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nextNEOpi.nf
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#!/usr/bin/env nextflow
// enable DSL 1
nextflow.enable.dsl = 1
import org.yaml.snakeyaml.Yaml
import java.nio.file.Files
log.info ""
log.info " NEXTFLOW ~ version ${workflow.nextflow.version} ${workflow.nextflow.build}"
log.info "-------------------------------------------------------------------------"
log.info " Nextfow NeoAntigen Prediction Pipeline - nextNEOpi "
log.info "-------------------------------------------------------------------------"
log.info ""
log.info " Features: "
log.info " - somatic variants from tumor + matched normal samples"
log.info " - CNV analysis"
log.info " - tumor muational burden"
log.info " - class I and class II HLA typing"
log.info " - gene fusion peptide prediction using RNAseq data"
log.info " - peptide MHC binding perdiction"
log.info " - clonality of neoantigens"
log.info " - expression of neoantigens"
log.info ""
log.info "-------------------------------------------------------------------------"
log.info "C O N F I G U R A T I O N"
log.info ""
log.info "Command Line: \t\t " + workflow.commandLine
log.info "Working Directory: \t " + workflow.workDir
log.info "Output Directory: \t " + params.outputDir
log.info ""
log.info "I N P U T"
log.info ""
log.info "batch file: \t\t " + params.batchFile
log.info ""
log.info "Please check --help for further instruction"
log.info "-------------------------------------------------------------------------"
// Check if License(s) were accepted
params.accept_license = false
if (params.accept_license) {
acceptLicense()
} else {
checkLicense()
}
/*
________________________________________________________________________________
C O N F I G U R A T I O N
________________________________________________________________________________
*/
if (params.help) exit 0, helpMessage()
// switch for enable/disable processes (debug/devel only: use if(params.RUNTHIS) { .... })
params.RUNTHIS = false
// default is not to get bams as input data
bamInput = false
// initialize RNA tag seq
have_RNA_tag_seq = params.RNA_tag_seq
// set and initialize the Exome capture kit
setExomeCaptureKit(params.exomeCaptureKit)
// check conda channels
if (params.enable_conda) {
checkCondaChannels()
}
// check IEDB dir
check_iedb_dir(params.databases.IEDB_dir)
// check MHCflurry dir
check_mhcflurry_dir(params.databases.MHCFLURRY_dir)
// set and check references and databases
reference = defineResources('references', params.WES, params.HLAHD_DIR)
database = defineResources('databases', params.WES, params.HLAHD_DIR)
// create tmp dir and make sure we have the realpath for it
tmpDir = mkTmpDir(params.tmpDir)
/*--------------------------------------------------
For workflow summary
---------------------------------------------------*/
// Has the run name been specified by the user?
// this has the bonus effect of catching both -name and --name
custom_runName = params.name
if ( !(workflow.runName ==~ /[a-z]+_[a-z]+/) ) {
custom_runName = workflow.runName
}
// Summary
def summary = [:]
summary['Pipeline Name'] = 'icbi/nextNEOpi'
summary['Pipeline Version'] = workflow.manifest.version
summary['Batch file'] = params.batchFile
summary['Read length'] = params.readLength
