pfu for cloning PCR

Matthew L. Brown, Ph.D gemini3 at mindspring.com
Wed Apr 30 17:54:55 EST 1997


Zhao,
A few days I posted a description of the A overhang addition reaction
that I use when I create PCR products with a proofreading enzyme like
Pfu. Anyway I want give all the details but you may e-mail me directly
if you can't find a similarl protocol in the literature or in the bionet
archives. Anyway you'll have to invest in a little Taq which you could
use after amplification with Pfu only on the desired PCR products. Add
additional dATP and small amount of Taq and incubate 5 min at 72 C. This
should add A's to most 3' ends.

Good Luck,

Matt Brown, Ph.D.
Research Fellow
Dept. of Medicine
Univ. of Alabama at Birmingham



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