diff --git a/.github/workflows/ci.yml b/.github/workflows/ci.yml index 6e833767..3996d01d 100644 --- a/.github/workflows/ci.yml +++ b/.github/workflows/ci.yml @@ -74,6 +74,7 @@ jobs: "test_contrast_list", "test_no_multiqc", "test_relevel", + "test_smrnaseq", "test_star_rsem", "test_star_salmon", "test_use_vst", diff --git a/.prettierignore b/.prettierignore index 437d763d..88a6ffb4 100644 --- a/.prettierignore +++ b/.prettierignore @@ -6,6 +6,7 @@ work/ data/ results/ .DS_Store +testdata/ testing/ testing* *.pyc diff --git a/CHANGELOG.md b/CHANGELOG.md index 2b5292b6..e07a79a6 100644 --- a/CHANGELOG.md +++ b/CHANGELOG.md @@ -7,6 +7,7 @@ and this project adheres to [Semantic Versioning](https://semver.org/spec/v2.0.0 ### Added +- [#213](https://github.com/qbic-pipelines/rnadeseq/pull/21) Added smrnaseq input support - [#212](https://github.com/qbic-pipelines/rnadeseq/pull/212) Added computational methods if no --software_versions - [#206](https://github.com/qbic-pipelines/rnadeseq/pull/206) Added logic to decide between rlog and vst, added tryCatch for heatmap saving because this only works unreliably - [#202](https://github.com/qbic-pipelines/rnadeseq/pull/202) Added background list to pathway analysis diff --git a/assets/RNAseq_report.Rmd b/assets/RNAseq_report.Rmd index 0fe78896..8d3d3135 100644 --- a/assets/RNAseq_report.Rmd +++ b/assets/RNAseq_report.Rmd @@ -86,6 +86,35 @@ label_size <- function(dimension_int) { val <- max(c(2, 10-dimension_int/15)) return(val) } + +# Function for creating a dataframe from smrnaseq files +build_smrnaseq_df <- function(files) { + merged_mat <- F + for (f in files) { + file_tab <- read.table(f, sep="\t", row.names=1) + # Usually, the last row is weird and has the rowname *, remove it + if (grep("*", tail(rownames(file_tab), n=1), fixed=T)) { + file_tab <- head(file_tab, -1) + } + + # If merged_mat already exists, add third matrix column to it (second column after rownames) + if (is.data.frame(merged_mat)) { + + # Check if rownames are the same + if (rownames(file_tab) != rownames(merged_mat)) { + stop(paste("Rownames of smrnaseq file", file_tab, "differ from the rownames of the other files; exiting...")) + } + merged_mat[[basename(f)]] <- file_tab[,2] + } else { + + # Otherwise initiate it from third matrix column and rownames + merged_mat <- data.frame(file_tab[,2]) + rownames(merged_mat) <- rownames(file_tab) + colnames(merged_mat) <- c(basename(f)) + } + } + merged_mat +} ``` ```{r param_validation, echo=FALSE, message=FALSE, warning=FALSE, results = 'hide'} @@ -159,8 +188,8 @@ write_error_msg <- function(counts, qbicCodes) { } #Check necessary params -if (!(params$input_type %in% c("featurecounts", "rsem", "salmon"))){ - stop(paste0("Wrong input type ", params$input_type, ", must be one of [featurecounts, rsem, salmon]!")) +if (!(params$input_type %in% c("featurecounts", "rsem", "salmon", "smrnaseq"))){ + stop(paste0("Wrong input type ", params$input_type, ", must be one of [featurecounts, rsem, salmon, smrnaseq]!")) } if (params$input_type %in% c("rsem", "salmon") & !isProvided(params$path_gtf)){ stop(paste0("For input type salmon, gtf file needs to be provided!\nIf using igenomes, please check that the entry for your genome contains a gtf file and otherwise provide one with `--gtf`.")) @@ -340,9 +369,60 @@ metadata$sampleName = paste(row.names(metadata),metadata$Secondary.Name,sep="_") row.names(metadata) = metadata$sampleName metadata_save <- metadata +if (params$input_type == "smrnaseq") { + + # Prepare files from smrnaseq, start with hairpins + hairpin_files <- file.path(gsub("/$", "", params$path_gene_counts), paste0(qbicCodes, "_mature_hairpin.sorted.idxstats")) + if (!(all(file.exists(hairpin_files)))) { + missing_files <- "Could not find all of the specified _mature_hairpin.sorted.idxstats files! Please check that for all QBiC codes in the metadata sheet, a _mature_hairpin.sorted.idxstats file is present in the gene_counts folder and make sure that no typos exist. The following file(s) could not be found: " + for (f in hairpin_files) { + if (!file.exists(f)) { + missing_files <- paste0(missing_files, f, ", ") + } + } + missing_files <- paste0(substring(missing_files, 1, nchar(missing_files)-2), ".") + stop(missing_files, call.=F) + } + + # Non-hairpins next + other_files <- file.path(gsub("/$", "", params$path_gene_counts), paste0(qbicCodes, "_mature.sorted.idxstats")) + if (!(all(file.exists(other_files)))) { + missing_files <- "Could not find all of the specified _mature.sorted.idxstats files! Please check that for all QBiC codes in the metadata sheet, a _mature.sorted.idxstats file is present in the gene_counts folder and make sure that no typos exist. The following file(s) could not be found: " + for (f in other_files) { + if (!file.exists(f)) { + missing_files <- paste0(missing_files, f, ", ") + } + } + missing_files <- paste0(substring(missing_files, 1, nchar(missing_files)-2), ".") + stop(missing_files, call.=F) + } + + # Combine files for later + files <- c(hairpin_files, other_files) + + # Merge hairpins and counts + hairpin_df <- build_smrnaseq_df(hairpin_files) + other_df <- build_smrnaseq_df(other_files) + colnames(other_df) <- colnames(hairpin_df) + count.table <- rbind(hairpin_df, other_df) + + # Need to order columns in count.table + count.table <- count.table[, order(names(count.table))] + + count.table$Ensembl_ID <- rownames(count.table) + count.table <- cbind(Ensembl_ID=rownames(count.table), gene_name=rownames(count.table), count.table) + write.table(count.table, paste("differential_gene_expression/gene_counts_tables/raw_gene_counts.tsv",sep=""), append = FALSE, quote = FALSE, sep = "\t",eol = "\n", na = "NA", dec = ".", row.names = F, qmethod = c("escape", "double")) + + +} + # Load count table for FeatureCounts if (params$input_type == "featurecounts"){ count.table <- read.table(params$path_gene_counts, header = T,sep = "\t",na.strings =c("","NA"),quote=NULL,stringsAsFactors=F,dec=".",fill=TRUE,row.names=1) +} + +# Processing is the same for FeatureCounts and the smrnaseq matrix that is built above +if (params$input_type %in% c("featurecounts", "smrnaseq")) { count.table$Ensembl_ID <- row.names(count.table) drop <- c("Ensembl_ID","gene_name") gene_names <- count.table[,drop] @@ -361,8 +441,8 @@ if (params$input_type == "featurecounts"){ count.table <- count.table[, order(names(count.table))] if (!identical(names(count.table),qbicCodes)) { - error_out <- write_error_msg(count.table, qbicCodes) - stop(paste0(error_out, "\n"), call.