If Cufflinks works without a reference GTF, then the problem is the mismatch b/t your GTF and your bam file. A couple things to check: (1) that your genome chrom names in F3.csfata match those in your GTF; if not, you'll need to modify your GTF to match the names in your fasta. (2) that your GTF is sorted as your BAM is sorted. If these issues don't solve your problem, it's best to use the Cufflinks help email that I noted in my previous email. Good luck, J. On Mar 31, 2011, at 10:32 AM, lishiyong wrote:
Hello,I don't use Galaxy.But I have uploaded the file(sorted file 20:test_44.bam refgene.gtf : ) It's works OK without the reference GTF file ,while with reference, I can't gain the right results, I do these in my computer. 2011-03-31 lishiyong 发件人： Jeremy Goecks 发送时间： 2011-03-31 21:36:52 收件人： lishiyong 抄送： galaxy-user 主题： Re: [galaxy-user] cufflinks FPKM Hello,
It's not clear whether you're using Galaxy. If you're using Galaxy, please share you history with me (History Options --> Share/Publish --> Share with User --> my email) and I'll take a look; otherwise, Cufflinks has an email list for questions: tophat.cufflinks@gmail.com
Best, J.
On Mar 31, 2011, at 3:39 AM, lishiyong wrote:
Hi:
I gain the SOLiD sequencing data.I used bowtie to map human genome then I sort the sam file .I used cuffinks to calculate FPKM with the sam file ,human gtf file .it gives 0 FPKM values and this is for all genes .what's the reason?
(1) bowtie -C human_hg18_color -f F3.csfasta -Q F3_QV.qual -v 2 -k 100 -p 6 --mapq --sam test.sam (2) samtools view -uS test.sam 2>/dev/null | samtools sort -m 2000000000 - test.bam (3) cufflinks -G refGene_hg18.gtf test.bam.bam 2011-03-31 lishiyong ___________________________________________________________ The Galaxy User list should be used for the discussion of Galaxy analysis and other features on the public server at usegalaxy.org. Please keep all replies on the list by using "reply all" in your mail client. For discussion of local Galaxy instances and the Galaxy source code, please use the Galaxy Development list:
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Jeremy Goecks