the [STAR and Salmon](#star-and-salmon) section, you can choose to pseudoalign and quantify your data with [Salmon](https://salmon.readthedocs.io/en/latest/salmon.html) or [Kallisto](https://pachterlab.github.io/kallisto/) by providing the `--pseudo_aligner` parameter. By default, Salmon is run in addition to the standard alignment workflow defined by `--aligner`, mainly because it allows you to obtain QC metrics with respect to the genomic alignments. However, you can provide the `--skip_alignment` parameter if you would like to run Salmon or Kallisto in isolation. If Salmon or Kallisto are run in isolation, the outputs mentioned above will be found in a folder named `salmon` or `kallisto`. If Salmon is run alongside STAR, the folder will be named `star_salmon`. diff --git a/docs/usage.md b/docs/usage.md index 9c5cd902c..869e5baea 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -93,9 +93,9 @@ The `--umitools_grouping_method` parameter affects [how similar, but non-identic #### Examples: -| UMI type | Source | Pipeline parameters | -| ------------ | ---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------- | -| In read name | [Illumina BCL convert >3.7.5](https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/software_documentation/bcl_convert/bcl-convert-v3-7-5-software-guide-1000000163594-00.pdf) | `--with_umi --skip_umi_extract --umitools_umi_separator ":"` | +| UMI type | Source | Pipeline parameters | +| ------------ | --------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------- | +| In read name | [Illumina BCL convert >3.7.5](https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/software_documentation/bcl_convert/bcl-convert-v3-7-5-software-guide-1000000163594-00.pdf) | `--with_umi --skip_umi_extract --umitools_umi_separator ":"` | | In sequence | [Lexogen QuantSeq® 3’ mRNA-Seq V2 FWD](https://www.lexogen.com/quantseq-3mrna-sequencing) + [UMI Second Strand Synthesis Module](https://faqs.lexogen.com/faq/how-can-i-add-umis-to-my-quantseq-libraries) | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern "^(?P.{6})(?P.{4}).*"` | | In sequence | [Lexogen CORALL® Total RNA-Seq V1](https://www.lexogen.com/corall-total-rna-seq/)
    > _mind [Appendix H](https://www.lexogen.com/wp-content/uploads/2020/04/095UG190V0130_CORALL-Total-RNA-Seq_2020-03-31.pdf) regarding optional trimming_ | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern "^(?P.{12}).*"`
    Optional: `--clip_r2 9 --three_prime_clip_r2 12` | | In sequence | [Takara Bio SMARTer® Stranded Total RNA-Seq Kit v3](https://www.takarabio.com/documents/User%20Manual/SMARTer%20Stranded%20Total%20RNA/SMARTer%20Stranded%20Total%20RNA-Seq%20Kit%20v3%20-%20Pico%20Input%20Mammalian%20User%20Manual-a_114949.pdf) | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern2 "^(?P.{8})(?P.{6}).*"` | From 2decfdc87b99dfdff87bf363146ce1ac626e3f30 Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Tue, 14 Nov 2023 14:28:10 +0000 Subject: [PATCH 152/184] [skip ci] Alphabetically order enum fields for salmon strandedness --- nextflow_schema.json | 18 +++++++++--------- 1 file changed, 9 insertions(+), 9 deletions(-) diff --git a/nextflow_schema.json b/nextflow_schema.json index 47171d860..26e1e2f5f 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -378,21 +378,21 @@ "help_text": "See [Salmon docs](https://salmon.readthedocs.io/en/latest/library_type.html).", "enum": [ "A", - "U", - "SF", - "SR", "IS", - "IU", "ISF", "ISR", + "IU", + "MS", + "MSF", + "MSR", + "MU", "OS", - "OU", "OSF", "OSR", - "MS", - "MU", - "MSF", - "MSR" + "OU", + "SF", + "SR", + "U" ] }, "min_mapped_reads": { From 5ea79acc448d5a85ed71bbdcb8ae6f22eabe04bc Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 14 Nov 2023 15:08:36 +0000 Subject: [PATCH 153/184] [skip ci] Bump module from branch --- modules.json | 4 ++-- .../nf-core/kallisto/quant/environment.yml | 1 + modules/nf-core/kallisto/quant/main.nf | 23 +++++++++++-------- modules/nf-core/kallisto/quant/meta.yml | 2 +- .../kallisto/quant/tests/main.nf.test.snap | 6 ++--- 5 files changed, 20 insertions(+), 16 deletions(-) diff --git a/modules.json b/modules.json index 3bd2a9132..d06dfe193 100644 --- a/modules.json +++ b/modules.json @@ -71,8 +71,8 @@ "installed_by": ["modules"] }, "kallisto/quant": { - "branch": "master", - "git_sha": "2680fb696b197fceb2113790b19364a8da35eab0", + "branch": "kallisto_updates", + "git_sha": "bc4719dcd079fcdb650ddeac05739c2f7dd58c84", "installed_by": ["modules"] }, "picard/markduplicates": { diff --git a/modules/nf-core/kallisto/quant/environment.yml b/modules/nf-core/kallisto/quant/environment.yml index d62586580..c2d6306bb 100644 --- a/modules/nf-core/kallisto/quant/environment.yml +++ b/modules/nf-core/kallisto/quant/environment.yml @@ -1,3 +1,4 @@ +name: kallisto_quant channels: - conda-forge - bioconda diff --git a/modules/nf-core/kallisto/quant/main.nf b/modules/nf-core/kallisto/quant/main.nf index 49ec2cee2..2ad0bb78b 100644 --- a/modules/nf-core/kallisto/quant/main.nf +++ b/modules/nf-core/kallisto/quant/main.nf @@ -16,10 +16,10 @@ process KALLISTO_QUANT { val fragment_length_sd output: - tuple val(meta), path("${prefix}") , emit: results - tuple val(meta), path("*info.json"), emit: json_info - tuple val(meta), path("*.log.txt") , emit: log - path "versions.yml" , emit: versions + tuple val(meta), path("${prefix}") , emit: results + tuple val(meta), path("*.run_info.json") , emit: json_info + tuple val(meta), path("*.log") , emit: log + path "versions.yml" , emit: versions when: task.ext.when == null || task.ext.when @@ -41,10 +41,14 @@ process KALLISTO_QUANT { single_end_params = "--single --fragment-length=${fragment_length} --sd=${fragment_length_sd}" } - def strandedness = (meta.strandedness == 'forward') ? '--fr-stranded' : + def strandedness = '' + if (!args.contains('--fr-stranded') && !args.contains('--rf-stranded')) { + strandedness = (meta.strandedness == 'forward') ? '--fr-stranded' : (meta.strandedness == 'reverse') ? '--rf-stranded' : '' + } + """ - kallisto quant \\ + mkdir -p $prefix && kallisto quant \\ --threads ${task.cpus} \\ --index ${index} \\ ${gtf_input} \\ @@ -53,11 +57,10 @@ process KALLISTO_QUANT { ${strandedness} \\ ${args} \\ -o $prefix \\ - ${reads} 2> >(tee -a ${prefix}.log.txt >&2) + ${reads} 2> >(tee -a ${prefix}/kallisto_quant.log >&2) - if [ -f $prefix/run_info.json ]; then - cp $prefix/run_info.json "${prefix}_run_info.json" - fi + cp ${prefix}/kallisto_quant.log ${prefix}.log + cp ${prefix}/run_info.json ${prefix}.run_info.json cat <<-END_VERSIONS > versions.yml "${task.process}": diff --git a/modules/nf-core/kallisto/quant/meta.yml b/modules/nf-core/kallisto/quant/meta.yml index 2dce7b639..d5100290f 100644 --- a/modules/nf-core/kallisto/quant/meta.yml +++ b/modules/nf-core/kallisto/quant/meta.yml @@ -11,7 +11,7 @@ tools: documentation: https://pachterlab.github.io/kallisto/manual tool_dev_url: https://github.com/pachterlab/kallisto doi: "10.1038/nbt.3519" - licence: "['BSD_2_clause']" + licence: ["BSD_2_clause"] input: - meta: type: map diff --git a/modules/nf-core/kallisto/quant/tests/main.nf.test.snap b/modules/nf-core/kallisto/quant/tests/main.nf.test.snap index b2a7beebd..d59d7f06f 100644 --- a/modules/nf-core/kallisto/quant/tests/main.nf.test.snap +++ b/modules/nf-core/kallisto/quant/tests/main.nf.test.snap @@ -7,7 +7,7 @@ "id": "test", "single_end": false }, - "test.log.txt:md5,8a5987f8e779cd12ca708e2212f771f5" + "test.log:md5,8a5987f8e779cd12ca708e2212f771f5" ] ] ], @@ -35,7 +35,7 @@ "id": "test", "single_end": true }, - "test.log.txt:md5,9c166f0c50cd4fdbdbf1bff9d5d8aba2" + "test.log:md5,9c166f0c50cd4fdbdbf1bff9d5d8aba2" ] ] ], @@ -55,4 +55,4 @@ ], "timestamp": "2023-11-02T09:16:05.168753684" } -} \ No newline at end of file +} From 83d9fb4ae341006bbd4594bd306aa686187c0782 Mon Sep 17 00:00:00 2001 From: nf-core-bot Date: Tue, 14 Nov 2023 17:29:17 +0000 Subject: [PATCH 154/184] [automated] Fix linting with Prettier --- docs/usage.md | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/docs/usage.md b/docs/usage.md index 869e5baea..9c5cd902c 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -93,9 +93,9 @@ The `--umitools_grouping_method` parameter affects [how similar, but non-identic #### Examples: -| UMI type | Source | Pipeline parameters | -| ------------ | --------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------- | -| In read name | [Illumina BCL convert >3.7.5](https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/software_documentation/bcl_convert/bcl-convert-v3-7-5-software-guide-1000000163594-00.pdf) | `--with_umi --skip_umi_extract --umitools_umi_separator ":"` | +| UMI type | Source | Pipeline parameters | +| ------------ | ---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------- | +| In read name | [Illumina BCL convert >3.7.5](https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/software_documentation/bcl_convert/bcl-convert-v3-7-5-software-guide-1000000163594-00.pdf) | `--with_umi --skip_umi_extract --umitools_umi_separator ":"` | | In sequence | [Lexogen QuantSeq® 3’ mRNA-Seq V2 FWD](https://www.lexogen.com/quantseq-3mrna-sequencing) + [UMI Second Strand Synthesis Module](https://faqs.lexogen.com/faq/how-can-i-add-umis-to-my-quantseq-libraries) | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern "^(?P.{6})(?P.{4}).*"` | | In sequence | [Lexogen CORALL® Total RNA-Seq V1](https://www.lexogen.com/corall-total-rna-seq/)
    > _mind [Appendix H](https://www.lexogen.com/wp-content/uploads/2020/04/095UG190V0130_CORALL-Total-RNA-Seq_2020-03-31.pdf) regarding optional trimming_ | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern "^(?P.{12}).*"`