=F) + error_out <- write_error_msg(count.table, qbicCodes) + stop(paste0(error_out, "\n"), call.=F) } names(count.table) = metadata$sampleName @@ -443,7 +523,7 @@ DT::datatable(df_QA_neat, options = list(scrollX = "100%", scrollY = "400px", p ################## RUN DESEQ2 ###################################### # Run DESeq function -if (params$input_type == "featurecounts") { +if (params$input_type %in% c("featurecounts", "smrnaseq")) { cds <- DESeqDataSetFromMatrix( countData =count.table, colData =metadata, design = eval(parse(text=as.character(design[[1]])))) } else if (params$input_type %in% c("rsem", "salmon")) { ## Create a dataframe which consists of both the gene id and the transcript name @@ -469,7 +549,7 @@ if (params$input_type == "featurecounts") { #-->the sample name is in the file name files <- file.path(gsub("/$", "", params$path_gene_counts), paste0(qbicCodes, ".genes.results")) if (!(all(file.exists(files)))) { - missing_files <- "DESeq2.R could not find all of the specified .genes.results files! Please check that for all QBiC codes in the metadata sheet, a .genes.results file is present in the gene_counts folder and make sure that no typos exist. The following file(s) could not be found: " + missing_files <- "Could not find all of the specified .genes.results files! Please check that for all QBiC codes in the metadata sheet, a .genes.results file is present in the gene_counts folder and make sure that no typos exist. The following file(s) could not be found: " for (f in files) { if (!file.exists(f)) { missing_files <- paste0(missing_files, f, ", ") @@ -507,7 +587,7 @@ if (params$input_type == "featurecounts") { #-->the sample name is in the subfolder name, not in the file name! files <- file.path(gsub("/$", "", params$path_gene_counts), qbicCodes, "quant.sf") if (!(all(file.exists(files)))) { - missing_files <- "DESeq2.R could not find all of the specified /quant.sf files! Please check that for all QBiC codes in the metadata sheet, a /quant.sf file is present in the gene_counts folder and subfolder and make sure that no typos exist. The following file(s) could not be found: " + missing_files <- "Could not find all of the specified /quant.sf files! Please check that for all QBiC codes in the metadata sheet, a /quant.sf file is present in the gene_counts folder and subfolder and make sure that no typos exist. The following file(s) could not be found: " for (f in files) { if (!file.exists(f)) { missing_files <- paste0(missing_files, f, ", ") @@ -533,7 +613,7 @@ if (params$input_type == "featurecounts") { cds <- DESeqDataSetFromTximport(txi=txi.salmon, colData =coldata, design = eval(parse(text=as.character(design[[1]])))) } } else { - stop(paste0("Invalid input type: ", params$input_type, "! Input type must be one of: featurecounts, rsem, salmon!")) + stop(paste0("Invalid input type: ", params$input_type, "! Input type must be one of: featurecounts, rsem, salmon, smrnaseq!")) } #Apply relevel if provided; this must be done before calling DESeq() @@ -550,7 +630,7 @@ if (isProvided(params$path_relevel)) { cds <- DESeq(cds) # sizeFactors(cds) as indicator of library sequencing depth -if (params$input_type == "featurecounts") { +if (params$input_type %in% c("featurecounts", "smrnaseq")) { sf <- sizeFactors(cds) } else { # If tximport was used, sizeFactors doesn't work, so use workaround as described here: https://support.bioconductor.org/p/97676/ @@ -562,6 +642,11 @@ write.table(sf,paste("differential_gene_expression/gene_counts_tables/sizeFactor # Write cds assay table to file write.table(counts(cds, normalized=T), paste0("differential_gene_expression/gene_counts_tables/deseq2_library_scaled_gene_counts.tsv"), append=F, quote = F, sep = "\t", eol = "\n", na = "NA", dec = ".", row.names = T, col.names = T, qmethod = c("escape", "double")) +# Or for smrnaseq save folder/file names to txt file +if (params$input_type == "smrnaseq") { + write(files, file="differential_gene_expression/metadata/smrnaseq_files.txt") +} + # Or for rsem/salmon save folder/file names to txt file if (params$input_type == "rsem") { write(files, file="differential_gene_expression/metadata/star_rsem_files.txt") @@ -1273,7 +1358,7 @@ if (isProvided(params$path_contrast_pairs)) { write.table(DE_genes_contrast_genename, file=paste("differential_gene_expression/allgenes/DE_contrast_",contname,".tsv",sep=""), sep="\t", quote=F, col.names = T, row.names = F) # Select only significantly DE - DE_genes_contrast <- subset(DE_genes_contrast_genename, padj <= params$adj_pval_threshold & abs(log2FoldChange) >= params$logFC_threshold) + DE_genes_contrast <- subset(DE_genes_contrast_genename, padj <= params$adj_pval_threshold & abs(log2FoldChange) >= params$logFC_threshold) DE_genes_contrast <- DE_genes_contrast[order(DE_genes_contrast$padj),] # Round table before writing @@ -1294,7 +1379,8 @@ if (isProvided(params$path_contrast_pairs)) { # Calculating DE genes for default contrasts (no contrast matrix or list or pairs provided) if (!isProvided(params$path_contrast_matrix) & !isProvided(params$path_contrast_list) & !isProvided(params$path_contrast_pairs)) { contrast_names <- coefficients[2:length(coefficients)] - contrast_names <- contrast_names[!startsWith(contrast_names, "batch_")] + contrast_names <- contrast_names[!startsWith(contrast_names, "batch")] + for (contname in contrast_names) { results_DEseq_contrast <- results(cds, name=contname) @@ -1313,7 +1399,7 @@ if (!isProvided(params$path_contrast_matrix) & !isProvided(params$path_contrast_ write.table(DE_genes_contrast_genename, file=paste("differential_gene_expression/allgenes/DE_contrast_",contname,".tsv",sep=""), sep="\t", quote=F, col.names = T, row.names = F) # Select only significantly DE - DE_genes_contrast <- subset(DE_genes_contrast_genename, padj <= params$adj_pval_threshold & abs(log2FoldChange) >= params$logFC_threshold) + DE_genes_contrast <- subset(DE_genes_contrast_genename, padj <= params$adj_pval_threshold & abs(log2FoldChange) >= params$logFC_threshold) DE_genes_contrast <- DE_genes_contrast[order(DE_genes_contrast$padj),] # Round table before writing if (isProvided(params$round_DE) && params$round_DE>-1) { @@ -1695,7 +1781,7 @@ for (file in contrast_files){ if (isProvided(params$set_background)) { gostres <- gost(query=q, organism=gost_id, - significant=TRUE, + significant=T, correction_method="fdr", sources=datasources, evcodes=TRUE, @@ -1711,7 +1797,7 @@ for (file in contrast_files){ # Also run without bg for Venn diagrams gostres_nobg <- gost(query=q, organism=gost_id, - significant=TRUE, + significant=T, correction_method="fdr", sources=datasources, evcodes=TRUE, @@ -1725,7 +1811,7 @@ for (file in contrast_files){ } else { gostres <- gost(query=q, organism=gost_id, - significant=TRUE, + significant=T, correction_method="fdr", sources=datasources, evcodes=TRUE, @@ -2109,6 +2195,7 @@ and the subfolder: `pathway_heatmaps`. # Summary and outlook ```{r quote, echo=FALSE, results='asis'} + # Only add hyperlink to quote file if it was provided in the params quote_string <- ifelse( isProvided(params$path_quote), @@ -2150,6 +2237,10 @@ if (software_versions_csv) { quant_tool <- "Salmon" quant_version <- as.character(version["SALMON_QUANT"][[1]]["salmon"]) quant_cite <- "[@patro2017salmon]" + } else if (params$input_type == "smrnaseq") { + quant_tool <- "SAMtools" + quant_version <- as.character(version["SAMTOOLS_IDXSTATS"][[1]]["samtools"]) + quant_cite <- "[@li2009sequence; @li2011statistical]" } } ``` @@ -2164,23 +2255,50 @@ cat("No software versions file was provided for primary analysis.") ```{r software_versions_RNASeq, echo=FALSE, results='asis', eval=isProvided(params$path_software_versions)} -# In the following part, I use newlines (in code) where possible whenever the paste0 function switches from a string to a variable call to better distinguish between the two so it is obvious in which lines quotes are needed and in which not --> Cannot always use newlines as this sometimes translates into the report.html - +# In the following part, I use newlines (in code) where possible whenever the paste0 function switches from a string to a variable call to better distinguish between the two so it is clearer in which lines quotes are needed and in which not --> Cannot always use newlines as this sometimes translates into the report.html +# TODO remove F! +if (params$input_type == "featurecounts" & F){ + cat(paste0( + "## RNAseq data analysis + + The Nextflow-based nf-core pipeline `rnaseq ", as.character(ifelse(software_versions_csv, version["nf-core/rnaseq"], version["Workflow"][[1]]["nf-core/rnaseq"])), + "` [^1] was used for the RNAseq Bioinfomatics analysis. `FASTQC ", + as.character(ifelse(software_versions_csv, version["FastQC"], version["FASTQC"][[1]]["fastqc"])), + "` [^3] [@andrews2010fastqc] was used to determine quality of the FASTQ files. + Subsequently, adapter trimming was conducted with `Trim Galore ", + as.character(ifelse(software_versions_csv, version["Trim Galore!"], version["TRIMGALORE"][[1]]["trimgalore"])), + "` [^4] [@krueger2012trim]. `STAR v", + as.character(ifelse(software_versions_csv, substring((version["STAR"]), 7, ), version["MAKE_TRANSCRIPTS_FASTA"][[1]]["star"])), + "` [@Dobin2013] aligner was used to map the reads that passed the quality control to the reference genome. + The RNA-seq data quality control was performed with `RSeQC ", + as.character(ifelse(software_versions_csv, version["RSeQC"], version["RSEQC_BAMSTAT"][[1]]["rseqc"])), + "` [@wang2012rseqc] and read quantification of the features (e.g. genes) with `", + quant_tool, + " ", + quant_version, + "` ", + quant_cite, + "." + )) +} else if (params$input_type == "smrnaseq"){ cat(paste0( -"## RNAseq data analysis - -The Nextflow-based nf-core pipeline `rnaseq ", as.character(ifelse(software_versions_csv, version["nf-core/rnaseq"], version["Workflow"][[1]]["nf-core/rnaseq"])), -"` [^1] was used for the RNAseq Bioinfomatics analysis. `FASTQC ", -as.character(ifelse(software_versions_csv, version["FastQC"], version["FASTQC"][[1]]["fastqc"])), -"` [^3] [@andrews2010fastqc] was used to determine quality of the FASTQ files. -Subsequently, adapter trimming was conducted with `Trim Galore ", -as.character(ifelse(software_versions_csv, version["Trim Galore!"], version["TRIMGALORE"][[1]]["trimgalore"])), -"` [^4] [@krueger2012trim]. `STAR v", -as.character(ifelse(software_versions_csv, substring((version["STAR"]), 7, ), version["MAKE_TRANSCRIPTS_FASTA"][[1]]["star"])), -"` [@Dobin2013] aligner was used to map the reads that passed the quality control to the reference genome. -The RNA-seq data quality control was performed with `RSeQC ", -as.character(ifelse(software_versions_csv, version["RSeQC"], version["RSEQC_BAMSTAT"][[1]]["rseqc"])), -"` [@wang2012rseqc] and read quantification of the features (e.g. genes) with `", +"## smRNA-seq data analysis + +The Nextflow-based nf-core pipeline `smrnaseq ", as.character(version["Workflow"][[1]]["nf-core/smrnaseq"]), +"` [^6] was used for the smRNA-seq Bioinfomatics analysis. `FASTQC ", +as.character(version["FASTQC"][[1]]["fastqc"]), +"` [^3] [@andrews2010fastqc] was