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Copy path7. HTSeq to obtain gene counts
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7. HTSeq to obtain gene counts
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####NOTE: The gff3 file was fist converted into GTF format file using following script####
## -- unzip gff3 file
gunzip -d CillinoinensisPawnee_588_v1.1.gene_exons.gff3.gz
## -- use gffread from CUFFLINKS to generate GTF file (http://cole-trapnell-lab.github.io/cufflinks/file_formats/#the-gffread-utility
ml gffread/0.11.6-GCCcore-8.3.0
gffread -E CillinoinensisPawnee_588_v1.1.gene_exons.gff3 -T -o CillinoinensisPawnee_588_v1.1.gene_exons.gtf
## Now run HTSeq (Note this script was separated into three subscripts to make the job run faster)
#!/bin/bash
#SBATCH --job-name=htseq # Job name
#SBATCH --partition=highmem_p # Partition (queue) name
#SBATCH --ntasks=1 # Run a single task
#SBATCH --mem=100gb # Job memory request
#SBATCH --time=100:00:00 # Time limit hrs:min:sec
#SBATCH --output=htseq.%j.out # Standard output log
#SBATCH --error=htseq.%j.err # Standard error log
#SBATCH --mail-type=END,FAIL # Mail events (NONE, BEGIN, END, FAIL, ALL)
#SBATCH [email protected] # Where to send mail
gtf="/scratch/gb92068/RNA_seq_updated/mapping/pawnee/genome_downloads/CillinoinensisPawnee_588_v1.1.gene_exons.gtf"
bam="/scratch/gb92068/RNA_seq_updated/mapping/pawnee/bowtie_STAR_mapping/starout_pecan/"
outdir="/scratch/gb92068/RNA_seq_updated/mapping/pawnee/bowtie_STAR_mapping/htseq/htseq_out/"
cd $SLURM_SUBMIT_DIR
ml HTSeq/0.13.5-foss-2019b-Python-3.7.4
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/Control_1_24hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/Control_1_24hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/Control_1_48hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/Control_1_48hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/Control_1_96hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/Control_1_96hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/Control_2_24hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/Control_2_24hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/Control_2_48hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/Control_2_48hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/Control_2_96hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/Control_2_96hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/Control_3_24hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/Control_3_24hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/Control_3_48hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/Control_3_48hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/Control_3_96hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/Control_3_96hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/De-Tif-11_1_24hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/De-Tif-11_1_24hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/De-Tif-11_1_48hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/De-Tif-11_1_48hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/De-Tif-11_1_96hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/De-Tif-11_1_96hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/De-Tif-11_2_24hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/De-Tif-11_2_24hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/De-Tif-11_2_48hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/De-Tif-11_2_48hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/De-Tif-11_2_96hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/De-Tif-11_2_96hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/De-Tif-11_3_24hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/De-Tif-11_3_24hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/De-Tif-11_3_48hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/De-Tif-11_3_48hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/De-Tif-11_3_96hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/De-Tif-11_3_96hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/Pa-OK-11_1_24hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/Pa-OK-11_1_24hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/Pa-OK-11_1_48hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/Pa-OK-11_1_48hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/Pa-OK-11_1_96hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/Pa-OK-11_1_96hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/Pa-OK-11_2_24hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/Pa-OK-11_2_24hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/Pa-OK-11_2_48hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/Pa-OK-11_2_48hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/Pa-OK-11_2_96hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/Pa-OK-11_2_96hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/Pa-OK-11_3_24hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/Pa-OK-11_3_24hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/Pa-OK-11_3_48hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/Pa-OK-11_3_48hr.htseq.counts
htseq-count -f bam -r pos -s reverse -t exon -i gene_id $bam/Pa-OK-11_3_96hrAligned.sortedByCoord.out.bam $gtf --minaqual 10 > $outdir/Pa-OK-11_3_96hr.htseq.counts