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URL: https://dx.doi.org/DOI:10.17504/protocols.io.7kkhkuw

Authors: Alba Balletbó

Group: iGEM Wageningen 2019

Summary: Thermo Scientific™ Phire™ Hot Start II DNA Polymerase is a novel DNA polymerase designed for use in all routine and high throughput PCR applications. A special DNA binding domain linked to the Phire Hot Start II DNA Polymerase enhances the processivity of the polymerase, enabling short extension times and improved yields. The polymerase is also capable of amplifying long DNA fragments, such as the 7.5 kb genomic DNA used in Thermo Scientific quality control assays. Phire Hot Start II DNA Polymerase provides 2-fold fidelity compared to Taq DNA polymerase. The hot start modification of the polymerase is based on the Affibody™ inactivation method1,2 . It inhibits DNA polymerase activity at ambient temperatures, thus preventing amplification of non-specific products. At polymerization temperatures the Affibody molecule is released, rendering the polymerase fully active. Phire Hot Start II DNA Polymerase generates blunt ends in the amplification products. It does not possess the 5’→3’ exonuclease activity needed for hydrolysis experiments.

Affiliations: Wageningen University

Version: 1

Publication Date: 2019

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