ThermalAce™ DNA Polymerase - FAQs

When using a proofreading polymerase such as Pfx, ThermalAce, Pfu, or Vent, is it necessary to neutralize or remove the enzyme after regenerating A-overhangs with a Taq polymerase?

It is best practice to remove the enzymes via phenol-chloroform extraction to prevent the proofreading enzyme from removing the A-overhang again after Taq incubation. If not done, TA cloning efficiencies could be 4-10 fold lower. Alternatively, the PCR product can be cleaned up by gel purification or PCR cleanup column.