Hi All, I am trying to analyse some human RNA-seq dataset which is in BAM format to generate FPKM values using cufflinks. The ensembl group have aligned the data using BWA 0.5.9 to generate these BAM datasets. I realise that TopHat alignments are optimal for Cufflinks, but is it possible for me to use cufflinks or some other tool to derive FPKM values from BWA aligned reads? I tried using cufflinks on one of the BAM files but got an error: Error running cufflinks. return code = -11 Command line: cufflinks -q --no-update-check -I 300000 -F 0.100000 -j 0.150000 -p 8 /galaxy-repl/main/files/006/853/dataset_6853120.dat The additional output was: cufflinks v2.1.1 cufflinks -q --no-update-check -I 300000 -F 0.100000 -j 0.150000 -p 8 I have sent it to the galaxy team and awaiting their reply. Any thoughts about how I can go about analysing this data. Best wishes Mervyn Dr. Mervyn G Thomas BSc, MBChB, PhD Academic Foundation Doctor School of Medicine University of Leicester UK