summary['Exome capture kit'] = params.exomeCaptureKit
summary['Fasta Ref'] = params.references.RefFasta
summary['MillsGold'] = params.databases.MillsGold
summary['hcSNPS1000G'] = params.databases.hcSNPS1000G
summary['HapMap'] = params.databases.HapMap
summary['Cosmic'] = params.databases.Cosmic
summary['DBSNP'] = params.databases.DBSNP
summary['GnomAD'] = params.databases.GnomAD
summary['GnomADfull'] = params.databases.GnomADfull
summary['KnownIndels'] = params.databases.KnownIndels
summary['BlastDB'] = params.references.ProteinBlastDBdir
summary['priority variant Caller'] = params.primaryCaller
summary['Mutect 1 and 2 minAD'] = params.minAD
summary['VarScan min_cov'] = params.min_cov
summary['VarScan min_cov_tumor'] = params.min_cov_tumor
summary['VarScan min_cov_normal'] = params.min_cov_normal
summary['VarScan min_freq_for_hom'] = params.min_freq_for_hom
summary['VarScan somatic_pvalue'] = params.somatic_pvalue
summary['VarScan somatic_somaticpvalue'] = params.somatic_somaticpvalue
summary['VarScan strand_filter'] = params.strand_filter
summary['VarScan processSomatic_pvalue'] = params.processSomatic_pvalue
summary['VarScan max_normal_freq'] = params.max_normal_freq
summary['VarScan min_tumor_freq'] = params.min_tumor_freq
summary['VarScan min_map_q'] = params.min_map_q
summary['VarScan min_base_q'] = params.min_base_q
summary['VEP assembly'] = params.vep_assembly
summary['VEP species'] = params.vep_species
summary['VEP options'] = params.vep_options
summary['Number of scatters'] = params.scatter_count
summary['Output dir'] = params.outputDir
summary['Working dir'] = workflow.workDir
summary['TMP dir'] = tmpDir
summary['Current home'] = "$HOME"
summary['Current user'] = "$USER"
summary['Current path'] = "$PWD"
summary['Picard maxRecordsInRam'] = params.maxRecordsInRam
summary['Script dir'] = workflow.projectDir
summary['Config Profile'] = workflow.profile
if(params.email) summary['E-mail Address'] = params.email
log.info summary.collect { k,v -> "${k.padRight(30)}: $v" }.join("\n")
log.info "-------------------------------------------------------------------------"
// End Summary
// determine the publishDirMode
def publishDirMode = get_publishMode(params.outputDir, params.publishDirMode)
// Check if we got a batch file
params.batchFile = null
if (params.batchFile == null) {
log.error "No sample sheet specified, please use --batchFile to pass the sample sheet"
exit(1)
}
def batchCSV = file(params.batchFile).splitCsv(header:true)
def validFQfields = ["sampleName",
"reads1",
"reads2",
"sampleType",
"HLAfile",
"sex"]
def validBAMfields = ["sampleName",
"bam",
"sampleType",
"HLAfile",
"sex"]
def validSampleTypes = ["tumor_DNA", "normal_DNA", "tumor_RNA"]
if (batchCSV.size() > 0) {
if (batchCSV[0].keySet().sort() == validFQfields.sort()) {
bamInput = false
} else if (batchCSV[0].keySet().sort() == validBAMfields.sort()) {
bamInput = true
} else {
exit 1, "Error: Incorrect fields in batch file, please check your batchFile"
}
} else {
exit 1, "Error: No samples found, please check your batchFile"
}
def raw_data = []
def custom_HLA_data = []
def t_map = [:]
def n_map = [:]
def r_map = [:]
def lt_map = [:]
for ( row in batchCSV ) {
def meta = [:]
def reads = []
meta.sampleName = row.sampleName.toString()
