A Comprehensive Review of Pfu DNA Polymerase: Extraction, Production and Applications

Pfu DNA Polymerase, a thermostable enzyme derived from the hyperthermophilic archaeon Pyrococcus furiosus, is widely recognized for its high fidelity and strong processivity. Its 3'-5' exonuclease activity makes it indispensable for the correct amplification of both short and complex DNA strands. These biochemical properties of Pfu DNA Polymerase have encouraged considerable advancements in its extraction and production methods. This review covers some of the traditional approaches for purification, including protein purification and affinity chromatography, and updates on recent advances in recombinant gene expression, automated systems of production, and membrane-based technologies. New ways of engineering enzymes have recently been developed, such as CRISPR-Cas9-mediated gene optimization, which raised the bar on extraction efficiency to meet emerging demand. Once challenging, Pfu DNA Polymerase production has been significantly streamlined through recombinant expression in E. coli both at a laboratory and commercial scale. The optimization techniques that are involved with IPTG concentration and Response Surface Methodology have increased the yield by up to 30%. Auto-induction means even higher biomass outputs are allowed. Today, applications of Pfu DNA Polymerase span from standard PCR up to advanced clinical diagnostics in the fields of molecular biology, forensic analysis, clinical microbiology, and biotechnology.