Fluorometric determination of microRNA using arched probe-mediated isothermal exponential amplification combined with DNA-templated silver nanoclusters

被引：22

作者：

Wu, Hao ^{[1
]}

Wu, Jun ^{[1
]}

Liu, Yaling ^{[1
]}

Wang, Hongyong ^{[1
]}

Zou, Pei ^{[1
]}

机构：

[1] Jiangsu Inst Nucl Med, Minist Hlth, Jiangsu Key Lab Mol Nucl Med, Key Lab Nucl Med, Wuxi 214063, Jiangsu, Peoples R China

来源：

MICROCHIMICA ACTA | 2019年 / 186卷 / 11期

关键词：

MicroRNA-141; Arched probe; Silver nanoclusters; Isothermal amplification; DNA polymerase; Nicking endonuclease; Human serum; QUANTIFICATION;

D O I：

10.1007/s00604-019-3836-4

中图分类号：

O65 [分析化学];

学科分类号：

070302 ; 081704 ;

摘要：

A highly sensitive fluorometric method is described for the determination of microRNA-141. It is based on the use of arched probe-mediated isothermal exponential amplification reaction (EXPAR) and of DNA-templated silver nanoclusters (DNA-AgNCs). The EXPAR utilizes microRNA-141 as the trigger, polymerases and endonucleases as amplification activators, and two arched probes as exponential amplification templates. This enables the conversion of microRNA to a large number of reporter sequences under isothermal conditions within minutes. The generated reporter sequences act as scaffolds for the synthesis of fluorescent DNA-AgNCs by reduction of Ag (I) with NaBH4. The DNA-AgNCs function as signalling fluorophores with excitation/emission maxima at 540/610 nm. The method exhibits high sensitivity for microRNA-141 with a detection limit as low as 0.87 fM and a dynamic range from 1 fM to 500 fM. The method can distinguish nucleotides in the microRNA-200 family.

引用

页数：8

共 33 条

[1] MicroRNAs: Genomics, biogenesis, mechanism, and function (Reprinted from Cell, vol 116, pg 281-297, 2004) [J].