When should I use proofreading enzymes such as Pfu, Pwo, Vent or Deep Vent?
If you need a short fragment from a plasmid, use a high template ( i.e ., ng of plasmid) and keep the cycle number low (10 or 15), then it is fine to use proofreading DNA polymerases. However, you can not assume that the PCR product will have 100% fidelity just by using a proofreading DNA polymerase. No DNA polymerase is perfect. That is, when fidelity is important you still have to sequence your clone no matter how accurate the proofreading DNA polymerase is.
