Autonomous Replication of Nucleic Acids by Polymerization/Nicking Enzyme/DNAzyme Cascades for the Amplified Detection of DNA and the Aptamer鈥揅ocaine Complex
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- 作者：Fuan Wang ; Lina Freage ; Ron Orbach ; Itamar Willner
- 刊名：Analytical Chemistry
- 出版年：2013
- 出版时间：September 3, 2013
- 年：2013
- 卷：85
- 期：17
- 页码：8196-8203
- 全文大小：524K
- 年卷期：v.85,no.17(September 3, 2013)
- ISSN：1520-6882
文摘
The progressive development of amplified DNA sensors and aptasensors using replication/nicking enzymes/DNAzyme machineries is described. The sensing platforms are based on the tailoring of a DNA template on which the recognition of the target DNA or the formation of the aptamer鈥搒ubstrate complex trigger on the autonomous isothermal replication/nicking processes and the displacement of a Mg2+-dependent DNAzyme that catalyzes the generation of a fluorophore-labeled nucleic acid acting as readout signal for the analyses. Three different DNA sensing configurations are described, where in the ultimate configuration the target sequence is incorporated into a nucleic acid blocker structure associated with the sensing template. The target-triggered isothermal autonomous replication/nicking process on the modified template results in the formation of the Mg2+-dependent DNAzyme tethered to a free strand consisting of the target sequence. This activates additional template units for the nucleic acid self-replication process, resulting in the ultrasensitive detection of the target DNA (detection limit 1 aM). Similarly, amplified aptamer-based sensing platforms for cocaine are developed along these concepts. The modification of the cocaine-detection template by the addition of a nucleic acid sequence that enables the autonomous secondary coupled activation of a polymerization/nicking machinery and DNAzyme generation path leads to an improved analysis of cocaine (detection limit 10 nM).