    Optional: `--clip_r2 9 --three_prime_clip_r2 12` | | In sequence | [Takara Bio SMARTer® Stranded Total RNA-Seq Kit v3](https://www.takarabio.com/documents/User%20Manual/SMARTer%20Stranded%20Total%20RNA/SMARTer%20Stranded%20Total%20RNA-Seq%20Kit%20v3%20-%20Pico%20Input%20Mammalian%20User%20Manual-a_114949.pdf) | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern2 "^(?P.{8})(?P.{6}).*"` | From 6c75322180c7743a68c6815fa46f91f4424721b8 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 14 Nov 2023 17:32:50 +0000 Subject: [PATCH 155/184] [skip ci] removed kallisto strandedness as a standalone parameter --- docs/usage.md | 8 ++++---- nextflow.config | 1 - nextflow_schema.json | 7 ------- 3 files changed, 4 insertions(+), 12 deletions(-) diff --git a/docs/usage.md b/docs/usage.md index 9c5cd902c..d53ff77a5 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -67,7 +67,7 @@ By default, the pipeline uses [STAR](https://github.com/alexdobin/STAR) (i.e. `- You also have the option to pseudoalign and quantify your data directly with [Salmon](https://salmon.readthedocs.io/en/latest/salmon.html) or [Kallisto](https://pachterlab.github.io/kallisto/) by specifying `salmon` or `kallisto` to the `--pseudo_aligner` parameter. The selected pseudoaligner will then be run in addition to the standard alignment workflow defined by `--aligner`, mainly because it allows you to obtain QC metrics with respect to the genomic alignments. However, you can provide the `--skip_alignment` parameter if you would like to run Salmon or Kallisto in isolation. By default, the pipeline will use the genome fasta and gtf file to generate the transcripts fasta file, and then to build the Salmon index. You can override these parameters using the `--transcript_fasta` and `--salmon_index` parameters, respectively. -The library preparation protocol (library type) used by Salmon quantification is inferred by the pipeline based on the information provided in the samplesheet, however, you can override it using the `--salmon_quant_libtype` parameter. You can find the available options in the [Salmon documentation](https://salmon.readthedocs.io/en/latest/library_type.html). Similarly, strandedness is taken from the sample sheet or calculated automatically, and passed to Kallisto on a per-library basis, but you can apply a global override by setting `--kallisto_quant_strandedness` see the [Kallisto documentation](https://pachterlab.github.io/kallisto/manual). +The library preparation protocol (library type) used by Salmon quantification is inferred by the pipeline based on the information provided in the samplesheet, however, you can override it using the `--salmon_quant_libtype` parameter. You can find the available options in the [Salmon documentation](https://salmon.readthedocs.io/en/latest/library_type.html). Similarly, strandedness is taken from the sample sheet or calculated automatically, and passed to Kallisto on a per-library basis, but you can apply a global override by setting the Kallisto strandedness parameters in `--extra_kallisto_quant_args` like `--extra_kallisto_quant_args '--fr-stranded'` see the [Kallisto documentation](https://pachterlab.github.io/kallisto/manual). When running Salmon in mapping-based mode via `--pseudo_aligner salmon` the entire genome of the organism is used by default for the decoy-aware transcriptome when creating the indices (see second bulleted option in [Salmon documentation](https://salmon.readthedocs.io/en/latest/salmon.html#preparing-transcriptome-indices-mapping-based-mode)). @@ -93,9 +93,9 @@ The `--umitools_grouping_method` parameter affects [how similar, but non-identic #### Examples: -| UMI type | Source | Pipeline parameters | -| ------------ | ---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------- | -| In read name | [Illumina BCL convert >3.7.5](https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/software_documentation/bcl_convert/bcl-convert-v3-7-5-software-guide-1000000163594-00.pdf) | `--with_umi --skip_umi_extract --umitools_umi_separator ":"` | +| UMI type | Source | Pipeline parameters | +| ------------ | --------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------- | +| In read name | [Illumina BCL convert >3.7.5](https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/software_documentation/bcl_convert/bcl-convert-v3-7-5-software-guide-1000000163594-00.pdf) | `--with_umi --skip_umi_extract --umitools_umi_separator ":"` | | In sequence | [Lexogen QuantSeq® 3’ mRNA-Seq V2 FWD](https://www.lexogen.com/quantseq-3mrna-sequencing) + [UMI Second Strand Synthesis Module](https://faqs.lexogen.com/faq/how-can-i-add-umis-to-my-quantseq-libraries) | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern "^(?P.{6})(?P.{4}).*"` | | In sequence | [Lexogen CORALL® Total RNA-Seq V1](https://www.lexogen.com/corall-total-rna-seq/)
    > _mind [Appendix H](https://www.lexogen.com/wp-content/uploads/2020/04/095UG190V0130_CORALL-Total-RNA-Seq_2020-03-31.pdf) regarding optional trimming_ | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern "^(?P.{12}).*"`
    Optional: `--clip_r2 9 --three_prime_clip_r2 12` | | In sequence | [Takara Bio SMARTer® Stranded Total RNA-Seq Kit v3](https://www.takarabio.com/documents/User%20Manual/SMARTer%20Stranded%20Total%20RNA/SMARTer%20Stranded%20Total%20RNA-Seq%20Kit%20v3%20-%20Pico%20Input%20Mammalian%20User%20Manual-a_114949.pdf) | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern2 "^(?P.{8})(?P.{6}).*"` | diff --git a/nextflow.config b/nextflow.config index bc38b6124..2c8b29691 100644 --- a/nextflow.config +++ b/nextflow.config @@ -69,7 +69,6 @@ params { extra_kallisto_quant_args = null kallisto_quant_fraglen = 200 kallisto_quant_fraglen_sd = 200 - kallisto_quant_strandedness = null save_merged_fastq = false save_unaligned = false save_align_intermeds = false diff --git a/nextflow_schema.json b/nextflow_schema.json index 26e1e2f5f..ddbe77de4 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -422,13 +422,6 @@ "description": "Extra arguments to pass to Salmon quant command in addition to defaults defined by the pipeline.", "fa_icon": "fas fa-plus" }, - "kallisto_quant_strandedness": { - "type": "string", - "fa_icon": "fas fa-ribbon", - "description": "Override Kallisto library type inferred based on strandedness defined in meta object.", - "enum": ["fr-stranded", "rf-stranded"], - "help_text": "See [Kallisto docs](https://pachterlab.github.io/kallisto/manual)" - }, "extra_kallisto_quant_args": { "type": "string", "description": "Extra arguments to pass to Kallisto quant command in addition to defaults defined by the pipeline.", From 5458b5c42b156ed1f3b951c75321ff653f29fd32 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 14 Nov 2023 18:08:28 +0000 Subject: [PATCH 156/184] [skip ci] Fix kallisto log file ext --- conf/modules.config | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/conf/modules.config b/conf/modules.config index 6061e3c76..5b46659cf 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -1105,7 +1105,7 @@ if (!params.skip_pseudo_alignment && params.pseudo_aligner == 'kallisto') { publishDir = [ path: { "${params.outdir}/${params.pseudo_aligner}" }, mode: params.publish_dir_mode, - saveAs: { filename -> filename.equals('versions.yml') || filename.endsWith('.run_info.json') || filename.endsWith('.log.txt') ? null : filename } + saveAs: { filename -> filename.equals('versions.yml') || filename.endsWith('.run_info.json') || filename.endsWith('.log') ? null : filename } ] } } From 68b897791573d8423c56a6b4c9549d7b2f1e6f50 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Tue, 14 Nov 2023 20:08:59 +0000 Subject: [PATCH 157/184] Fix tiny issue --- docs/output.md | 1 - 1 file changed, 1 deletion(-) diff --git a/docs/output.md b/docs/output.md index f5d272c6b..a4c559b3b 100644 --- a/docs/output.md +++ b/docs/output.md @@ -44,7 +44,6 @@ The pipeline is built using [Nextflow](https://www.nextflow.io/) and processes d - [Pseudoalignment and quantification](#pseudoalignment-and-quantification) - [Salmon](#pseudoalignment) - Wicked fast gene and isoform quantification relative to the transcriptome - [Kallisto](#pseudoalignment) - Near-optimal probabilistic RNA-seq quantification - Wicked fast gene and isoform quantification relative to the transcriptome - [Workflow reporting and genomes](#workflow-reporting-and-genomes) - [Reference genome files](#reference-genome-files) - Saving reference genome indices/files - [Pipeline information](#pipeline-information) - Report metrics generated during the workflow execution From 5ad263a6a4ea7afd8b91b37887933f8383786dff Mon Sep 17 00:00:00 2001 From: nf-core-bot Date: Tue, 14 Nov 2023 20:10:35 +0000 Subject: [PATCH 158/184] [automated] Fix linting with Prettier --- docs/usage.md | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/docs/usage.md b/docs/usage.md index d53ff77a5..0f3050907 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -93,9 +93,9 @@ The `--umitools_grouping_method` parameter affects [how similar, but non-identic #### Examples: -| UMI type | Source | Pipeline parameters | -| ------------ | --------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------- | -| In read name | [Illumina BCL convert >3.7.5](https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/software_documentation/bcl_convert/bcl-convert-v3-7-5-software-guide-1000000163594-00.pdf) | `--with_umi --skip_umi_extract --umitools_umi_separator ":"` | +| UMI type | Source | Pipeline parameters | +| ------------ | ---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------- | +| In read name | [Illumina BCL convert >3.7.5](https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/software_documentation/bcl_convert/bcl-convert-v3-7-5-software-guide-1000000163594-00.pdf) | `--with_umi --skip_umi_extract --umitools_umi_separator ":"` | | In sequence | [Lexogen QuantSeq® 3’ mRNA-Seq V2 FWD](https://www.lexogen.com/quantseq-3mrna-sequencing) + [UMI Second Strand Synthesis Module](https://faqs.lexogen.com/faq/how-can-i-add-umis-to-my-quantseq-libraries) | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern "^(?P.{6})(?P.{4}).*"` | | In sequence | [Lexogen CORALL® Total RNA-Seq V1](https://www.lexogen.com/corall-total-rna-seq/)
