diff --git a/CHANGELOG.md b/CHANGELOG.md index c5c417dd3..f85fb7c6c 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -17,6 +17,7 @@ Special thanks to the following for their contributions to the release: - [PR #1369](https://github.com/nf-core/rnaseq/pull/1369) - Add umicollapse as an alternative to umi-tools - [PR #1461](https://github.com/nf-core/rnaseq/pull/1461) - Add FASTQ linting during preprocessing - [PR #1463](https://github.com/nf-core/rnaseq/pull/1463) - Move channel operations outside of the onComplete() block +- [PR #1467](https://github.com/nf-core/rnaseq/pull/1467) - Add test suite for UMI handling functionality ### Software dependencies diff --git a/tests/.nftignore b/tests/.nftignore index a01770b1e..7f3fad699 100644 --- a/tests/.nftignore +++ b/tests/.nftignore @@ -13,6 +13,7 @@ star_rsem/*.{genes,isoforms}.results star_rsem/log/*.log star_salmon/log/*.Log.{final.out,out,progress.out} trimgalore/*fastq.gz_trimming_report.txt +umitools/*.umi_extract.log {hisat2,star_rsem,star_salmon}/*.{bam,bam.bai} {hisat2,star_rsem,star_salmon}/bigwig/*.{forward,reverse}.bigWig {hisat2,star_rsem,star_salmon}/dupradar/box_plot/*_duprateExpBoxplot.pdf @@ -31,6 +32,7 @@ trimgalore/*fastq.gz_trimming_report.txt {hisat2,star_rsem,star_salmon}/stringtie/*.ballgown/t_data.ctab {hisat2,star_rsem,star_salmon}/stringtie/*.gene.abundance.txt {hisat2,star_rsem,star_salmon}/stringtie/*.{coverage,transcripts}.gtf +{hisat2,star_rsem,star_salmon}/umitools/genomic_dedup_log/*_UMICollapse.log {multiqc,multiqc/**}/multiqc_report.html {multiqc,multiqc/**}/multiqc_report_data/fastqc_{raw,trimmed}_top_overrepresented_sequences_table.txt {multiqc,multiqc/**}/multiqc_report_data/hisat2_pe_plot.txt @@ -81,6 +83,8 @@ trimgalore/*fastq.gz_trimming_report.txt {salmon,star_rsem,star_salmon}/deseq2_qc/deseq2.sample.dists.txt {salmon,star_rsem,star_salmon}/deseq2_qc/size_factors/*.txt {salmon,star_rsem,star_salmon}/deseq2_qc/size_factors/deseq2.size_factors.RData +{salmon,star_rsem,star_salmon}/umitools/{genomic,transcriptomic}_dedup_log/* +{salmon,star_rsem,star_salmon}/umitools/prepare_for_salmon_log/* {salmon,star_salmon}/*/aux_info/fld.gz {salmon,star_salmon}/*/aux_info/meta_info.json {salmon,star_salmon}/*/libParams/flenDist.txt diff --git a/tests/umi.nf.test b/tests/umi.nf.test new file mode 100644 index 000000000..dba4c07fe --- /dev/null +++ b/tests/umi.nf.test @@ -0,0 +1,110 @@ +nextflow_pipeline { + + name "Test pipeline with dummy UMI settings" + script "../main.nf" + + test("Params: --aligner hisat2 --umi_dedup_tool 'umicollapse'") { + + when { + params { + with_umi = true + umitools_extract_method = "regex" + umitools_bc_pattern = "^(?PCGA.{8}){s<=2}.*" + umitools_dedup_stats = true + skip_bbsplit = true + umi_dedup_tool = 'umicollapse' + aligner = 'hisat2' + outdir = "$outputDir" + } + } + + then { + // stable_name: All files + folders in ${params.outdir}/ with a stable name + def stable_name = getAllFilesFromDir(params.outdir, relative: true, includeDir: true, ignore: ['pipeline_info/*.