NimbleGen SeqCap and empPCR lib L

[Zaag]

Yeah we do that but on the 454-FLX+ so no junior here. I am not sure what exactly are the differences regarding emPCR and such, but if you have questions just mail me, or ask them here (mail is faster answer).

Ok.Thank you very much for your reply. I was wondering if you also perform SeqCap EZ Choice...I have a lot of question regarding this method but I don't think many people of this forum perform SeqCap and then run on a GS Junior platform

[Zaag]

That would be a small volume emulsion. They come in three sizes, small medium and large.

Can I ask you what is SVE?

[Zaag]

After sequence capture I usually end up between 1 and 2 cpb; so if you measured your sample with a Qubit before calculating the nr of molecules my guess would be that 1cpb would be a safe number.

Usually I do one SVE with 1 cpb and one with 2 cpb and after calculating enrichment I choose a cpb for my bulk emulsion.

Ok, thank you Zaag. May I ask you how many copy per bead do you use in emPCR lib L? I would start trying 1 cpb since it is the first time that I perform this emPCR. Do you have any suggestion?

[Zaag]

No you don't, after LM-PCR only 1 of the strands will be able to get PCRed on the bead.

Dear all, I performed sequence capture of target exons using NimbleGen SeqCap EZ strategy. Now I have to proceed to emPCR (Lib L). Should I denature my for 95 C degrees for 2 minutes?