HiFi PCR and RE:TritonX100

Markus Schneemann schneema at cmgm.Stanford.EDU
Mon Nov 30 16:18:34 EST 1998


Hi all:

1) Hi-Fi PCR:
For PCR you can mix Taq with a proofreading polymerase to increase
fidelity.
Are Vent, Pfu, Pwo equivalent?
How much of those do you mix with Taq - 1:1 or 1:10 or even 1:100?
Can you clone the products into TA-vectors or do you get blunt-ends from
the proofreading enzyme?

Commercially available kits (Stratagene, Invitrogen) don't tell how they
'blend' the taq with their proofreader.

2)RE:TRITON X-100
I posted about Triton X-100 as a buffer ingredient for PCR and my feeling
that it would decrease sensitivity about 10fold. We tested it against
laureth12 side by side, and the sensitivity is 10x lower with Triton. Most
likely, some peroxides in the triton harm the Taq.
Boehringer sells their buffers with TritonX-100, but apparently their
Triton doesn't do it...

Cheers,

Markus

Markus Schneemann
schneema at cmgm.stanford.edu




More information about the Methods mailing list