Highly Efficient Isothermal DNA Amplification System Using Three Elements of 5'-DNA-RNA-3' Chimeric Primers, RNaseH and Strand-displacing DNA Polymerase

We developed an efficient method of isothermally amplifying DNA termed ICAN, Isothermal and Chimeric primer-initiated Amplification of Nucleic acids. This method allows the amplification of target DNA under isothermal conditions at around 55°C using only a pair of 5'-DNA-RNA-3' chimeric pr...

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Bibliographic Details
Published in:Journal of biochemistry (Tokyo) 2007-08, Vol.142 (2), p.273-281
Main Authors: Mukai, Hiroyuki, Uemori, Takashi, Takeda, Osamu, Kobayashi, Eiji, Yamamoto, Junko, Nishiwaki, Kazue, Enoki, Tatsuji, Sagawa, Hiroaki, Asada, Kiyozo, Kato, Ikunoshin
Format: Article
Language:English
Subjects:
chimeric primer
detection
DNA - metabolism
DNA polymerase
DNA Primers - chemistry
DNA-Directed DNA Polymerase - metabolism
isothermal DNA amplification
Nucleic Acid Amplification Techniques - methods
Ribonuclease H - chemistry
Ribonuclease H - metabolism
RNA - chemistry
RNaseH
Temperature
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Summary:We developed an efficient method of isothermally amplifying DNA termed ICAN, Isothermal and Chimeric primer-initiated Amplification of Nucleic acids. This method allows the amplification of target DNA under isothermal conditions at around 55°C using only a pair of 5'-DNA-RNA-3' chimeric primers, a thermostable RNaseH and a DNA polymerase with strong strand-displacing activity. ICAN is capable of amplifying DNA at least several times greater than the amount produced with PCR by increasing primer concentration. This method would be applicable for on-site DNA detection including gene diagnosis, and would also be suitable for 'real time' detection when combined with a cycling probe.
ISSN:0021-924X
1756-2651
DOI:10.1093/jb/mvm138