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omnivore 03-01-2011 08:50 PM

Non-specific band after gel purification of PCR products
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I'm trying to purify my PCR products (obtained by using barcoded fusion primes) by gel purification to increase the quality of amplicon pyrosequencing.

My problem is that non-specific band of about 1200 bp is not removed by gel purification process (please see the attached pdf file. My product size is about 600 bp and I usually cut out gel slices between 550 and 650 bp).

Do you know how it can occur? And, If I run these samples without further purification, how badly does the non-specific band affect the overall sequencing results?

Thanks in advance.

PatrickJ 03-02-2011 09:24 PM

You might try a nested PCR reaction on the gel-purified product

cali4nia 10-21-2013 10:44 PM

any progress omnivore? I got the same problem now. ty. the sequencing worked fine tho.

GA-J 04-05-2017 07:19 PM

It will not affect your seq, or limited affect. It may be a dimer of your product.

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