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Merged
Juke34 merged 8 commits into from
Feb 5, 2026
+302 −123
Merged

Cram #41

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1c73281
increment version
Juke34 Feb 3, 2026
bc8613e
fix missing publishDir output for subread
Juke34 Feb 4, 2026
1cf0ef6
add new processes to make cram output available through --cram parameter
Juke34 Feb 4, 2026
48f945c
update samtools to latest version
Juke34 Feb 4, 2026
2c3b052
add apt-get update before downloading java
Juke34 Feb 5, 2026
38643a2
fix tag
Juke34 Feb 5, 2026
00f9b11
add test with cram output
Juke34 Feb 5, 2026
30651f1
fix option order
Juke34 Feb 5, 2026
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6 changes: 4 additions & 2 deletions .github/workflows/main.yml
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Expand Up @@ -40,7 +40,7 @@ jobs:
${{ runner.os }}-openjdk-11
- name: Install openjdk
if: ${{ steps.cache-openjdk.outputs.cache-hit != 'true' }}
run: sudo apt-get install openjdk-11-jdk
run: sudo apt-get update && sudo apt-get install -y openjdk-11-jdk
# Install/cache nextflow
- name: Cache nextflow
id: cache-nextflow
Expand All @@ -52,13 +52,15 @@ jobs:
${{ runner.os }}-nextflow
- name: Install Nextflox
if: ${{ steps.cache-nextflow.outputs.cache-hit != 'true' }}
run: curl -s https://get.nextflow.io | bash && mv nextflow /usr/local/bin && chmod +x /usr/local/bin/nextflow
run: export NXF_VER=24.10.6 && curl -s https://get.nextflow.io | bash && mv nextflow /usr/local/bin && chmod +x /usr/local/bin/nextflow

# Checks-out your repository under $GITHUB_WORKSPACE, so your job can access it
- uses: actions/checkout@v3
# Run tests sequentialy
- name: test short single
run: nextflow run -ansi-log -profile docker,test_illumina_single aline.nf
- name: test short single cram
run: nextflow run -ansi-log -profile docker,test_illumina_single aline.nf --cram
- name: test short paired
run: nextflow run -ansi-log -profile docker,test_illumina_paired aline.nf
- name: test ont
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5 changes: 3 additions & 2 deletions README.md
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Expand Up @@ -8,7 +8,7 @@ AliNe (Alignment in Nextflow) - RNAseq DNAseq
AliNe is a pipeline written in Nextflow that aims to efficiently align reads against a reference using the tools of your choice.

Input: file, list of file, folder or csv
Output: Coordinate sorted BAM file.
Output: Coordinate sorted BAM/CRAM file.

## Table of Contents

Expand Down Expand Up @@ -337,6 +337,7 @@ On success you should get a message looking like this:
--reference path to the reference file (fa, fa.gz, fasta or fasta.gz)
--aligner aligner(s) to use among this list (comma or space separated) [bbmap, bowtie, bowtie2, bwaaln, bwamem, bwamem2, bwasw, graphmap2, hisat2, kallisto, minimap2, novoalign, nucmer, ngmlr, star, subread, sublong]
--outdir path to the output directory (default: alignment_results)
--cram output alignment files in sorted CRAM format instead of sorted BAM (default: false). This saves disk space but disables FastQC on alignment files.
--annotation [Optional][used by graphmap2, STAR, subread] Absolute path to the annotation file (gtf or gff3)

Type of input reads
Expand All @@ -351,7 +352,7 @@ On success you should get a message looking like this:

Extra steps
--trimming_fastp run fastp for trimming (default: false)
--fastqc run fastqc on raw and aligned reads (default: false)
--fastqc run fastqc on raw and aligned reads (default: false). Note: FastQC will be automatically disabled for alignment files when --cram is enabled.
--samtools_stats run samtools stats on aligned reads (default: false)
--multiqc_config path to the multiqc config file (default: config/multiqc_conf.yml)

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278 changes: 176 additions & 102 deletions aline.nf
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2 changes: 1 addition & 1 deletion config/softwares.config
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Expand Up @@ -58,7 +58,7 @@ process {
container = 'quay.io/biocontainers/salmon:1.10.3--h6dccd9a_2'
}
withLabel: 'samtools' {
container = 'quay.io/biocontainers/samtools:1.3.1--h0cf4675_11'
container = 'quay.io/biocontainers/samtools:1.23--h96c455f_0'
}
withLabel: 'seqkit' {
container = 'quay.io/biocontainers/seqkit:2.8.2--h9ee0642_1'
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2 changes: 1 addition & 1 deletion modules/bwamem2.nf
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Expand Up @@ -35,7 +35,7 @@ process bwamem2_index {
*/
process bwamem2 {
label 'bwamem2'
tag "$sample"
tag "${meta.id}"
publishDir "${params.outdir}/${outpath}", pattern: "*bwamem2.log", mode: 'copy'

input:
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27 changes: 26 additions & 1 deletion modules/fastqc.nf
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Expand Up @@ -21,5 +21,30 @@ process fastqc {
mkdir fastqc_${sample_id}${add_suffix}logs
fastqc -t ${task.cpus} -o fastqc_${sample_id}${add_suffix}logs -q ${reads}
"""

