I am trying to perform a reference-based analysis of transcriptomic data in Galaxy Australia. I have downloaded the reference genome FASTA file and GTF file for Arabidopsis thaliana from NCBI. I have successfully mapped the raw reads to the reference genome and obtained the results in a BAM file.
However, when I try to run the FeatureCounts tool on the BAM file using the GTF file, I am unable to find any variability in the gene counts. Instead, I am only getting '0' values for each gene count.
Could you please suggest the possible reasons behind this issue? I would appreciate if you could provide some guidance on how I can troubleshoot and resolve this problem to obtain meaningful gene expression results from the reference-based mapping.
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Galaxy specific questions are best posted to their support site: https://help.galaxyproject.org/
That said, problems with counting are likely because of some mismatch between the genome and the annotation used (e.g. chromosome names mismatch).