Taq polymerase trivia

Dr. Duncan Clark Duncan at nospam.demon.co.uk
Sun Nov 28 05:48:33 EST 1999


In article <tchappell-2811990007020001 at sdts10-77.znet.net>, tom chappell
<tchappell at znet.com> writes
>Vent

I hate to be picky and awkward but Vent has 3-5' exo activity and
therefore shouldn't put on the overhang.


Quick overview.

The pols currently used in PCR applications fall into two main groups:

1.      Thermophilic prokaryotes - polA like polymerases.

Type A i.e. Taq, Tth, Tfl, Tth, Tbr

All isolated from Thermus sp. Have 5-3' exo activity and are missing a
few crucial aa's in their 3-5' exo domain (hence they are 3-5' exo
minus) and pol. Activity. Some have RT activity. All have 3' terminal
transferase activity. Cut down variants like Stoffel, are deleted for
the 5-3' exo activity. 

Type B  ie. Tma.

Isolated from Thermotoga sp, Thermosipho sp., Carboxydothermus
hydrogenoformans  etc. Have both 5-3' and 3-5' exo activity and pol
activity. Ultma, as sold by PE Biosystems, is not a full length enzyme
of Tma. It retains 3-5' exo but no 5-3' exo. It's fidelity has been
reported however to be very poor. C.hydrogenoformans has good RT
activity

2.      Hyperthermophilic archaebacteria - polB like polymerases

i.e. Pfu, Pwo, Vent, 9N7, Deep Vent.

Isolated from Pyrococcus sp. or Thermococcus sp.  All have 3-5' exo and
pol activity. No associated RT. Hate dUTP in primers or in PCR. No 5-3'
exo activity. 3-5' exo minus derivatives. Debate over which have 3'
terminal transferase activity. 


Not a group but mixes of both Group 1 and Group 2 are sold so you get
all the activities in one enzyme mix. 

Finally don't expect too much from these enzymes. They are most likeley
only repair type enzymes after all and are therefore limited in speed
and fidelity.

Duncan
-- 
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
http://www.dnamp.com
http://www.genesys.demon.co.uk




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