used to determine the quality of the miRNA samples. ", +ifelse("TRIMGALORE" %in% names(version), paste0("Subsequently, adapter trimming was conducted with `Trim Galore ", +as.character(version["TRIMGALORE"][[1]]["trimgalore"]), +"` [^4] [@krueger2012trim]."), + ifelse("FASTP" %in% names(version), paste0("Subsequently, adapter trimming was conducted with `fastP ", + as.character(version["FASTP"][[1]]["fastp"]), + "` [^7] [@chen2018fastp; @chen2023ultrafast]."), + "")), +" `Bowtie v", as.character(version["BOWTIE_MAP_HAIRPIN"][[1]]["bowtie"]), "` [@langmead2009ultrafast] aligner was used to map the reads that passed the quality control to the reference database. +The smRNA-seq data quality control was performed with `miRTrace ", +as.character(version["MIRTRACE_RUN"][[1]]["mirtrace"]), +"` [^8] [@kang2018mirtrace] and read quantification of the features (e.g. genes) with `", quant_tool, " ", quant_version, @@ -2188,15 +2306,14 @@ quant_version, quant_cite, "." )) +} +``` -```{r MQC_cite, echo=FALSE, results='asis', eval=software_versions_csv&isProvided(params$path_software_versions)} - +```{r MQC_cite, echo=FALSE, results='asis', eval=dir.exists(paste0(wd,"/QC/multiqc_data/"))&software_versions_csv&isProvided(params$path_software_versions)} cat(paste0("\nAn aggregation of the quality control for the RNAseq analysis was performed with `MultiQC ", as.character(version["MultiQC"]), "` [^2] [@ewels2016multiqc].")) - ``` ```{r software_versions_DE, echo=FALSE, results='asis'} - cat(paste0( "## Differential gene expression analysis @@ -2263,3 +2380,6 @@ sessioninfo::session_info()%>% [^4]: https://www.bioinformatics.babraham.ac.uk/projects/trim_galore/ [^5]: https://github.com/qbic-pipelines/rnadeseq +[^6]: https://github.com/nf-core/smrnaseq +[^7]: https://github.com/OpenGene/fastp +[^8]: https://github.com/friedlanderlab/mirtrace diff --git a/assets/references.bibtex b/assets/references.bibtex index fdf26cc2..92bccdc4 100644 --- a/assets/references.bibtex +++ b/assets/references.bibtex @@ -1,95 +1,95 @@ @article{ewels2016multiqc, - title={MultiQC: summarize analysis results for multiple tools and samples in a single report}, - author={Ewels, Philip and Magnusson, M{\aa}ns and Lundin, Sverker and K{\"a}ller, Max}, - journal={Bioinformatics}, - volume={32}, - number={19}, - pages={3047--3048}, - year={2016}, - publisher={Oxford University Press} +title={MultiQC: summarize analysis results for multiple tools and samples in a single report}, +author={Ewels, Philip and Magnusson, M{\aa}ns and Lundin, Sverker and K{\"a}ller, Max}, +journal={Bioinformatics}, +volume={32}, +number={19}, +pages={3047--3048}, +year={2016}, +publisher={Oxford University Press} } @article{kim2015hisat, - title={HISAT: a fast spliced aligner with low memory requirements}, - author={Kim, Daehwan and Langmead, Ben and Salzberg, Steven L}, - journal={Nature methods}, - volume={12}, - number={4}, - pages={357}, - year={2015}, - publisher={Nature Publishing Group} +title={HISAT: a fast spliced aligner with low memory requirements}, +author={Kim, Daehwan and Langmead, Ben and Salzberg, Steven L}, +journal={Nature methods}, +volume={12}, +number={4}, +pages={357}, +year={2015}, +publisher={Nature Publishing Group} } @article{wang2012rseqc, - title={RSeQC: quality control of RNA-seq experiments}, - author={Wang, Liguo and Wang, Shengqin and Li, Wei}, - journal={Bioinformatics}, - volume={28}, - number={16}, - pages={2184--2185}, - year={2012}, - publisher={Oxford University Press} +title={RSeQC: quality control of RNA-seq experiments}, +author={Wang, Liguo and Wang, Shengqin and Li, Wei}, +journal={Bioinformatics}, +volume={28}, +number={16}, +pages={2184--2185}, +year={2012}, +publisher={Oxford University Press} } @article{liao2014featurecounts, - title={featureCounts: an efficient general purpose program for assigning sequence reads to genomic features}, - author={Liao, Yang and Smyth, Gordon K and Shi, Wei}, - journal={Bioinformatics}, - volume={30}, - number={7}, - pages={923--930}, - year={2014} +title={featureCounts: an efficient general purpose program for assigning sequence reads to genomic features}, +author={Liao, Yang and Smyth, Gordon K and Shi, Wei}, +journal={Bioinformatics}, +volume={30}, +number={7}, +pages={923--930}, +year={2014} } @article{love2014differential, - title={Differential analysis of count data--the DESeq2 package}, - author={Love, Michael and Anders, Simon and Huber, Wolfgang}, - journal={Genome Biol}, - volume={15}, - pages={550}, - year={2014} +title={Differential analysis of count data--the DESeq2 package}, +author={Love, Michael and Anders, Simon and Huber, Wolfgang}, +journal={Genome Biol}, +volume={15}, +pages={550}, +year={2014} } @article{Li2009, - author = {Li, Heng and Durbin, Richard}, - title = {Fast and Accurate Short Read Alignment with Burrows--Wheeler Transform}, - journal = {Bioinformatics}, - issue_date = {July 2009}, - volume = {25}, - number = {14}, - month = jul, - year = {2009}, - issn = {1367-4803}, - pages = {1754--1760}, - numpages = {7}, - url = {http://dx.doi.org/10.1093/bioinformatics/btp324}, - doi = {10.1093/bioinformatics/btp324}, - acmid = {1576130}, - publisher = {Oxford University Press}, - address = {Oxford, UK}, +author = {Li, Heng and Durbin, Richard}, +title = {Fast and Accurate Short Read Alignment with Burrows--Wheeler Transform}, +journal = {Bioinformatics}, +issue_date = {July 2009}, +volume = {25}, +number = {14}, +month = jul, +year = {2009}, +issn = {1367-4803}, +pages = {1754--1760}, +numpages = {7}, +url = {http://dx.doi.org/10.1093/bioinformatics/btp324}, +doi = {10.1093/bioinformatics/btp324}, +acmid = {1576130}, +publisher = {Oxford University Press}, +address = {Oxford, UK}, } @article{Lex2014, - 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title={FastQC: a quality control tool for high throughput sequence data}, - author={Andrews, Simon