native vs cloned Pfu

Vladimir Svetlov svetlov at oncology.wisc.edu
Fri Sep 12 17:34:32 EST 1997


In article <dboyd-1209971024330001 at cc-hsc-k-box26.cc.umanitoba.ca>,
dboyd at ccu.umanitoba.ca (DAB) wrote:

> I was doing a PCR on some bacterial DNA with native Pfu. DNA is about 64%
> A+T, PCR product is about 6kb, primers anneal at about 52.5 C. I used
 > annealing temps from 50-52, and extensions at 72 for 12 min. I was getting
> product and pretty good yeilds. I ran out of Pfu and ordered the cloned
> stuff from Stratagene. All of a sudden the PCR isn't working so good. I
 > notice the rx buffers are different. What about efficiency, robustness
> etc? I'm also starting to get these bright bands at about 100-200 bp. I'm
> basically a novice at PCR so any idea on how to improve the performance
 > will be much appreciated.
 
Quite a few people reported problems with cloned Pfu not seen with the
native. A lot of hand-waving transpired but that's about it. Personally I
had similar problems and eventually dropped Pfu altogether since I found
Pwo both reliable and robust with no loss in fidelity. Stratagene in its
own kits like QuickChange uses native rather than recombinant Pfu. So there
you go, sailor. Go back to nature or switch to Pwo. 
Regards,
V.



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