Hello,
I'm trying to perform a restriction digest on single fibers. I'm using 2 enzymes BanII and Cviki-1. What I do is I cut 1 fiber out of muscle tissue on a slide (LCM) and perform than a dna extraction. Then i do a PCR with a fam labbeled primer. In the next step I do the restriction digest. The problem I experience is that not the whole sample is digested. about 10% stays undigested. I tried already using different restriction buffers, adding more enzyme, longer incubation times, adding BSA but nothing seems to help.
If I start with 200 ng DNA in a pcr reaction and perform than the restriction digest everything is digested. So I assume it has something to do with the small amount of DNA.
Does anybody have experience with restrction digest starting from small amounts of DNA?
Restriction digest in single fibers
Started by KimV, May 23 2012 12:19 AM
restriction restriction digest
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Also tagged with one or more of these keywords: restriction, restriction digest
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