    > _mind [Appendix H](https://www.lexogen.com/wp-content/uploads/2020/04/095UG190V0130_CORALL-Total-RNA-Seq_2020-03-31.pdf) regarding optional trimming_ | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern "^(?P.{12}).*"`
    Optional: `--clip_r2 9 --three_prime_clip_r2 12` | | In sequence | [Takara Bio SMARTer® Stranded Total RNA-Seq Kit v3](https://www.takarabio.com/documents/User%20Manual/SMARTer%20Stranded%20Total%20RNA/SMARTer%20Stranded%20Total%20RNA-Seq%20Kit%20v3%20-%20Pico%20Input%20Mammalian%20User%20Manual-a_114949.pdf) | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern2 "^(?P.{8})(?P.{6}).*"` | From fb70fdf3d9191d4f80899a3e7927d3959288deb9 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Wed, 15 Nov 2023 10:30:15 +0000 Subject: [PATCH 159/184] Simplify GTF filtering --- bin/filter_gtf.py | 28 ++++++++--------------- modules/local/gtf_filter/main.nf | 3 +-- subworkflows/local/prepare_genome/main.nf | 16 ++++++------- workflows/rnaseq.nf | 8 +++---- 4 files changed, 22 insertions(+), 33 deletions(-) diff --git a/bin/filter_gtf.py b/bin/filter_gtf.py index 345d4a68e..dd8e1866e 100755 --- a/bin/filter_gtf.py +++ b/bin/filter_gtf.py @@ -21,7 +21,7 @@ def tab_delimited(file: str) -> float: data = f.read(1024) return statistics.median(line.count("\t") for line in data.split("\n")) -def filter_gtf(fasta: str, gtf_in: str, gtf_in_genome_out: str, gtf_transcript_out: str, skip_transcript_id_check: bool) -> None: +def filter_gtf(fasta: str, gtf_in: str, filtered_gtf_out: str, skip_transcript_id_check: bool) -> None: """Filter GTF file based on FASTA sequence names.""" if tab_delimited(gtf_in) != 8: raise ValueError("Invalid GTF file: Expected 8 tab-separated columns.") @@ -32,34 +32,26 @@ def filter_gtf(fasta: str, gtf_in: str, gtf_in_genome_out: str, gtf_transcript_o seq_names_in_gtf = set() try: - with open(gtf_in) as gtf, open(gtf_in_genome_out, "w") as out, open(gtf_transcript_out, "w") as out2: - line_count_all, line_count_transcript = 0, 0 + with open(gtf_in) as gtf, open(filtered_gtf_out, "w") as out: + line_count = 0 for line in gtf: seq_name = line.split("\t")[0] seq_names_in_gtf.add(seq_name) # Add sequence name to the set if seq_name in seq_names_in_genome: - out.write(line) - line_count_all += 1 + if skip_transcript_id_check or re.search(r'transcript_id "([^"]+)"', line): + out.write(line) + line_count += 1 - if not skip_transcript_id_check and re.search(r'transcript_id "([^"]+)"', line): - with open(gtf_transcript_out, "a") as out2: - out2.write(line) - line_count_transcript += 1 - - if line_count_all == 0: - raise ValueError("No overlapping scaffolds found.") - - if not skip_transcript_id_check and line_count_transcript == 0: - raise ValueError("No transcript_id values found in the GTF file.") + if line_count == 0: + raise ValueError("All GTF lines removed by filters") except IOError as e: logger.error(f"File operation failed: {e}") return logger.debug("All sequence IDs from GTF: " + ", ".join(sorted(seq_names_in_gtf))) - logger.info(f"Extracted {line_count_all} matching sequences from {gtf_in} into {gtf_in_genome_out}") - logger.info(f"Wrote {line_count_transcript} lines with transcript IDs to {gtf_transcript_out}") + logger.info(f"Extracted {line_count} matching sequences from {gtf_in} into {filtered_gtf_out}") if __name__ == "__main__": parser = argparse.ArgumentParser(description="Filters a GTF file based on sequence names in a FASTA file.") @@ -69,5 +61,5 @@ def filter_gtf(fasta: str, gtf_in: str, gtf_in_genome_out: str, gtf_transcript_o parser.add_argument("--skip_transcript_id_check", action='store_true', help="Skip checking for transcript IDs in the GTF file") args = parser.parse_args() - filter_gtf(args.fasta, args.gtf, args.prefix + "_in_genome.gtf", args.prefix + "_with_transcript_ids.gtf", args.skip_transcript_id_check) + filter_gtf(args.fasta, args.gtf, args.prefix + ".filtered.gtf", args.skip_transcript_id_check) diff --git a/modules/local/gtf_filter/main.nf b/modules/local/gtf_filter/main.nf index c12cf4116..cf3536f95 100644 --- a/modules/local/gtf_filter/main.nf +++ b/modules/local/gtf_filter/main.nf @@ -11,8 +11,7 @@ process GTF_FILTER { path gtf output: - path "*_in_genome.gtf" , emit: genome_gtf - path "*_with_transcript_ids.gtf" , emit: transcript_id_gtf + path "*.filtered.gtf" , emit: genome_gtf path "versions.yml" , emit: versions when: diff --git a/subworkflows/local/prepare_genome/main.nf b/subworkflows/local/prepare_genome/main.nf index e743c1dab..d639adf6c 100644 --- a/subworkflows/local/prepare_genome/main.nf +++ b/subworkflows/local/prepare_genome/main.nf @@ -90,6 +90,8 @@ workflow PREPARE_GENOME { // // Apply filtering we may need for GTFs // + + ch_filtered_gtf = ch_gtf filtering_useful = ( @@ -109,13 +111,9 @@ workflow PREPARE_GENOME { !transcript_fasta ) - if (params.skip_gtf_filter || ! filtering_useful){ - ch_gtf_with_transcript_ids = ch_gtf - ch_gtf_genome = ch_gtf - } else { + if (filtering_useful){ GTF_FILTER ( ch_fasta, ch_gtf ) - ch_gtf_with_transcript_ids = GTF_FILTER.out.transcript_id_gtf - ch_gtf_genome = GTF_FILTER.out.genome_gtf + ch_filtered_gtf = GTF_FILTER.out.genome_gtf } } @@ -166,7 +164,7 @@ workflow PREPARE_GENOME { ch_versions = ch_versions.mix(PREPROCESS_TRANSCRIPTS_FASTA_GENCODE.out.versions) } } else { - ch_transcript_fasta = MAKE_TRANSCRIPTS_FASTA ( ch_fasta, ch_gtf_genome ).transcript_fasta + ch_transcript_fasta = MAKE_TRANSCRIPTS_FASTA ( ch_fasta, ch_filtered_gtf ).transcript_fasta ch_versions = ch_versions.mix(GTF_FILTER.out.versions) ch_versions = ch_versions.mix(MAKE_TRANSCRIPTS_FASTA.out.versions) } @@ -292,10 +290,10 @@ workflow PREPARE_GENOME { emit: fasta = ch_fasta // channel: path(genome.fasta) - gtf = ch_gtf_genome // channel: path(genome.gtf) + gtf = ch_gtf // channel: path(genome.gtf) + filtered_gtf = ch_filtered_gtf // channel: path(genome.gtf) fai = ch_fai // channel: path(genome.fai) gene_bed = ch_gene_bed // channel: path(gene.bed) - gtf_with_transcript_ids = ch_gtf_with_transcript_ids // channel: path(gtf) transcript_fasta = ch_transcript_fasta // channel: path(transcript.fasta) chrom_sizes = ch_chrom_sizes // channel: path(genome.sizes) splicesites = ch_splicesites // channel: path(genome.splicesites.txt) diff --git a/workflows/rnaseq.nf b/workflows/rnaseq.nf index 0912c661c..529d8e7c5 100755 --- a/workflows/rnaseq.nf +++ b/workflows/rnaseq.nf @@ -383,7 +383,7 @@ workflow RNASEQ { ALIGN_STAR ( ch_filtered_reads, PREPARE_GENOME.out.star_index.map { [ [:], it ] }, - PREPARE_GENOME.out.gtf.map { [ [:], it ] }, + PREPARE_GENOME.out.filtered_gtf.map { [ [:], it ] }, params.star_ignore_sjdbgtf, '', params.seq_center ?: '', @@ -473,7 +473,7 @@ workflow RNASEQ { ch_transcriptome_bam, ch_dummy_file, PREPARE_GENOME.out.transcript_fasta, - PREPARE_GENOME.out.gtf_with_transcript_ids, + PREPARE_GENOME.out.filtered_gtf, true, params.salmon_quant_libtype ?: '' ) @@ -648,7 +648,7 @@ workflow RNASEQ { if (!params.skip_alignment && !params.skip_stringtie) { STRINGTIE_STRINGTIE ( ch_genome_bam, - PREPARE_GENOME.out.gtf_with_transcript_ids + PREPARE_GENOME.out.filtered_gtf ) ch_versions = ch_versions.mix(STRINGTIE_STRINGTIE.out.versions.first()) } @@ -798,7 +798,7 @@ workflow RNASEQ { ch_filtered_reads, PREPARE_GENOME.out.salmon_index, ch_dummy_file, - PREPARE_GENOME.out.gtf_with_transcript_ids, + PREPARE_GENOME.out.filtered_gtf, false, params.salmon_quant_libtype ?: '' ) From 5c213c5d4f675283297fcfb44a41680ae77f0bf9 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Wed, 15 Nov 2023 10:33:02 +0000 Subject: [PATCH 160/184] Blackify --- bin/filter_gtf.py | 11 ++++++++--- 1 file changed, 8 insertions(+), 3 deletions(-) diff --git a/bin/filter_gtf.py b/bin/filter_gtf.py index dd8e1866e..3d09bd859 100755 --- a/bin/filter_gtf.py +++ b/bin/filter_gtf.py @@ -10,18 +10,21 @@ logger = logging.getLogger("fasta_gtf_filter") logger.setLevel(logging.INFO) + def extract_fasta_seq_names(fasta_name: str) -> Set[str]: """Extracts the sequence names from a FASTA file.""" with open(fasta_name) as fasta: return {line[1:].split(None, 1)[0] for line in fasta if line.startswith(">")} + def tab_delimited(file: str) -> float: """Check if file is tab-delimited and return median number of tabs.""" with open(file, "r") as f: data = f.read(1024) return statistics.median(line.count("\t") for line in data.split("\n")) -def filter_gtf(fasta: str, gtf_in: str, filtered_gtf_out: str, skip_transcript_id_check: bool) -> None: + +def filter_gtf(fasta: str, gtf_in: str, filtered_gtf_out: str, skip_transcript_id_check: bool) -> None: """Filter GTF file based on FASTA sequence names.""" if tab_delimited(gtf_in) != 8: raise ValueError("Invalid GTF file: Expected 8 tab-separated columns.") @@ -53,13 +56,15 @@ def filter_gtf(fasta: str, gtf_in: str, filtered_gtf_out: str, skip_transcript_ logger.debug("All sequence IDs from GTF: " + ", ".join(sorted(seq_names_in_gtf))) logger.info(f"Extracted {line_count} matching sequences from {gtf_in} into {filtered_gtf_out}") + if __name__ == "__main__": parser = argparse.ArgumentParser(description="Filters a GTF file based on sequence names in a FASTA file.") parser.add_argument("--gtf", type=str, required=True, help="GTF file") parser.add_argument("--fasta", type=str, required=True, help="Genome fasta file") parser.add_argument("--prefix", dest="prefix", default="genes", type=str, help="Prefix for output GTF files") - parser.add_argument("--skip_transcript_id_check", action='store_true', help="Skip checking for transcript IDs in the GTF file") + parser.add_argument( + "--skip_transcript_id_check", action="store_true", help="Skip checking for transcript IDs in the GTF file" + ) args = parser.parse_args() filter_gtf(args.fasta, args.gtf, args.prefix + ".filtered.gtf", args.skip_transcript_id_check) - From 6d55142efacc07ee3ac173045e766d971c8e713f Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Wed, 15 Nov 2023 10:54:57 +0000 Subject: [PATCH 161/184] Document GTF filtering --- docs/usage.md | 10 +++++++--- 1 file changed, 7 insertions(+), 3 deletions(-) diff --git a/docs/usage.md b/docs/usage.md index 9748aa5de..3df13b073 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -91,9 +91,9 @@ The `--umitools_grouping_method` parameter affects [how similar, but non-identic #### Examples: -| UMI type | Source | Pipeline parameters | -| ------------ | ---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------- | -| In read name | [Illumina BCL convert >3.7.5](https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/software_documentation/bcl_convert/bcl-convert-v3-7-5-software-guide-1000000163594-00.pdf) | `--with_umi --skip_umi_extract --umitools_umi_separator ":"` | +| UMI type | Source | Pipeline parameters | +| ------------ | --------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------- | +| In read name | [Illumina BCL convert >3.7.5](https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/software_documentation/bcl_convert/bcl-convert-v3-7-5-software-guide-1000000163594-00.pdf) | `--with_umi --skip_umi_extract --umitools_umi_separator ":"` | | In sequence | [Lexogen QuantSeq® 3’ mRNA-Seq V2 FWD](https://www.lexogen.com/quantseq-3mrna-sequencing) + [UMI Second Strand Synthesis Module](https://faqs.lexogen.com/faq/how-can-i-add-umis-to-my-quantseq-libraries) | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern "^(?P.