Nicking Enzyme Amplification Reaction

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The Nicking Enzyme Amplification Reaction (German, single strand break enzyme amplification reaction , NEAR) is a biochemical method for the duplication ( amplification ) of DNA . It is a variant of isothermal DNA amplification .

principle

The nicking enzyme amplification reaction uses a DNA polymerase with DNA double strand separating properties at constant temperature (isothermal), which during the synthesis of a second DNA strand separates the existing double strand and displaces the existing second strand. Furthermore, an endonuclease such as Nt.BstNBI (a nicking enzyme ) is used to generate single-strand breaks in double-stranded DNA.

Alternative methods of amplifying DNA are e.g. B. the polymerase chain reaction , the multidisplacement amplification , the isothermal assembly , the loop-mediated isothermal amplification (LAMP), nucleic acid sequence-based amplification ( NASBA ), the helicase-dependent amplification (HDA), the recombinase polymerase amplification , the rolling circle replication (RCA). Further detection methods are e.g. B. Nicking endonuclease signal amplification ( NESA ) and nicking endonuclease assisted nanoparticle activation ( NENNA ), exonuclease-aided target recycling , junction or Y-probes , split DNAZyme and deoxyribozyme amplification , non-covalent DNA catalysis and the hybridization chain reaction ( HCR).

Applications

NEAR is used to amplify DNA, including labeled oligonucleotides .

Individual evidence

  1. ↑ Patent US7972820 : Isothermal amplification of nucleic acids on a solid support. Published on January 13, 2011 . ‌
  2. ↑ EC Oriero, J. Jacobs, JP Van Geertruyden, D. Nwakanma, U. D'Alessandro: Molecular-based isothermal tests for field diagnosis of malaria and their potential contribution to malaria elimination. In: Journal of Antimicrobial Chemotherapy . [Electronic publication before going to press] September 2014, doi: 10.1093 / jac / dku343 , PMID 25223973 .
  3. ↑ L. Yan, J. Zhou, Y. Zheng, AS Gamson, BT Roembke, S. Nakayama, HO Sintim: Isothermal amplified detection of DNA and RNA. In: Molecular bioSystems. Volume 10, Number 5, May 2014, pp. 970-1003, doi: 10.1039 / c3mb70304e , PMID 24643211 .
  4. ↑ P. Ménová, V. Raindlová, M. Hocek: Scope and limitations of the nicking enzyme amplification reaction for the synthesis of base-modified oligonucleotides and primers for PCR. In: Bioconjugate Chemistry . Volume 24, Number 6, June 2013, pp. 1081-1093, doi: 10.1021 / bc400149q , PMID 23682869 .
  5. ↑ D. Dziuba, R. Pohl, M. Hocek: Bodipy-Labeled Nucleoside Triphosphates for Polymerase Synthesis of Fluorescent DNA. In: Bioconjugate Chemistry. Volume 25, Number 11, November 2014, pp. 1984-1995, doi: 10.1021 / bc5003554 , PMID 25290695 .