Why use Klenow on a PCR product to clone?
Brain Foley
brianf at med.uvm.edu
Tue Nov 9 16:38:35 EST 1993
Dear BioNetters:
I was recently asked why so many of the protocols people have
supplied to this list for cloning a PCR product include treating
the PCR product with Klenow fragment to "clean up" the PCR
product ends prior to blunt-end ligation.
The PCR product has a 3' overhang, usually an A, added by the
polymerase. Klenow lacks 3'-5' exonuclease activity, and is only
a 5'-3' polymerase (unless I'm mistaken). Thus it does not seem
to us that Klenow should actaully do anything to "clean up" the
3' overhanging A.
Are we missing something???
--
********************************************************************
* Brian Foley * If we knew what we were doing *
* Molecular Genetics Dept. * it wouldn't be called research *
* University of Vermont * *
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