How can I get counts from transgenic expression in RNA-seq data?

Question

[erik.burchard]

Hello all,

This is probably a pretty simple question, but I've never done an analysis with transgenic expression involved before and I want to make sure I do it correctly. Basically, I have RNA-seq data, a genome and annotation files for the host, and the sequence of a transgene and its expression vector. The transgene is exogenous and therefore not in the host genome. Mapping the reads to the host transcriptome is straightforward enough, but what would be the best method by which to get counts data for the exogenous transgene (along with any portions of the vector itself which may be present)?

Thanks,

Erik

Answer 1

[benformatics]

You should add the transgene as a chromosome to the FASTA you used to generate your reference index. You could also add it to your GTF but I think it would be considerably more work. Hopefully you are mapping with something like STAR which should accommodate this well.

[Trivas]

Alternatively, you can take the unmapped reads from the host genome and map it to a reference genome of your expression vector.

[erik.burchard]

Great, thank you! I'm using STAR so this should work just fine. Again, thanks so much!!

[erik.burchard]

Oh, and one more question about this. I have to add it to the GFF file if I want STAR to map it, don't I?

[benformatics]

Not necessarily unless you are using STAR to do your gene/exon counts. Otherwise, you could just count the alignment to the specific chromosome.