New high fidelity polymerases from Thermococcus species

Griffiths, Kate; Nayak, Sunil; Park, Kyusung; Mandelman, David; Modrell, Brett; Lee, Jun; Ng, Bernie; Gibbs, Moreland D.; Bergquist, Peter L.

doi:10.1016/j.pep.2006.07.022

Cited by 17 publications

(8 citation statements)

References 34 publications

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“…Other family B archaeal DNA pols from Thermococcus species are claimed to be used in routine PCR: Thermococcus 9°N-7 DNA pol (see below) (Southworth et al 1996), Thermococcus JDF-3 DNA pol (see below), Thermococcus celericrescens (Tcel) DNA pol (Kim et al 2011b), Thermococcus fumicolans (Tfu) DNA pol (CambonBonavita et al 2000), Thermococcus sp. NA1 (TNA1) DNA pol (Kim et al 2007), Thermococcus peptonophilus (Tpe) DNA pol (Lee et al 2010), Thermococcus zilligii (Tzi) DNA pol (Griffiths et al 2007), Thermococcus aggregans (Tag) DNA pol (Bohlke et al. 2000), Thermococcus celer (Tce) DNA pol (Kim et al 2011a), Thermococcus marinus (Tma) DNA pol (Bae et al 2009), Thermococcus pacificus (Tpa) DNA pol (Ppyun et al 2012), Thermococcus thioreducens (Tth) DNA pol (Marsic et al 2008), and Thermococcus waiotapuensis (Twa) DNA pol Cho et al 2014).…”

Section: Routine Pcrmentioning

confidence: 99%

Archaeal DNA polymerases in biotechnology

Zhang

Kang

et al. 2015

Appl Microbiol Biotechnol

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DNA polymerase (pol) is a ubiquitous enzyme that synthesizes DNA strands in all living cells. In vitro, DNA pol is used for DNA manipulation, including cloning, PCR, site-directed mutagenesis, sequencing, and several other applications. Family B archaeal DNA pols have been widely used for molecular biological methods. Biochemical and structural studies reveal that each archaeal DNA pol has different characteristics with respect to fidelity, processivity and thermostability. Due to their high fidelity and strong thermostability, family B archaeal DNA pols have the extensive application on high-fidelity PCR, DNA sequencing, and site-directed mutagenesis while family Y archaeal DNA pols have the potential for error-prone PCR and random mutagenesis because of their low fidelity and strong thermostability. This information combined with mutational analysis has been used to construct novel DNA pols with altered properties that enhance their use as biotechnological reagents. In this review, we focus on the development and use of family B archaeal DNA pols.

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Section: Routine Pcrmentioning

confidence: 99%

Archaeal DNA polymerases in biotechnology

Zhang

Kang

et al. 2015

Appl Microbiol Biotechnol

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“…The importance of proofreading activity has been demonstrated for Thermococcus litoralis (Vent) and Pfu DNA polymerases, which exhibit 5-fold and 7-40-fold increase in error rate, respectively, when 3′ → 5′ exonuclease activity is inactivated [13,21]. The significant increase in error frequency is also observed when 3′ → 5′ exonuclease activity is inactivated in Thermococcus zilligii (Tzi) DNA polymerase [10].…”

Section: Characterization Of Tpe Dna Polymerasementioning

confidence: 99%

“…Among the various applications, the most outstanding is the polymerase chain reaction (PCR) [5], based upon the use of thermostable Taq and Pfu DNA polymerases. Thus, various DNA polymerases from hyperthermophilic archaea have previously isolated and characterized [4,[6][7][8][9][10][11][12][13][14][15][16][17][18][19]. Archaeal DNA polymerases have been mostly identified as members of family B, similar to eukaryotic replicative DNA polymerases [20].…”

mentioning

confidence: 99%

“…Archaeal DNA polymerases have been mostly identified as members of family B, similar to eukaryotic replicative DNA polymerases [20]. Common traits of the family B polymerases include remarkable accuracy during replication and strong 3′ → 5′ exonuclease activity [6,[10][11][12][13][14][15]. The contribution of proofreading activity to DNA polymerase fidelity is also evident when the error rates of proofreading and non-proofreading enzymes are compared [21].…”

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confidence: 99%

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Biochemical Properties and PCR Performance of a Family B DNA Polymerase from Hyperthermophilic Euryarchaeon Thermococcus peptonophilus

Lee

Kim

Bae

et al. 2009

Appl Biochem Biotechnol

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The Thermococcus peptonophilus (Tpe) DNA polymerase gene was expressed under the control of the T7lac promoter on pET-22b(+) in Escherichia coli BL21-CodonPlus(DE3)-RIL in order to fully elucidate its biochemical properties and evaluate its feasibility in polymerase chain reaction (PCR) application. The expressed enzyme was then purified by heat treatment followed by two steps of column chromatography after which optimum pH and temperature of the enzyme were evaluated to be 7.0 and 75 degrees C, respectively. The optimal buffer for PCR with Tpe DNA polymerase consisted of 50 mM Tris-HCl (pH 8.0), 2 mM MgCl(2), 80 mM KCl, and 0.02% Triton X-100. Tpe DNA polymerase revealed a 3.6-fold higher fidelity (3.37 x 10(-6)) than Taq DNA polymerase (12.13 x 10(-6)) and performed significantly more efficiently in PCR amplification than both Taq and Pfu DNA polymerases. Ratios of 31:1 of Taq to Tpe DNA polymerases allowed PCR amplification of targets up to 15 kb in length with a 2.2-fold higher fidelity than Taq DNA polymerase. The results of the PCR experiments indicate that Tpe DNA polymerase may provide a higher fidelity DNA amplification in a shorter reaction time.

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“…The majority of archaeal DNA polymerases have been identified as a member of family B, which are enzymes possessing 3 → 5 exonuclease activity (proofreading activity; editing ability) (Griffiths et al, 2007;Lundberg et al, 1991;Takagi et al, 1997) and which have the ability to correct errors introduced during polymerization. To improve the low fidelity of PCR with Taq polymerase, archaeal family B DNA polymerases have been utilized.…”

Section: Introductionmentioning

confidence: 99%