Mining and characterization of new enzymes based on Phi29 DNA polymerase.

Mengyao HAO; Lingling HU; Minghao HAN; Congyu LI; Hong CHANG; Jianmei LUO; Huifeng JIANG

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Mining and characterization of new enzymes based on Phi29 DNA polymerase.

Author: Mengyao HAO ¹ ; Lingling HU ¹ ; Minghao HAN ¹ ; Congyu LI ¹ ; Hong CHANG ¹ ; Jianmei LUO ¹ ; Huifeng JIANG ²
Author Information

1. School of Biological Engineering, Tianjin University of Science & Technology, Tianjin 300457, China.
2. Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.
Publication Type:Journal Article
Keywords: Phi29 DNA polymerase; exonuclease activity; heterologous expression; mining of new enzymes; rolling circle amplification
MeSH: DNA-Directed DNA Polymerase/metabolism*; Bacillus Phages/genetics*; Metagenome
From: Chinese Journal of Biotechnology 2025;41(1):427-436
CountryChina
Language:Chinese
Abstract: In recent years, the bacteriophage Φ29 (Phi29) DNA polymerase has garnered increasing attention due to its high-fidelity amplification capacity at constant temperatures. To advance the industrial application of this type of isothermal polymerases, this study mined and characterized new enzymes from the microbial metagenome based on the known Phi29 DNA polymerase sequence. The results revealed that a new enzyme, Php29 DNA polymerase, was identified in the microbial metagenome with plants as the hosts. This enzyme exhibited higher strand displacement activity, with a 59.5% similarity to bacteriophage Φ29. Experimental validation demonstrated that the enzyme had 3'→5' exonuclease activity, and its amplification products can serve as substrates for further catalytic reactions. The discovery and validation of Php29 DNA polymerase gives insights into the future industrial application of isothermal polymerases.