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LBMC
RMI2
RMI2 pipelines
Commits
c2bf2271
Commit
c2bf2271
authored
5 years ago
by
elabaron
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add RNAseq pipeline
parent
4d9aa8e9
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src/RNAseq.config
+112
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112 additions, 0 deletions
src/RNAseq.config
src/RNAseq.nf
+217
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217 additions, 0 deletions
src/RNAseq.nf
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329 additions
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0 deletions
src/RNAseq.config
0 → 100644
+
112
−
0
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c2bf2271
profiles
{
sge
{
process
{
withName
:
trimming
{
beforeScript
=
"source $baseDir/.conda_psmn.sh"
conda
=
"$baseDir/.conda_envs/cutadapt_2.1"
executor
=
"sge"
clusterOptions
=
"-cwd -V"
cpus
=
1
memory
=
"20GB"
time
=
"12h"
queue
=
'monointeldeb128,monointeldeb48,h48-E5-2670deb128,h6-E5-2667v4deb128'
}
withName
:
rRNA_removal
{
beforeScript
=
"source $baseDir/.conda_psmn.sh"
conda
=
"$baseDir/.conda_envs/bowtie2_2.3.4.1"
executor
=
"sge"
clusterOptions
=
"-cwd -V"
cpus
=
16
memory
=
"30GB"
time
=
"24h"
queue
=
'E5-2670deb128A,E5-2670deb128B,E5-2670deb128C,E5-2670deb128D,E5-2670deb128E,E5-2670deb128F'
penv
=
'openmp16'
}
withName
:
hisat2_human
{
beforeScript
=
"source /usr/share/lmod/lmod/init/bash; module use ~/privatemodules"
module
=
"hisat2/2.1.0:samtools/1.7"
executor
=
"sge"
clusterOptions
=
"-cwd -V"
memory
=
"20GB"
cpus
=
16
time
=
"12h"
queue
=
'E5-2670deb128A,E5-2670deb128B,E5-2670deb128C,E5-2670deb128D,E5-2670deb128E,E5-2670deb128F'
penv
=
'openmp16'
}
withName
:
sort_bam
{
beforeScript
=
"source $baseDir/.conda_psmn.sh"
conda
=
"$baseDir/.conda_envs/samtools_1.7"
executor
=
"sge"
clusterOptions
=
"-cwd -V"
cpus
=
1
memory
=
"20GB"
time
=
"12h"
queue
=
'monointeldeb128,monointeldeb48,h48-E5-2670deb128,h6-E5-2667v4deb128'
}
withName
:
index_bam
{
beforeScript
=
"source $baseDir/.conda_psmn.sh"
conda
=
"$baseDir/.conda_envs/samtools_1.7"
executor
=
"sge"
clusterOptions
=
"-cwd -V"
cpus
=
1
memory
=
"20GB"
time
=
"12h"
queue
=
'monointeldeb128,monointeldeb48,h48-E5-2670deb128,h6-E5-2667v4deb128'
}
withName
:
dedup
{
beforeScript
=
"source /usr/share/lmod/lmod/init/bash; module use ~/privatemodules"
module
=
"umi_tools_1.0.0"
executor
=
"sge"
clusterOptions
=
"-cwd -V"
cpus
=
1
memory
=
"20GB"
time
=
"12h"
queue
=
'monointeldeb128,monointeldeb48,h48-E5-2670deb128,h6-E5-2667v4deb128'
}
withName
:
counting
{
beforeScript
=
"source /usr/share/lmod/lmod/init/bash; module use ~/privatemodules"
module
=
"htseq/0.11.2"
executor
=
"sge"
clusterOptions
=
"-cwd -V"
cpus
=
1
memory
=
"20GB"
time
=
"12h"
queue
=
'monointeldeb128,monointeldeb48,h48-E5-2670deb128,h6-E5-2667v4deb128'
}
}
}
docker
{
docker
.
temp
=
'auto'
docker
.
enabled
=
true
process
{
withName
:
adaptor_removal
{
container
=
"lbmc/cutadapt:2.1"
cpus
=
1
}
withName
:
rRNA_removal
{
container
=
"lbmc/bowtie2:2.3.4.1"
cpus
=
4
}
withName
:
hisat2_human
{
cpus
=
4
container
=
"lbmc/hisat2:2.1.0"
}
withName
:
sort_bam
{
container
=
"lbmc/samtools:1.7"
cpus
=
1
}
withName
:
index_bam
{
container
=
"lbmc/samtools:1.7"
cpus
=
1
}
withName
:
dedup
{
container
=
"lbmc/umi_tools:1.0.0"
cpus
=
1
}
withName
:
counting
{
container
=
"lbmc/htseq:0.11.2"
cpus
=
1
}
}
}
}
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src/RNAseq.nf
0 → 100644
+
217
−
0
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c2bf2271
/*
* RNAseq Analysis pipeline
*/
params
.
input
=
"data/demultiplexed/*{_R1,_R2}.fastq.gz"
Channel
.
fromFilePairs
(
params
.
input
)
.
ifEmpty
{
error
"Cannot find any file matching: ${params.input}"
}
.
set
{
input_channel
}
/* Trimming by quality */
process
trimming
{
tag
"$file_id"
cpus
4
publishDir
"results/RNAseq/01_cutadapt/"
,
mode:
'copy'
echo
true
input:
set
file_id
,
file
(
reads
)
from
input_channel
output:
set
file_id
,
"*cut_{R1,R2}.fastq.gz"
into
fastq_files_cut
file
"*.txt"
into
rapport_UrQt
script:
"""
cutadapt -a AGATCGGAAGAGC -A AGATCGGAAGAGC \
-o ${pair_id}_cut_R1.fastq.gz -p ${pair_id}_tmp_R2.fastq.gz \
${reads[0]} ${reads[1]} > ${pair_id}_report.txt
cutadapt -u -14 -o ${pair_id}_cut_R2.fastq.gz ${pair_id}_tmp_R2.fastq.gz \
> ${pair_id}_cut_report.txt
"""
}
/* rRNA and tRNA filtering */
params
.
