Question: DEXseq count reads tool obtaining only zero counts for all exons
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gravatar for f.carlisle
5 days ago by
f.carlisle0
f.carlisle0 wrote:

I'd like to run DEXseq on my RNA-seq dataset to identify any potential differences in exon inclusion between samples. I used HISAT2 to generate BAM files of aligned reads, and then DEXseq-Count mode-prepare to generate an annotation GFF file. My reference genome for alignments was hg38, and I used the ENSEMBL hg38 gene annotation GTF to prepare my GFF file. However, whenever I try to run DEXseq-Count in count mode, I obtain zero counts for all exons. I have separately run transcript-level analyses on these BAM files, so I know they're okay and I should be getting non-zero exon counts. Any ideas what I might be doing wrong?

Thanks for your help.

ADD COMMENTlink modified 5 days ago by Jennifer Hillman Jackson23k • written 5 days ago by f.carlisle0
0
gravatar for Jennifer Hillman Jackson
5 days ago by
United States
Jennifer Hillman Jackson23k wrote:

Hello,

Compare your analysis methods to those in our tutorial utilizing the DEXseq tool and it should reveal where the problem is: https://galaxyproject.github.io/training-material/topics/transcriptomics/tutorials/ref-based/tutorial.html#inference-of-the-differential-exon-usage

A data mismatch could also be a factor (mismatched chromosome identifiers between the inputs) - even if the prior transcript-level analysis was successful. How to check: https://galaxyproject.org/support/chrom-identifiers/

For reference, should you need more help:

Thanks! Jen, Galaxy team

ADD COMMENTlink written 5 days ago by Jennifer Hillman Jackson23k

Hi Jen, Thanks for the suggestions. My analysis methods are identical, so it looks like a data mismatch may be the problem.

ADD REPLYlink written 4 days ago by f.carlisle0
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