}

// To take in consideration the index coming along when aligned files are provided
process fastqc_ali {
label 'fastqc'
tag "${meta.id}"
publishDir "${params.outdir}/${outpath}", mode: 'copy'

input:
tuple val(meta), path(reads), path(index)
val outpath
val suffix

output:
path ("*logs")

script:

// Suffix to separate different runs
def sample_id = meta.id
def add_suffix = suffix ? "_${suffix}_" : '_'

"""
mkdir fastqc_${sample_id}${add_suffix}logs
fastqc -t ${task.cpus} -o fastqc_${sample_id}${add_suffix}logs -q ${reads}
"""
}
3 changes: 2 additions & 1 deletion modules/graphmap2.nf
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Expand Up @@ -87,11 +87,12 @@ process graphmap2 {
rm ${fileName}_graphmap2.sam
awk '!/^@HD/ && !/^@SQ/ && !/^@RG/ && !/^@PG/ && !/^@CO/ && NF' ${reads[1].baseName}_graphmap2.sam >> ${fileName}_graphmap2_concatR1R2.sam
rm ${reads[1].baseName}_graphmap2.sam
sed -i '/^\$/d' ${fileName}_graphmap2_concatR1R2.sam # remove empty lines that can be generated by graphmap2 in header
"""
} else {
"""
graphmap2 ${graphmap2_options} -i ${graphmap2_index_files} -t ${task.cpus} -r ${genome} -d ${reads[0]} -o ${fileName}_graphmap2.sam 2> ${fileName}_graphmap2.log
sed -i '\$!N; /^\\(.*\\)\\n\\1\$/!P; D' ${fileName}_graphmap2.sam
sed -i '/^\$/d' ${fileName}_graphmap2.sam # remove empty lines that can be generated by graphmap2 in header
"""
}
}
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4 changes: 2 additions & 2 deletions modules/kallisto.nf
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Expand Up @@ -26,7 +26,7 @@ process kallisto_index {
process kallisto {
label 'kallisto'
tag "${meta.id}"
publishDir "${params.outdir}/${outpath}", pattern: "${filename}/*.bam", mode: 'copy'
publishDir "${params.outdir}/${outpath}", mode: 'copy', pattern: "*.log"

input:
tuple val(meta), path(reads)
Expand All @@ -42,7 +42,7 @@ process kallisto {
def kallisto_options = meta.kallisto_options ?: ""

// catch filename
filename = AlineUtils.getCleanName(reads) + "_kallisto_sorted"
filename = AlineUtils.getCleanName(reads) + "_kallisto"

// For paired-end reads, Kallisto automatically estimates the fragment length distribution from the data and does not require you to specify it manually
if (meta.paired){
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2 changes: 1 addition & 1 deletion modules/ngmlr.nf
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Expand Up @@ -15,7 +15,7 @@ process ngmlr {
val outpath

output:
tuple val(meta), path ("*ngmlr.sam"), emit: tuple_sample_sam, optional:true
tuple val(meta), path ("*.sam"), emit: tuple_sample_sam, optional:true
path "*.log", emit: ngmlr_summary

script:
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61 changes: 56 additions & 5 deletions modules/samtools.nf
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Expand Up @@ -70,26 +70,77 @@ process samtools_merge_bam_if_paired {
/*
http://www.htslib.org/doc/samtools-sort.html
Sort alignments by leftmost coordinates
And convert to cram if needed (when samtools_bam2cram can be avoided) to save disk space
*/
process samtools_sort {
label 'samtools'
tag "${meta.id}"
publishDir "${params.outdir}/${outpath}", mode: 'copy'

input:
tuple val(meta), path(bam)
val outpath
path(genome_fasta)

output:
tuple val(meta), path ("*_sorted.{bam,cram}"), emit: tuple_sample_ali

script:

if (params.cram) {
"""
samtools sort -@ ${task.cpus} --reference ${genome_fasta} -o ${bam.baseName}_sorted.cram ${bam}
"""
} else {
"""
samtools sort -@ ${task.cpus} -o ${bam.baseName}_sorted.bam ${bam}
"""
}
}


