A high-fidelity polymerase with gap DNA preference and its application

Abstract

The invention discloses a gap DNA-preferential high-fidelity polymerase and its application. The invention discloses that the high-fidelity polymerization characteristic of the Klenow fragment (KlenDr) derived from the DNA polymerase I of Deinococcus radiodurans is not dependent on the 3'-5' corrective excision activity, and has a preference for binding to nick DNA, different Compatible with existing commercial high-fidelity polymerases. Due to the specific affinity of KlenDr to the gap DNA substrate, the 3' end of the upstream primer will not be excised, and the downstream nucleotide chain will rarely be replaced. It will have a very broad application prospect in genetic engineering operations such as gap filling and the construction of sequencing libraries.

Description

technical field

[0001] The invention belongs to the field of genetic engineering and relates to a gap DNA-preferential high-fidelity polymerase and its application. Background technique

[0002] DNA polymerase has a wide range of applications in genetic engineering operations such as molecular cloning, DNA sequencing, and library construction, and its ability to accurately replicate DNA sequences is crucial. Most of the high-fidelity DNA polymerases currently commercially used rely on the 3'-5' exo-correction activity: when the inserted nucleotide does not conform to the Watson-Crick base pairing principle, the inappropriate spatial conformation will force a new insertion The nucleotides are transferred from the polymerization active center to the 3'-5' excision active center, thereby excising the wrongly inserted nucleotides to ensure the accuracy of DNA replication. At present, there is a lack of high-fidelity polymerases that are not dependent on 3'-5' corrective excisio...

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