cuffdiff replicates empty results

[fangquan]

I use cuffdiff to detect differential expressed genes in two samples, since each has 2 replicates, so i use commas to separate replicates.

cuffdiff -m 40 genes.gtf  /60PO/accepted_hits.bam , /6124/accepted_hits.bam /3584/accepted_hits.bam , /4189/accepted_hits.bam

It gives empty results;

But if I run:

cuffdiff -m 40 genes.gtf  /60PO/accepted_hits.bam /3584/accepted_hits.bam

It works fine.

I have no idea why the replicates doesn't work.

[Jane M]

Unfortunately, I can't provide you a solution, I'm interested in the answer, I also have a problem with the replicates, but I don't get empty resuts (http://seqanswers.com/forums/showthread.php?t=13868). The only difference is that I didn't use the option -m. Did you try without?

[dpryan]

A version of the following seems to have worked for me:

cuffdiff mm9_ensembl.combined.gtf -N -u -p 8 -L Label1,Label2 -b ~/bowtie/indexes/mm9.fa -o Label1_vs_Label2_ensembl L1/L1_accepted_hits.bam,L4/L4_accepted_hits.bam L2/L2_accepted_hits.bam,L3/L3_accepted_hits.bam

It looks like you've added a space in your comma separated list. I wonder if that's the cause of your problem (I've not looked through the code for cuffdiff to see how it reads in its arguments, but it's not uncommon to take a space as a separator between positional arguments).

[Jane M]

Sorry, where did I add a space in my list? I don't see differences between our commands, except the options:

cuffdiff -L Raman,HM1 -N --FDR 0.05 /path/fichier.gtf /path/Cufflinks/L2/accepted_hits.bam,/path/Cufflinks/L4/accepted_hits.bam,/path/Cufflinks/L6/accepted_hits.bam /path/Cufflinks/L3/accepted_hits.bam,/path/Cufflinks/L7/accepted_hits.bam,/path/Cufflinks/L8/accepted_hits.bam

A version of the following seems to have worked for me:

cuffdiff mm9_ensembl.combined.gtf -N -u -p 8 -L Label1,Label2 -b ~/bowtie/indexes/mm9.fa -o Label1_vs_Label2_ensembl L1/L1_accepted_hits.bam,L4/L4_accepted_hits.bam L2/L2_accepted_hits.bam,L3/L3_accepted_hits.bam

It looks like you've added a space in your comma separated list. I wonder if that's the cause of your problem (I've not looked through the code for cuffdiff to see how it reads in its arguments, but it's not uncommon to take a space as a separator between positional arguments).

[dpryan]

It's in the initial post ("/60PO/accepted_hits.bam , /6124/accepted_hits.bam" has two spaces in it surrounding the comma), not the one you just placed. Perhaps it's just a artifact of the forum software, then.

[Jane M]

Ok, I see...
Which version of Cuffdiff do you use?
I'm using the version 0.9.3 and I just read that the version 1.0.0 allows to handle the replicates. This could explain the problem...

[dpryan]

Ah, that'll probably do it then. I'm using the most recent (1.0.3).

[Jane M]

I confirm, the version was the problem ! So to handle replicates, the version 1.0.0 at least is necessary