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A Novel Low Temperature PCR Assured High-Fidelity DNA Amplification
State Key Laboratory of Molecular Biology, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 320 Yueyang Road, Shanghai 200031, China
Milford Hospital and Milford Molecular Laboratory, 2044 Bridgeport Avenue, Milford, CT 06460, USA
These authors contributed equally to this work.
* Author to whom correspondence should be addressed.
Received: 17 May 2013; in revised form: 5 June 2013 / Accepted: 7 June 2013 / Published: 20 June 2013
Abstract: As previously reported, a novel low temperature (LoTemp) polymerase chain reaction (PCR) catalyzed by a moderately heat-resistant (MHR) DNA polymerase with a chemical-assisted denaturation temperature set at 85 °C instead of the conventional 94–96 °C can achieve high-fidelity DNA amplification of a target DNA, even after up to 120 PCR thermal cycles. Furthermore, such accurate amplification is not achievable with conventional PCR. Now, using a well-recognized L1 gene segment of the human papillomavirus (HPV) type 52 (HPV-52) as the template for experiments, we demonstrate that the LoTemp high-fidelity DNA amplification is attributed to an unusually high processivity and stability of the MHR DNA polymerase whose high fidelity in template-directed DNA synthesis is independent of non-existent 3'–5' exonuclease activity. Further studies and understanding of the characteristics of the LoTemp PCR technology may facilitate implementation of DNA sequencing-based diagnostics at the point of care in community hospital laboratories.
Keywords: low temperature PCR; high-fidelity DNA amplification; LoTemp PCR; high processivity
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Hong, G.; Lee, S.H.; Ge, S.; Zhou, S. A Novel Low Temperature PCR Assured High-Fidelity DNA Amplification. Int. J. Mol. Sci. 2013, 14, 12853-12862.
Hong G, Lee SH, Ge S, Zhou S. A Novel Low Temperature PCR Assured High-Fidelity DNA Amplification. International Journal of Molecular Sciences. 2013; 14(6):12853-12862.
Hong, Guofan; Lee, Sin H.; Ge, Shichao; Zhou, Shaoxia. 2013. "A Novel Low Temperature PCR Assured High-Fidelity DNA Amplification." Int. J. Mol. Sci. 14, no. 6: 12853-12862.