Question: cuffdiff does not accept BAM files produced by HISAT2
gravatar for abryants
2.9 years ago by
abryants0 wrote:

I ran HISAT2 on my several  RNAseq samples and then wanted to process the resulting BAMs with cuffdiff for splicing differences. The cuffdiff does not accept these BAM files and states the need for the files to be in the same order. I sorted the files with BAMtools, but it didn't fix the issue. I will appreciate a solution to this problem or any useful comment.




rna-seq cufflinks bam • 1.9k views
ADD COMMENTlink modified 2.9 years ago • written 2.9 years ago by abryants0


Well, I guess i don't do it right. TopHat BAMs are giving the same error:

Fatal error: Exit code 1 () Error: sort order of reads in BAMs must be the same

ADD REPLYlink written 2.9 years ago by abryants0
gravatar for Jennifer Hillman Jackson
2.9 years ago by
United States
Jennifer Hillman Jackson25k wrote:


As a quick guess, the sort order of the reference annotation used could potentially be a factor. Sort the GTF dataset by chromosome and start coordinate and re-run to see if that makes a difference (use the tool "Sort").

If that does not work, would you like to submit the problematic run as a bug report? This will allow us to look over the data and provide the best feedback. 

Should you be executing the job on a local/cloud, if some portion of it can be replicated on and submitted as a bug, that will be enough. It the problem cannot be replicated on Main, then that is also useful information to know about.

This question was a bit difficult to track because of the update submitted as a reply to the original post (indicating a solution) instead of as a comment. I'll adjust that now.

Thanks, Jen, Galaxy team


ADD COMMENTlink written 2.9 years ago by Jennifer Hillman Jackson25k
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