A-tailing with Taq
Patrick F.H. Lai
pfhlai at LOOKSMART.COM
Mon Aug 10 20:29:58 EST 1998
---- Begin Original Message ----
> I have always read that Taq has a preference for adding an extra >"=
A" at the end of a PCR product, however, does anyone know >semi-quantita=
>(for example, a number between 25% and 100%) how often this happens? >=
I was also wondering if I should add some extra dATP at the end of my >P=
CR reaction, and let it tail for 10' at 72C, rather than just >letting t=
he reaction "tail" for 10' at 72C at the end of the PCR.
> Any suggestions?
><kunitake at salk.edu>
---- End Original Message ----
Dear Dr. Koichi Kunitake,
For the purpose of TA-cloning into pGEM-T Easy, I never had to do
extra work. The 10 min of 72 deg C incubation after 25~35 cycles
was sufficient for my cloning purposes.
I tried Taq polymerase from BM and Pharmacia, and both worked for me.
However, if your DNA fragment is really AT-rich, adding extra dATP
does not sound like a bad idea.
Don't worry, my fellow researcher.
TA cloning works, especially when you are using kits.
As long as you put enough inserts in your ligation reaction,
I am sure you will get more big, white colonies than you want on your
(However, if you are T-tailing your own vector,
then it may be another story.)
Good luck. :-)
Hope this is useful info. :-)
Patrick F.H. Lai < PFHLai at looksmart.com >
University of Toronto
Toronto, Ontario, Canada
LookSmart =85 or keep looking.
More information about the Methods