{html,json,txt}']) + // stable_path: All files in ${params.outdir}/ with stable content + def stable_path = getAllFilesFromDir(params.outdir, ignoreFile: 'tests/.nftignore') + assertAll( + { assert workflow.success}, + { assert snapshot( + // Number of successful tasks + workflow.trace.succeeded().size(), + // pipeline versions.yml file for multiqc from which Nextflow version is removed because we tests pipelines on multiple Nextflow versions + removeNextflowVersion("$outputDir/pipeline_info/nf_core_rnaseq_software_mqc_versions.yml"), + // All stable path name, with a relative path + stable_name, + // All files with stable contents + stable_path + ).match() } + ) + } + } + + test("--umi_dedup_tool 'umitools'") { + + when { + params { + with_umi = true + umitools_extract_method = "regex" + umitools_bc_pattern = "^(?PCGA.{8}){s<=2}.*" + umitools_dedup_stats = true + skip_bbsplit = true + outdir = "$outputDir" + } + } + + then { + // stable_name: All files + folders in ${params.outdir}/ with a stable name + def stable_name = getAllFilesFromDir(params.outdir, relative: true, includeDir: true, ignore: ['pipeline_info/*.{html,json,txt}']) + // stable_path: All files in ${params.outdir}/ with stable content + def stable_path = getAllFilesFromDir(params.outdir, ignoreFile: 'tests/.nftignore') + assertAll( + { assert workflow.success}, + { assert snapshot( + // Number of successful tasks + workflow.trace.succeeded().size(), + // pipeline versions.yml file for multiqc from which Nextflow version is removed because we tests pipelines on multiple Nextflow versions + removeNextflowVersion("$outputDir/pipeline_info/nf_core_rnaseq_software_mqc_versions.yml"), + // All stable path name, with a relative path + stable_name, + // All files with stable contents + stable_path + ).match() } + ) + } + } + + test("--umi_dedup_tool 'umitools - stub") { + + options "-stub" + + when { + params { + with_umi = true + umitools_extract_method = "regex" + umitools_bc_pattern = "^(?PCGA.{8}){s<=2}.*" + umitools_dedup_stats = true + outdir = "$outputDir" + } + } + + then { + // stable_name: All files + folders in ${params.outdir}/ with a stable name + def stable_name = getAllFilesFromDir(params.outdir, relative: true, includeDir: true, ignore: ['pipeline_info/*.{html,json,txt}']) + // stable_path: All files in ${params.outdir}/ with stable content + def stable_path = getAllFilesFromDir(params.outdir, ignoreFile: 'tests/.nftignore') + assertAll( + { assert workflow.success}, + { assert snapshot( + // Number of successful tasks + workflow.trace.succeeded().size(), + // pipeline versions.yml file for multiqc from which Nextflow version is removed because we tests pipelines on multiple Nextflow versions + removeNextflowVersion("$outputDir/pipeline_info/nf_core_rnaseq_software_mqc_versions.yml"), + // All stable path name, with a relative path + stable_name, + // All files with stable contents + stable_path + ).match() } + ) + } + } +} diff --git a/tests/umi.nf.test.snap b/tests/umi.nf.test.snap new file mode 100644 index 000000000..b12fa48d0 --- /dev/null +++ b/tests/umi.nf.test.snap @@ -0,0 +1,2812 @@ +{ + "--umi_dedup_tool 'umitools'": { + "content": [ + 265, + { + "BEDTOOLS_GENOMECOV_FW": { + "bedtools": "2.31.1" + }, + "CAT_FASTQ": { + "cat": 9.5 + }, + "CUSTOM_CATADDITIONALFASTA": { + "python": "3.12.2" + }, + "CUSTOM_GETCHROMSIZES": { + "getchromsizes": 1.21 + }, + "CUSTOM_TX2GENE": { + "python": "3.10.4" + }, + "DESEQ2_QC_PSEUDO": { + "r-base": "4.0.3", + "bioconductor-deseq2": "1.28.0" + }, + "DESEQ2_QC_STAR_SALMON": { + "r-base": "4.0.3", + "bioconductor-deseq2": "1.28.0" + }, + "DUPRADAR": { + "bioconductor-dupradar": "1.32.0" + }, + "FASTQC": { + "fastqc": "0.12.1" + }, + "FQ_LINT": { + "fq": "0.12.0 (2024-07-08)" + }, + "FQ_SUBSAMPLE": { + "fq": "0.12.0 (2024-07-08)" + }, + "GTF2BED": { + "perl": "5.26.2" + }, + "GTF_FILTER": { + "python": "3.9.5" + }, + "GUNZIP_ADDITIONAL_FASTA": { + "gunzip": 1.1 + }, + "GUNZIP_GTF": { + "gunzip": 1.1 + }, + "MULTIQC_CUSTOM_BIOTYPE": { + "python": "3.9.5" + }, + "QUALIMAP_RNASEQ": { + "qualimap": 2.3 + }, + "RSEQC_BAMSTAT": { + "rseqc": "5.0.2" + }, + "RSEQC_INFEREXPERIMENT": { + "rseqc": "5.0.2" + }, + "RSEQC_INNERDISTANCE": { + "rseqc": "5.0.2" + }, + "RSEQC_JUNCTIONANNOTATION": { + "rseqc": "5.0.2" + }, + "RSEQC_JUNCTIONSATURATION": { + "rseqc": "5.0.2" + }, + "RSEQC_READDISTRIBUTION": { + "rseqc": "5.0.2" + }, + "RSEQC_READDUPLICATION": { + "rseqc": "5.0.2" + }, + "SALMON_QUANT": { + "salmon": "1.10.3" + }, + "SAMTOOLS_FLAGSTAT": { + "samtools": 1.21 + }, + "SAMTOOLS_IDXSTATS": { + "samtools": 1.21 + }, + "SAMTOOLS_INDEX": { + "samtools": 1.21 + }, + "SAMTOOLS_SORT": { + "samtools": 1.21 + }, + "SAMTOOLS_STATS": { + "samtools": 1.21 + }, + "SE_GENE": { + "bioconductor-summarizedexperiment": "1.32.0" + }, + "STAR_ALIGN": { + "star": "2.7.11b", + "samtools": 1.2, + "gawk": "5.1.0" + }, + "STAR_GENOMEGENERATE": { + "star": "2.7.11b", + "samtools": 1.2, + "gawk": "5.1.0" + }, + "STRINGTIE_STRINGTIE": { + "stringtie": "2.2.3" + }, + "SUBREAD_FEATURECOUNTS": { + "subread": "2.0.6" + }, + "TRIMGALORE": { + "trimgalore": "0.6.10", + "cutadapt": 4.9 + }, + "TXIMETA_TXIMPORT": { + "bioconductor-tximeta": "1.20.1" + }, + "UCSC_BEDCLIP": { + "ucsc": 377 + }, + "UCSC_BEDGRAPHTOBIGWIG": { + "ucsc": 469 + }, + "UMITOOLS_DEDUP": { + "umitools": "1.1.5" + }, + "UMITOOLS_EXTRACT": { + "umitools": "1.1.5" + }, + "UMITOOLS_PREPAREFORSALMON": { + "umitools": "1.1.5" + }, + "UNTAR_SALMON_INDEX": { + "untar": 1.34 + }, + "Workflow": { + "nf-core/rnaseq": "v3.18.0dev" + } + }, + [ + "custom", + "custom/out", + "custom/out/genome_gfp.fasta", + "custom/out/genome_gfp.gtf", + "fastqc", + "fastqc/raw", + "fastqc/raw/RAP1_IAA_30M_REP1_raw_1_fastqc.html", + "fastqc/raw/RAP1_IAA_30M_REP1_raw_1_fastqc.zip", + 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"fastqc/trim/RAP1_UNINDUCED_REP1_trimmed_trimmed_fastqc.zip", + "fastqc/trim/RAP1_UNINDUCED_REP2_trimmed_trimmed_fastqc.html", + "fastqc/trim/RAP1_UNINDUCED_REP2_trimmed_trimmed_fastqc.zip", + "fastqc/trim/WT_REP1_trimmed_1_val_1_fastqc.html", + "fastqc/trim/WT_REP1_trimmed_1_val_1_fastqc.zip", + "fastqc/trim/WT_REP1_trimmed_2_val_2_fastqc.html", + "fastqc/trim/WT_REP1_trimmed_2_val_2_fastqc.zip", + "fastqc/trim/WT_REP2_trimmed_1_val_1_fastqc.html", + "fastqc/trim/WT_REP2_trimmed_1_val_1_fastqc.zip", + "fastqc/trim/WT_REP2_trimmed_2_val_2_fastqc.html", + "fastqc/trim/WT_REP2_trimmed_2_val_2_fastqc.zip", + "fq_lint", + "fq_lint/raw", + "fq_lint/raw/RAP1_IAA_30M_REP1.fq_lint.txt", + "fq_lint/raw/RAP1_UNINDUCED_REP1.fq_lint.txt", + "fq_lint/raw/RAP1_UNINDUCED_REP2.fq_lint.txt", + "fq_lint/raw/WT_REP1.fq_lint.txt", + "fq_lint/raw/WT_REP2.fq_lint.txt", + "fq_lint/trimmed", + "fq_lint/trimmed/RAP1_IAA_30M_REP1.fq_lint.txt", + "fq_lint/trimmed/RAP1_UNINDUCED_REP1.fq_lint.txt", + 