Problem with Deseq2
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6 weeks ago
yoosefyud ▴ 40

Hello every one I'm trying to obtain differentially expressed genes in galaxy using Deseq2. But unfortunately, I get the following error. I need to mention that my data is pair end and the stringtie results are in a list.

Fatal error: An undefined error occurred, please check your input carefully and contact your administrator.
Tool generated the following standard error:

estimating size factors
Error in estimateSizeFactorsForMatrix(counts(object), locfunc = locfunc,  : 
  every gene contains at least one zero, cannot compute log geometric means
Calls: DESeq ... estimateSizeFactors -> .local -> estimateSizeFactorsForMatrix
galaxy Deseq2 sequencing RNA-seq • 221 views
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every gene contains at least one zero, cannot compute log geometric means

How can this be, so what kind of data is this?

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How do you generate your countData? Maybe the problem is in the upstream steps, because the matrix seems to have no counts for your genes.

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6 weeks ago

It seems that you have data with zero counts. Filter to get rid of them. DESeq2 best practices recommend a filtering of low counts reads before the actual analysis

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I do not think filtering is a general best practice. From the vignette:

While it is not necessary to pre-filter low count genes before running the DESeq2 functions, there are two reasons which make pre-filtering useful: by removing rows in which there are very few reads, we reduce the memory size of the dds data object, and we increase the speed of the transformation and testing functions within DESeq2.

https://bioconductor.org/packages/release/bioc/vignettes/DESeq2/inst/doc/DESeq2.html#pre-filtering

In any case it will not help here I guess as every gene contains a zero so removal of lowly-expressed genes will not change that. So either one or many corrupted samples or unusual data. @OP, please elaborate.

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