Entering edit mode
6.6 years ago
garimav89
•
0
Hi, My cuffdiff is hanging everytime, im trying to run at this point:
[11:02:35] Inspecting maps and determining fragment length distributions.
My command is
./cuffdiff -o cuffdiff_data2 \
-b ../../RNA/Ref_transcriptome/index/Homo_sapiens.GRCh37.75.cdna.all.fa \
../../RNA/data2/merged/merged.gtf ../../RNA/data2/bunny/tophat/accepted_hits.bam, \
../../RNA/data2/lal/tophat/accepted_hits.bam, ../../RNA/data2/nameless/tophat/accepted_hits.bam \
../../RNA/data2/control/tophat/accepted_hits.bam
Does it consume CPU cycles (check
top) or does it just sit there for hours consuming no resources and doing nothing?There's little reason to use cuffdiff on human samples, unless you're really expecting to find new isoforms.
As Devon pointed, there is no reason to use cuffdiff unless you're really expecting to find new isoforms. In fact, there is no need to even map the reads against the reference genome, just quantify the counts against a reference transcriptome. If you use kallisto + sleuth, you will get the results really quickly.
Not a good idea. "Members of a discussion forum are usually unpaid volunteers who offer their time and expertise by volition and not by obligation. They are therefore not obliged to answer any questions at all." (http://journals.plos.org/ploscompbiol/article?id=10.1371/journal.pcbi.1002202 ).
Unfortunately, most forums cannot guarantee you an answer within a time period. We respond quickly, especially to questions posted on weekdays in the morning (which yours is), but this cannot be guaranteed. I think it'd be better to manage expectations and give a question 24-36 hours, within which you can be confident you'd get a response unless the topic is super niche.
I have removed the term conveying urgency from your post for the above reason. Thank you for understanding.
You should know that the old 'Tuxedo' pipeline of Tophat(2) and Cufflinks is no longer the "advisable" tool for RNA-seq analysis. The software is deprecated/ in low maintenance and should be replaced by HISAT2, StringTie and ballgown. See this paper: Transcript-level expression analysis of RNA-seq experiments with HISAT, StringTie and Ballgown. There are also other alternatives, including alignment with STAR and bbmap, or pseudo-alignment using salmon.
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Thanku so much for this gesture. Actually I have troubleshooted this problem a lot, still cant find the solution
And have you also read the other comments?
Could you once please try without frag bias correction (-b) and the fasta , something like below and see if it works
Yes I did try, but it wasn't working. But then I tried with -M option in cuffdiff and masked the human gtf file. It worked. Hence, It was inferred that human gtf file is having some problem.