diff --git a/src/.docker_modules/star/2.7.8a/Dockerfile b/src/.docker_modules/star/2.7.8a/Dockerfile
new file mode 100644
index 0000000000000000000000000000000000000000..32d712d67e2905fc28b9c7114d5acfcc3560b60f
--- /dev/null
+++ b/src/.docker_modules/star/2.7.8a/Dockerfile
@@ -0,0 +1,2 @@
+FROM quay.io/biocontainers/star:2.7.8a--0
+MAINTAINER Xavier Grand
diff --git a/src/.docker_modules/star/2.7.8a/docker_init.sh b/src/.docker_modules/star/2.7.8a/docker_init.sh
new file mode 100755
index 0000000000000000000000000000000000000000..f39c19203c5cbba0f64b91c6cab13f4936814a5d
--- /dev/null
+++ b/src/.docker_modules/star/2.7.8a/docker_init.sh
@@ -0,0 +1,5 @@
+#!/bin/sh
+docker pull xgrand/star:2.7.8a
+# docker build src/.docker_modules/star/2.7.8a/ -t 'xgrand/star:2.7.8a'
+# docker push xgrand/star:2.7.8a
+docker buildx build --platform linux/amd64,linux/arm64 -t "xgrand/star:2.7.8a" --push src/.docker_modules/star/2.7.8a
\ No newline at end of file
diff --git a/src/RNAseq_XGR.nf b/src/RNAseq_XGR.nf
index a5bc73ee3644efe3acbb4ea32732ae7257b9b3c0..84e426aa452751a4b71eb432fab46ae301f745af 100644
--- a/src/RNAseq_XGR.nf
+++ b/src/RNAseq_XGR.nf
@@ -22,7 +22,7 @@ def helpMessage() {
Usage:
The typical command for running the pipeline is as follows:
- nextflow ./src/star_fusion.nf -c ./src/nextflow.config -profile singularity
+ nextflow ./src/RNAseq_XGR.nf -c ./src/nextflow.config -profile singularity
Mandatory arguments:
--project [path] Path to the project folder. Results are saved in this folder.
diff --git a/src/arriba_fusion.nf b/src/arriba_fusion.nf
index c46b936689428c78e7abe878d294fb44a5ba9133..7c1b29aa2e0a0b53d8ab9e5738a2e7abb37894d6 100644
--- a/src/arriba_fusion.nf
+++ b/src/arriba_fusion.nf
@@ -30,10 +30,10 @@ def helpMessage() {
Available: docker, singularity, podman, psmn, ccin2p3
Input:
- --fastq [path] Path to fastq folder.
- --bam [path] Path to the bam-containing folder.
+ --fastq [path] Path to fastq files.
+ --bam [path] Path to the bam files.
- References:
+ References: Can be downloaded with download_references.sh (not implemented in pipeline).
--genome [path] Path to genome reference fasta file.
--gtf [path] Path to genome annotation gtf file.
@@ -61,10 +61,11 @@ if (params.help || params.h) {
*/
params.project = ""
+params.bam_folder = ""
+params.genome = ""
+params.gtf = ""
params.bam = ""
params.fastq = ""
-if (params.genome) { params.genome = path(params.genome, checkIfExists: true) } else { exit 1, "No genome specified." }
-if (params.gtf) { params.gtf = path(params.gtf, checkIfExists: true) } else { exit 1, "No annotation specified." }
/* Params out */
params.fastp_out = "$params.project/fastp/"
@@ -80,6 +81,12 @@ params.index_bam_out = "$params.project/Bam_filt_sort_indexed/"
log.info "Reference genome : ${params.genome}"
log.info "Genome annotation : ${params.gtf}"
+if(params.bam_folder != "") {
+ log.info "bam files (--bam): ${bam}"
+}
+else {
+ log.info "fastq files (--fastq): ${params.fastq}"
+}
/*
****************************************************************
@@ -87,23 +94,28 @@ log.info "Genome annotation : ${params.gtf}"
****************************************************************
*/
-if(params.bam != "") {
+if(params.bam_folder != "") {
Channel
.fromPath( params.bam )
+ .ifEmpty { error "Cannot find any bam files in: ${params.bam}" }
+ .map { it -> [it.simpleName, it]}
.set { bam_files }
}
else {
Channel
- .fromFilePairs( params.fastq, size = -1 )
- .set(fastq_files)
+ .fromFilePairs( params.fastq, size: -1)
+ .set { fastq_files }
}
Channel
.fromPath( params.genome )
+ .ifEmpty { error "Cannot find any fasta files in: ${params.genome}" }