meta.sampleType = row.sampleType.toString()
t_map[meta.sampleName] = (t_map[meta.sampleName] == null) ? 0 : t_map[meta.sampleName]
n_map[meta.sampleName] = (n_map[meta.sampleName] == null) ? 0 : n_map[meta.sampleName]
if(row.sex) {
if (row.sex.toLowerCase() in ["xx", "female"]) {
meta.sex = "XX"
} else if (row.sex.toLowerCase() in ["xy", "male"]) {
meta.sex = "XY"
} else if (row.sex.toLowerCase() in ["none", "na"]) {
meta.sex = "None"
println("WARNING: " + row.sampleName + " sex not specified will infer from data")
} else {
exit 1, "sex should be one of: XX, xx, XY, xy, Female, female, Male, male, None, none, NA, got: " + row.sex
}
meta.maleRef = (meta.sex == "XY") ? true : false
} else {
println("WARNING: " + row.sampleName + " sex not specified will infer from data")
meta.sex = "None"
meta.maleRef = true
}
if (row.sampleType in validSampleTypes) {
t_map[meta.sampleName] += (row.sampleType == "tumor_DNA") ? 1 : 0
n_map[meta.sampleName] += (row.sampleType == "normal_DNA") ? 1 : 0
} else {
exit 1, "Error: Incorrect sampleType [got: " + row.sampleType + " - allowed: " + validSampleTypes + "]: please check your batchFile"
}
// remove stuff that is not required in the HLAfile channel
def meta_min = meta.clone()
meta_min.keySet().removeAll(['sampleType', 'libType'])
if (row.HLAfile) {
custom_HLA_data.add([meta_min, file(row.HLAfile, checkIfExists: true)])
} else {
custom_HLA_data.add([meta_min, []])
}
if (row.sampleType == "tumor_RNA") {
meta.have_RNA = true
r_map[row.sampleName] = true
}
if (! row.bam) {
meta.libType = "SE"
if (row.reads1) { reads.add(file(row.reads1, checkIfExists: true)) }
if (row.reads2) {
reads.add(file(row.reads2, checkIfExists: true))
meta.libType = "PE"
}
if (meta.sampleType != "tumor_RNA") {
if (lt_map[meta.sampleName] == null) {
lt_map[meta.sampleName] = meta.libType
} else {
if (lt_map[meta.sampleName] != meta.libType) {
exit 1, "Please do not mix pe and se for tumor/normal pairs: " + meta.sampleName + " - Not supported"
}
}
}
raw_data.add([meta, reads])
} else {
raw_data.add([meta, file(row.bam, checkIfExists: true)])
}
}
// check if we have T/N DNA pairs for all patients
raw_data.each {
record ->
if (t_map[record[0].sampleName] == 0) {
exit 1, "NO tumor DNA sample specified for: " + record[0].sampleName
}
if (n_map[record[0].sampleName] == 0) {
exit 1, "NO normal DNA sample specified for: " + record[0].sampleName
}
}
// update meta for RNA
raw_data.each {
record ->
record[0].have_RNA = (r_map[record[0].sampleName]) ? true : false
}
custom_HLA_data = custom_HLA_data.each {
record ->
record[0].have_RNA = (r_map[record[0].sampleName]) ? true : false
}.unique()
use_custom_hlas = (custom_HLA_data.size() > 0) ? true : false
batch_raw_data_ch = Channel.fromList(raw_data)
batch_custom_HLA_data_ch = Channel.fromList(custom_HLA_data)
if (bamInput == false) {
batch_raw_data_ch.map {
meta, fastq ->
[ meta, fastq ]
}
.groupTuple(by: [0])
.branch {
meta, fastq ->
single : fastq.unique().size() == 1
return [ meta, fastq.flatten().unique() ]
multiple: fastq.unique().size() > 1
return [ meta, fastq.flatten().unique() ]
}
.set { fastq_ch }
} else {
fastq_ch = Channel.of().branch{single: []; multiple: []}
}
// optional panel of normals file
pon_file = file(params.mutect2ponFile)
