Question: Error HISAT2 output for HTseq-count

Question

[11yj3312]

Dear all, I am analyzing paired-end stranded RNAseq data . I would be interested in using HISAT2 for the alignment, and HTseq-count to count the features of the aligned reads. When I used HTseq-count to count the reads:

htseq-count -r pos -f bam SRR3589957_sorted.bam ../reference/gencode.v26lift37.annotation.gtf > SRR3589956.count

However,I got this error :

Error occured when processing SAM input (record #69114540 in file SRR3589957_sorted.bam): my_showwarning() takes from 3 to 5 positional arguments but 6 were given [Exception type: TypeError, raised in warnings.py:99]

I converted sam file to bam and sorted the reads by pos using samtools

Does anyone has an idea of the problem with the reads aligned with HISAT2? Are they compatible with HTseq-count?

Thank you very much for any help. Jing Yu

[anc.informatics]

Not an answer to your question, but consider featureCounts which is an ultra fast read count program and work well with Hisat2 output SAM file. Also, it rearrange read itself and final table of read count matrix for all paired end SAM can be obtained. For ex,

featureCounts -p -t exon -g gene_id -a genes.gtf -o counts.txt  con1.sam con2.sam treat1.sam treat2.sam

[11yj3312]

Think you !I will try it

[WouterDeCoster]

That's an odd warning, are you using the latest htseq-count version?

But as said by anc.informatics: featureCounts is an excellent choice.

[11yj3312]

I have tried the latest htseq-count，No previous error occurred.Think you!