and others}, - year={2010} +title={FastQC: a quality control tool for high throughput sequence data}, +author={Andrews, Simon and others}, +year={2010} } @Book{ggplot, - author = {Hadley Wickham}, - title = {ggplot2: Elegant Graphics for Data Analysis}, - publisher = {Springer-Verlag New York}, - year = {2016}, - isbn = {978-3-319-24277-4}, - url = {http://ggplot2.org}, +author = {Hadley Wickham}, +title = {ggplot2: Elegant Graphics for Data Analysis}, +publisher = {Springer-Verlag New York}, +year = {2016}, +isbn = {978-3-319-24277-4}, +url = {http://ggplot2.org}, } @article{anders2012detecting, - title={Detecting differential usage of exons from RNA-seq data}, - author={Anders, Simon and Reyes, Alejandro and Huber, Wolfgang}, - journal={Genome research}, - volume={22}, - number={10}, - pages={2008--2017}, - year={2012}, - publisher={Cold Spring Harbor Lab} +title={Detecting differential usage of exons from RNA-seq data}, +author={Anders, Simon and Reyes, Alejandro and Huber, Wolfgang}, +journal={Genome research}, +volume={22}, +number={10}, +pages={2008--2017}, +year={2012}, +publisher={Cold Spring Harbor Lab} } @article{ewels2020nf, - title={The nf-core framework for community-curated bioinformatics pipelines}, - author={Ewels, Philip A and Peltzer, Alexander and Fillinger, Sven and Patel, Harshil and Alneberg, Johannes and Wilm, Andreas and Garcia, Maxime Ulysse and Di Tommaso, Paolo and Nahnsen, Sven}, - journal={Nature biotechnology}, - volume={38}, - number={3}, - pages={276--278}, - year={2020}, - publisher={Nature Publishing Group} +title={The nf-core framework for community-curated bioinformatics pipelines}, +author={Ewels, Philip A and Peltzer, Alexander and Fillinger, Sven and Patel, Harshil and Alneberg, Johannes and Wilm, Andreas and Garcia, Maxime Ulysse and Di Tommaso, Paolo and Nahnsen, Sven}, +journal={Nature biotechnology}, +volume={38}, +number={3}, +pages={276--278}, +year={2020}, +publisher={Nature Publishing Group} +} +@article{kang2018mirtrace, +title={miRTrace reveals the organismal origins of microRNA sequencing data}, +author={Kang, Wenjing and Eldfjell, Yrin and Fromm, Bastian and Estivill, Xavier and Biryukova, Inna and Friedl{\"a}nder, Marc R}, +journal={Genome biology}, +volume={19}, +number={1}, +pages={1--15}, +year={2018}, +publisher={BioMed Central} +} +@article{langmead2009ultrafast, +title={Ultrafast and memory-efficient alignment of short DNA sequences to the human genome}, +author={Langmead, Ben and Trapnell, Cole and Pop, Mihai and Salzberg, Steven L}, +journal={Genome biology}, +volume={10}, +number={3}, +pages={1--10}, +year={2009}, +publisher={BioMed Central} +} +@article{chen2018fastp, +title={fastp: an ultra-fast all-in-one FASTQ preprocessor}, +author={Chen, Shifu and Zhou, Yanqing and Chen, Yaru and Gu, Jia}, +journal={Bioinformatics}, +volume={34}, +number={17}, +pages={i884--i890}, +year={2018}, +publisher={Oxford University Press} +} +@article{chen2023ultrafast, +title={Ultrafast one-pass FASTQ data preprocessing, quality control, and deduplication using fastp}, +author={Chen, Shifu}, +journal={iMeta}, +pages={e107}, +year={2023}, +publisher={Wiley Online Library} +} +@article{li2009sequence, +title={The sequence alignment/map format and SAMtools}, +author={Li, Heng and Handsaker, Bob and Wysoker, Alec and Fennell, Tim and Ruan, Jue and Homer, Nils and Marth, Gabor and Abecasis, Goncalo and Durbin, Richard and 1000 Genome Project Data Processing Subgroup}, +journal={bioinformatics}, +volume={25}, +number={16}, +pages={2078--2079}, +year={2009}, +publisher={Oxford University Press} +} +@article{li2011statistical, +title={A statistical framework for SNP calling, mutation discovery, association mapping and population genetical parameter estimation from sequencing data}, +author={Li, Heng}, +journal={Bioinformatics}, +volume={27}, +number={21}, +pages={2987--2993}, +year={2011}, +publisher={Oxford University Press} } diff --git a/conf/test_smrnaseq.config b/conf/test_smrnaseq.config new file mode 100644 index 00000000..797fa332 --- /dev/null +++ b/conf/test_smrnaseq.config @@ -0,0 +1,32 @@ +/* + * ------------------------------------------------- + * Nextflow config file for running tests + * ------------------------------------------------- + * Defines bundled input files and everything required + * to run a fast and simple test. Use as follows: + * nextflow run qbic-pipelines/rnadeseq -profile test_smrnaseq + */ + +params { + config_profile_name = 'Test smrnaseq' + config_profile_description = 'Minimal test dataset to check pipeline function' + // Limit resources so that this can run on Travis + max_cpus = 2 + max_memory = 6.GB + max_time = 48.h + // Input data + input = 'testdata/smrnaseq/samplesheet.tsv' + gene_counts = 'testdata/smrnaseq/counts' + model = 'testdata/smrnaseq/design.txt' + logFC_threshold = 1 + project_summary = 'https://raw.githubusercontent.com/qbic-pipelines/rnadeseq/dev/testdata/summary.tsv' + software_versions = 'https://raw.githubusercontent.com/qbic-pipelines/rnadeseq/dev/testdata/software_versions_smrnaseq.yml' + genelist = 'testdata/smrnaseq/requested_genes.txt' + input_type = 'smrnaseq' + run_pathway_analysis = true + genome = 'GRCh37' + batch_effect = true + set_background = false // This is solely to get some enriched pathways so that the github tests also check the pathway analysis output + // species_library = "org.Hs.eg.db" + // organism = "hsapiens" +} diff --git a/docs/output.md b/docs/output.md index bdc278d0..3c30fd18 100644 --- a/docs/output.md +++ b/docs/output.md @@ -27,8 +27,9 @@ This directory contains the zipped results. When unzipping them, the following s - `linear_model.txt`: linear model used for DE analysis. - `gene_list.txt`: provided list of interesting genes (if provided). - `star_[rsem/salmon]_files.txt`: list of counts files that were provided as input (only