scatter_count = Channel.from(params.scatter_count)
padding = params.readLength + 100
MiXMHC2PRED = ( params.MiXMHC2PRED != "" ) ? file(params.MiXMHC2PRED) : ""
// check HLAHD & OptiType
have_HLAHD = false
run_OptiType = (params.disable_OptiType) ? false : true
if (params.HLAHD_DIR != "") {
HLAHD = file(params.HLAHD_DIR + "/bin/hlahd.sh")
if (checkToolAvailable(HLAHD, "exists", "warn")) {
HLAHD_DIR = file(params.HLAHD_DIR)
HLAHD_PATH = HLAHD_DIR + "/bin"
if(params.HLAHD_module != "") {
if (checkToolAvailable("bowtie2", "inPath", "warn", module=params.HLAHD_module)) {
have_HLAHD = true
}
} else {
if (checkToolAvailable("bowtie2", "inPath", "warn")) {
have_HLAHD = true
}
}
}
}
if (! have_HLAHD && run_OptiType) {
log.warn "WARNING: HLAHD not available - can not predict Class II neoepitopes"
} else if (! have_HLAHD && ! run_OptiType && use_custom_hlas) {
log.warn "WARNING: HLAHD not available and OptiType disabled - using only user supplied HLA types"
} else if (! have_HLAHD && ! run_OptiType && ! use_custom_hlas) {
exit 1, "ERROR: HLAHD not available and OptiType disabled - can not predict HLA types"
}
// check if all tools are installed when not running conda or singularity
have_vep = false
if (! workflow.profile.contains('conda') && ! workflow.profile.contains('singularity')) {
def execTools = ["fastqc", "fastp", "bwa", "samtools", "sambamba", "gatk", "vep", "bam-readcount",
"perl", "bgzip", "tabix", "bcftools", "yara_mapper", "python", "cnvkit.py",
"OptiTypePipeline.py", "alleleCounter", "freec", "Rscript", "java", "multiqc",
"sequenza-utils"]
for (tool in execTools) {
checkToolAvailable(tool, "inPath", "error")
}
VARSCAN = "java -jar " + file(params.VARSCAN)
have_vep = true
} else {
VARSCAN = "varscan "
}
// check if we have mutect1 installed
have_Mutect1 = false
if (params.MUTECT1 != "" && file(params.MUTECT1) && params.JAVA7 != "" && file(params.JAVA7)) {
if(checkToolAvailable(params.JAVA7, "exists", "warn") && checkToolAvailable(params.MUTECT1, "exists", "warn")) {
JAVA7 = file(params.JAVA7)
MUTECT1 = file(params.MUTECT1)
have_Mutect1 = true
}
}
// check if we have GATK3 installed
have_GATK3 = false
if (params.GATK3 != "" && file(params.GATK3) && params.JAVA8 != "" && file(params.JAVA8) && ! workflow.profile.contains('conda') && ! workflow.profile.contains('singularity')) {
if(checkToolAvailable(params.JAVA8, "inPath", "warn") && checkToolAvailable(params.GATK3, "exists", "warn")) {
JAVA8 = file(params.JAVA8)
GATK3 = file(params.GATK3)
have_GATK3 = true
}
} else if (workflow.profile.contains('singularity')) {
JAVA8 = "java"
GATK3 = "/usr/local/opt/gatk-3.8/GenomeAnalysisTK.jar"
have_GATK3 = true
} else if (workflow.profile.contains('conda')) {
JAVA8 = "java"
GATK3 = "\$CONDA_PREFIX/opt/gatk-3.8/GenomeAnalysisTK.jar"
have_GATK3 = true
}
// check MIXCR licence
if (params.TCR && params.MIXCR_lic != "") {
checkToolAvailable(params.MIXCR_lic, "exists", "error")
} else if (params.TCR && params.MIXCR_lic == "") {
exit 1, "ERROR: no MiXCR license file specified, please provide a MiXCR license file in params.config or by using the --MIXCR_lic option.\nIf you do not have a MiXCR license you may:\n\ta) run nextNEOpi with --TCR false\n\tb) request one at https://licensing.milaboratories.com"
}
/*
