Merge branch 'CW-3998' into 'dev'
Handle annotations with no strand CW-3998 Closes CW-3998 See merge request epi2melabs/workflows/wf-transcriptomes!170
This commit is contained in:
Sarah Griffiths
commit
d5df11cce4
@ -53,7 +53,8 @@ docker-run:
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"fusions", "differential_expression", "isoforms", "isoforms_bam",
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"only_differential_expression", "differential_expression_gff3",
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"ncbi_gzip", "ncbi_no_gene_id", "ensembl_with_versions",
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"differential_expression_mouse", "no_ref_annotation"
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"differential_expression_mouse", "no_ref_annotation",
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"unstranded_annotation_error"
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]
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rules:
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# NOTE As we're overriding the rules block for the included docker-run
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@ -67,21 +68,21 @@ docker-run:
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NF_WORKFLOW_OPTS: "--fastq ${CI_PROJECT_NAME}/data/ERR6053095_chr20.fastq --transcriptome-source reference-guided \
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--ref_genome ${CI_PROJECT_NAME}/data/chr20/hg38_chr20.fa --ref_annotation ${CI_PROJECT_NAME}/data/chr20/gencode.v22.annotation.chr20.gtf --pychopper_backend phmm \
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-c ${CI_PROJECT_NAME}/data/demo.nextflow.config"
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NF_IGNORE_PROCESSES: preprocess_reads,merge_transcriptomes,decompress_annotation,decompress_ref,decompress_transcriptome,preprocess_ref_transcriptome
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NF_IGNORE_PROCESSES: preprocess_reads,check_annotation_strand,merge_transcriptomes,decompress_annotation,decompress_ref,decompress_transcriptome,preprocess_ref_transcriptome
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- if: $MATRIX_NAME == "isoforms_bam"
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variables:
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NF_BEFORE_SCRIPT: mkdir -p ${CI_PROJECT_NAME}/data/ && wget -O ${CI_PROJECT_NAME}/data/wf-isoforms_test_data.tar.gz https://ont-exd-int-s3-euwst1-epi2me-labs.s3.amazonaws.com/wf-isoforms/wf-isoforms_test_data.tar.gz && tar -xzvf ${CI_PROJECT_NAME}/data/wf-isoforms_test_data.tar.gz -C ${CI_PROJECT_NAME}/data/ && wget -O ${CI_PROJECT_NAME}/data/demo.nextflow.config https://ont-exd-int-s3-euwst1-epi2me-labs.s3.amazonaws.com/wf-isoforms/demo.nextflow.config
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NF_WORKFLOW_OPTS: "--bam ${CI_PROJECT_NAME}/data/ERR6053095_chr20.bam --transcriptome-source reference-guided \
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--ref_genome ${CI_PROJECT_NAME}/data/chr20/hg38_chr20.fa --ref_annotation ${CI_PROJECT_NAME}/data/chr20/gencode.v22.annotation.chr20.gtf --pychopper_backend phmm \
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-c ${CI_PROJECT_NAME}/data/demo.nextflow.config"
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NF_IGNORE_PROCESSES: preprocess_reads,merge_transcriptomes,decompress_annotation,decompress_ref,decompress_transcriptome,preprocess_ref_transcriptome
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NF_IGNORE_PROCESSES: preprocess_reads,check_annotation_strand,merge_transcriptomes,decompress_annotation,decompress_ref,decompress_transcriptome,preprocess_ref_transcriptome
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- if: $MATRIX_NAME == "no_ref_annotation"
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variables:
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NF_BEFORE_SCRIPT: mkdir -p ${CI_PROJECT_NAME}/data/ && wget -O ${CI_PROJECT_NAME}/data/wf-isoforms_test_data.tar.gz https://ont-exd-int-s3-euwst1-epi2me-labs.s3.amazonaws.com/wf-isoforms/wf-isoforms_test_data.tar.gz && tar -xzvf ${CI_PROJECT_NAME}/data/wf-isoforms_test_data.tar.gz -C ${CI_PROJECT_NAME}/data/ && wget -O ${CI_PROJECT_NAME}/data/demo.nextflow.config https://ont-exd-int-s3-euwst1-epi2me-labs.s3.amazonaws.com/wf-isoforms/demo.nextflow.config
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NF_WORKFLOW_OPTS: "--fastq ${CI_PROJECT_NAME}/data/ERR6053095_chr20.fastq --transcriptome-source reference-guided \
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--ref_genome ${CI_PROJECT_NAME}/data/chr20/hg38_chr20.fa \
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-c ${CI_PROJECT_NAME}/data/demo.nextflow.config"
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NF_IGNORE_PROCESSES: run_gffcompare,preprocess_reads,merge_transcriptomes,decompress_annotation,decompress_ref,decompress_transcriptome,preprocess_ref_transcriptome
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NF_IGNORE_PROCESSES: run_gffcompare,check_annotation_strand,preprocess_reads,merge_transcriptomes,decompress_annotation,decompress_ref,decompress_transcriptome,preprocess_ref_transcriptome
