I'm trying to learn nextflow but it's not going very well. I used NGS-based double-end sequencing data to build an analysis flow from fastq files to vcf files using Nextflow. However I got stuck right at the beginning, as shown in the code. The first process and the second porcess sworks fine, but when passing the files to the third process there is an ERROR and I can't execute the whole process anymore. What should I do? Thanks for a help.
Following is my code:
#! /usr/bin/env nextflow
params.fq1 = "/home/duxu/project/data/*1.fq.gz"
params.fq2 = "/home/duxu/project/data/*2.fq.gz"
params.index = "/home/duxu/project/result/index.list"
params.primer1 = "/home/duxu/project/data/primer_1.fasta"
params.primer2 = "/home/duxu/project/data/primer_2.fasta"
params.ref = "/home/duxu/project/data/hg19.p13.plusMT.no_alt_analysis_set/"
params.output='results'
params.refhg19 = "/home/duxu/project/data/hg19.p13.plusMT.no_alt_analysis_set/hg19.p13.plusMT.no_alt_analysis_set.fa"
params.Mills = "/home/duxu/project/data/Mills_and_1000G_gold_standard.indels.hg19.sites.vcf"
params.1000G = "/home/duxu/project/data/1000G_phase1.indels.hg19.sites.vcf"
params.dbsnp = "/home/duxu/project/data/dbsnp_138.hg19.vcf
fq2 = Channel.fromPath(params.fq2)
fq2 = Channel.fromPath(params.fq2)
index = Channel.fromPath(params.index)
index.into { index_1; index_2; index_3 }
primer1 = Channel.fromPath(params.primer1)
primer2 = Channel.fromPath(params.primer2)
ref = Channel.fromPath(params.ref)
refhg19 = Channel.fromPath(params.refhg19)
refhg19.into { refhg19_1; refhg19_2 ; refhg19_3; refhg19_4; refhg19_5}
Mills = Channel.fromPath(params.Mills)
1000G = Channel.fromPath(params.1000G)
dbsnp = Channel.fromPath(params.dbsnp)
This is first process:
process soapnuke{
conda'soapnuke'
tag{"soapnuk ${fq1} ${fq2}"}
publishDir "${params.outdir}/SOAPnuke", mode: 'copy'
input:
file rawfq1 from fq1
file rawfq2 from fq2
output:
file 'clean1.fastq.gz' into clean_fq1
file 'clean2.fastq.gz' into clean_fq2
script:
"""
SOAPnuke filter -1 $rawfq1 -2 $rawfq2 -l 12 -q 0.5 -Q 2 -o . \
-C clean1.fastq.gz -D clean2.fastq.gz --trim 8,0,8,0
"""
}
The second process
process barcode_splitter{
tag{"barcode_splitter"}
publishDir "${params.output}/barcode_splitter", mode: 'copy'
input:
file split1 from clean_fq1
file split2 from clean_fq2
file index from index_1
output:
file '*-read-1.fastq.gz' into trimmed_index1
file '*-read-2.fastq.gz' into trimmed_index2
script:
"""
barcode_splitter --bcfile $index $split1 $split2 --idxread 1 2 --mismatches 1 --suffix .fastq --gzipout
mv multimatched-read-1.fastq.gz multicatched.1.fastq.gz
mv multimatched-read-2.fastq.gz multicatched.2.fastq.gz
mv untimatched-read-1.fastq.gz multicatched.1.fastq.gz
mv untimatched-read-2.fastq.gz multicatched.2.fastq.gz
"""
}
The third ,and I got an error from this step. In fact, this process has multiple samples, since the previous process baicode_splitter output multiple files. This process cutadapt is designed to excise the first few bases of multiple samples.
process cutadapt{
tag{"cutadapt"}
publishDir "${params.output}/cut_primer", mode: 'copy'
input:
val sample from sample
file primer_1 from primer1
file primer_2 from primer2
file ${sample}-read-1.fastq.gz from trimmed_index1.collect()
file ${sample}-read-2.fastq.gz from trimmed_index2.collect()
output:
file '*.trim.1.fastq.gz' into trimmed_primer1
file '*.trim.2.fastq.gz' into trimmed_primer2
script:
"""
cutadapt -g file:$primer_1 -G file:$primer_2 -j 64 --discard-untrimmed -o \${sample}.trim.1.fastq.gz -p \$(sample}.trim.2.fastq.gz ${sample}-read-1.fastq.gz ${sample}-read-2.fastq.gz
"""
}
The fourth process is designed to match multiple samples to a reference genome
process bwa_mapping{
tag{bwa_maping}
publishDir "${params.output}/barcode_splitter", mode: 'copy'
input:
file pair1 from trimmed_primer1
file pair2 from trimmed_primer2
file refhg19 from refhg19_1
output:
file '*_R1_R2.bam' into addheader
script:
"""
bwa mem -t 64 $refhg19 trimmed_primer_*-read-1.fastq trimmed_primer_*-read-2.fastq | samtools view -@8 -b | samtools sort -m 2G -@64 > * _R1_R2.bam
"""
}
The next remaining processes are all multisample-based operations
process {BaseRecalibrator
tag{"BaseRecalibrator"}
publishDir "${params.output}/BQSR/BaseRecalibratoraddheader", mode: 'copy'
input:
file BaseRecalibrator from BaseRecalibrator_1
file refhg19 from refhg19_2
file Mills from Mills
file 1000G from 1000G
file dbsnp from dbsnp
output:
file '*.recal.table' into ApplyBQSR
script:
"""
gatk --java-options "-XX:ParallelGCThreads=2" BaseRecalibrator -I BaseRecalibrator -R $refhg19_2 --known-sites $Mills --known-sites $1000G --known-sites $dbsnp -O *.recal.table
"""
}
process {ApplyBQSR
tag{"ApplyRecalibrator"}
publishDir "${params.output}/BQSR/ApplyRecalibrator", mode: 'copy'
input:
file ApplyBQSR from ApplyBQSR
file refhg19 from refhg19_3
file BaseRecalibrator from BaseRecalibrator_2
output:
file '*.bam' into HaplotypeCaller
script:
"""
gatk --java-options "-XX:ParallelGCThreads=2" ApplyBQSR -I BaseRecalibrator -R $refhg19_3 --bqsr-recal-file $AppleBQSR -O *.bam
"""
}
process {HaplotypeCaller
tag{"HaplotypeCallerr"}
publishDir "${params.output}/GATK/HaplotypeCaller", mode: 'copy'
input:
file HaplotypeCaller from HaplotypeCaller
file refhg19 from refhg19_4
file BaseRecalibrator from BaseRecalibrator_3
output:
file '*.g.vcf.gz' into GenotypeGVCFs
script:
"""
gatk --java-options "-XX:ParallelGCThreads=2" HaplotypeCaller -R $refhg19_4 -I BaseRecalibrator -O *.g.vcf.gz -ERC GVCF
"""
}
process {GenotypeGVCFs
tag{"GenotypeGVCFs"}
publishDir "${params.output}/GATK/GenotypeGVCFs", mode: 'copy'
input:
file GenotypeGVCFs from GenotypeGVCFs
file refhg19 from refhg19_5
output:
file '*.vcf.gz' into end
script:
"""
gatk --java-options "-XX:ParallelGCThreads=2" GenotypeGVCFs -R $refhg19_5 -V $GenotypeGVCFs -O *.vcf.gz
"""