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"multiqc/star_salmon/multiqc_report_data/fastqc_raw_per_sequence_gc_content_plot_Counts.txt", + "multiqc/star_salmon/multiqc_report_data/fastqc_raw_per_sequence_gc_content_plot_Percentages.txt", + "multiqc/star_salmon/multiqc_report_data/fastqc_raw_per_sequence_quality_scores_plot.txt", + "multiqc/star_salmon/multiqc_report_data/fastqc_raw_sequence_counts_plot.txt", + "multiqc/star_salmon/multiqc_report_data/fastqc_raw_sequence_duplication_levels_plot.txt", + "multiqc/star_salmon/multiqc_report_data/fastqc_raw_top_overrepresented_sequences_table.txt", + "multiqc/star_salmon/multiqc_report_data/fastqc_sequence_length_distribution_plot.txt", + "multiqc/star_salmon/multiqc_report_data/fastqc_trimmed-status-check-heatmap.txt", + "multiqc/star_salmon/multiqc_report_data/fastqc_trimmed_overrepresented_sequences_plot.txt", + "multiqc/star_salmon/multiqc_report_data/fastqc_trimmed_per_base_n_content_plot.txt", + 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"multiqc/star_salmon/multiqc_report_data/samtools-idxstats-mapped-reads-plot_Raw_Counts.txt", + "multiqc/star_salmon/multiqc_report_data/samtools-stats-dp.txt", + "multiqc/star_salmon/multiqc_report_data/samtools_alignment_plot.txt", + "multiqc/star_salmon/multiqc_report_data/star_alignment_plot.txt", + "multiqc/star_salmon/multiqc_report_data/star_summary_table.txt", + "multiqc/star_salmon/multiqc_report_data/umitools_deduplication_barplot.txt", + "multiqc/star_salmon/multiqc_report_data/umitools_stats_violin.txt", + "multiqc/star_salmon/multiqc_report_plots", + "multiqc/star_salmon/multiqc_report_plots/pdf", + "multiqc/star_salmon/multiqc_report_plots/pdf/cutadapt_filtered_reads_plot-cnt.pdf", + "multiqc/star_salmon/multiqc_report_plots/pdf/cutadapt_filtered_reads_plot-pct.pdf", + "multiqc/star_salmon/multiqc_report_plots/pdf/cutadapt_trimmed_sequences_plot_3_Counts.pdf", + "multiqc/star_salmon/multiqc_report_plots/pdf/cutadapt_trimmed_sequences_plot_3_Obs_Exp.pdf", + 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ch_multiqc_files.mix(ALIGN_STAR.out.idxstats.collect{it[1]}) ch_multiqc_files = ch_multiqc_files.mix(ch_star_log.collect{it[1]}) if (params.bam_csi_index) { @@ -300,6 +297,14 @@ workflow RNASEQ { .single_end .mix(UMITOOLS_PREPAREFORSALMON.out.bam) .set { ch_transcriptome_bam } + } else { + // The deduplicated stats should take priority for MultiQC, but use + // them straight out of the aligner otherwise + + ch_multiqc_files = ch_multiqc_files + .mix(ALIGN_STAR.out.stats.collect{it[1]}) + .mix(ALIGN_STAR.out.flagstat.collect{it[1]}) + .mix(ALIGN_STAR.out.idxstats.collect{it[1]}) } // @@ -381,9 +386,6 @@ workflow RNASEQ { ch_genome_bam = FASTQ_ALIGN_HISAT2.out.bam ch_genome_bam_index = FASTQ_ALIGN_HISAT2.out.bai ch_unaligned_sequences = FASTQ_ALIGN_HISAT2.out.fastq - ch_multiqc_files = ch_multiqc_files.mix(FASTQ_ALIGN_HISAT2.out.stats.collect{it[1]}) - ch_multiqc_files = ch_multiqc_files.mix(FASTQ_ALIGN_HISAT2.out.flagstat.collect{it[1]}) - ch_multiqc_files = ch_multiqc_files.mix(FASTQ_ALIGN_HISAT2.out.idxstats.collect{it[1]}) ch_multiqc_files = ch_multiqc_files.mix(FASTQ_ALIGN_HISAT2.out.summary.collect{it[1]}) if (params.bam_csi_index) { @@ -420,6 +422,13 @@ workflow RNASEQ { ch_genome_bam_index = UMI_DEDUP_GENOME.out.csi } ch_versions = ch_versions.mix(UMI_DEDUP_GENOME.out.versions) + } else { + // The deduplicated stats should take priority for MultiQC, but use + // them straight out of the aligner otherwise + ch_multiqc_files = ch_multiqc_files + .mix(FASTQ_ALIGN_HISAT2.out.stats.collect{it[1]}) + .mix(FASTQ_ALIGN_HISAT2.out.flagstat.collect{it[1]}) + .mix(FASTQ_ALIGN_HISAT2.out.idxstats.collect{it[1]}) } }