________________________________________________________________________________
P R O C E S S E S
________________________________________________________________________________
*/
gatk4_chck_file = file(baseDir + "/assets/.gatk4_install_ok.chck")
gatk4_chck_file.append()
nextNEOpiENV_setup_ch0 = Channel.value("OK")
/*
*********************************************
** P R E P R O C E S S I N G **
*********************************************
*/
// Handle BAM input files. We need to convert BAMs to Fastq
if(bamInput) {
process check_PE {
label 'nextNEOpiENV'
tag "$meta.sampleName : $meta.sampleType"
input:
tuple(
val(meta),
path(bam)
) from batch_raw_data_ch
val (nextNEOpiENV_setup) from nextNEOpiENV_setup_ch0
output:
tuple(
val(meta),
path(bam),
stdout
) into bam_ch
script:
"""
check_pe.py $bam
"""
}
(bam_ch, check_seqLib_ch) = bam_ch.into(2)
seqLibTypes_ok = Channel.value(check_seqLibTypes_ok(check_seqLib_ch))
process bam2fastq {
label 'nextNEOpiENV'
tag "$meta.sampleName : $meta.sampleType"
publishDir "${params.outputDir}/analyses/${meta.sampleName}/01_preprocessing",
mode: publishDirMode
input:
tuple(
val(meta),
path(bam),
val(libType),
val(libOK)
) from bam_ch
.combine(seqLibTypes_ok)
val (nextNEOpiENV_setup) from nextNEOpiENV_setup_ch0
output:
tuple(
val(meta),
path("${prefix}_R*.fastq.gz"),
) into (
bam_fastq_ch
)
script:
def STperThreadMem = (int) Math.max(((int) Math.floor((task.memory.toGiga() - 4) / task.cpus)), 1)
prefix = meta.sampleName + "_" + meta.sampleType
meta.libType = libType
if (libType == "PE")
"""
samtools sort -@ ${task.cpus} -m ${STperThreadMem}G -l 0 -n ${bam} | \\
samtools fastq \\
-@ ${task.cpus} \\
-c 5 \\
-1 ${prefix}_R1.fastq.gz \\
-2 ${prefix}_R2.fastq.gz \\
-0 /dev/null -s /dev/null \\
-n \\
/dev/stdin
"""
else if (libType == "SE")
"""
samtools fastq \\
-@ ${task.cpus} \\
-n \\
${bam} | \\
bgzip -@ ${task.cpus} -c /dev/stdin > ${prefix}_R1.fastq.gz
"""
}
} else {
bam_fastq_ch = channel.empty()
}
// END BAM input handling
// merge multi run or multi lane sample fastq
process merge_fastq {
tag "$meta.sampleName : $meta.sampleType"
publishDir "${params.outputDir}/${meta.sampleName}/01_preprocessing/",
mode: publishDirMode,
enabled: params.fullOutput
input:
tuple meta, path("R1/R1.*"), path("R2/R2.*") from fastq_ch.multiple.map{
meta, r -> {
def r1 = []
def r2 = []
r.eachWithIndex{ v, ix -> ( ix & 1 ? r2 : r1 ) << v }
tuple( meta, r1.sort(), r2.sort())
}
}
output:
tuple val(meta), path("*_R{1,2}.merged.fastq.gz") into merged_fastq_ch
script:
def prefix = meta.sampleName + "_" + meta.sampleType
"""
cat R1/* > ${prefix}_R1.merged.fastq.gz
cat R2/* > ${prefix}_R2.merged.fastq.gz
"""
}
// here we have our final raw fastq files:
// merged if multi lane runs are used
// bam2fastq if bam files are used
(raw_reads_ch, fastqc_reads_ch) = merged_fastq_ch.mix(fastq_ch.single, bam_fastq_ch).into(2)
// Common region files preparation for faster processing
if (params.WES) {
process 'RegionsBedToIntervalList' {
label 'nextNEOpiENV'
tag 'RegionsBedToIntervalList'
publishDir "$params.outputDir/supplemental/00_prepare_Intervals/",
mode: publishDirMode
input:
tuple(
path(RefDict),
path(RegionsBed)
) from Channel.value(
[ reference.RefDict,
reference.RegionsBed ]
)
val (nextNEOpiENV_setup) from nextNEOpiENV_setup_ch0
output:
path(
"${RegionsBed.baseName}.interval_list"
) into (
RegionsBedToIntervalList_out_ch0,
RegionsBedToIntervalList_out_ch1
)
script:
def JAVA_Xmx = '-Xmx' + task.memory.toGiga() + "G"
"""
gatk --java-options ${JAVA_Xmx} BedToIntervalList \\
-I ${RegionsBed} \\
-O ${RegionsBed.baseName}.interval_list \\
-SD $RefDict
"""
}
process 'BaitsBedToIntervalList' {
label 'nextNEOpiENV'
tag 'BaitsBedToIntervalList'
publishDir "$params.outputDir/supplemental/00_prepare_Intervals/",
mode: publishDirMode
input:
tuple(
path(RefDict),
path(BaitsBed)
) from Channel.value(
[ reference.RefDict,
reference.BaitsBed ]
)
val (nextNEOpiENV_setup) from nextNEOpiENV_setup_ch0
output:
path(
"${BaitsBed.baseName}.interval_list"
) into BaitsBedToIntervalList_out_ch0
script:
def JAVA_Xmx = '-Xmx' + task.memory.toGiga() + "G"
"""
gatk --java-options ${JAVA_Xmx} BedToIntervalList \\
-I ${BaitsBed} \\
-O ${BaitsBed.baseName}.interval_list \\
-SD $RefDict
"""
}
} else {
RegionsBedToIntervalList_out_ch0 = Channel.fromPath('NO_FILE')
RegionsBedToIntervalList_out_ch1 = Channel.empty()
BaitsBedToIntervalList_out_ch0 = Channel.empty()
}
process 'preprocessIntervalList' {
label 'nextNEOpiENV'
tag 'preprocessIntervalList'
publishDir "$params.outputDir/supplemental/00_prepare_Intervals/",
mode: publishDirMode
input:
tuple(
path(RefFasta),
path(RefIdx),
path(RefDict)
) from Channel.value(
[ reference.RefFasta,
reference.RefIdx,
reference.RefDict ]
)
path(interval_list) from RegionsBedToIntervalList_out_ch0
val (nextNEOpiENV_setup) from nextNEOpiENV_setup_ch0
output:
path(
outFileName
) into (
preprocessIntervalList_out_ch0,
preprocessIntervalList_out_ch1,
preprocessIntervalList_out_ch2,
preprocessIntervalList_out_ch3,
preprocessIntervalList_out_ch4,
preprocessIntervalList_out_ch5,
preprocessIntervalList_out_ch6,
preprocessIntervalList_out_ch7,
preprocessIntervalList_out_ch8
)
script:
outFileName = (params.WES) ? interval_list.baseName + "_merged_padded.interval_list" : "wgs_ScatterIntervalsByNs.interval_list"
def JAVA_Xmx = '-Xmx' + task.memory.toGiga() + "G"
if(params.WES)
"""
gatk PreprocessIntervals \\
-R $RefFasta \\
-L ${interval_list} \\
--bin-length 0 \\
--padding ${padding} \\
--interval-merging-rule OVERLAPPING_ONLY \\
-O ${outFileName}
"""
else
"""
gatk --java-options ${JAVA_Xmx} ScatterIntervalsByNs \\
--REFERENCE $RefFasta \\
--OUTPUT_TYPE ACGT \\
--OUTPUT ${outFileName}
"""
}
// Splitting interval file in 20(default) files for scattering Mutect2
process 'SplitIntervals' {
label 'nextNEOpiENV'
tag "SplitIntervals"
publishDir "$params.outputDir/supplemental/00_prepare_Intervals/SplitIntervals/",
mode: publishDirMode,
enabled: params.fullOutput
input:
tuple(
path(RefFasta),
path(RefIdx),
path(RefDict)
) from Channel.value(
[ reference.RefFasta,
reference.RefIdx,
reference.RefDict ]
)
path(IntervalsList) from preprocessIntervalList_out_ch0
val x from scatter_count
val (nextNEOpiENV_setup) from nextNEOpiENV_setup_ch0
output:
path(
"${IntervalName}/*-scattered.interval_list"
) into (
SplitIntervals_out_ch0,
SplitIntervals_out_ch1,
SplitIntervals_out_ch2,
SplitIntervals_out_scatterBaseRecalTumorGATK4_ch,
SplitIntervals_out_scatterTumorGATK4applyBQSRS_ch,
SplitIntervals_out_scatterBaseRecalNormalGATK4_ch,
SplitIntervals_out_scatterNormalGATK4applyBQSRS_ch,