/*
http://www.htslib.org/doc/samtools-view.html
Convert BAM to CRAM format
*/
process samtools_bam2cram {
label 'samtools'
tag "${meta.id}"

input:
tuple val(meta), path(bam)
path(genome_fasta)

output:
tuple val(meta), path ("*_sorted.bam"), emit: tuple_sample_sortedbam
tuple val(meta), path ("*.cram"), emit: tuple_sample_ali

script:

"""
samtools sort -@ ${task.cpus} ${bam} -o ${bam.baseName}_sorted.bam
samtools view -C -T ${genome_fasta} -o ${bam.baseName}.cram ${bam}
"""
}

/*
http://www.htslib.org/doc/samtools-index.html
Index BAM or CRAM files
*/
process samtools_index {
label 'samtools'
tag "${meta.id}"
publishDir "${params.outdir}/${outpath}", mode: 'copy', pattern: "{*.bam,*.cram,*.crai,*.bai}"

input:
tuple val(meta), path(alignment)
val outpath

output:
tuple val(meta), path(alignment), path ("*.{bai,crai}"), emit: tuple_sample_ali

script:

"""
samtools index ${alignment}
"""
}

/*
http://www.htslib.org/doc/samtools-stats.html
Expand All @@ -101,7 +152,7 @@ process samtools_stats {
publishDir "${params.outdir}/${outpath}", mode: 'copy'

input:
tuple val(meta), path(bam)
tuple val(meta), path(bam), path(index)
path(genome_fasta)
val outpath
val suffix
Expand Down
22 changes: 22 additions & 0 deletions modules/seqkit.nf
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Expand Up @@ -60,3 +60,25 @@ process seqkit_convert {
"""
}
}

/*
* Clean FASTA headers by removing everything after the first space
* and create samtools index
*/
process seqkit_clean_fasta_headers {
label 'seqkit'
tag "${fasta.baseName}"
publishDir "${params.outdir}/${outpath}", mode: 'copy'

input:
path(fasta)
val outpath

output:
path("*.clean.fa"), emit: clean_fasta

script:
"""
seqkit replace -p " .*" -r "" ${fasta} > ${fasta.baseName}.clean.fa
"""
}
6 changes: 4 additions & 2 deletions modules/star.nf
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Expand Up @@ -73,7 +73,7 @@ process star_index {
process star {
label 'star'
tag "${meta.id}"
publishDir "${params.outdir}/${outpath}", mode: 'copy'
publishDir "${params.outdir}/${outpath}", mode: 'copy', pattern: "{*.out,*SJ.out.tab}"

input:
tuple val(meta), path(reads)
Expand Down Expand Up @@ -137,7 +137,7 @@ For a study with multiple samples, it is recommended to collect 1st pass junctio
process star2pass{
label 'star'
tag "${meta.id}"
publishDir "${params.outdir}/${outpath}", pattern: "*.log", mode: 'copy'
publishDir "${params.outdir}/${outpath}", mode: 'copy', pattern: "{*.out,*SJ.out.tab}"

input:
tuple val(meta), path(reads)
Expand All @@ -149,6 +149,8 @@ process star2pass{
output:
tuple val(meta), path ("*.bam"), emit: tuple_sample_bam
path "*.out", emit: star_summary
path "*SJ.out.tab", emit: splice_junctions


script:
// options for STAR
Expand Down
5 changes: 3 additions & 2 deletions modules/subread.nf
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Expand Up @@ -26,11 +26,12 @@ process subread_index {

/*
* To align with subread
* Particularity it is directly sorted by coordinates
*/
process subread {
label 'subread'
tag "${meta.id}"
publishDir "${params.outdir}/${outpath}", pattern: "*subread.vcf", mode: 'copy'
publishDir "${params.outdir}/${outpath}", mode: 'copy', pattern: "*.{vcf,log}"

input:
tuple val(meta), path(fastq)
Expand All @@ -42,7 +43,7 @@ process subread {
output:
tuple val(meta), path ("*.bam"), emit: tuple_sample_bam, optional:true
path "*subread.vcf", emit: subread_vcf, optional:true
path "*.log", emit: sublong_log
path "*.log", emit: subread_log

script:
// options for subread
Expand Down
2 changes: 1 addition & 1 deletion nextflow.config
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Expand Up @@ -5,7 +5,7 @@ manifest {
description = 'Nextflow alignment pipeline'
mainScript = 'aline.nf'
nextflowVersion = '>=22.04.0'
version = '1.4.0'
version = '1.5.3'
}


Expand Down