T/A cloning again. To Kinase or Not

Rafael Maldonado rafael at howard.genetics.utah.edu
Wed Jan 17 19:41:09 EST 1996


On 17 Jan 1996, HIRANYA ROYCHOWDHURY wrote:

> On 16 Jan 1996, Francois-Xavier Maquart wrote:
> 
> > Dear Netters.
> > 
> > I have just discover this newsgroup. Fascinating.
> > At the begining I have a naive question: Should I kinase PCR products
> > (or oligos)
> Yes, phosphorylating will definitely better the chance.
> 
>  for T/A cloning? Either Promega pGEM-T vector system kit
> > or Invitrogen TA cloning system kit do not discuss this problem.


I don't get the point. Kinase what? PCR product, as it is, is already 
kinased (I mean that it has a phosphate at the 5' end).

Oligos are different. You have to make sure your oligos are kinased for 
cloning, but it is not strictily necesary if your vector is kinased (or, 
it is not dephosphatased). But only in the case of DIRECT cloning of 
oligos, not through PCR. 


Rafa

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