gravatar for s.l.lee

2 hours ago by

Hi,

I have 40 samples which I have sent for RNA-seq to the depth of 30-60M reads per sample. The sequencing company have sequenced my samples in 3 different batches to achieve the required depth.

My questions are:

1 is there a 'best practice' for how to combine the data from 3 batches into one before DGE analysis? I am aware that there's a method to merge bam files using samtools but I also know of those who convert individual bam files into counts and combine the counts after. Both sounds equally reasonable to me. Is there a different in each method? if so, which is better?

  1. At present, I have decided to try out the second option - convert bam files to counts and combine them. For DGE - should I add all the counts from 3 batches together or use the average? (The sum seems to be the more logical method to appreciate the DGE at the required depth.)

Thank you. Appreciate all advice as I am new to this.



Source link