if `--input_type` is `rsem` or `salmon`). + - `smrnaseq_files.txt`: list of counts files that were provided as input (only if `--input_type` is `smrnaseq`). - `gene_counts_tables/` - - `raw_gene_counts.txt`: raw gene counts table from the nf-core/rnaseq pipeline and used for the differential gene expression analysis (only if `--input_type` is `featurecounts`). + - `raw_gene_counts.txt`: raw gene counts table from the nf-core/rnaseq pipeline and used for the differential gene expression analysis (only if `--input_type` is `featurecounts` or `smrnaseq`; in the second case, will internally build a counts table from the input files, taking the third column of each .sorted.idxstats file (second column if counting rownames) and combining all values from both hairpin and non-hairpin mature files into one table). - `rlog_transformed_gene_counts.tsv`: normalized gene counts with the "regularized logarithm" approach. This normalization is used prior to PCA analysis and heatmap plotting of the gene counts (only when rlog transformation is used). - `vst_transformed_gene_counts.tsv`: normalized gene counts with the "variance stabilizing" transformation (only when vst transformation is used). - `sizeFactor_libraries.tsv`: size factors for each sample. @@ -47,7 +48,7 @@ Pathway analysis with [gProfileR](https://biit.cs.ut.ee/gprofiler/gost) R packag **Output directory: `results/pathway_analysis/`** -This directory contains the zipped pathway analysis results (`gProfileR.zip`). When unzipping them, a subfolder for each contrast used for the differential gene expression analysis is found. Inside each contrast folder, there is the following output: +If pathway analysis was run, this directory contains the zipped pathway analysis results (`gProfileR.zip`). When unzipping them, a subfolder for each contrast used for the differential gene expression analysis is found. Inside each contrast folder, there is the following output: - `*_gost_pathway_venn_diagram.pdf/png` - Venn diagrams showing the numbers of enriched pathways when using a background gene list vs when not using a bg list. @@ -66,8 +67,6 @@ QBiC report for RNAseq analysis. In this directory the zipped report is contained. This file is ready for upload to the project in the QBiC portal. Once unzipped, this directory contains: - - - `Report.html` - QBiC report describing the RNAseq results. - `DESeq2/` @@ -75,6 +74,6 @@ In this directory the zipped report is contained. This file is ready for upload - `QC/` - Quality control results. Contains: - `fastqc.zip`: zipped fastqc results for each sample. - - `multiqc_data`: data from the multiQC process of the RNAseq pipeline which is incorporated into the report. - - `multiqc_plots`: plots from the multiQC process of the RNAseq pipeline which are incorporated into the report. - - `qc_summary.tsv`: summary of the multiQC process in a table format. + - `multiqc_data`: If multiQC input was provided: data from the multiQC process of the RNAseq pipeline which is incorporated into the report. + - `multiqc_plots`: If multiQC input was provided: plots from the multiQC process of the RNAseq pipeline which are incorporated into the report. + - `qc_summary.tsv`: If multiQC input was provided: summary of the multiQC process in a table format. diff --git a/docs/usage.md b/docs/usage.md index 13587b1f..2f6719e4 100644 --- a/docs/usage.md +++ b/docs/usage.md @@ -82,7 +82,7 @@ NXF_OPTS='-Xms1g -Xmx4g' ## Pre-requisites -The `qbic-pipelines/rnadeseq` pipeline relies on the output from the `nf-core/rnaseq` pipeline. To be able to match the results of the `nf-core/rnaseq` pipeline with the metadata samplesheet containing the experimental design for the differential expression analysis, **the filenames of the fastq files used as input to the `qbic-pipelines/rnadeseq` pipeline, need to start with the corresponding QBiC codes!**. _E.g. QBICKXXXXX_original_file_name.fastq_. Once the filenames are corrected if necessary, you can run the `qbic-pipelines/rnadeseq` pipeline as usual. +The `qbic-pipelines/rnadeseq` pipeline relies on the output from the `nf-core/rnaseq` pipeline or the `nf-core/smrnaseq` pipeline. To be able to match the results of the initial pipeline with the metadata samplesheet containing the experimental design for the differential expression analysis, **the filenames of the fastq files used as input to the `qbic-pipelines/rnadeseq` pipeline need to start with the corresponding QBiC codes!**. _E.g. QBICKXXXXX_original_file_name.fastq_. Once the filenames are corrected if necessary, you can run the `qbic-pipelines/rnadeseq` pipeline as usual. ## Running the pipeline @@ -218,6 +218,36 @@ QDESQ083AC Sample3 K562 QDESQ084AK Sample4 K562 ``` +- OR a folder containing files with smrnaseq output (folder/sampleXXX_mature_hairpin.sorted.idxstats, folder/sampleXXX_mature_sorted.idxstats). + For smrnaseq, the --input_type parameter must be set to 'smrnaseq'. + For example: + +```bash +--gene_counts 'path/to/smrnaseq_folder' --metadata 'path/to/smrnaseq_metadata.tsv' + +tree testdata/smrnaseq/counts/ +testdata/smrnaseq/counts/ +├── Clone1_NN1_mature.sorted.idxstats +├── Clone1_NN1_mature_hairpin.sorted.idxstats +├── Clone1_NN3_mature.sorted.idxstats +├── Clone1_NN3_mature_hairpin.sorted.idxstats +├── Clone9_NN1_mature.sorted.idxstats +├── Clone9_NN1_mature_hairpin.sorted.idxstats +. +. +. + +head 'path/to/smrnaseq_metadata.tsv' +``` + +```tsv +QBiC Code Secondary Name Condition: cellline +QDESQ081AU Sample1 GM12878 +QDESQ082A4 Sample2 GM12878 +QDESQ083AC Sample3 K562 +QDESQ084AK Sample4 K562 +``` + ### `--input` Metadata table/samplesheet (TSV) is the "Sample_preparations_sheet.tsv" that can be directly downloaded from the qPortal --> Browser. Rows are samples and columns contain sample grouping. Important columns are: @@ -344,7 +374,7 @@ Set this flag to run pathway analysis, otherwise, this step will be