I have aligned my reads using HISAT, filtered and merged my data into a BAM file. I have tried reading counts using HT-seq but failed to get any results. What other tools should i use to read counts for my data(RNA-seq,PE)? My data is required for differential gene expression(DESeq)
FeatureCounts is an alternative tool. Example usage is in the tutorials linked below.
However, getting no results from HTseq-count usually means there is a problem with the reference annotation and if present, can impact FeatureCounts results as well.
- Verify the content and format. GFF/GTF formatted data is accepted, GFF3 is not. Use the tool gffread to convert, if needed.
- Also check to make sure the reference annotation is an exact match for the genome build/source used for mapping, Mismatches will effectively result in the annotation not actually being used when counting up the input BAM's mapped reads.
Galaxy support FAQs: https://galaxyproject.org/support/#getting-inputs-right
Galaxy Tutorials, including those for RNA-seq: https://galaxyproject.org/learn/
Thanks, Jen, Galaxy team