Pfu DNA Polymerase - Proofreading Ability of Pfu Polymerase

Proofreading Ability of Pfu Polymerase

Pfu DNA polymerase has superior thermostability and proofreading properties compared to Taq DNA polymerase. Unlike Taq DNA polymerase, Pfu DNA polymerase possesses 3' to 5' exonuclease proofreading activity, meaning that it works its way along the DNA from the 5' end to the 3' end and corrects nucleotide-misincorporation errors. This means that Pfu DNA polymerase-generated PCR fragments will have fewer errors than Taq-generated PCR inserts.

Commercially available Pfu typically results in an error rate of 1 in 1.3 million base pairs and can yield 2.6% mutated products when amplifying 1 kb fragments using PCR. However, Pfu is slower and typically requires 1–2 minutes per cycle to amplify 1kb of DNA at 72 °C. Using Pfu DNA polymerase in PCR reactions also results in blunt-ended PCR products.

Pfu DNA polymerase is hence superior to Taq DNA polymerase for techniques that require high-fidelity DNA synthesis, but can also be used in conjunction with Taq polymerase to obtain the fidelity of Pfu with the speed of Taq polymerase activity.