+ .map { it -> [it.simpleName, it]}
.set { genome }
Channel
.fromPath( params.gtf )
+ .ifEmpty { error "Cannot find any annotation files in: ${params.gtf}" }
.set { gtf }
/*
@@ -113,9 +125,11 @@ Channel
*/
include { fastp } from './nf_modules/fastp/main.nf'
-include { fastqc_fastq as fastqc_raw } from fastqc_mod addParams(fastqc_fastq_out: "$params.project/01_fastqc_raw/")
-include { fastqc_fastq as fastqc_preprocessed } from fastqc_mod addParams(fastqc_fastq_out: "$params.project/02_fastqc_preprocessed/")
+include { fastqc_fastq as fastqc_raw } from './nf_modules/fastqc/main.nf' addParams(fastqc_fastq_out: "$params.project/01_fastqc_raw/")
+include { fastqc_fastq as fastqc_preprocessed } from './nf_modules/fastqc/main.nf' addParams(fastqc_fastq_out: "$params.project/02_fastqc_preprocessed/")
include { multiqc } from './nf_modules/multiqc/main.nf' addParams(multiqc_out: "$params.project/QC/")
+include { index_with_gtf } from './nf_modules/star/main_2.7.8a.nf' addParams(star_mapping_fastq_out: "$params.project/STAR_index/")
+include { mapping_fastq_withChimeric } from './nf_modules/star/main_2.7.8a.nf' addParams(star_mapping_fastq_out: "$params.project/STAR/")
include { arriba } from "./nf_modules/arriba/main.nf"
/*
@@ -127,26 +141,20 @@ include { arriba } from "./nf_modules/arriba/main.nf"
workflow {
if(params.bam == ""){
- fastp()
- fastqc_raw()
- fastqc_preprocessed()
- multiqc()
- .mix(
- fastqc_preprocessed.out.report
- ).collect()
- index_fasta()
- mapping_fastq()
- filter_bam_quality()
- sort_bam()
- index_bam()
+ fastp(fastq_files)
+ // fastqc_raw(fastq_files.collect())
+ // fastqc_preprocessed(fastp_out.fastq.collect())
+ // multiqc(fastqc_raw_out.report)
+ // .mix(
+ // fastqc_preprocessed.out.report
+ // ).collect()
+ index_with_gtf(genome, gtf)
+ // mapping_fastq_withChimeric(index_fasta_out.index, fastp_out.fastq)
+ // filter_bam_quality(mapping_fastq_withChimeric_out.bam)
+ // arriba()
+ }
+ else {
+ arriba(bam_files, gtf, genome)
}
-
-
-
- //###################### ARRIBA FUSION ########################
-
- arriba(fastq_files, gtf, genome)
-
- //################ GRAPHICAL REPRESENTATIONS ##################
}
\ No newline at end of file
diff --git a/src/nextflow.config b/src/nextflow.config
index d954a5318b14d575366ce5a3e87963138f0549b8..5758b7d165b14e2970d85c7bf53b33a80d86d865 100644
--- a/src/nextflow.config
+++ b/src/nextflow.config
@@ -18,7 +18,7 @@ profiles {
docker.enabled = true
process {
errorStrategy = 'finish'
- memory = '16GB'
+ memory = '15GB'
withLabel: big_mem_mono_cpus {
cpus = 1
}
@@ -47,7 +47,7 @@ profiles {
podman.enabled = true
process {
errorStrategy = 'finish'
- memory = '16GB'
+ memory = '15GB'
withLabel: big_mem_mono_cpus {
cpus = 1
}
@@ -77,7 +77,7 @@ profiles {
singularity.cacheDir = "./bin/"
process {
errorStrategy = 'finish'
- memory = '16GB'
+ memory = '15GB'
withLabel: big_mem_mono_cpus {
cpus = 1
}
diff --git a/src/nf_modules/star/main_2.7.8a.nf b/src/nf_modules/star/main_2.7.8a.nf
new file mode 100644
index 0000000000000000000000000000000000000000..a711426d86db32b46866e40b75daaadf6ef3049e
--- /dev/null
+++ b/src/nf_modules/star/main_2.7.8a.nf
@@ -0,0 +1,183 @@
+version = "2.7.8a"
+container_url = "xgrand/star:${version}"
+
+params.star_mapping_fastq_out = ""
+
+
+process gff3_2_gtf {
+ container = "dceoy/cufflinks"
+ label "small_mem_mono_cpus"
+
+ input:
+ tuple val(genome_id), path(gff3_file)
+ output:
+ path "${genome_id}.gtf", emit: gtf
+ script:
+"""
+gffread ${gff3_file} -T -o ${genome_id}.gtf
+"""
+}
+
+
+process index_with_gtf {
+ container = "${container_url}"
