RNAseq: differences when shifting from 100bp to 150bp?

Question

[fr]

Our colleagues have previously sequenced data using NovaSeq 100bp PE reads. However, now they changed to 150bp PE reads and I'm wondering whether results will be comparable with those of the 100bp PE if:

• they proceed as usual with the 150bp PE reads
• they trim the results from 150bp to 100bp

Is there anything I should pay attention in this situation? The idea is simply to align to reference, get counts and proceed with transcript comparison.

Thanks in advance

Answer 1

[ATpoint]

Yes, results will be comparable. Still, there will always be some few genes in regions that are diffcult to map that will have different counts depeding on the read length, but this is typically a very limited number. Take a few of these 150bp samples, copy them and trim it to 100bp, then perform DGE between 150 vs 100bp, you will see the effect is very limited. Still, this is of course dependent on the library prep method an cellular context etc. I suggest to be sure you try it out.