I am trying to perform RNA-seq analysis using Cufflinks at Galaxy. In order to use the "Reference Annotation" option, I uploaded the GTF file for zebrafish GRCz10 genome assembly from Ensembl ( ftp://ftp.ensembl.org/pub/current_gtf/danio_rerio/ ). However this GTF format dataset cannot be recognized by Cufflinks, as it does not appear in the drop-down list for "Reference Annotation".
I also tried the GFF format annotation from NCBI ( http://www.ncbi.nlm.nih.gov/genome/50 ). It can be recognized by Cufflinks (shown in the drop-down), but results in FPKM values of zero for all genes. Since the Tophat mapping was done with the reference genome from Ensembl, maybe only the reference annotation from Ensembl fits?
Please give me some advice on how to get the "Reference Annotation" to work.
Thanks a lot,