How to analyse RNA-SEQ duplicated data coming from a low cluster denstiy experiment

Question

[ghlopez]

Hello all,

I have recently received data from a RNA-Seq experiment. Apparently there has been a drop in the cluster density and samples had to be repeated in order to achieve the minimum number of reads and for that reason I have received two fastq files from two different lanes per sample. When I analyse the fastq files, both have a similar number of reads (15m).

Hiow should I proceed with the analysis of these files? Should I analyse them individually? Should I combine them some how? Any ideas will be greatly appreciated

Thanks in advance and kind regards

Answer 1

[Devon Ryan]

Start by treating them separately and make a PCA or MDS plot. If the two replicates of each sample cluster together (they probably will, but it's best to look for an obvious batch effect first), then go ahead and merge them for further use.

[ghlopez]

Thanks for your suggestion. I will initially treat the samples as two technical replicates and check how they do behave

Cheers and thanks one more time