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nano commands.bedtools
#enter below commands into the file
singularity exec ${BIOCONTAINER_DIR}/biocontainers/bedtools/bedtools-2.27.1--1.simg bedtools multicov -bams hisat_results/C1_R1.sorted.bam hisat_results/C1_R2.sorted.bam hisat_results/C1_R3.sorted.bam hisat_results/C2_R1.sorted.bam hisat_results/C2_R2.sorted.bam hisat_results/C2_R3.sorted.bam -bed ../reference/genes.formatted.gtf > gene_counts.gff
#use ctrl+x to quit out of nano |
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launcher_creator.py -j commands.bedtools -n multicov -q normal -t 02:00:00 -a UT-2015-05-18OTH21164 -l bedtools_launcher.slurm -m "module load biocontainers; module load bedtools"
sbatch --reservation=RNAseq bedtools_launcher.slurm |
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HTseq
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Image from:http://www-huber.embl.de/users/anders/HTSeq/
Htseq is a module on stampede2lonestar6, but not part of biocontainers. Module spider htseq shows us the other modules (intel/17.0.4) we need in order to load htseq.
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nano commands.htseq
#put these lines in the commands file
htseq-count -m intersection-nonempty --stranded reverse -i gene_id hisat_results/C1_R1.sam ../reference/genes.formatted.gtf > C1_count1.gff
htseq-count -m intersection-nonempty --stranded reverse -i gene_id hisat_results/C1_R2.sam ../reference/genes.formatted.gtf > C1_count2.gff
htseq-count -m intersection-nonempty --stranded reverse -i gene_id hisat_results/C1_R3.sam ../reference/genes.formatted.gtf > C1_count3.gff
htseq-count -m intersection-nonempty --stranded reverse -i gene_id hisat_results/C2_R1.sam ../reference/genes.formatted.gtf > C2_count4.gff
htseq-count -m intersection-nonempty --stranded reverse -i gene_id hisat_results/C2_R2.sam ../reference/genes.formatted.gtf > C2_count5.gff
htseq-count -m intersection-nonempty --stranded reverse -i gene_id hisat_results/C2_R3.sam ../reference/genes.formatted.gtf > C2_count6.gff |
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launcher_creator.py -j commands.htseq -n htseq -q normal -t 02:00:00 -a UT-2015-05-18OTH21164 -l htseq_launcher.slurm -m "module load htseq"
sbatch --reservation=RNAseq htseq_launcher.slurm |
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join C1_count1.gff C1_count2.gff| join - C1_count3.gff | join - C2_count4.gff |join - C2_count5.gff|join - C2_count6.gff > gene_counts_HTseq.gff
#if you have many samples, use for-loop and join |
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nano commands.express
#put these commands into the file
singularity exec ${BIOCONTAINER_DIR}/biocontainers/express/express-1.5.1--h2d50403_1.simg express -o C1_R1_counts.express ../reference/transcripts.fasta hisat_results/C1_R1.sam
singularity exec ${BIOCONTAINER_DIR}/biocontainers/express/express-1.5.1--h2d50403_1.simg express -o C1_R2_counts.express ../reference/transcripts.fasta hisat_results/C1_R2.sam
singularity exec ${BIOCONTAINER_DIR}/biocontainers/express/express-1.5.1--h2d50403_1.simg express -o C1_R3_counts.express ../reference/transcripts.fasta hisat_results/C1_R3.sam
singularity exec ${BIOCONTAINER_DIR}/biocontainers/express/express-1.5.1--h2d50403_1.simg express -o C2_R1_counts.express ../reference/transcripts.fasta hisat_results/C2_R1.sam
singularity exec ${BIOCONTAINER_DIR}/biocontainers/express/express-1.5.1--h2d50403_1.simg express -o C2_R2_counts.express ../reference/transcripts.fasta hisat_results/C2_R2.sam
singularity exec ${BIOCONTAINER_DIR}/biocontainers/express/express-1.5.1--h2d50403_1.simg express -o C2_R3_counts.express ../reference/transcripts.fasta hisat_results/C2_R3.sam |
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launcher_creator.py -j commands.express -n express -q normal -t 04:00:00 -a UT-2015-05-18OTH21164 -l express_launcher.slurm -m "module load biocontainers;module load express"
sbatch --reservation=RNAseq express_launcher.slurm |
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Other Gene Counting Options
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