Helicase-dependent isothermal DNA amplification

被引：649

作者：

Myriam Vincent ^{[1
]}

Yan Xu ^{[1
]}

Huimin Kong ^{[1
]}

机构：

[1] New England Biolabs, Beverly, MA 01915

来源：

The EMBO Reports | 2004年 / 5卷 / 8期

关键词：

DNA amplification; DNA polymerase; Helicae; Isothermal; UvrD;

D O I：

10.1038/sj.embor.7400200

中图分类号：

学科分类号：

摘要：

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. © 2004 EUROPEAN MOLECULAR BIOLOGY ORGANIZATION.

引用

页码：795 / 800

页数：5

共 21 条

[1]

Ali J.A., Maluf N.K., Lohman T.M., An oligomeric form of E. coli UvrD is required for optimal helicase activity, J. Mol. Biol., 293, pp. 815-834, (1999)

[2]

Andras S.C., Power J.B., Cocking E.C., Davey M.R., Strategies for signal amplification in nucleic acid detection, Mol. Biotechnol., 19, pp. 29-44, (2001)

[3]

Brownie J., Shawcross S., Theaker J., Whitcombe D., Ferrie R., Newton C., Little S., The elimination of primer-dimer accumulation in PCR, Nucleic Acids Res., 25, pp. 3235-3241, (1997)

[4]

Bujalowski W., Lohman T.M., Negative co-operativity in Escherichia coli single strand binding protein-oligonucleotide interactions. II. Salt, temperature and oligonucleotide length effects, J. Mol. Biol., 207, pp. 269-288, (1989)

[5]

Caruthers J.M., McKay D.B., Helicase structure and mechanism, Curr. Opin. Struct. Biol., 12, pp. 123-133, (2002)

[6]

Casas-Finet J.R., Karpel R.L., Bacteriophage T4 gene 32 protein: Modulation of protein-nucleic acid and protein-protein association by structural domains, Biochemistry, 32, pp. 9735-9744, (1993)

[7]

Chong S., Et al., Utilizing the C-terminal cleavage activity of a protein splicing element to purify recombinant proteins in a single chromatographic step, Nucleic Acids Res., 26, pp. 5109-5115, (1998)

[8]

Desplats C., Dez C., Tetart F., Eleaume H., Krisch H.M., Snapshot of the genome of the pseudo-T-even bacteriophage RB49, J. Bacteriol., 184, pp. 2789-2804, (2002)

[9]

Fire A., Xu S.Q., Rolling replication of short DNA circles, Proc. Natl. Acad. Sci. USA, 92, pp. 4641-4645, (1995)

[10]

Ghedin E., Wang S., Foster J.M., Slatkon B.E., First sequenced genome of a parasitic nematode, Trends Parasitol., 20, pp. 151-153, (2004)

← 1 2 3 →