{6})(?P.{4}).*"` | | In sequence | [Lexogen CORALL® Total RNA-Seq V1](https://www.lexogen.com/corall-total-rna-seq/)
    > _mind [Appendix H](https://www.lexogen.com/wp-content/uploads/2020/04/095UG190V0130_CORALL-Total-RNA-Seq_2020-03-31.pdf) regarding optional trimming_ | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern "^(?P.{12}).*"`
    Optional: `--clip_r2 9 --three_prime_clip_r2 12` | | In sequence | [Takara Bio SMARTer® Stranded Total RNA-Seq Kit v3](https://www.takarabio.com/documents/User%20Manual/SMARTer%20Stranded%20Total%20RNA/SMARTer%20Stranded%20Total%20RNA-Seq%20Kit%20v3%20-%20Pico%20Input%20Mammalian%20User%20Manual-a_114949.pdf) | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern2 "^(?P.{8})(?P.{6}).*"` | @@ -196,6 +196,10 @@ Notes: - As of v3.7 of the pipeline, if you are using a genome downloaded from AWS iGenomes and using `--aligner star_salmon` (default) the version of STAR to use for the alignment will be auto-detected (see [#808](https://github.com/nf-core/rnaseq/issues/808)). +### GTF filtering + +By default, the input GTF file will be filtered to ensure that sequence names correspond to those in the genome, and to remove rows with empty transcript identifiers. Filtering can be bypassed completely where you are confident it is not necessary, using the `--skip_gtf_filter` flag. The transcript identifer filter can be disabled specifically using `skip_gtf_transcript_filter`. + ## Running the pipeline The typical command for running the pipeline is as follows: From 6e1454b363e1d9fb599d30047cb97a80e6782873 Mon Sep 17 00:00:00 2001 From: nf-core-bot Date: Wed, 15 Nov 2023 10:56:28 +0000 Subject: [PATCH 162/184] [automated] Fix linting with Prettier --- docs/usage.md | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/docs/usage.md b/docs/usage.md index 3df13b073..82319c466 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -91,9 +91,9 @@ The `--umitools_grouping_method` parameter affects [how similar, but non-identic #### Examples: -| UMI type | Source | Pipeline parameters | -| ------------ | --------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------- | -| In read name | [Illumina BCL convert >3.7.5](https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/software_documentation/bcl_convert/bcl-convert-v3-7-5-software-guide-1000000163594-00.pdf) | `--with_umi --skip_umi_extract --umitools_umi_separator ":"` | +| UMI type | Source | Pipeline parameters | +| ------------ | ---------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------- | ---------------------------------------------------------------------------------------------------------------------------------------------- | +| In read name | [Illumina BCL convert >3.7.5](https://emea.support.illumina.com/content/dam/illumina-support/documents/documentation/software_documentation/bcl_convert/bcl-convert-v3-7-5-software-guide-1000000163594-00.pdf) | `--with_umi --skip_umi_extract --umitools_umi_separator ":"` | | In sequence | [Lexogen QuantSeq® 3’ mRNA-Seq V2 FWD](https://www.lexogen.com/quantseq-3mrna-sequencing) + [UMI Second Strand Synthesis Module](https://faqs.lexogen.com/faq/how-can-i-add-umis-to-my-quantseq-libraries) | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern "^(?P.{6})(?P.{4}).*"` | | In sequence | [Lexogen CORALL® Total RNA-Seq V1](https://www.lexogen.com/corall-total-rna-seq/)
    > _mind [Appendix H](https://www.lexogen.com/wp-content/uploads/2020/04/095UG190V0130_CORALL-Total-RNA-Seq_2020-03-31.pdf) regarding optional trimming_ | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern "^(?P.{12}).*"`
    Optional: `--clip_r2 9 --three_prime_clip_r2 12` | | In sequence | [Takara Bio SMARTer® Stranded Total RNA-Seq Kit v3](https://www.takarabio.com/documents/User%20Manual/SMARTer%20Stranded%20Total%20RNA/SMARTer%20Stranded%20Total%20RNA-Seq%20Kit%20v3%20-%20Pico%20Input%20Mammalian%20User%20Manual-a_114949.pdf) | `--with_umi --umitools_extract_method "regex" --umitools_bc_pattern2 "^(?P.{8})(?P.{6}).*"` | From aff1d2e02717247831644769fc3ba84868c3fdde Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Wed, 15 Nov 2023 13:06:09 +0000 Subject: [PATCH 163/184] Bump modules config for merged module --- modules.json | 4 ++-- modules/nf-core/kallisto/quant/main.nf | 2 +- 2 files changed, 3 insertions(+), 3 deletions(-) diff --git a/modules.json b/modules.json index d06dfe193..84d48d87f 100644 --- a/modules.json +++ b/modules.json @@ -71,8 +71,8 @@ "installed_by": ["modules"] }, "kallisto/quant": { - "branch": "kallisto_updates", - "git_sha": "bc4719dcd079fcdb650ddeac05739c2f7dd58c84", + "branch": "master", + "git_sha": "bdc2a97ced7adc423acfa390742db83cab98c1ad", "installed_by": ["modules"] }, "picard/markduplicates": { diff --git a/modules/nf-core/kallisto/quant/main.nf b/modules/nf-core/kallisto/quant/main.nf index 2ad0bb78b..f5444d791 100644 --- a/modules/nf-core/kallisto/quant/main.nf +++ b/modules/nf-core/kallisto/quant/main.nf @@ -59,7 +59,7 @@ process KALLISTO_QUANT { -o $prefix \\ ${reads} 2> >(tee -a ${prefix}/kallisto_quant.log >&2) - cp ${prefix}/kallisto_quant.log ${prefix}.log + cp ${prefix}/kallisto_quant.log ${prefix}.log cp ${prefix}/run_info.json ${prefix}.run_info.json cat <<-END_VERSIONS > versions.yml From f7d078cc159e3ae0a2b722c465ae0a20fcfe66e6 Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Wed, 15 Nov 2023 14:19:01 +0000 Subject: [PATCH 164/184] Bump pipeline version to 3.13.0 --- CHANGELOG.md | 13 ++++++------- assets/multiqc_config.yml | 4 ++-- nextflow.config | 2 +- 3 files changed, 9 insertions(+), 10 deletions(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index 357dd24db..80c51ea61 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -3,7 +3,7 @@ The format is based on [Keep a Changelog](https://keepachangelog.com/en/1.0.0/) and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0.html). -## v3.13.0dev - [date] +## [[3.13.0](https://github.com/nf-core/rnaseq/releases/tag/3.13.0)] - 2023-11-17 ### Credits @@ -14,7 +14,6 @@ Special thanks to the following for their contributions to the release: - [Júlia Mir Pedrol](https://github.com/mirpedrol) - [Matthias Zepper](https://github.com/MatthiasZepper) - [Maxime Garcia](https://github.com/maxulysse) -- [Jonathan Manning](https://github.com/pinin4fjords) Thank you to everyone else that has contributed by reporting bugs, enhancements or in any other way, shape or form. @@ -30,13 +29,13 @@ Thank you to everyone else that has contributed by reporting bugs, enhancements - [PR #1083](https://github.com/nf-core/rnaseq/pull/1083) - Move local modules and subworkflows to subfolders - [PR #1088](https://github.com/nf-core/rnaseq/pull/1088) - Updates contributing and code of conduct documents with nf-core template 2.10 - [PR #1091](https://github.com/nf-core/rnaseq/pull/1091) - Reorganise parameters in schema for better usability -- [PR #1107](https://github.com/nf-core/rnaseq/pull/1107) - Expand GTF filtering to remove rows with empty transcript ID when required, fix STAR GTF usage -- [#1082](https://github.com/nf-core/rnaseq/issues/1082) - More informative error message for filter_gtf_for_genes_in_genome.py -- [#1102](https://github.com/nf-core/rnaseq/issues/1102) - gene entries with empty transcript_id fields -- [#1074](https://github.com/nf-core/rnaseq/issues/1074) - Enable quantification using StringTie AND a custom Ensembl genome - [PR #1106](https://github.com/nf-core/rnaseq/pull/1106) - Kallisto quantification -- [PR #1106](https://github.com/nf-core/rnaseq/pull/1106) - MultiQC [version bump](https://github.com/nf-core/rnaseq/pull/1106/commits/aebad067a10a45510a2b421da852cb436ae65fd8) +- [PR #1107](https://github.com/nf-core/rnaseq/pull/1107) - Expand GTF filtering to remove rows with empty transcript ID when required, fix STAR GTF usage - [#1050](https://github.com/nf-core/rnaseq/issues/1050) - Provide custom prefix/suffix for summary files to avoid overwriting +- [#1074](https://github.com/nf-core/rnaseq/issues/1074) - Enable quantification using StringTie AND a custom +- [#1082](https://github.com/nf-core/rnaseq/issues/1082) - More informative error message for `filter_gtf_for_genes_in_genome.py` +- [#1102](https://github.com/nf-core/rnaseq/issues/1102) - gene entries with empty transcript_id fields +Ensembl genome ### Software dependencies diff --git a/assets/multiqc_config.yml b/assets/multiqc_config.yml index 100536f4e..4085d95e3 100644 --- a/assets/multiqc_config.yml +++ b/assets/multiqc_config.yml @@ -1,7 +1,7 @@ report_comment: > - This report has been generated by the
