核酸恒温扩增技术(HDA).doc

EMBO Rep. 2004 August; 5(8): 795–800. Published online 2004 July 9. doi: 10.1038/sj.embor.7400200. PMCID: PMC1249482 Copyright ? 2004, European Molecular Biology Organization Scientific Report Helicase-dependent isothermal DNA amplification Myriam Vincent,1* Yan Xu,1* and Huimin Kong1a 1New England Biolabs, 32 Tozer Road, Beverly, Massachusetts 01915, USA aTel: +1 978 927 5054; Fax: +1 978 921 1350; E-mail: kong@ *These authors contributed equally to this work Received January 14, 2004; Revised May 24, 2004; Accepted June 14, 2004. This article has been cited by other articles in PMC. ?Other Sections▼ Abstract Introduction Results Discussion Methods Supplementary Material References Abstract Polymerase chain reaction is the most widely used method for in vitro DNA amplification. However, it requires thermocycling to separate two DNA strands. In vivo, DNA is replicated by DNA polymerases with various accessory proteins, including a DNA helicase that acts to separate duplex DNA. We have devised a new in vitro isothermal DNA amplification method by mimicking this in vivo mechanism. Helicase-dependent amplification (HDA) utilizes a DNA helicase to generate single-stranded templates for primer hybridization and subsequent primer extension by a DNA polymerase. HDA does not require thermocycling. In addition, it offers several advantages over other isothermal DNA amplification methods by having a simple reaction scheme and being a true isothermal reaction that can be performed at one temperature for the entire process. These properties offer a great potential for the development of simple portable DNA diagnostic devices to be used in the field and at the point-of-care. Keywords: DNA amplification, isothermal, helicase, DNA polymerase, UvrD ?Other Sections▼ Abstract Introduction Results Discussion Methods Supplementary Material References Introduction The polymerase chain reaction (PCR) revolutionized our capabilities to do biological research, and it has b