Does anyone here use HTSeq to aggregate read counts per gene for their Bowtie2 generated .SAM files? If not, what other method do you use?
I have found that PE Illumina reads put through Bowtie2 create a .SAM that breaks HTSeq. It seems that Bowtie2 tries multiple alignments, and only reports the best possible alignment. When HTSeq sees a read's flag variable that indicates it had multiple alignments, it throws a warning and drops the read. Therefore, I believe HTSeq is only reporting counts for uniquely mapping read pairs in Bowtie2 .SAM output.
Does anyone have any thoughts on this?
I have found that PE Illumina reads put through Bowtie2 create a .SAM that breaks HTSeq. It seems that Bowtie2 tries multiple alignments, and only reports the best possible alignment. When HTSeq sees a read's flag variable that indicates it had multiple alignments, it throws a warning and drops the read. Therefore, I believe HTSeq is only reporting counts for uniquely mapping read pairs in Bowtie2 .SAM output.
Does anyone have any thoughts on this?
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