Recently, I run cufflink in galaxy on the internet. I want to compare two samples, However, I found no transcript or gene passed the significant level, even many of them have large FPKM in one sample and 0 FPKM in another sample.
Below is my cufflink process:
I have four samples belong to two group. the test have three samples, and the control has one sample.
First, using accept_hit.bam from tophat, I run cufflink without annotation on each sample.
Then, for the four "gtf" files from four samples, I run cuffcompare to combine these transcript and compare to the annotation genome. However, at this step, I found the transcript accuracy is very low.
See one example:
Missed exons: 10673/11776 ( 90.6%)
Wrong exons: 1254/2007 ( 62.5%)
Missed introns: 8529/8637 ( 98.7%)
Wrong introns: 2/5 ( 40.0%)
Missed loci: 0/504 ( 0.0%)
Wrong loci: 1248/2002 ( 62.3%)
at last, I run cufdiff between this two group sample.