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cplinka 10-05-2020 01:33 PM

NGS library enrichment question
I have a Hi-C library with P5 and P7 adapters. I would like to enrich for fragments containing a locus-of-interest by capture and extension with a biotinylated bait primer, and then amplification with a nested primer and either of the P5/P7 adapters.

Is is practical to add another sequencing primer adapter to the nested primer, so that I can have the new, enriched library paired-end sequenced? Or will I realistically have to clone the products from the enriched library?

I apologize for being new to NGS, and thanks in advance for any advice!

nucacidhunter 10-05-2020 04:19 PM

It would be more cost effective to add adapter sequences to the nested primer for sequencing on cheapest Illumina flow cell.

cplinka 10-06-2020 10:48 AM

But if the library already has P5 and P7 adapters, what adapter could I add to the nested primer?

Also, thank you for your response!

nucacidhunter 10-06-2020 05:39 PM

If you add a PCR handle to 5 end of probe and then amplify captured target with P5 or P7 primer and the handle primer followed by nested primer containing the other adapter sequences, there will be very small residues of initial library.

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