Strange problems with Invitrogen TOPO vectors Reply-2
Dr. Duncan Clark
Duncan at nospam.demon.co.uk
Wed Oct 6 04:25:21 EST 1999
In article <37FA7843.852D3541 at osuunx.ucc.okstate.edu>, Earl Blewett
<blewett at osuunx.ucc.okstate.edu> writes
>We use rTth for the amplification, gel purify the product, add A residues with
>Taq and wizard-prep it. The final product is a clean band when run on a gel.
Why add Taq?
Tth like all other Thermus polymerase (Tfl, Tub, Taq, TseC, TspB, Tbr,
Tfi etc.) has an integral single base terminal transferase activity,
providing you do the extra 5-10mins final extension. Without that extra
extension a lower proportion of the final product has the overhang (a
quite old NAR single page tech tips reference - and I know I don't
remember which one).
The problem with being on the cutting edge is that you occasionally get
sliced from time to time....
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