Home-Made TA cloning

Aydemir Akin akin at sage.cc.purdue.edu
Wed Jul 12 02:19:42 EST 1995


In article <3ttsks$6mf at gazette.bcm.tmc.edu>, Brian Desany
<bd033054 at condor.mbcr.bcm.tmc.edu> wrote:

> There are restriction enzymes that leave single base overhangs in the 
> middle of their site, XcmI (ccannnnn/nnnntgg) for example.  Two of these 
> sites with the appropriate bases chosen for the "n"'s should make it 
> really easy to TA clone - you don't have to ligate anything to the 
> cleaved vector before you do your actual TA cloning reaction.  Although 
> I haven't got around to trying it yet.

===============================================

There is a published article on this.  You may want to take a look at it.
[Kovalic, D., et al.  General method for direct cloning of DNA fragments
generated by the polymerase chain reaction.  NAR 19 (16):4560, 1991.]

In their paper, they engineered pGEm5fZ(+) to have XcmI site. Digestion of
their vector with XcmI produces 'T' overhangs enabling PCR product cloning.

-- 
Aydemir Akin, D.V.M.

akin at sage.cc.purdue.edu

1175 Animal Disease Diagnostic Lab
Purdue University
West Lafayette, IN 47907-1175




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