    nf-core/rnaseq + This report has been generated by the nf-core/rnaseq analysis pipeline. For information about how to interpret these results, please see the - documentation. + documentation. report_section_order: "nf-core-rnaseq-methods-description": order: -1000 diff --git a/nextflow.config b/nextflow.config index 7837c0a29..ab539161f 100644 --- a/nextflow.config +++ b/nextflow.config @@ -317,7 +317,7 @@ manifest { description = """RNA sequencing analysis pipeline for gene/isoform quantification and extensive quality control.""" mainScript = 'main.nf' nextflowVersion = '!>=23.04.0' - version = '3.13.0dev' + version = '3.13.0' doi = 'https://doi.org/10.5281/zenodo.1400710' } From 1c6012ecbb087014ea4b8f0f3d39b874850277a8 Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Wed, 15 Nov 2023 14:38:26 +0000 Subject: [PATCH 165/184] [skip ci] Changes from local review --- conf/modules.config | 2 -- docs/usage.md | 2 +- modules/local/gtf_filter/main.nf | 6 ++--- nextflow_schema.json | 4 +-- subworkflows/local/prepare_genome/main.nf | 30 +++++++++++------------ 5 files changed, 21 insertions(+), 23 deletions(-) diff --git a/conf/modules.config b/conf/modules.config index 5b4742df0..6e28f549b 100644 --- a/conf/modules.config +++ b/conf/modules.config @@ -165,7 +165,6 @@ process { ext.args = '--record-count 1000000 --seed 1' ext.prefix = { "${meta.id}.subsampled" } publishDir = [ - mode: params.publish_dir_mode, enabled: false ] } @@ -173,7 +172,6 @@ process { withName: '.*:FASTQ_SUBSAMPLE_FQ_SALMON:SALMON_QUANT' { ext.args = '--skipQuant' publishDir = [ - mode: params.publish_dir_mode, enabled: false ] } diff --git a/docs/usage.md b/docs/usage.md index dd4cb12b4..524acba9b 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -200,7 +200,7 @@ Notes: ### GTF filtering -By default, the input GTF file will be filtered to ensure that sequence names correspond to those in the genome, and to remove rows with empty transcript identifiers. Filtering can be bypassed completely where you are confident it is not necessary, using the `--skip_gtf_filter` flag. The transcript identifer filter can be disabled specifically using `skip_gtf_transcript_filter`. +By default, the input GTF file will be filtered to ensure that sequence names correspond to those in the genome fasta file, and to remove rows with empty transcript identifiers. Filtering can be bypassed completely where you are confident it is not necessary, using the `--skip_gtf_filter` parameter. If you just want to skip the 'transcript_id' checking component of the GTF filtering script used in the pipeline this can be disabled specifically using the `--skip_gtf_transcript_filter` parameter. ## Running the pipeline diff --git a/modules/local/gtf_filter/main.nf b/modules/local/gtf_filter/main.nf index cf3536f95..d14e8ff42 100644 --- a/modules/local/gtf_filter/main.nf +++ b/modules/local/gtf_filter/main.nf @@ -11,13 +11,13 @@ process GTF_FILTER { path gtf output: - path "*.filtered.gtf" , emit: genome_gtf - path "versions.yml" , emit: versions + path "*.filtered.gtf", emit: genome_gtf + path "versions.yml" , emit: versions when: task.ext.when == null || task.ext.when - script: // filter_gtf_for_genes_in_genome.py is bundled with the pipeline, in nf-core/rnaseq/bin/ + script: // filter_gtf.py is bundled with the pipeline, in nf-core/rnaseq/bin/ """ filter_gtf.py \\ --gtf $gtf \\ diff --git a/nextflow_schema.json b/nextflow_schema.json index d350a90fa..f7c04d5ab 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -89,12 +89,12 @@ "type": "boolean", "fa_icon": "fas fa-forward", "description": "Skip filtering of GTF for valid scaffolds and/ or transcript IDs.", - "help_text": "If you're confident on the validity of the GTF with respect to the genome file, or wish to disregard failures thriggered by the filtering module, activate this option." + "help_text": "If you're confident on the validity of the GTF with respect to the genome fasta file, or wish to disregard failures thriggered by the filtering module, activate this option." }, "skip_gtf_transcript_filter": { "type": "boolean", "fa_icon": "fas fa-forward", - "description": "Skip just the transcript_id check of GTF filtering." + "description": "Skip the 'transcript_id' checking component of the GTF filtering script used in the pipeline." }, "gene_bed": { "type": "string", diff --git a/subworkflows/local/prepare_genome/main.nf b/subworkflows/local/prepare_genome/main.nf index 1df79b107..e86612001 100644 --- a/subworkflows/local/prepare_genome/main.nf +++ b/subworkflows/local/prepare_genome/main.nf @@ -310,20 +310,20 @@ workflow PREPARE_GENOME { } emit: - fasta = ch_fasta // channel: path(genome.fasta) - gtf = ch_gtf // channel: path(genome.gtf) - filtered_gtf = ch_filtered_gtf // channel: path(genome.gtf) - fai = ch_fai // channel: path(genome.fai) - gene_bed = ch_gene_bed // channel: path(gene.bed) - transcript_fasta = ch_transcript_fasta // channel: path(transcript.fasta) - chrom_sizes = ch_chrom_sizes // channel: path(genome.sizes) - splicesites = ch_splicesites // channel: path(genome.splicesites.txt) - bbsplit_index = ch_bbsplit_index // channel: path(bbsplit/index/) - star_index = ch_star_index // channel: path(star/index/) - rsem_index = ch_rsem_index // channel: path(rsem/index/) - hisat2_index = ch_hisat2_index // channel: path(hisat2/index/) - salmon_index = ch_salmon_index // channel: path(salmon/index/) - kallisto_index = ch_kallisto_index // channel: [ meta, path(kallisto/index/) ] + fasta = ch_fasta // channel: path(genome.fasta) + gtf = ch_gtf // channel: path(genome.gtf) + filtered_gtf = ch_filtered_gtf // channel: path(genome.gtf) + fai = ch_fai // channel: path(genome.fai) + gene_bed = ch_gene_bed // channel: path(gene.bed) + transcript_fasta = ch_transcript_fasta // channel: path(transcript.fasta) + chrom_sizes = ch_chrom_sizes // channel: path(genome.sizes) + splicesites = ch_splicesites // channel: path(genome.splicesites.txt) + bbsplit_index = ch_bbsplit_index // channel: path(bbsplit/index/) + star_index = ch_star_index // channel: path(star/index/) + rsem_index = ch_rsem_index // channel: path(rsem/index/) + hisat2_index = ch_hisat2_index // channel: path(hisat2/index/) + salmon_index = ch_salmon_index // channel: path(salmon/index/) + kallisto_index = ch_kallisto_index // channel: [ meta, path(kallisto/index/) ] - versions = ch_versions.ifEmpty(null) // channel: [ versions.yml ] + versions = ch_versions.ifEmpty(null) // channel: [ versions.yml ] } From e7126c80966e6d2999cc5db3f9b5bbe0e98a9ba5 Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Wed, 15 Nov 2023 15:53:32 +0000 Subject: [PATCH 166/184] Move --skip_* parameters to the same section --- nextflow_schema.json | 22 +++++++++++----------- 1 file changed, 11 insertions(+), 11 deletions(-) diff --git a/nextflow_schema.json b/nextflow_schema.json index f7c04d5ab..60e6585cf 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -85,17 +85,6 @@ "description": "Path to GFF3 annotation file.", "help_text": "This parameter must be specified if `--genome` or `--gtf` are not specified." }, - "skip_gtf_filter": { - "type": "boolean", - "fa_icon": "fas fa-forward", - "description": "Skip filtering of GTF for valid scaffolds and/ or transcript IDs.", - "help_text": "If you're confident on the validity of the GTF with respect to the genome fasta file, or wish to disregard failures thriggered by the filtering module, activate this option." - }, - "skip_gtf_transcript_filter": { - "type": "boolean", - "fa_icon": "fas fa-forward", - "description": "Skip the 'transcript_id' checking component of the GTF filtering script used in the pipeline." - }, "gene_bed": { "type": "string", "format": "file-path", @@ -531,6 +520,17 @@ "fa_icon": "fas fa-fast-forward", "description": "Options to skip various steps within the workflow.", "properties": { + "skip_gtf_filter": { + "type": "boolean", + "fa_icon": "fas fa-forward", + "description": "Skip filtering of GTF for valid scaffolds and/ or transcript IDs.", + "help_text": "If you're confident on the validity of the GTF with respect to the genome fasta file, or wish to disregard failures thriggered by the filtering module, activate this option." + }, + "skip_gtf_transcript_filter": { + "type": "boolean", + "fa_icon": "fas fa-forward", + "description": "Skip the 'transcript_id' checking component of the GTF filtering script used in the pipeline." + }, "skip_bbsplit": { "type": "boolean", "default": true, From 13cc0e98b957a2b842857c75ef3fdf51977a6697 Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Wed, 15 Nov 2023 16:00:19 +0000 Subject: [PATCH 167/184] [skip ci] Tidy up code block --- subworkflows/local/prepare_genome/main.nf | 7 +++---- 1 file changed, 3 insertions(+), 4 deletions(-) diff --git a/subworkflows/local/prepare_genome/main.nf b/subworkflows/local/prepare_genome/main.nf index e86612001..b81005b8a 100644 --- a/subworkflows/local/prepare_genome/main.nf +++ b/subworkflows/local/prepare_genome/main.nf @@ -93,9 +93,7 @@ workflow PREPARE_GENOME { // // Apply filtering we may need for GTFs // - - ch_filtered_gtf = ch_gtf - + ch_filtered_gtf = ch_gtf filtering_useful = ( // Condition 1: Alignment is required and aligner is set to 'star_salmon' @@ -114,9 +112,10 @@ workflow PREPARE_GENOME { !transcript_fasta ) - if (filtering_useful){ + if (filtering_useful) { GTF_FILTER ( ch_fasta, ch_gtf ) ch_filtered_gtf = GTF_FILTER.out.genome_gtf + ch_versions = ch_versions.mix(GTF_FILTER.out.versions) } } From 3ac4a3afc7aac982866e56cbc92b329c0bcd312c Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Wed, 15 Nov 2023 17:58:18 +0000 Subject: [PATCH 168/184] Move filter_gtf logic into main workflow --- subworkflows/local/prepare_genome/main.nf | 31 +++-------------------- workflows/rnaseq.nf | 30 ++++++++++++++++++---- 2 files changed, 29 insertions(+), 