This is the first time im trying this but I keep running into issues. I'm running the htseq-count on my aligned BAM files to create count tables that I can then feed into Deseq2. I run the htseq-count using the default settings and the UCSC Main on Mouse: refGene (Genome) as the GFF file. The job finishes but there are no counts on any of the genes that I can see.
Just out of curiosity I fed the files into Deseq2 and the job crushed with this comment :
Fatal error: An undefined error occured, please check your intput carefully and contact your administrator. Creating a generic function for 'nchar' from package 'base' in package 'S4Vectors' all samples have 0 counts for all genes. check the counting script. all genes have equal values for all samples. will not be able to perform differential analysis 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
Any Help would be much appreciated Thanks :)
Hello,
There may be a reference genome mismatch problem. This is how to check: https://wiki.galaxyproject.org/Support#Reference_genomes
If that is not the problem, please send in a bug report so that we can review. https://wiki.galaxyproject.org/Support#Reporting_tool_errors
Thanks for reporting the problem, Jen, Galaxy team
Dear Jennifer thanks for the help,
The reads were mapped by the people doing the sequencing back to the mouse Grcm38.ERCC reference genome. What I did is import the GTF from the UCSC table browser Dec.2011(GRCm38/mm10) for the "Genes & Gene Predictions", shouldn't this be the correct reference?
Thanks