What is Fidelity?

New England Biolabs has been involved in the discovery and development of high fidelity polymerases for many years (Table 1). Beginning with the discovery of Vent® DNA Polymerase from Thermococcus litoralis in 1990 (1,2), NEB researchers have identified and engineered several high fidelity polymerases – all with differing processivities, speeds and degrees of accuracy.

New England Biolabs' High-Fidelity DNA Polymerase History

The fidelity of a DNA polymerase refers to its ability to accurately replicate a template. A critical aspect of this is the ability of the DNA polymerase to read a template strand, select the appropriate nucleoside triphosphate and insert the correct nucleotide at the 3′ primer terminus, such that canonical Watson-Crick base pairing is maintained. The rate of misincorporation (incorporating the incorrect nucleotide) is known as the polymerase's "error rate". In addition to effective discrimination for correct over incorrect nucleotide incorporation, some DNA polymerases possess a 3′→5′ exonuclease activity. This activity, also termed "proofreading", is used to excise incorrectly incorporated mononucleotides that are then replaced with the correct nucleotide. High-fidelity PCR utilizes DNA polymerases that couple low misincorporation rates with proofreading activity to give faithful replication of the DNA target of interest.

(1) Mattila, P., et al. (1991) Nucleic Acids Research, 19, 4967–4973. PMID: 1923765
(2) Eckert, K. A., & Kunkel, T. A. (1991) Genome Research, 1, 17–24. PMID: 1842916

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