Hi all,
I am looking for a method to compare the significance of the expression difference of two different transcripts within a specific condition.
with that said, i will try to explain a bit more. We have a RNA-Seq experiment with three conditions. We are comparing each pair of conditions to find significant differentially regulated genes on gene (deseq2,cuffdiff) and transcript level (kallisto,cuffdiff,dexseq).
For some of the transcripts (isoforms of the same gene) we would like not only to know whether or not they are significant DE between the two conditions, but also if transcriptA is significantly higher/lower expressed than transcriptB.
About the data - we have 10 replicates for each of the three conditions.
For the first question I have done done the calculations with the mentioned tools. I am struggling to find a way to calculate the second problem. Is there a method to calculate such significances?
thanks for the help,
Assa
thanks Devon for the fast reply.
I have already the TPMs from kallisto. Does it make sense to use them, or would it be better to run salmon with its own options? or for that matter any counts ( I can use voom to transform them into log-counts per millions)
by munging the data, do you mean taking the results of each "transcriptA.condition1" and "transcriptB.condition1" and consider them as two separate data sets for the comparison?
Yes, you'll need to remove transcripts so you have things like major and minor transcript columns. So something like
You only care about major vs. minor isoform differences. You don't need all of the treatments if you don't care about them, though you can leave them all in if it's useful.