Creating T-cloning vector

Dr. Duncan Clark Duncan at
Thu Feb 18 04:36:52 EST 1999

In article <36CC4D6D.761A at>, Dr Antony Halsall
<halsalla at> writes
>I am trying to produce a T-cloning vector for the direct ligation of a 
>PCR product into a reporter plasmid (pGL3-basic). I know it is possible 
>to create the necessary 3' T overhangs by incorporating two adjacent 
>restriction sites for either HphI, AspEI or XcmI, however all these 
>enzymes will cut pGL3-basic. So my question is does anyone know of 
>another enzyme that will allow the creation of 2 3' T overhangs or a 
>method for cutting at a site present in the pGL3-basic and filling in to 
>create the appropriate overhang?
>Thanks in advance
>Dr A. Halsall
>Dept. Psychological Medicine

Blunt your vector. Either incubate with Taq pol and just dTTP or use
terminal transferase and ddTTP. All in appropriate buffers for the
enzyme in question


The problem with being on the cutting edge is that you occasionally get 
sliced from time to time....

Duncan Clark
DNAmp Ltd.
Tel: +44(0)1252376288
FAX: +44(0)8701640382

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