Using HTSeq for paired-end files

Question

[Whoknows]

Hi friends

I have 3 RNA-Seq paired end samples which I want to generate differentially expressed gene list, I used these commands:

samtools sort -n accepted_hits.bam accepted_hits_bam_sorted
samtools view -h accepted_hits_bam_sorted.bam >accepted_hits_sorted.sam

But I'm not sure about the following line that must create count table for DESeq input, is it fine (for paired-end)?

htseq-counts --stranded=no --mode=intersection-nonempty -t exon -i gene_id my.sort.sam annotation.gtf > output.txt

Thanks

Answer 1

[Devon Ryan]

That's fine.

For future reference, you can also just use htseq-counts --stranded=no --mode=intersection-nonempty -r pos accepted_hits.bam annotation.gtf > output.txt and save the hassle of sorting and/or conversion to SAM.