indexrRNA
=
"/Xnfs/lbmcdb/Ricci_team/shared_data/genomes/human_rRNA_tRNA/*.bt2"
log
.
info
"index files : ${params.indexrRNA}"
Channel
.
fromPath
(
params
.
indexrRNA
)
.
ifEmpty
{
error
"Cannot find any index files matching: ${params.indexrRNA}"
}
.
set
{
rRNA_index_files
}
process
rRNA_removal
{
tag
"$file_id"
cpus
8
publishDir
"results/RNAseq/U937/02_rRNA_depletion/"
,
mode:
'copy'
input:
set
file_id
,
file
(
reads
)
from
fastq_files_cut
file
index
from
rRNA_index_files
.
toList
()
output:
set
file_id
,
"*.fastq.gz"
into
rRNA_removed_files
file
"*.txt"
into
bowtie_report
script:
"""
bowtie2 --sensitive -p ${task.cpus} -x human_rRNA_tRNA \
-1 ${reads[0]} -2 ${reads[1]} --un-conc-gz ${file_id}_R%.fastq.gz 2> \
${file_id}_bowtie2_report.txt > /dev/null
if grep -q "Error " ${file_id}_bowtie2_report.txt; then
exit 1
fi
"""
}
/* mapping against human genome with hisat2 */
params
.
index_hg38
=
"/media/adminmanu/Stockage/HISAT2_index_hg38_tran/*.ht2"
log
.
info
"index : ${params.index_hg38}"
Channel
.
fromPath
(
params
.
index_hg38
)
.
ifEmpty
{
error
"Cannot find any hg38 index files matching: ${params.index_hg38}"
}
.
set
{
index_file_hg38
}--
paired
process
hisat2_human
{
tag
"$file_id"
input:
set
file_id
,
file
(
fastq_filtred
)
from
rRNA_removed_reads
file
index
from
index_file_hg38
.
toList
()
output:
set
file_id
,
"*.fastq.gz"
into
reads_non_aligned_hg38
set
file_id
,
"*.bam"
into
reads_aligned_hg38
file
"*.txt"
into
hisat_report
script:
"""
hisat2 -x genome_tran -p ${task.cpus} \
-1 ${fastq_filtred[0]} -2 ${fastq_filtred[1]} \
--un-conc-gz ${file_id}_notaligned_hg38_R%.fastq.gz \
--rna-strandness 'F' \
2> ${file_id}_hisat2_hg38.txt | samtools view -bS -F 4 -o ${file_id}.bam
"""
}
/* sorting */
process
index_bam
{
tag
"$file_id"
publishDir
"${params.output}/03_hisat2_hg38/"
,
mode:
'copy'
input:
set
file_id
,
file
(
bam
)
from
reads_aligned_hg38
output:
set
file_id
,
"*_sorted.{bam,bam.bai}"
into
sorted_bam_files
script:
"""
samtools sort -@ ${task.cpus} -O BAM -o ${file_id}_sorted.bam ${bam}
samtools index ${file_id}_sorted.bam
"""
}
sorted_bam_files
.
into
{
for_dedup
;
for_htseq
}
/* deduplicating reads
params.dedup_options = "--paired"
process dedup {
tag "$file_id"
input:
set file_id, file(bam) from for_dedup
output:
set file_id, "*dedup.bam" into dedup_bam
file "*.txt" into dedup_report
script:
"""
umi_tools dedup -I ${bam[0]} \
${params.dedup_options} \
-S ${file_id}_dedup.bam > report.txt
"""
}
process sort_bam {
tag "$file_id"
publishDir "${params.output}/03_hisat2_hg38_dedup/", mode: 'copy'
input:
set file_id, file(bam) from dedup_bam
file dedup from dedup_report
output:
set file_id, "*_sorted.{bam,bam.bai}" into sorted_bam_files_2
file "*.txt" into report_dedup
script:
"""
samtools sort -@ ${task.cpus} -O BAM -o ${file_id}_sorted.bam ${bam}
samtools index ${file_id}_sorted.bam
cat ${dedup} > ${file_id}_dedup_report.txt
"""
} */
/* HTseq */
params
.
gtf
=
"$baseDir/data/annotation/*.gtf"
log
.
info
"gtf files : ${params.gtf}"
Channel
.
fromPath
(
params
.
gtf
)
.
ifEmpty
{
error
"Cannot find any gtf file matching: ${params.gtf}"
}
.
set
{
gtf_file
}
process
counting
{
tag
"$file_id"
publishDir
"${params.output}/04_HTseq/"
,
mode:
'copy'
input:
set
file_id
,
file
(
bam
)
from
for_htseq
file
gtf
from
gtf_file
.
toList
()
output:
file
"*.count"
into
count_files
script:
"""
htseq-count ${bam[0]} ${gtf} \
--mode=intersection-nonempty \
-a 10 \
-s yes \
-t CDS \
-i gene_id \
-r pos \
-f bam \
> ${file_id}_CDS.count
htseq-count ${bam[0]} ${gtf} \
--mode=intersection-nonempty \
-a 10 \
-s yes \
-t exon \
-i gene_id \
-r pos \
-f bam \
> ${file_id}_exon.count
"""
}
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