+ label "big_mem_multi_cpus"
+
+ input:
+ tuple val(genome_id), path(genome_fasta)
+ path gtf_file
+
+ output:
+ tuple val(genome_id), path ("*"), emit: index
+
+ script:
+"""
+STAR --runThreadN ${task.cpus} --runMode genomeGenerate \
+--genomeDir ./ \
+--genomeFastaFiles ${genome_fasta} \
+--sjdbGTFfile ${gtf_file} \
+--genomeSAindexNbases 13 # min(14, log2(GenomeLength)/2 - 1)
+"""
+}
+
+workflow index_with_gff {
+ take:
+ genome_fasta
+ gff_file
+ main:
+ gff3_2_gtf(gff_file)
+ index_with_gtf(genome_fasta,gff3_2_gtf.out.gtf)
+ emit:
+ report = index_with_gtf.out.index
+}
+
+
+process index_without_gff {
+ container = "${container_url}"
+ label "big_mem_multi_cpus"
+
+ input:
+ tuple val(genome_id), path(genome_fasta)
+
+ output:
+ tuple val(genome_id), path ("*"), emit: index
+
+ script:
+"""
+STAR --runThreadN ${task.cpus} --runMode genomeGenerate \
+--genomeDir ./ \
+--genomeFastaFiles ${genome_fasta} \
+--genomeSAindexNbases 13 # min(14, log2(GenomeLength)/2 - 1)
+"""
+}
+
+
+process mapping_fastq {
+ container = "${container_url}"
+ label "big_mem_multi_cpus"
+ if (params.star_mapping_fastq_out != "") {
+ publishDir "results/${params.star_mapping_fastq_out}", mode: 'copy'
+ }
+
+ input:
+ tuple val(index_id), path(index)
+ tuple val(reads_id), path(reads)
+
+ output:
+ path "*.Log.final.out", emit: report
+ tuple val(reads_id), path("*.bam"), emit: bam
+
+ script:
+if (reads_id instanceof List){
+ file_prefix = reads_id[0]
+ } else {
+ file_prefix = reads_id
+ }
+
+if (reads.size() == 2)
+"""
+mkdir -p index
+mv ${index} index/
+STAR --runThreadN ${task.cpus} \
+--genomeDir index/ \
+--readFilesCommand zcat \
+--readFilesIn ${reads[0]} ${reads[1]} \
+--outFileNamePrefix ${reads_id}. \
+--alignIntronMax 10000 \
+--outSAMtype BAM SortedByCoordinate \
+--outSAMstrandField intronMotif
+
+mv ${reads_id}.Aligned.sortedByCoord.out.bam ${reads_id}.bam
+"""
+else
+"""
+mkdir -p index
+mv ${index} index/
+STAR --runThreadN ${task.cpus} \
+--genomeDir index/ \
+--readFilesCommand zcat \
+--readFilesIn ${reads} \
+--outFileNamePrefix ${reads_id}. \
+--alignIntronMax 10000 \
+--outSAMtype BAM SortedByCoordinate \
+--outSAMstrandField intronMotif
+
+mv ${reads_id}.Aligned.sortedByCoord.out.bam ${reads_id}.bam
+"""
+}
+
+process mapping_fastq_withChimeric {
+ container = "${container_url}"
+ label "big_mem_multi_cpus"
+ if (params.star_mapping_fastq_out != "") {
+ publishDir "results/${params.star_mapping_fastq_out}", mode: 'copy'
+ }
+
+ input:
+ tuple val(index_id), path(index)
+ tuple val(reads_id), path(reads)
+
+ output:
+ path "*.Log.final.out", emit: report
+ tuple val(reads_id), path("*.bam"), emit: bam
+
+ script:
+if (reads_id instanceof List){
+ file_prefix = reads_id[0]
+ } else {
+ file_prefix = reads_id
+ }
+
+if (reads.size() == 2)
+"""
+mkdir -p index
+mv ${index} index/
+STAR --runThreadN ${task.cpus} \
+--genomeDir index/ \
+--readFilesCommand zcat \
+--readFilesIn ${reads[0]} ${reads[1]} \
+--outFileNamePrefix ${reads_id}. \
+--alignIntronMax 10000 \
+--outSAMtype BAM SortedByCoordinate \
+--outSAMstrandField intronMotif \
+--chimOutType WithinBAM
+
+mv ${reads_id}.Aligned.sortedByCoord.out.bam ${reads_id}.bam
+"""
+else
+"""
+mkdir -p index
+mv ${index} index/
+STAR --runThreadN ${task.cpus} \
+--genomeDir index/ \
+--readFilesCommand zcat \
+--readFilesIn ${reads} \
+--outFileNamePrefix ${reads_id}. \
+--alignIntronMax 10000 \
+--outSAMtype BAM SortedByCoordinate \
+--outSAMstrandField intronMotif \
+--chimOutType WithinBAM
+
+mv ${reads_id}.Aligned.sortedByCoord.out.bam ${reads_id}.bam
+"""
+}
\ No newline at end of file