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Old 03-01-2011, 07:50 PM   #1
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Location: Korea

Join Date: Oct 2010
Posts: 4
Unhappy Non-specific band after gel purification of PCR products

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.
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File Type: pdf Gel.pdf (86.3 KB, 104 views)
omnivore is offline   Reply With Quote
Old 03-02-2011, 08:24 PM   #2
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You might try a nested PCR reaction on the gel-purified product
PatrickJ is offline   Reply With Quote
Old 10-21-2013, 09:44 PM   #3
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any progress omnivore? I got the same problem now. ty. the sequencing worked fine tho.
cali4nia is offline   Reply With Quote
Old 04-05-2017, 06:19 PM   #4
Location: USA

Join Date: Jul 2015
Posts: 28

It will not affect your seq, or limited affect. It may be a dimer of your product.
GA-J is offline   Reply With Quote

amplicon, gel purification, non-specific band, pyrosequencing

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