Merging RNA-Seq files run at two different times with identical setup

[adrian]

Hi group:
I am sure this question has been asked in many flavors but after pouring over those, I still could not get what exactly I am asking for. I thought members could provide an answer.

My question:

I have two samples with 3 RNA-Seq replicates ( by combining all 3 replicates from two lanes I get 30Mil reads 100bp length PE reads).

Obviously this is not a good read coverage for identifying splicing differences.

I am aiming now for 70Mil reads to identify the splicing differences.

My question:
1. I am going for another sequencing run shooting for another 40 Mil read coverage.

Can I combine the new 40 Mil reads with previous 30 Mil reads and do analysis for identifying splicing patterns. Is there any hidden surprises that one cannot combine two batches like these (concerning insert sizes etc..). I will use MATS for splicing.

Obviously I will choose the same design (100bp and paired-end design) and I have the same RNA library left from previous sequencing run which I will use for new sequencing.


thanks a lot.
Adrian

[adrian]

Any suggestions I appreciate..
thanks

[GenoMax]

As long as other things remain constant (no of cycles, sequencing kit version, machine) it should be ok.

If it has been a while since you ran the first batch .... You may be able to leverage new technology (e.g. V2 rapid kits) and get all new data that can give you the coverage you desire.