vent and pwo

Dr. Peter Gegenheimer PGegen at UKans.nolospamare.edu
Sat Dec 5 22:30:31 EST 1998


On Thu, 3 Dec 1998 20:16:25, cbookste at MEDICINE.BSD.UCHICAGO.EDU (Cres Bookstein)
wrote:

> Mark,  Vent adds 3'-A overhangs on PCR product.

OK, I realize that several posters have gotten VENT-amplified DNA to clone into a
TA-vector. (VENT-amplified products can also be cloned into blunt-ended sites, or
self-circularized. This doesn't mean that some fraction of the product doesn't have
overhangs.)

However, I'm sure that NEB would like to know about this, as they have determined
that there is little or no 3'-overhang addition. No enzyme with 3' to 5' proofreading
ability should allow addition of single-stranded bases, since these are substrates
for the proofreading exonuclease activity!

If anyone has sequenced termini of VENT-amplified products, or of such products
ligated into TA vectors, s/he should publish it pronto in BioTechniques so we can all
see it.


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