Confirming intron length

Question

[Apex92]

Hi all,

I have RNA-Seq data (ctrl vs kd) and performed a bunch of analyses. One of the analyses I performed was via rMATS tool to find possible retained introns. In the rMATS output, I do get coordinates for individual retained introns but I think depending on the rMATS output, the exact positions listed might be including the last/first exonic nucleotide, or excluding it.

I want the true length of each retained intron to perform possible frameshift analyses (intron length divided by 3), thus even one nucleotide offset can make my analyses go wrong. With that said, how can I extract intron sequence so then I can calculate the length of it?

Thank you.

Answer 1

[colindaven]

I would use STAR to align the raw reads against the dataset, and check this manually in detail.

Get the BAMs, and use IGV to inspect. I think this is the only method if you need that accuracy.

[Apex92]

Great - thank you for your input.