How To Avoid Mixing Intronic Region When Sequencing Non-Poly-A Rna Using Rna-Seq?

Question

[Hanfei Sun]

The data source is here:

http://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE30567

I was wondering how it could avoid mixing intronic RNA (result of pre-mRNA splicing) when using non poly A RNA sequencing. Anyone has ideas about that?

Thanks!

Answer 1

[Steve Lianoglou]

First: where does it say that there are no intronic reads in any of these files? In fact, in one of the cited papers, they say:

We simply count the number of reads mapping across the exon boundaries into the adjacent intron sequence (which originate from primary, unspliced mRNA molecules) ...

So, clearly, they still have intronic reads.

That having been said -- since this data is from rna-seq conducted in different cellular compartments, you can analyze only the rna from the cytosol in order to find non-polyA RNAs w/o intronic reads (assuming no nulcear RNA contaminated the cytoplasmic RNA fraction).