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- if: $MATRIX_NAME == "fusions"
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variables:
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NF_BEFORE_SCRIPT: mkdir -p ${CI_PROJECT_NAME}/data/ && wget -O ${CI_PROJECT_NAME}/data/wf-isoforms_test_data.tar.gz https://ont-exd-int-s3-euwst1-epi2me-labs.s3.amazonaws.com/wf-isoforms/wf-isoforms_test_data.tar.gz && tar -xzvf ${CI_PROJECT_NAME}/data/wf-isoforms_test_data.tar.gz -C ${CI_PROJECT_NAME}/data/ && wget -O ${CI_PROJECT_NAME}/data/demo.nextflow.config https://ont-exd-int-s3-euwst1-epi2me-labs.s3.amazonaws.com/wf-isoforms/demo.nextflow.config
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@ -89,7 +90,7 @@ docker-run:
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--ref_genome ${CI_PROJECT_NAME}/data/chr20/hg38_chr20.fa --ref_annotation ${CI_PROJECT_NAME}/data/chr20/gencode.v22.annotation.chr20.gtf \
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--jaffal_refBase ${CI_PROJECT_NAME}/data/chr20/ --jaffal_genome hg38_chr20 --jaffal_annotation genCode22 \
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-c ${CI_PROJECT_NAME}/data/demo.nextflow.config"
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NF_IGNORE_PROCESSES: preprocess_reads,merge_transcriptomes,decompress_annotation,decompress_ref,decompress_transcriptome,preprocess_ref_transcriptome
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NF_IGNORE_PROCESSES: preprocess_reads,check_annotation_strand,merge_transcriptomes,decompress_annotation,decompress_ref,decompress_transcriptome,preprocess_ref_transcriptome
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- if: $MATRIX_NAME == "differential_expression"
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variables:
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NF_BEFORE_SCRIPT: mkdir -p ${CI_PROJECT_NAME}/data/ && wget -O ${CI_PROJECT_NAME}/data/differential_expression.tar.gz https://ont-exd-int-s3-euwst1-epi2me-labs.s3.amazonaws.com/wf-isoforms/differential_expression.tar.gz && tar -xzvf ${CI_PROJECT_NAME}/data/differential_expression.tar.gz -C ${CI_PROJECT_NAME}/data/ && wget -O ${CI_PROJECT_NAME}/data/demo.nextflow.config https://ont-exd-int-s3-euwst1-epi2me-labs.s3.amazonaws.com/wf-isoforms/demo.nextflow.config
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@ -187,3 +188,15 @@ docker-run:
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NF_IGNORE_PROCESSES: >
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preprocess_reads,merge_transcriptomes,assemble_transcripts,decompress_annotation,
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build_minimap_index,get_transcriptome,merge_gff_bundles,run_gffcompare,build_minimap_index,split_bam
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- if: $MATRIX_NAME == "unstranded_annotation_error"
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variables:
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NF_BEFORE_SCRIPT: mkdir -p ${CI_PROJECT_NAME}/data/ && wget -O ${CI_PROJECT_NAME}/data/differential_expression.tar.gz https://ont-exd-int-s3-euwst1-epi2me-labs.s3.amazonaws.com/wf-transcriptomes/differential_expression.tar.gz && tar -xzvf ${CI_PROJECT_NAME}/data/differential_expression.tar.gz -C ${CI_PROJECT_NAME}/data/ && wget -O ${CI_PROJECT_NAME}/data/demo.nextflow.config https://ont-exd-int-s3-euwst1-epi2me-labs.s3.amazonaws.com/wf-isoforms/demo.nextflow.config
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NF_WORKFLOW_OPTS: "--fastq ${CI_PROJECT_NAME}/data/differential_expression/differential_expression_fastq \
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--de_analysis \
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--ref_genome ${CI_PROJECT_NAME}/data/differential_expression/hg38_chr20.fa --transcriptome-source reference-guided \
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--ref_annotation ${CI_PROJECT_NAME}/data/differential_expression/unstranded_annotation.gtf \
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--direct_rna --minimap2_index_opts '-k 15' --sample_sheet ${CI_PROJECT_NAME}/data/differential_expression/sample_sheet.csv \
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-c ${CI_PROJECT_NAME}/data/demo.nextflow.config"
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NF_IGNORE_PROCESSES: preprocess_reads,merge_transcriptomes,decompress_annotation,decompress_ref,decompress_transcriptome,preprocess_ref_transcriptome
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ASSERT_NEXTFLOW_FAILURE: "test_fail" # set to any non-zero length str to allow the nextflow CMD to fail
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ASSERT_NEXTFLOW_FAILURE_REXP: "In ref_annotation, transcript features must have a strand of either '+' or '-'"
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@ -9,6 +9,8 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0
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- Workflow now accepts BAM or FASTQ files as input (using the --bam or --fastq parameters, respectively).
