T/A cloning

martin LEACH leach at bu.edu
Mon Jan 15 16:23:45 EST 1996


check out the Jan 96 issue of biotechniques...

there is a nice article on increasing the efficiency of T-vector ligations...

basically in the preparation...

1. digest the vector with a blunt cutter (we use EcorV)
2. do the T'ing reaction using Taq pol
3. Set up a ligation reaction...self-ligating the t'ing reaction
   any non-t'd vector will ligate to other non-t'd vector forming either
circular plasmids or concatamers of it.
4. Run the ligation reaction on the gel and excise the linear form...that with
a T at each end of the vector.

....increases the efficacy of cloning since the t-vector prep is now enriched
for the correct construct.


.....          Martin Leach                Email:leach at bu.edu
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