skipped. ### `--input_type` -This tells the pipeline which type of input dataset is provided. Must be one of 'featurecounts', 'rsem', 'salmon', default: featurecounts. +This tells the pipeline which type of input dataset is provided. Must be one of 'featurecounts', 'rsem', 'salmon', 'smrnaseq', default: featurecounts. ### `--multiqc` @@ -393,7 +423,7 @@ Which genome to use for analysis, e.g. GRCh37; see `/conf/igenomes.config` for w ### `--gtf` -GTF file to be used for DESeq if input is rsem or salmon, not necessary for featurecounts. +GTF file to be used for DESeq if input is rsem or salmon, not necessary for featurecounts or smrnaseq. ### `--organism` diff --git a/nextflow.config b/nextflow.config index fd1595d6..866a45e4 100644 --- a/nextflow.config +++ b/nextflow.config @@ -198,6 +198,7 @@ profiles { test_contrast_list { includeConfig 'conf/test_contrast_list.config' } test_relevel { includeConfig 'conf/test_relevel.config' } test_use_vst { includeConfig 'conf/test_use_vst.config' } + test_smrnaseq { includeConfig 'conf/test_smrnaseq.config' } test_star_salmon { includeConfig 'conf/test_star_salmon.config' } test_star_rsem { includeConfig 'conf/test_star_rsem.config' } test_batcheffect { includeConfig 'conf/test_batcheffect.config' } diff --git a/nextflow_schema.json b/nextflow_schema.json index 3aec2e6a..e992d9cd 100644 --- a/nextflow_schema.json +++ b/nextflow_schema.json @@ -13,11 +13,11 @@ "input_type": { "type": "string", "default": "featurecounts", - "description": "Which type of input is provided, one of featurecounts, rsem, salmon" + "description": "Which type of input is provided, one of featurecounts, rsem, salmon, smrnaseq." }, "gene_counts": { "type": "string", - "description": "If input_type = featurecounts: Raw count table (TSV). Columns are samples and rows are genes. 1st column Ensembl_ID, 2nd column gene_name. If input_type = rsem or input_type = salmon: Path to rsem/salmon output folder." + "description": "If input_type = featurecounts: Raw count table (TSV). Columns are samples and rows are genes. 1st column Ensembl_ID, 2nd column gene_name. If input_type = rsem or input_type = salmon: Path to rsem/salmon output folder. If input_type = smrnaseq: Path to smrnaseq output folder containing hairpin and non-hairpin mature .sorted.idxstats files." }, "input": { "type": "string", diff --git a/testdata/smrnaseq/counts/Clone1_NN1_mature.sorted.idxstats b/testdata/smrnaseq/counts/Clone1_NN1_mature.sorted.idxstats new file mode 100644 index 00000000..8ccd1462 --- /dev/null +++ b/testdata/smrnaseq/counts/Clone1_NN1_mature.sorted.idxstats @@ -0,0 +1,2589 @@ +hsa-let-7a-5p 22 0 0 +hsa-let-7a-3p 21 0 0 +hsa-let-7a-2-3p 22 0 0 +hsa-let-7b-5p 22 0 0 +hsa-let-7b-3p 22 0 0 +hsa-let-7c-5p 22 0 0 +hsa-let-7c-3p 22 0 0 +hsa-let-7d-5p 22 0 0 +hsa-let-7d-3p 22 0 0 +hsa-let-7e-5p 22 0 0 +hsa-let-7e-3p 22 0 0 +hsa-let-7f-5p 22 0 0 +hsa-let-7f-1-3p 22 0 0 +hsa-let-7f-2-3p 22 0 0 +hsa-miR-15a-5p 22 0 0 +hsa-miR-15a-3p 22 0 0 +hsa-miR-16-5p 22 0 0 +hsa-miR-16-1-3p 22 0 0 +hsa-miR-17-5p 23 0 0 +hsa-miR-17-3p 22 0 0 +hsa-miR-18a-5p 23 0 0 +hsa-miR-18a-3p 23 0 0 +hsa-miR-19a-5p 22 0 0 +hsa-miR-19a-3p 23 0 0 +hsa-miR-19b-1-5p 23 0 0 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68 0 0 +hsa-mir-5701-3 82 0 0 +* 0 0 2651242 diff --git a/testdata/smrnaseq/design.txt b/testdata/smrnaseq/design.txt new file mode 100644 index 00000000..4983798f --- /dev/null +++ b/testdata/smrnaseq/design.txt @@ -0,0 +1 @@ +~ batch + condition_treatment diff --git a/testdata/smrnaseq/requested_genes.txt b/testdata/smrnaseq/requested_genes.txt new file mode 100644 index 00000000..22e59997 --- /dev/null +++ b/testdata/smrnaseq/requested_genes.txt @@ -0,0 +1,5 @@ +hsa-mir-100 +hsa-mir-125b-1 +hsa-mir-200a +hsa-mir-505 +hsa-let-7c diff --git a/testdata/smrnaseq/samplesheet.tsv b/testdata/smrnaseq/samplesheet.tsv new file mode 100644 index 00000000..ddd07d13 --- /dev/null +++ b/testdata/smrnaseq/samplesheet.tsv @@ -0,0 +1,9 @@ +QBiC Code Condition: treatment batch Secondary Name +Clone1_NN1 clone1 1 1 +Clone1_NN3 clone1 3 2 +Clone9_NN1 clone9 1 3 +Clone9_NN2 clone9 2 4 +Clone9_NN3 clone9 3 5 +Control_N1 control 1 6 +Control_N2 control 2 7 +Control_N3 control 3 8 diff --git a/testdata/software_versions_smrnaseq.yml b/testdata/software_versions_smrnaseq.yml new file mode 100644 index 00000000..2c314188 --- /dev/null +++ b/testdata/software_versions_smrnaseq.yml @@ -0,0 +1,64 @@ +BOWTIE_MAP_GENOME: + bowtie: 1.3.1 + samtools: '1.14' +BOWTIE_MAP_HAIRPIN: + bowtie: 1.3.1 + samtools: '1.14' +BOWTIE_MAP_MATURE: + bowtie: 1.3.1 + samtools: '1.14' +BOWTIE_MAP_SEQCLUSTER: + bowtie: 1.3.1 + samtools: '1.14' +CUSTOM_DUMPSOFTWAREVERSIONS: + python: 3.11.0 + yaml: '6.0' +FASTP: + fastp: 0.23.2 +FASTQC_RAW: + fastqc: 0.11.9 +FASTQC_TRIM: + fastqc: 0.11.9 +FORMAT_HAIRPIN: + fastx_toolkit: 0.0.14 +FORMAT_MATURE: + fastx_toolkit: 0.0.14 +INDEX_GENOME: + bowtie: 1.3.1 +INDEX_HAIRPIN: + bowtie: 1.3.1 +INDEX_MATURE: + bowtie: 1.3.1 +MIRDEEP2_MAPPER: + mapper: 2.0.1 +MIRDEEP2_PIGZ": + pigz: 2.3.4 +MIRDEEP2_RUN": + mirdeep2: 2.0.1 +MIRTOP_QUANT": + mirtop: 0.4.25 +MIRTRACE_RUN": + mirtrace: 1.0.1 +PARSE_HAIRPIN": + seqkit: 2.3.0 +PARSE_MATURE": + seqkit: 2.3.0 +SAMPLESHEET_CHECK: + python: 3.8.3 +SAMTOOLS_FLAGSTAT: + samtools: '1.17' +SAMTOOLS_IDXSTATS: + samtools: '1.17' +SAMTOOLS_INDEX: + samtools: '1.17' +SAMTOOLS_SORT: + samtools: '1.17' +SAMTOOLS_STATS: + samtools: '1.17' +SEQCLUSTER_SEQUENCES": + seqcluster: 1.2.9 +TABLE_MERGE: + r-base: 3.6.2 +Workflow: + Nextflow: 22.10.2 + nf-core/smrnaseq: 2.2.1 diff --git a/tests/test_smrnaseq.yml b/tests/test_smrnaseq.yml new file mode 100644 index 00000000..31beedd6 --- /dev/null +++ b/tests/test_smrnaseq.yml @@ -0,0 +1,89 @@ +- name: Run test profile with smrnaseq + command: nextflow run main.nf -profile test_smrnaseq,docker --citest true --round_DE 3 + tags: + - test_smrnaseq + files: + - path: results_test/differential_gene_expression/metadata/smrnaseq_files.txt + md5sum: ce3cc117fc953a8ff4d17d0118123faf + - path: results_test/differential_gene_expression/DE_genes_tables/DE_contrast_condition_treatment_clone9_vs_clone1.tsv + md5sum: 46ca6c2ec1046e088853c96b28cd30e3 + - path: 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md5sum: b5c4c68a304f08a54b2b9d8969334e8c + - path: results_test/pathway_analysis/DE_contrast_condition_treatment_clone9_vs_clone1/DE_contrast_condition_treatment_clone9_vs_clone1_pathway_enrichment_results.tsv + md5sum: ab088ff01a4e3991be8a424e676afcdf + - path: results_test/pathway_analysis/DE_contrast_condition_treatment_control_vs_clone1/DE_contrast_condition_treatment_control_vs_clone1_KEGG_pathway_enrichment_plot.pdf + - path: results_test/pathway_analysis/DE_contrast_condition_treatment_control_vs_clone1/DE_contrast_condition_treatment_control_vs_clone1_KEGG_pathway_enrichment_plot.png + - path: results_test/pathway_analysis/DE_contrast_condition_treatment_control_vs_clone1/DE_contrast_condition_treatment_control_vs_clone1_KEGG_pathway_enrichment_plot.svg + - path: results_test/pathway_analysis/DE_contrast_condition_treatment_control_vs_clone1/DE_contrast_condition_treatment_control_vs_clone1_KEGG_pathway_enrichment_results.tsv + md5sum: 4a2ed390ae3e9fea628bd9c014e88cc6 + - path: results_test/pathway_analysis/DE_contrast_condition_treatment_control_vs_clone1/DE_contrast_condition_treatment_control_vs_clone1_pathway_enrichment_results.tsv + md5sum: db2149ba5f2dbb6d4aed3f6284072f39 + - path: results_test/RNAseq_report.html diff --git a/tests/test_star_salmon.yml b/tests/test_star_salmon.yml index 20b498cc..dbe49c9a 100644 --- a/tests/test_star_salmon.yml +++ b/tests/test_star_salmon.yml @@ -19,15 +19,15 @@ - path: results_test/differential_gene_expression/plots/boxplots_requested_genes/IL10_ENSG00000136634.pdf - path: results_test/differential_gene_expression/plots/boxplots_requested_genes/IL10_ENSG00000136634.png - path: results_test/differential_gene_expression/plots/boxplots_requested_genes/IL10_ENSG00000136634.svg - - path: results_test/differential_gene_expression/plots/further_diagnostics_plots//all_results_MA_plot_condition_cellline_H1_vs_GM12878.pdf - - path: results_test/differential_gene_expression/plots/further_diagnostics_plots//all_results_MA_plot_condition_cellline_H1_vs_GM12878.png - - path: results_test/differential_gene_expression/plots/further_diagnostics_plots//all_results_MA_plot_condition_cellline_H1_vs_GM12878.svg - - path: results_test/differential_gene_expression/plots/further_diagnostics_plots//all_results_MA_plot_condition_cellline_K562_vs_GM12878.pdf - - path: results_test/differential_gene_expression/plots/further_diagnostics_plots//all_results_MA_plot_condition_cellline_K562_vs_GM12878.png - - path: results_test/differential_gene_expression/plots/further_diagnostics_plots//all_results_MA_plot_condition_cellline_K562_vs_GM12878.svg - - path: results_test/differential_gene_expression/plots/further_diagnostics_plots//all_results_MA_plot_condition_cellline_MCF7_vs_GM12878.pdf - - path: results_test/differential_gene_expression/plots/further_diagnostics_plots//all_results_MA_plot_condition_cellline_MCF7_vs_GM12878.png - - path: results_test/differential_gene_expression/plots/further_diagnostics_plots//all_results_MA_plot_condition_cellline_MCF7_vs_GM12878.svg + - path: results_test/differential_gene_expression/plots/further_diagnostics_plots/all_results_MA_plot_condition_cellline_H1_vs_GM12878.pdf + - path: results_test/differential_gene_expression/plots/further_diagnostics_plots/all_results_MA_plot_condition_cellline_H1_vs_GM12878.png + - path: results_test/differential_gene_expression/plots/further_diagnostics_plots/all_results_MA_plot_condition_cellline_H1_vs_GM12878.svg + - path: results_test/differential_gene_expression/plots/further_diagnostics_plots/all_results_MA_plot_condition_cellline_K562_vs_GM12878.pdf + - path: results_test/differential_gene_expression/plots/further_diagnostics_plots/all_results_MA_plot_condition_cellline_K562_vs_GM12878.png + - path: results_test/differential_gene_expression/plots/further_diagnostics_plots/all_results_MA_plot_condition_cellline_K562_vs_GM12878.svg + - path: results_test/differential_gene_expression/plots/further_diagnostics_plots/all_results_MA_plot_condition_cellline_MCF7_vs_GM12878.pdf + - path: results_test/differential_gene_expression/plots/further_diagnostics_plots/all_results_MA_plot_condition_cellline_MCF7_vs_GM12878.png + - path: results_test/differential_gene_expression/plots/further_diagnostics_plots/all_results_MA_plot_condition_cellline_MCF7_vs_GM12878.svg - path: results_test/differential_gene_expression/plots/further_diagnostics_plots/histogram_of_p.values_condition_cellline_H1_vs_GM12878.pdf - path: results_test/differential_gene_expression/plots/further_diagnostics_plots/histogram_of_p.values_condition_cellline_H1_vs_GM12878.png - path: results_test/differential_gene_expression/plots/further_diagnostics_plots/histogram_of_p.values_condition_cellline_H1_vs_GM12878.svg diff --git a/workflows/rnadeseq.nf b/workflows/rnadeseq.nf index f9b73a9c..ffbbde2c 100644 --- a/workflows/rnadeseq.nf +++ b/workflows/rnadeseq.nf @@ -15,7 +15,7 @@ def checkPathParamList = [ for (param in checkPathParamList) { if (param) { file(param, checkIfExists: true) } } // Check mandatory parameters -if (!(params.input_type in ["featurecounts", "salmon", "rsem"])) { exit 1, 'Wrong input type ' + params.input_type + ', must be one of "featurecounts", "salmon", "rsem"!' } +if (!(params.input_type in ["featurecounts", "salmon", "rsem", "smrnaseq"])) { exit 1, 'Wrong input type ' + params.input_type + ', must be one of "featurecounts", "salmon", "rsem", "smrnaseq"!' } ch_counts_path = Channel.fromPath(params.gene_counts) ch_metadata_file = Channel.fromPath(params.input)