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Old 05-15-2017, 04:30 PM   #8
Location: Honolulu, HI

Join Date: Jul 2015
Posts: 40

Originally Posted by ymilesz View Post
Hi I am also following the ddRAD protocol from Peterson et al 2012, and I too am having trouble with PCR after size selection. Are you doing the 20uL reactions as suggested as well?
Hi ymilesz,

Sorry for the late response. At first I actually ran 50uL reactions (due to low DNA concentrations in the elute), and then I tried running 25uL reactions.

Interestingly, the ones that tended to fail were the ones that showed higher DNA concentrations post-size-selection. I added water to these to dilute them down to 20ng total DNA content. I'm starting to suspect that this may have affected my results, but I'm not sure.
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