Question: cuffdiff splicing.diff 'notest' - interpretation?
gravatar for fing
4.6 years ago by
United Kingdom
fing0 wrote:

Hi everyone,

I guess this is more of a general query about interpretation of cuffdiff output, since (thanks to some help from biostars) I'm pretty happy that my analysis is running correctly. I hope someone will be happy to help (again) as I'm an RNA-seq newbie.

I have ran cuffdiff on a set of RNA-seq data, 2 conditions with 30 replicate samples each (drosophila). When I look at the splicing.diff output file, approx. 80-90% of the gene IDs have the status 'notest'. I consulted the manual here, which states that this means 'not enough alignments for testing'. Now I could be naive here, but this surprised me given the number of replicates I have, and the RNA data is good quality from my previous checks. I've seen an example splicing.diff output from a colleague's previous analysis, and they have a similar low proportion of successful tests. So is this a pretty normal output, or is there a problem I'm missing that could explain this?

Thanks in advance for your help!

ADD COMMENTlink written 4.6 years ago by fing0

This would seem to be pretty normal, since most splicing events don't occur in most cells/conditions/developmental stages (not to mention that most genes aren't expressed in most conditions). Have a look at a few of these positions in a few BAM files in IGV or another browser. This should confirm whether this is the case or not.

ADD REPLYlink written 4.6 years ago by Devon Ryan95k

Thanks - this makes sense, good to know the output isn't too weird, and after having a look at some of the data it seems to make sense.

ADD REPLYlink modified 6 months ago by RamRS27k • written 4.6 years ago by fing0
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