Dear DESeq2 community,
I am just wondering whether I can use DESeq2 to perform transcript-level differential expression with Salmon quantification data? And if I can, is that essential to include bootstrap when using Salmon? I saw a tutorial said DESeq2 will not work if one want to do transcript-level differential expression, just curious whether it is true or not.
Thanks.
Thanks Michael. So what you suggested is using fishpond to do transcript-level differential expression instead of DESeq2?
Yes, it is exactly designed for this. Give it a shot and let me know if you have any questions.
Thanks Michael! I'll try it out. One more question, can I use DESeq2 for transcript-level differential expression if the read quantification data were generated by StringTie since this is an alignment-based aligner? Or I still have to use fishpond?
With StringTie they don’t have a measure of quantification uncertainty so you can’t use Swish. You can use DESeq2 then.
Hi Michael!
1)I have done quantification using Kallisto. Can fishpond be used with txi.kallisto$counts as an input to perform transcript-level differential expression.
2)In the vignette, https://bioconductor.org/packages/devel/bioc/vignettes/tximport/inst/doc/tximport.html#kallisto, abundance.h5 is used for reading transcript-level information. If I do not use tx2gene and follow the DEseq2 protocol, will I get transcript-level differential expression?
thanks ekta
You can use tximeta to import kallisto bootstraps to run Swish as well. You will get transcript level counts for performing DTE.
Hi Michael .. I used kallisto bootstraps to run Swish and performed the Differential transcript expression.
sum(mcols(y)$qvalue < .05) gives 1317
But surprisingly the qvalue for all these 1317 differentially expressed transcripts is 7.59E-06.
And after this, 220 transcripts have qvalue 0.916510084, 39337 transcripts have qvalue of 0.9442869.
What could be the possible reasons for it?
Ekta
This is not an issue, see the vignette which talks about this phenomenon.