Determining how many alleles/gene duplicates

[michael-steffen]

I am working with a few genes which I know have undergone a variable amount of gene duplication events within and between species.

The trouble I have been having though it determining the best way to figure out how many duplicates are being expressed in my RNA seq runs. When I do a denovo assembly, I can obviously see multiples, but determining the exact amount is something I can not figure out.

Any suggestions on who to deal with this? Our lab has access to geneious and clc genomics workbench if there programs are of any help.

Thanks,
Mike

[dpryan]

I don't think RNAseq will allow you to determine the exact number being expressed. Why don't you first determine the number in the genome (e.g., with a Southern blot or qPCR), determine how identical the sequence of the copies is, and then move forward accordingly. If some of the copies have identical sequence, then it'd be impossible to determine how many of them are being expressed without directly manipulating them (i.e., modifying their sequence to make them distinguishable).