T/A cloning: efficient enough for library-making?

Peter Wang plw at mrc-lmb.cam.ac.uk
Tue Oct 15 20:06:27 EST 1996


Question to those of you that know a lot about T/A cloning (which is 
often used to clone PCR products - taking advantage of the single 
untemplated A's that are added by Taq polymerase, these are ligated to a 
vector that has complementary single T overhangs):

How efficient is it?  Not just does it give a high percentage of clones 
with inserts, but is the absolute number of clones with inserts as high 
as you would get with sticky-end or blunt-end ligations?

I wonder about T/A ligation efficiency because several times in the New 
England Biolabs catalog it is said that fragments generated by 
restriction enzymes leaving a single base overhang are difficult to 
re-ligate.

I was thinking about using a T/A cloning strategy to shotgun clone a 
library of quite small DNA fragments, and would like to avoid the use of 
linkers/adaptors.

Thanks in advance for any insights or suggestions; please email direct 
as well as posting, if you don't mind.
 
---------------------------------------------------------
Peter Wang, M.D., Ph.D.
MRC Centre for Protein Engineering,
Hills Road, Cambridge, CB2 2QH, England

Tel (01223) 402104  (international calls +44-1223-402104)
Fax (01223) 402140  (     "          "   +44-1223-402140)
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