32 deletions(-) diff --git a/subworkflows/local/prepare_genome/main.nf b/subworkflows/local/prepare_genome/main.nf index b81005b8a..0be947954 100644 --- a/subworkflows/local/prepare_genome/main.nf +++ b/subworkflows/local/prepare_genome/main.nf @@ -53,6 +53,7 @@ workflow PREPARE_GENOME { is_aws_igenome // boolean: whether the genome files are from AWS iGenomes biotype // string: if additional fasta file is provided biotype value to use when appending entries to GTF file prepare_tool_indices // list: tools to prepare indices for + filter_gtf // boolean: whether to filter GTF file main: @@ -90,31 +91,9 @@ workflow PREPARE_GENOME { ch_versions = ch_versions.mix(GFFREAD.out.versions) } - // - // Apply filtering we may need for GTFs - // - ch_filtered_gtf = ch_gtf - filtering_useful = - ( - // Condition 1: Alignment is required and aligner is set to 'star_salmon' - !params.skip_alignment && params.aligner == 'star_salmon' - ) || - ( - // Condition 2: Pseudo-alignment is required and pseudo-aligner is set to 'salmon' - !params.skip_pseudo_alignment && params.pseudo_aligner == 'salmon' - ) || - ( - // Condition 3: Neither alignment nor stringtie are to be skipped - !params.skip_alignment && !params.skip_stringtie - ) || - ( - // Condition 4: Transcript FASTA file is not provided - !transcript_fasta - ) - - if (filtering_useful) { + if (filter_gtf) { GTF_FILTER ( ch_fasta, ch_gtf ) - ch_filtered_gtf = GTF_FILTER.out.genome_gtf + ch_gtf = GTF_FILTER.out.genome_gtf ch_versions = ch_versions.mix(GTF_FILTER.out.versions) } } @@ -166,7 +145,7 @@ workflow PREPARE_GENOME { ch_versions = ch_versions.mix(PREPROCESS_TRANSCRIPTS_FASTA_GENCODE.out.versions) } } else { - ch_transcript_fasta = MAKE_TRANSCRIPTS_FASTA ( ch_fasta, ch_filtered_gtf ).transcript_fasta + ch_transcript_fasta = MAKE_TRANSCRIPTS_FASTA ( ch_fasta, ch_gtf ).transcript_fasta ch_versions = ch_versions.mix(GTF_FILTER.out.versions) ch_versions = ch_versions.mix(MAKE_TRANSCRIPTS_FASTA.out.versions) } @@ -311,7 +290,6 @@ workflow PREPARE_GENOME { emit: fasta = ch_fasta // channel: path(genome.fasta) gtf = ch_gtf // channel: path(genome.gtf) - filtered_gtf = ch_filtered_gtf // channel: path(genome.gtf) fai = ch_fai // channel: path(genome.fai) gene_bed = ch_gene_bed // channel: path(gene.bed) transcript_fasta = ch_transcript_fasta // channel: path(transcript.fasta) @@ -323,6 +301,5 @@ workflow PREPARE_GENOME { hisat2_index = ch_hisat2_index // channel: path(hisat2/index/) salmon_index = ch_salmon_index // channel: path(salmon/index/) kallisto_index = ch_kallisto_index // channel: [ meta, path(kallisto/index/) ] - versions = ch_versions.ifEmpty(null) // channel: [ versions.yml ] } diff --git a/workflows/rnaseq.nf b/workflows/rnaseq.nf index 95a6b783d..403194ac2 100755 --- a/workflows/rnaseq.nf +++ b/workflows/rnaseq.nf @@ -39,6 +39,25 @@ if (!params.skip_bbsplit) { prepareToolIndices << 'bbsplit' } if (!params.skip_alignment) { prepareToolIndices << params.aligner } if (!params.skip_pseudo_alignment && params.pseudo_aligner) { prepareToolIndices << params.pseudo_aligner } +// Determine whether to filter the GTF or not +def filterGtf = + (( + // Condition 1: Alignment is required and aligner is set + !params.skip_alignment && params.aligner + ) || + ( + // Condition 2: Pseudoalignment is required and pseudoaligner is set + !params.skip_pseudo_alignment && params.pseudo_aligner + ) || + ( + // Condition 3: Transcript FASTA file is not provided + !params.transcript_fasta + )) && + ( + // Condition 4: --skip_gtf_filter is not provided + !params.skip_gtf_filter + ) + // Get RSeqC modules to run def rseqc_modules = params.rseqc_modules ? params.rseqc_modules.split(',').collect{ it.trim().toLowerCase() } : [] if (params.bam_csi_index) { @@ -175,7 +194,8 @@ workflow RNASEQ { params.gencode, is_aws_igenome, biotype, - prepareToolIndices + prepareToolIndices, + filterGtf ) ch_versions = ch_versions.mix(PREPARE_GENOME.out.versions) @@ -384,7 +404,7 @@ workflow RNASEQ { ALIGN_STAR ( ch_filtered_reads, PREPARE_GENOME.out.star_index.map { [ [:], it ] }, - PREPARE_GENOME.out.filtered_gtf.map { [ [:], it ] }, + PREPARE_GENOME.out.gtf.map { [ [:], it ] }, params.star_ignore_sjdbgtf, '', params.seq_center ?: '', @@ -474,7 +494,7 @@ workflow RNASEQ { ch_transcriptome_bam, ch_dummy_file, PREPARE_GENOME.out.transcript_fasta, - PREPARE_GENOME.out.filtered_gtf, + PREPARE_GENOME.out.gtf, 'salmon', true, params.salmon_quant_libtype ?: '', @@ -652,7 +672,7 @@ workflow RNASEQ { if (!params.skip_alignment && !params.skip_stringtie) { STRINGTIE_STRINGTIE ( ch_genome_bam, - PREPARE_GENOME.out.filtered_gtf + PREPARE_GENOME.out.gtf ) ch_versions = ch_versions.mix(STRINGTIE_STRINGTIE.out.versions.first()) } @@ -810,7 +830,7 @@ workflow RNASEQ { ch_filtered_reads, ch_pseudo_index, ch_dummy_file, - PREPARE_GENOME.out.filtered_gtf, + PREPARE_GENOME.out.gtf, params.pseudo_aligner, false, params.salmon_quant_libtype ?: '', From b67558d2757cf5c1f05528b328d2381a1f42d7cb Mon Sep 17 00:00:00 2001 From: nf-core-bot Date: Wed, 15 Nov 2023 18:06:06 +0000 Subject: [PATCH 169/184] [automated] Fix linting with Prettier --- CHANGELOG.md | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index 80c51ea61..5b5e0e7ef 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -32,10 +32,10 @@ Thank you to everyone else that has contributed by reporting bugs, enhancements - [PR #1106](https://github.com/nf-core/rnaseq/pull/1106) - Kallisto quantification - [PR #1107](https://github.com/nf-core/rnaseq/pull/1107) - Expand GTF filtering to remove rows with empty transcript ID when required, fix STAR GTF usage - [#1050](https://github.com/nf-core/rnaseq/issues/1050) - Provide custom prefix/suffix for summary files to avoid overwriting -- [#1074](https://github.com/nf-core/rnaseq/issues/1074) - Enable quantification using StringTie AND a custom +- [#1074](https://github.com/nf-core/rnaseq/issues/1074) - Enable quantification using StringTie AND a custom - [#1082](https://github.com/nf-core/rnaseq/issues/1082) - More informative error message for `filter_gtf_for_genes_in_genome.py` - [#1102](https://github.com/nf-core/rnaseq/issues/1102) - gene entries with empty transcript_id fields -Ensembl genome + Ensembl genome ### Software dependencies From 635ab21bc4751826eb46c601597a9af4b59906df Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Wed, 15 Nov 2023 18:34:41 +0000 Subject: [PATCH 170/184] Update CHANGELOG.md --- CHANGELOG.md | 2 ++ 1 file changed, 2 insertions(+) diff --git a/CHANGELOG.md b/CHANGELOG.md index 5b5e0e7ef..b5bfd28e6 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -14,6 +14,8 @@ Special thanks to the following for their contributions to the release: - [Júlia Mir Pedrol](https://github.com/mirpedrol) - [Matthias Zepper](https://github.com/MatthiasZepper) - [Maxime Garcia](https://github.com/maxulysse) +- [Steffen Möller](https://github.com/maxulysse) +- [hmehlan](https://github.com/hmehlan) Thank you to everyone else that has contributed by reporting bugs, enhancements or in any other way, shape or form. From cc3bc72be3d6deafaaa5925dca10bd540a3685d6 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Wed, 15 Nov 2023 22:30:24 +0000 Subject: [PATCH 171/184] fix credits --- CHANGELOG.md | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index b5bfd28e6..ddeda910f 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -10,12 +10,12 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 Special thanks to the following for their contributions to the release: - [Adam Talbot](https://github.com/adamrtalbot) +- [hmehlan](https://github.com/hmehlan) - [Jonathan Manning](https://github.com/pinin4fjords) - [Júlia Mir Pedrol](https://github.com/mirpedrol) - [Matthias Zepper](https://github.com/MatthiasZepper) - [Maxime Garcia](https://github.com/maxulysse) -- [Steffen Möller](https://github.com/maxulysse) -- [hmehlan](https://github.com/hmehlan) +- [Steffen Möller](https://github.com/smoe) Thank you to everyone else that has contributed by reporting bugs, enhancements or in any other way, shape or form. From de0c007d70a3c4d97532a1918c31b8377b0e3462 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 16 Nov 2023 09:52:01 +0000 Subject: [PATCH 172/184] Update test data links --- conf/test.config | 22 +++++++++++----------- 1 file changed, 11 insertions(+), 11 deletions(-) diff --git a/conf/test.config b/conf/test.config index d0430eacb..f9154ba31 100644 --- a/conf/test.config +++ b/conf/test.config @@ -21,19 +21,19 @@ params { // Input data - input = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/samplesheet/v3.10/samplesheet_test.csv" + input = "https://raw.githubusercontent.com/nf-core/test-datasets/7f1614baeb0ddf66e60be78c3d9fa55440465ac8/samplesheet/v3.10/samplesheet_test.csv" // Genome references - fasta = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/reference/genome.fasta" - gtf = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/reference/genes.gtf.gz" - gff = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/reference/genes.gff.gz" - transcript_fasta = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/reference/transcriptome.fasta" - additional_fasta = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/reference/gfp.fa.gz" - - bbsplit_fasta_list = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/reference/bbsplit_fasta_list.txt" - hisat2_index = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/reference/hisat2.tar.gz" - salmon_index = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/reference/salmon.tar.gz" - rsem_index = "https://raw.githubusercontent.com/nf-core/test-datasets/rnaseq/reference/rsem.tar.gz" + fasta = "https://raw.githubusercontent.com/nf-core/test-datasets/7f1614baeb0ddf66e60be78c3d9fa55440465ac8/reference/genome.fasta" + gtf = "https://raw.githubusercontent.com/nf-core/test-datasets/7f1614baeb0ddf66e60be78c3d9fa55440465ac8/reference/genes_with_empty_tid.gtf.gz" + gff = "https://raw.githubusercontent.com/nf-core/test-datasets/7f1614baeb0ddf66e60be78c3d9fa55440465ac8/reference/genes.gff.gz" + transcript_fasta = "https://raw.githubusercontent.com/nf-core/test-datasets/7f1614baeb0ddf66e60be78c3d9fa55440465ac8/reference/transcriptome.fasta" + additional_fasta = "https://raw.githubusercontent.com/nf-core/test-datasets/7f1614baeb0ddf66e60be78c3d9fa55440465ac8/reference/gfp.fa.gz" + + bbsplit_fasta_list = "https://raw.githubusercontent.com/nf-core/test-datasets/7f1614baeb0ddf66e60be78c3d9fa55440465ac8/reference/bbsplit_fasta_list.txt" + hisat2_index = "https://raw.githubusercontent.com/nf-core/test-datasets/7f1614baeb0ddf66e60be78c3d9fa55440465ac8/reference/hisat2.tar.gz" + salmon_index = "https://raw.githubusercontent.com/nf-core/test-datasets/7f1614baeb0ddf66e60be78c3d9fa55440465ac8/reference/salmon.tar.gz" + rsem_index = "https://raw.githubusercontent.com/nf-core/test-datasets/7f1614baeb0ddf66e60be78c3d9fa55440465ac8/reference/rsem.tar.gz" // Other parameters skip_bbsplit = false From 14c29a660e2ed00e18245c0dfb6e07981f9b346a Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 16 Nov 2023 10:30:06 +0000 Subject: [PATCH 173/184] Add authors and licenses to scripts in bin/ where missing --- bin/deseq2_qc.r | 2 ++ bin/dupradar.r | 2 ++ bin/fasta2gtf.py | 3 +++ bin/fastq_dir_to_samplesheet.py | 2 ++ bin/filter_gtf.py | 3 +++ bin/mqc_features_stat.py | 2 ++ bin/summarizedexperiment.r | 2 ++ bin/tx2gene.py | 3 +++ bin/tximport.r | 2 ++ 9 files changed, 21 insertions(+) diff --git a/bin/deseq2_qc.r b/bin/deseq2_qc.r index d99bc7c2b..9397f5f30 100755 --- a/bin/deseq2_qc.r +++ b/bin/deseq2_qc.r @@ -1,5 +1,7 @@ #!/usr/bin/env Rscript +# Written by Harshil Patel and released under the MIT license. + ################################################ ################################################ ## REQUIREMENTS ## diff --git a/bin/dupradar.r b/bin/dupradar.r index daf5a5798..d58b0eee1 100755 --- a/bin/dupradar.r +++ b/bin/dupradar.r @@ -1,5 +1,7 @@ #!/usr/bin/env Rscript +# Written by Phil Ewels and released under the MIT license. + # Command line argument processing args = commandArgs(trailingOnly=TRUE) if (length(args) < 5) { diff --git a/bin/fasta2gtf.py b/bin/fasta2gtf.py index 052322b24..716ee06ce 100755 --- a/bin/fasta2gtf.py +++ b/bin/fasta2gtf.py @@ -1,4 +1,7 @@ #!/usr/bin/env python3 + +# Written by Pranathi Vemuri and released under the MIT license. + """ Read a custom fasta file and create a custom GTF containing each entry """ diff --git a/bin/fastq_dir_to_samplesheet.py b/bin/fastq_dir_to_samplesheet.py index 1eb3657c6..7ceebe525 100755 --- a/bin/fastq_dir_to_samplesheet.py +++ b/bin/fastq_dir_to_samplesheet.py @@ -1,5 +1,7 @@ #!/usr/bin/env python3 +# Written by Harshil Patel and released under the MIT license. + import os import sys import glob diff --git a/bin/filter_gtf.py b/bin/filter_gtf.py index 3d09bd859..265250627 100755 --- a/bin/filter_gtf.py +++ b/bin/filter_gtf.py @@ -1,4 +1,7 @@ #!/usr/bin/env python + +# Written by Olga Botvinnik with subsequent reworking by Jonathan Manning. Released under the MIT license. + import logging import argparse import re diff --git a/bin/mqc_features_stat.py b/bin/mqc_features_stat.py index 689a3f215..fa69b231c 100755 --- a/bin/mqc_features_stat.py +++ b/bin/mqc_features_stat.py @@ -1,5 +1,7 @@ #!/usr/bin/env python +# Written by Senthilkumar Panneerselvam and released under the MIT license. + import argparse import logging import os diff --git a/bin/summarizedexperiment.r b/bin/summarizedexperiment.r index d55101ecf..767d542c7 100755 --- a/bin/summarizedexperiment.r +++ b/bin/summarizedexperiment.r @@ -1,5 +1,7 @@ #!/usr/bin/env Rscript +# Written by Lorena Pantano and released under the MIT license. + library(SummarizedExperiment) ## Create SummarizedExperiment (se) object from counts diff --git a/bin/tx2gene.py b/bin/tx2gene.py index 0a418920e..8e0c1c6a5 100755 --- a/bin/tx2gene.py +++ b/bin/tx2gene.py @@ -1,4 +1,7 @@ #!/usr/bin/env python + +# Written by Lorena Pantano with subsequent reworking by Jonathan Manning. Released under the MIT license. + import logging import argparse import glob diff --git a/bin/tximport.r b/bin/tximport.r index e1bad7511..ee3d8b212 100755 --- a/bin/tximport.r +++ b/bin/tximport.r @@ -1,5 +1,7 @@ #!/usr/bin/env Rscript +# Written by Lorena Pantano and released under the MIT license. + library(SummarizedExperiment) library(tximport) From 6ff89c4b98b2a0bf1b386441bd2f3106cf5671dc Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 16 Nov 2023 10:37:26 +0000 Subject: [PATCH 174/184] Add release fix to changelog --- CHANGELOG.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index ddeda910f..9eb25e856 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -37,7 +37,7 @@ Thank you to everyone else that has contributed by reporting bugs, enhancements - [#1074](https://github.com/nf-core/rnaseq/issues/1074) - Enable quantification using StringTie AND a custom - [#1082](https://github.com/nf-core/rnaseq/issues/1082) - More informative error message for `filter_gtf_for_genes_in_genome.py` - [#1102](https://github.com/nf-core/rnaseq/issues/1102) - gene entries with empty transcript_id fields - Ensembl genome +- [#976](https://github.com/nf-core/rnaseq/issues/976) - Add author and licenses for all custom scripts ### Software dependencies From 2ea2ad1948b701c394987ae7690cbc8598778038 Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Thu, 16 Nov 2023 12:05:44 +0100 Subject: [PATCH 175/184] [skip-ci] Update CHANGELOG.md --- CHANGELOG.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index 9eb25e856..5df7b82b8 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -33,11 +33,11 @@ Thank you to everyone else that has contributed by reporting bugs, enhancements - [PR #1091](https://github.com/nf-core/rnaseq/pull/1091) - Reorganise parameters in schema for better usability - [PR #1106](https://github.com/nf-core/rnaseq/pull/1106) - Kallisto quantification - [PR #1107](https://github.com/nf-core/rnaseq/pull/1107) - Expand GTF filtering to remove rows with empty transcript ID when required, fix STAR GTF usage +- [#976](https://github.com/nf-core/rnaseq/issues/976) - Add author and licenses for all custom scripts - [#1050](https://github.com/nf-core/rnaseq/issues/1050) - Provide custom prefix/suffix for summary files to avoid overwriting - [#1074](https://github.com/nf-core/rnaseq/issues/1074) - Enable quantification using StringTie AND a custom - [#1082](https://github.com/nf-core/rnaseq/issues/1082) - More informative error message for `filter_gtf_for_genes_in_genome.py` - [#1102](https://github.com/nf-core/rnaseq/issues/1102) - gene entries with empty transcript_id fields -- [#976](https://github.com/nf-core/rnaseq/issues/976) - Add author and licenses for all custom scripts ### Software dependencies From 5070c5094d664c28044c0ff495c682732a24c1f9 Mon Sep 17 00:00:00 2001 From: Harshil Patel Date: Thu, 16 Nov 2023 12:07:23 +0100 Subject: [PATCH 176/184] [skip-ci] Update bin/deseq2_qc.r --- bin/deseq2_qc.r | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/bin/deseq2_qc.r b/bin/deseq2_qc.r index 9397f5f30..14bd48f70 100755 --- a/bin/deseq2_qc.r +++ b/bin/deseq2_qc.r @@ -1,6 +1,6 @@ #!/usr/bin/env Rscript -# Written by Harshil Patel and released under the MIT license. +# Written by Harshil Patel and Gavin Kelly and released under the MIT license. ################################################ ################################################ From 05d4a6607b662446f7fd8e06906404ae2ad3804e Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 16 Nov 2023 11:43:53 +0000 Subject: [PATCH 177/184] [skip ci] address maxime feedback --- CHANGELOG.md | 9 +++++++++ assets/slackreport.json | 2 +- 2 files changed, 10 insertions(+), 1 deletion(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index 5df7b82b8..8a5d20bd6 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -53,6 +53,15 @@ Thank you to everyone else that has contributed by reporting bugs, enhancements > > **NB:** Dependency has been **removed** if new version information isn't present. +### Modules / Subworkflows + +| Script | Old name | New name | +| ----------------------------- | ----------------------------- | ----------------------------- | +| `local/gtf_filter` | `GTF_GENE_FILTER` | `GTF_FILTER` | +| `local/tx2gene` | `SALMON_TX2GENE` | `TX2GENE` | +| `local/tximport` | `SALMON_TXIMPORT` | `TXIMPORT` | +| `local/quantify_salmon` | `QUANTIFY_SALMON` | `QUANTIFY_PSEUDO_ALIGNMENT` | + ## [[3.12.0](https://github.com/nf-core/rnaseq/releases/tag/3.12.0)] - 2023-06-02 ### Credits diff --git a/assets/slackreport.json b/assets/slackreport.json index b03fc11c5..fc84fa5e5 100644 --- a/assets/slackreport.json +++ b/assets/slackreport.json @@ -3,7 +3,7 @@ { "fallback": "Plain-text summary of the attachment.", "color": "<% if (success) { %>good<% } else { %>danger<%} %>", - "author_name": "nf-core/rnaseq v${version} - ${runName}", + "author_name": "nf-core/rnaseq ${version} - ${runName}", "author_icon": "https://www.nextflow.io/docs/latest/_static/favicon.ico", "text": "<% if (success) { %>Pipeline completed successfully!