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### Changed
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- MA plot in the `results_dge.pdf` has been updated to match the MA plot in the report.
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### Added
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- Error message when running in `de_analysis` mode and `ref_annotation` input file contains unstranded annotations.
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## [v1.1.1]
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### Changed
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@ -117,7 +117,7 @@ input_reads.fastq ─── input_directory ─── input_directory
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| transcriptome_source | string | Select how the transcriptome used for analysis should be prepared. | To analyse only gene fusions and differential expression use of an existing transcriptome may be preferred and so 'precomputed' should be selected. In this case the 'ref_transcriptome' parameter should be specified. To create a reference transcriptome using an existing reference genome, select 'reference guided' and specify the 'ref_genome' parameter. | reference-guided |
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| ref_genome | string | Path to reference genome sequence [.fa/.fq/.fa.gz/fq.gz]. Required for reference-based workflow. | A reference genome is required for reference-based assembly of a transcriptome. | |
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| ref_transcriptome | string | Transcriptome reference file. Required for precomputed transcriptome calculation and for differential expression analysis. | A reference transcriptome related to the sample under study. Must be supplied when the 'Transcriptome source' parameter has been set to 'precomputed' or to perform differential expression. | |
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| ref_annotation | string | A reference annotation in GFF2 or GFF3 format (extensions .gtf(.gz), .gff(.gz), .gff3(.gz)). Only annotation files from [Encode](https://www.encodeproject.org), [Ensembl](https://www.ensembl.org/index.html) and [NCBI](https://www.ncbi.nlm.nih.gov/) are supported. | This will be used for guiding the transcriptome assembly and to label transcripts with their corresponding gene identifiers. | |
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| ref_annotation | string | A reference annotation in GFF2 or GFF3 format (extensions .gtf(.gz), .gff(.gz), .gff3(.gz)). Only annotation files from [Encode](https://www.encodeproject.org), [Ensembl](https://www.ensembl.org/index.html) and [NCBI](https://www.ncbi.nlm.nih.gov/) are supported. | This will be used for guiding the transcriptome assembly and to label transcripts with their corresponding gene identifiers. Note: If in de_analysis mode transcript strands must be only + or -. | |
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| direct_rna | boolean | Set to true for direct RNA sequencing. | Omits the pychopper step. | False |
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| analyse_unclassified | boolean | Analyse unclassified reads from input directory. By default the workflow will not process reads in the unclassified directory. | If selected and if the input is a multiplex directory the workflow will also process the unclassified directory. | False |
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@ -7,7 +7,7 @@
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| transcriptome_source | string | Select how the transcriptome used for analysis should be prepared. | To analyse only gene fusions and differential expression use of an existing transcriptome may be preferred and so 'precomputed' should be selected. In this case the 'ref_transcriptome' parameter should be specified. To create a reference transcriptome using an existing reference genome, select 'reference guided' and specify the 'ref_genome' parameter. | reference-guided |
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| ref_genome | string | Path to reference genome sequence [.fa/.fq/.fa.gz/fq.gz]. Required for reference-based workflow. | A reference genome is required for reference-based assembly of a transcriptome. | |
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| ref_transcriptome | string | Transcriptome reference file. Required for precomputed transcriptome calculation and for differential expression analysis. | A reference transcriptome related to the sample under study. Must be supplied when the 'Transcriptome source' parameter has been set to 'precomputed' or to perform differential expression. | |
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| ref_annotation | string | A reference annotation in GFF2 or GFF3 format (extensions .gtf(.gz), .gff(.gz), .gff3(.gz)). Only annotation files from [Encode](https://www.encodeproject.org), [Ensembl](https://www.ensembl.org/index.html) and [NCBI](https://www.ncbi.nlm.nih.gov/) are supported. | This will be used for guiding the transcriptome assembly and to label transcripts with their corresponding gene identifiers. | |
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| ref_annotation | string | A reference annotation in GFF2 or GFF3 format (extensions .gtf(.gz), .gff(.gz), .gff3(.gz)). Only annotation files from [Encode](https://www.encodeproject.org), [Ensembl](https://www.ensembl.org/index.html) and [NCBI](https://www.ncbi.nlm.nih.gov/) are supported. | This will be used for guiding the transcriptome assembly and to label transcripts with their corresponding gene identifiers. Note: If in de_analysis mode transcript strands must be only + or -. | |