SplitIntervals_out_ch3,
SplitIntervals_out_ch4,
SplitIntervals_out_ch5,
SplitIntervals_out_ch6,
)
val("${IntervalName}") into SplitIntervals_out_ch0_Name
script:
IntervalName = IntervalsList.baseName
"""
mkdir -p ${tmpDir}
gatk SplitIntervals \\
--tmp-dir ${tmpDir} \\
-R ${RefFasta} \\
-scatter ${x} \\
--interval-merging-rule ALL \\
--subdivision-mode BALANCING_WITHOUT_INTERVAL_SUBDIVISION_WITH_OVERFLOW \\
-L ${IntervalsList} \\
-O ${IntervalName}
"""
}
// convert padded interval list to Bed file (used by varscan)
// generate a padded tabix indexed region BED file for strelka
process 'IntervalListToBed' {
label 'nextNEOpiENV'
tag 'BedFromIntervalList'
publishDir "$params.outputDir/supplemental/00_prepare_Intervals/",
mode: publishDirMode
input:
path(paddedIntervalList) from preprocessIntervalList_out_ch1
val (nextNEOpiENV_setup) from nextNEOpiENV_setup_ch0
output:
path("${paddedIntervalList.baseName}.{bed.gz,bed.gz.tbi}") into (
RegionsBedToTabix_out_ch0,
RegionsBedToTabix_out_ch1
)
script:
def JAVA_Xmx = '-Xmx' + task.memory.toGiga() + "G"
"""
gatk --java-options ${JAVA_Xmx} IntervalListToBed \\
-I ${paddedIntervalList} \\
-O ${paddedIntervalList.baseName}.bed
bgzip -c ${paddedIntervalList.baseName}.bed > ${paddedIntervalList.baseName}.bed.gz &&
tabix -p bed ${paddedIntervalList.baseName}.bed.gz
"""
}
// convert scattered padded interval list to Bed file (used by varscan)
process 'ScatteredIntervalListToBed' {
label 'nextNEOpiENV'
tag 'ScatteredIntervalListToBed'
publishDir "$params.outputDir/supplemental/00_prepare_Intervals/SplitIntervals/${IntervalName}",
mode: publishDirMode,
enabled: params.fullOutput
input:
tuple(
val(IntervalName),
file(IntervalsList)
) from SplitIntervals_out_ch0_Name
.combine(
SplitIntervals_out_ch0.flatten()
)
val (nextNEOpiENV_setup) from nextNEOpiENV_setup_ch0
output:
path(
"*.bed"
) into (
ScatteredIntervalListToBed_out_ch0,
ScatteredIntervalListToBed_out_ch1
)
script:
def JAVA_Xmx = '-Xmx' + task.memory.toGiga() + "G"
"""
gatk --java-options ${JAVA_Xmx} IntervalListToBed \\
-I ${IntervalsList} \\
-O ${IntervalsList.baseName}.bed
"""
}
// FastQC
process FastQC {
label 'fastqc'
tag "$meta.sampleName : $meta.sampleType"
publishDir "${params.outputDir}/analyses/${meta.sampleName}/QC/fastqc",
mode: publishDirMode,
saveAs: { filename -> filename.indexOf(".zip") > 0 ? "zips/$filename" : "$filename"}
input:
tuple val(meta), path(reads) from fastqc_reads_ch
output:
tuple val(meta), path("*_fastqc.zip") into ch_fastqc // multiQC
script:
def reads_R1_ext = (reads[0].getExtension() == "gz") ? "fastq.gz" : reads[0].getExtension()
def reads_R1 = meta.sampleName + "_" + meta.sampleType + "_R1." + reads_R1_ext
// do we have PE reads?
def reads_R2 = "_missing_"
if(meta.libType == "PE") {
def reads_R2_ext = (reads[1].getExtension() == "gz") ? "fastq.gz" : reads[1].getExtension()
reads_R2 = meta.sampleName + "_" + meta.sampleType + "_R2." + reads_R2_ext
}
"""
if [ ! -e ${reads_R1} ]; then
ln -s ${reads[0]} ${reads_R1}
fi
if [ "${reads_R2}" != "_missing_" ] && [ ! -e ${reads_R2} ]; then
ln -s ${reads[1]} ${reads_R2}
fi
fastqc --quiet --threads ${task.cpus} \\
${reads_R1} ${reads_R2}
"""
}
// adapter trimming
//
// We first check if we need to trim DNA and RNA or only one of them.