<% } else { %>Pipeline completed with errors<% } %>", "fields": [ From 424e78bb085e093acf140f8f520d490df80e9ff1 Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 16 Nov 2023 11:51:24 +0000 Subject: [PATCH 178/184] Bump dumpsoftwareversions module --- modules.json | 2 +- .../nf-core/custom/dumpsoftwareversions/main.nf | 6 +++--- .../nf-core/custom/dumpsoftwareversions/meta.yml | 7 ++++--- .../custom/dumpsoftwareversions/tests/main.nf.test | 5 +++-- .../dumpsoftwareversions/tests/main.nf.test.snap | 14 +++++++------- 5 files changed, 18 insertions(+), 16 deletions(-) diff --git a/modules.json b/modules.json index 84d48d87f..46df8bb3f 100644 --- a/modules.json +++ b/modules.json @@ -17,7 +17,7 @@ }, "custom/dumpsoftwareversions": { "branch": "master", - "git_sha": "1b372269755a5c4a13c23bc130ebada8cb9d4cd0", + "git_sha": "bba7e362e4afead70653f84d8700588ea28d0f9e", "installed_by": ["modules"] }, "custom/getchromsizes": { diff --git a/modules/nf-core/custom/dumpsoftwareversions/main.nf b/modules/nf-core/custom/dumpsoftwareversions/main.nf index c9d014b11..7685b33cd 100644 --- a/modules/nf-core/custom/dumpsoftwareversions/main.nf +++ b/modules/nf-core/custom/dumpsoftwareversions/main.nf @@ -2,10 +2,10 @@ process CUSTOM_DUMPSOFTWAREVERSIONS { label 'process_single' // Requires `pyyaml` which does not have a dedicated container but is in the MultiQC container - conda "bioconda::multiqc=1.15" + conda "${moduleDir}/environment.yml" container "${ workflow.containerEngine == 'singularity' && !task.ext.singularity_pull_docker_container ? - 'https://depot.galaxyproject.org/singularity/multiqc:1.15--pyhdfd78af_0' : - 'biocontainers/multiqc:1.15--pyhdfd78af_0' }" + 'https://depot.galaxyproject.org/singularity/multiqc:1.17--pyhdfd78af_0' : + 'biocontainers/multiqc:1.17--pyhdfd78af_0' }" input: path versions diff --git a/modules/nf-core/custom/dumpsoftwareversions/meta.yml b/modules/nf-core/custom/dumpsoftwareversions/meta.yml index c32657de7..5f15a5fde 100644 --- a/modules/nf-core/custom/dumpsoftwareversions/meta.yml +++ b/modules/nf-core/custom/dumpsoftwareversions/meta.yml @@ -1,4 +1,4 @@ -# yaml-language-server: $schema=https://raw.githubusercontent.com/nf-core/modules/master/modules/yaml-schema.json +# yaml-language-server: $schema=https://raw.githubusercontent.com/nf-core/modules/master/modules/meta-schema.json name: custom_dumpsoftwareversions description: Custom module used to dump software versions within the nf-core pipeline template keywords: @@ -16,7 +16,6 @@ input: type: file description: YML file containing software versions pattern: "*.yml" - output: - yml: type: file @@ -30,7 +29,9 @@ output: type: file description: File containing software versions pattern: "versions.yml" - authors: - "@drpatelh" - "@grst" +maintainers: + - "@drpatelh" + - "@grst" diff --git a/modules/nf-core/custom/dumpsoftwareversions/tests/main.nf.test b/modules/nf-core/custom/dumpsoftwareversions/tests/main.nf.test index d528d3cb8..eec1db10a 100644 --- a/modules/nf-core/custom/dumpsoftwareversions/tests/main.nf.test +++ b/modules/nf-core/custom/dumpsoftwareversions/tests/main.nf.test @@ -1,12 +1,13 @@ nextflow_process { name "Test Process CUSTOM_DUMPSOFTWAREVERSIONS" - script "modules/nf-core/custom/dumpsoftwareversions/main.nf" + script "../main.nf" process "CUSTOM_DUMPSOFTWAREVERSIONS" + tag "modules" + tag "modules_nfcore" tag "custom" tag "dumpsoftwareversions" tag "custom/dumpsoftwareversions" - tag "modules_nfcore" test("Should run without failures") { when { diff --git a/modules/nf-core/custom/dumpsoftwareversions/tests/main.nf.test.snap b/modules/nf-core/custom/dumpsoftwareversions/tests/main.nf.test.snap index 8713b9216..4274ed57a 100644 --- a/modules/nf-core/custom/dumpsoftwareversions/tests/main.nf.test.snap +++ b/modules/nf-core/custom/dumpsoftwareversions/tests/main.nf.test.snap @@ -3,25 +3,25 @@ "content": [ { "0": [ - "software_versions.yml:md5,a027f820f30b8191a20ca16465daaf37" + "software_versions.yml:md5,1c851188476409cda5752ce971b20b58" ], "1": [ - "software_versions_mqc.yml:md5,ee4a1d028ad29987f9ac511f4668f17c" + "software_versions_mqc.yml:md5,2570f4ba271ad08357b0d3d32a9cf84d" ], "2": [ - "versions.yml:md5,f47ebd22aba1dd987b7e5d5247b766c3" + "versions.yml:md5,3843ac526e762117eedf8825b40683df" ], "mqc_yml": [ - "software_versions_mqc.yml:md5,ee4a1d028ad29987f9ac511f4668f17c" + "software_versions_mqc.yml:md5,2570f4ba271ad08357b0d3d32a9cf84d" ], "versions": [ - "versions.yml:md5,f47ebd22aba1dd987b7e5d5247b766c3" + "versions.yml:md5,3843ac526e762117eedf8825b40683df" ], "yml": [ - "software_versions.yml:md5,a027f820f30b8191a20ca16465daaf37" + "software_versions.yml:md5,1c851188476409cda5752ce971b20b58" ] } ], - "timestamp": "2023-10-11T17:10:02.930699" + "timestamp": "2023-11-03T14:43:22.157011" } } From 2081ee4bb95419f2a8382a369d7f82c242b7fefb Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 16 Nov 2023 11:58:14 +0000 Subject: [PATCH 179/184] Bump kallisto index module for env name --- modules.json | 2 +- modules/nf-core/kallisto/index/environment.yml | 1 + 2 files changed, 2 insertions(+), 1 deletion(-) diff --git a/modules.json b/modules.json index 46df8bb3f..74bd0d197 100644 --- a/modules.json +++ b/modules.json @@ -67,7 +67,7 @@ }, "kallisto/index": { "branch": "master", - "git_sha": "bdbbd9a849f9dd2b6078b303d36bf26e6adb8a9e", + "git_sha": "3f5420aa22e00bd030a2556dfdffc9e164ec0ec5", "installed_by": ["modules"] }, "kallisto/quant": { diff --git a/modules/nf-core/kallisto/index/environment.yml b/modules/nf-core/kallisto/index/environment.yml index d62586580..471b006a8 100644 --- a/modules/nf-core/kallisto/index/environment.yml +++ b/modules/nf-core/kallisto/index/environment.yml @@ -1,3 +1,4 @@ +name: kallisto_index channels: - conda-forge - bioconda From 86aa289f2f49888c73a3f5a6e76ccdf01f5b3a10 Mon Sep 17 00:00:00 2001 From: nf-core-bot Date: Thu, 16 Nov 2023 12:02:16 +0000 Subject: [PATCH 180/184] [automated] Fix linting with Prettier --- CHANGELOG.md | 12 ++++++------ 1 file changed, 6 insertions(+), 6 deletions(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index 8a5d20bd6..ba044a05a 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -55,12 +55,12 @@ Thank you to everyone else that has contributed by reporting bugs, enhancements ### Modules / Subworkflows -| Script | Old name | New name | -| ----------------------------- | ----------------------------- | ----------------------------- | -| `local/gtf_filter` | `GTF_GENE_FILTER` | `GTF_FILTER` | -| `local/tx2gene` | `SALMON_TX2GENE` | `TX2GENE` | -| `local/tximport` | `SALMON_TXIMPORT` | `TXIMPORT` | -| `local/quantify_salmon` | `QUANTIFY_SALMON` | `QUANTIFY_PSEUDO_ALIGNMENT` | +| Script | Old name | New name | +| ----------------------- | ----------------- | --------------------------- | +| `local/gtf_filter` | `GTF_GENE_FILTER` | `GTF_FILTER` | +| `local/tx2gene` | `SALMON_TX2GENE` | `TX2GENE` | +| `local/tximport` | `SALMON_TXIMPORT` | `TXIMPORT` | +| `local/quantify_salmon` | `QUANTIFY_SALMON` | `QUANTIFY_PSEUDO_ALIGNMENT` | ## [[3.12.0](https://github.com/nf-core/rnaseq/releases/tag/3.12.0)] - 2023-06-02 From 508f1da948b7021da01b8bbe256067983564042b Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 16 Nov 2023 12:03:50 +0000 Subject: [PATCH 181/184] Add module components I forgot to git add --- .../nf-core/custom/dumpsoftwareversions/environment.yml | 7 +++++++ modules/nf-core/custom/dumpsoftwareversions/tests/tags.yml | 2 ++ 2 files changed, 9 insertions(+) create mode 100644 modules/nf-core/custom/dumpsoftwareversions/environment.yml create mode 100644 modules/nf-core/custom/dumpsoftwareversions/tests/tags.yml diff --git a/modules/nf-core/custom/dumpsoftwareversions/environment.yml b/modules/nf-core/custom/dumpsoftwareversions/environment.yml new file mode 100644 index 000000000..f0c63f698 --- /dev/null +++ b/modules/nf-core/custom/dumpsoftwareversions/environment.yml @@ -0,0 +1,7 @@ +name: custom_dumpsoftwareversions +channels: + - conda-forge + - bioconda + - defaults +dependencies: + - bioconda::multiqc=1.17 diff --git a/modules/nf-core/custom/dumpsoftwareversions/tests/tags.yml b/modules/nf-core/custom/dumpsoftwareversions/tests/tags.yml new file mode 100644 index 000000000..405aa24ae --- /dev/null +++ b/modules/nf-core/custom/dumpsoftwareversions/tests/tags.yml @@ -0,0 +1,2 @@ +custom/dumpsoftwareversions: + - modules/nf-core/custom/dumpsoftwareversions/** From 18fb672072d41094400973e3a44d4007dc46b26f Mon Sep 17 00:00:00 2001 From: Jonathan Manning Date: Thu, 16 Nov 2023 12:13:15 +0000 Subject: [PATCH 182/184] Add new modules to changelog --- CHANGELOG.md | 14 ++++++++------ 1 file changed, 8 insertions(+), 6 deletions(-) diff --git a/CHANGELOG.md b/CHANGELOG.md index ba044a05a..78429e553 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -55,12 +55,14 @@ Thank you to everyone else that has contributed by reporting bugs, enhancements ### Modules / Subworkflows -| Script | Old name | New name | -| ----------------------- | ----------------- | --------------------------- | -| `local/gtf_filter` | `GTF_GENE_FILTER` | `GTF_FILTER` | -| `local/tx2gene` | `SALMON_TX2GENE` | `TX2GENE` | -| `local/tximport` | `SALMON_TXIMPORT` | `TXIMPORT` | -| `local/quantify_salmon` | `QUANTIFY_SALMON` | `QUANTIFY_PSEUDO_ALIGNMENT` | +| Script | Old name | New name | +| ------------------------ | ----------------- | --------------------------- | +| `local/gtf_filter` | `GTF_GENE_FILTER` | `GTF_FILTER` | +| `local/tx2gene` | `SALMON_TX2GENE` | `TX2GENE` | +| `local/tximport` | `SALMON_TXIMPORT` | `TXIMPORT` | +| `local/quantify_salmon` | `QUANTIFY_SALMON` | `QUANTIFY_PSEUDO_ALIGNMENT` | +| `nf-core/kallisto_index` | | `KALLISTO_INDEX` | +| `nf-core/kallisto_quant` | | `KALLISTO_QUANT` | ## [[3.12.0](https://github.com/nf-core/rnaseq/releases/tag/3.12.0)] - 2023-06-02 From cef00038555cfad56c633c83da582bf788c0b452 Mon Sep 17 00:00:00 2001 From: maxulysse Date: Thu, 16 Nov 2023 16:07:48 +0100 Subject: [PATCH 183/184] fix font and centering for text in file icons --- docs/images/nf-core-rnaseq_metro_map_grey.png | Bin 307103 -> 306481 bytes docs/images/nf-core-rnaseq_metro_map_grey.svg | 81 +++++++++--------- 2 files changed, 41 insertions(+), 40 deletions(-) diff --git a/docs/images/nf-core-rnaseq_metro_map_grey.png b/docs/images/nf-core-rnaseq_metro_map_grey.png index f2b5e4e878715c0720357cac59169594213829ea..da491829da67a9349409e6f0e2b27e8c6436d183 100644 GIT binary patch literal 306481 zcmeFZg;!PG7d3oER0PCE8bwN^k&qSvrMpwQyW2pd1xZQikd_t@knWN$kuH&x{MNC4 z&pW=q;2q;V!$%+B-gD31d+oL6nsaV^r6nI-z$Cy#p->k@9t+B%P#D7~)agQuv+y0k 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Pre-processing4. Post-processing
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