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| direct_rna | boolean | Set to true for direct RNA sequencing. | Omits the pychopper step. | False |
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| analyse_unclassified | boolean | Analyse unclassified reads from input directory. By default the workflow will not process reads in the unclassified directory. | If selected and if the input is a multiplex directory the workflow will also process the unclassified directory. | False |
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24
main.nf
24
main.nf
@ -374,7 +374,7 @@ process merge_transcriptomes {
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path "final_non_redundant_transcriptome.fasta", emit: fasta
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path "stringtie.gtf", emit: gtf
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"""
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stringtie --merge -G $ref_annotation -p ${task.cpus} -o stringtie.gtf query_annotations/*
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stringtie --rf --merge -G $ref_annotation -p ${task.cpus} -o stringtie.gtf query_annotations/*
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seqkit subseq --feature "transcript" --gtf-tag "transcript_id" --gtf stringtie.gtf $ref_genome > temp_transcriptome.fasta
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seqkit rmdup -s < temp_transcriptome.fasta > temp_del_repeats.fasta
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cat temp_del_repeats.fasta | sed 's/>.* />/' | sed -e 's/_[0-9]* \\[/ \\[/' > temp_rm_empty_seq.fasta
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@ -519,6 +519,22 @@ process output {
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}
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// Check ref_annotation transcript strand column for "." if in de_analysis mode
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process check_annotation_strand {
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label "isoforms"
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cpus 1
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memory "2 GB"
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input:
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path "ref_annotation.gtf"
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output:
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tuple stdout, path("ref_annotation.gtf")
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"""
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awk '{if (\$3=="transcript" && \$7 != "+" && \$7 != "-") print \$3, \$7}' "ref_annotation.gtf"
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"""
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}
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// workflow module
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workflow pipeline {
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take:
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@ -648,6 +664,12 @@ workflow pipeline {
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if (params.de_analysis){
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sample_sheet = file(params.sample_sheet, type:"file")
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// check ref annotation contains only + or - strand as DE analysis will error on .
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check_annotation_strand(ref_annotation).map { stdoutput, annotation ->
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// check if there was an error message
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if (stdoutput) error "In ref_annotation, transcript features must have a strand of either '+' or '-'."
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stdoutput
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}
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if (!params.ref_transcriptome){
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merge_transcriptomes(run_gffcompare.output.gtf.collect(), ref_annotation, ref_genome)
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transcriptome = merge_transcriptomes.out.fasta
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@ -60,7 +60,7 @@
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"format": "file-path",
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"demo_data": "${projectDir}/test_data/SIRV_isoforms.gtf",
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"description": "A reference annotation in GFF2 or GFF3 format (extensions .gtf(.gz), .gff(.gz), .gff3(.gz)). Only annotation files from [Encode](https://www.encodeproject.org), [Ensembl](https://www.ensembl.org/index.html) and [NCBI](https://www.ncbi.nlm.nih.gov/) are supported.",
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"help_text": "This will be used for guiding the transcriptome assembly and to label transcripts with their corresponding gene identifiers."
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"help_text": "This will be used for guiding the transcriptome assembly and to label transcripts with their corresponding gene identifiers. Note: If in de_analysis mode transcript strands must be only + or -."
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},
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"direct_rna": {
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"type": "boolean",
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