// If it is only one of them we need to combine the trimmed and raw
// reads again
def trim_adapters = false
if (params.trim_adapters || params.trim_adapters_RNAseq) {
trim_adapters = true
reads_to_trim = raw_reads_ch
if (params.trim_adapters && params.trim_adapters_RNAseq) {
reads_to_trim_ch = raw_reads_ch
reads_to_keep_ch = Channel.empty()
}
if (params.trim_adapters && ! params.trim_adapters_RNAseq) {
raw_reads_ch.branch {
DNA: it[0].sampleType != "tumor_RNA"
RNA: it[0].sampleType == "tumor_RNA"
}
.set{ raw_reads_ch }
reads_to_trim_ch = raw_reads_ch.DNA
reads_to_keep_ch = raw_reads_ch.RNA
}
if (! params.trim_adapters && params.trim_adapters_RNAseq) {
raw_reads_ch.branch {
DNA: it[0].sampleType != "tumor_RNA"
RNA: it[0].sampleType == "tumor_RNA"
}
.set{ raw_reads_ch }
reads_to_trim_ch = raw_reads_ch.RNA
reads_to_keep_ch = raw_reads_ch.DNA
}
}
if (trim_adapters) {
process fastp {
label 'fastp'
tag "$meta.sampleName : $meta.sampleType"
publishDir "$params.outputDir/analyses/${meta.sampleName}/",
mode: publishDirMode,
saveAs: {
filename ->
if(filename.indexOf(".json") > 0) {
return "QC/fastp/$filename"
} else if(filename.indexOf("NO_FILE") >= 0) {
return null
} else {
return "01_preprocessing/$filename"
}
}
input:
tuple val(meta), path(reads) from reads_to_trim_ch
output:
tuple val(meta), path("*_trimmed_R{1,2}.fastq.gz") into reads_trimmed_ch, fastqc_reads_trimmed_ch
tuple val(meta), path("*.json") into ch_fastp // multiQC
script:
def reads_R1 = "--in1 " + reads[0]
def trimmed_reads_R1 = "--out1 " + meta.sampleName + "_" + meta.sampleType + "_trimmed_R1.fastq.gz"
// do we have PE reads?
def reads_R2 = ""
def trimmed_reads_R2 = ""
if(meta.libType == "PE") {
reads_R2 = "--in2 " + reads[1]
trimmed_reads_R2 = "--out2 " + meta.sampleName + "_" + meta.sampleType + "_trimmed_R2.fastq.gz"
}
def fastpAdapter = ''
def adapterSeqFile
def aseq = false
def aseqR2 = false
def afile = false
if (meta.sampleType.indexOf("DNA") > 0) {
afile = params.adapterSeqFile
aseq = params.adapterSeq
aseqR2 = params.adapterSeqR2
} else {
afile = params.adapterSeqFileRNAseq
aseq = params.adapterSeqRNAseq
aseqR2 = params.adapterSeqR2RNAseq
}
if(afile != false) {
adapterSeqFile = Channel.fromPath(afile)
fastpAdapter = "--adapter_fasta " + adapterSeqFile
} else {
if(aseq != false) {
adapterSeq = Channel.value(aseq)
fastpAdapter = "--adapter_sequence " + aseq.getVal()
if(aseqR2 != false && meta.libType == "PE") {
adapterSeqR2 = Channel.value(aseqR2)
fastpAdapter += " --adapter_sequence_r2 " + adapterSeqR2.getVal()
}
}
}
"""
fastp --thread ${task.cpus} \\
${reads_R1} \\
${reads_R2} \\
${trimmed_reads_R1} \\
${trimmed_reads_R2} \\
--json ${meta.sampleName}_${meta.sampleType}_fastp.json \\
${fastpAdapter} \\
${params.fastpOpts}
"""
}
// FastQC after adapter trimming
process FastQC_trimmed {
label 'fastqc'
tag "$meta.sampleName : $meta.sampleType"
publishDir "${params.outputDir}/analyses/${meta.sampleName}/QC/fastqc",
mode: publishDirMode,
saveAs: { filename -> filename.indexOf(".zip") > 0 ? "zips/$filename" : "$filename"}