High-fidelity PCR is important for several applications including cloning, genetic profiling, and next-generation sequencing. Routine high-fidelity amplification of longer and difficult targets with wild-type proofreading polymerases is challenging due to the low processivity and lack of robustness of these enzymes. Researchers often resort to blended enzyme systems (Taq polymerase combined with a low proportion of a proofreading DNA polymerase), with which high yields can be obtained from complex and/or low copy templates.
However, the robustness of enzyme blends comes at the cost of fidelity: error rates are typically only 2X – 4X lower than of wild-type Taq, and PCR-induced mutations are common. Single enzyme, wild-type proofreading polymerases are capable of error rates ~20X lower than Taq; however, many are notorious for low sensitivity, poor priming specificity, long extension times, and low yield.
KAPA HiFi DNA Polymerase is a novel enzyme engineered specifically for high-fidelity PCR, offering true high fidelity of single-enzyme proofreading polymerases combined with the robustness, sensitivity and ability to amplify long fragments of enzyme blends. KAPA HiFi DNA Polymerase was evolved to have an increased affinity for DNA without the need for accessory protein domains. The intrinsic high processivity of KAPA HiFi DNA Polymerase results in significant improvements to yield, sensitivity, target length, and the ability to amplify difficult templates.