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how to get my Pfu Turbo going?
Posted by: mastermix (IP Hidden, New member, 1)
Date: August 10, 2005 01:36AM
Hi Guys,
I'm using Pfu Turbo mastermix for cloning. My template is about 2.7kb, in pcDNA3.1, and not GC rich. I tried Pfu Turbo several times but it never worked, while normal Taq could give me a not very high yield. My PCR reaction was set as: 94C 2min, 94C 30sec, 52C 30sec, 72C 4min (or 6min, 35 cycles), and 72C 7min. I tried different concentrations of templates, primers, magnisium and annealing temperature, all gave me nothing, not even primer dimers. I am totally freaked out. Can somebody give me any suggestions? Thanks in advance
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