Zhang Xiao-Long, Yang Zhe-Han, Chang Yuan-Yuan, Liu Di, Li Yun-Rui, Chai Ya-Qin, Zhuo Ying, Yuan Ruo
Key Laboratory of Luminescent and Real-Time Analytical Chemistry , Ministry of Education , College of Chemistry and Chemical Engineering , Southwest University , Chongqing 400715 , China . Email:
Chem Sci. 2019 Nov 7;11(1):148-153. doi: 10.1039/c9sc05084a. eCollection 2020 Jan 7.
Herein, by directly introducing mismatched reactant DNA, high-reactivity and high-threshold enzyme-free target recycling amplification (EFTRA) is explored. The developed high-efficiency EFTRA (HEEFTRA) was applied as a programmable DNA signal converter, possessing higher conversion efficiency than the traditional one with perfect complement owing to the more negative reaction standard free energy (Δ). Once traces of input target miRNA interact with the mismatched reactant DNA, amounts of ferrocene (Fc)-labeled output DNA could be converted the EFTRA. Impressively, the Fc-labeled output DNA could be easily captured by the DNA tetrahedron nanoprobes (DTNPs) on the electrode surface to form triplex-forming oligonucleotide (TFO) at pH = 7.0 for sensitive electrochemical signal generation and the DTNPs could be regenerated at pH = 10.0, from which the conversion efficiency () will be accurately obtained, benefiting the selection of suitable mismatched bases to obtain high-efficiency EFTRA (HEEFTRA). As a proof of concept, the HEEFTRA as an evolved DNA signal converter is successfully applied for the ultrasensitive detection of miRNA-21, which gives impetus to the design of other signal converters with excellent efficiency for ultimate applications in sensing analysis, clinical diagnosis, and other areas.
在此，通过直接引入错配的反应物DNA，探索了高反应性和高阈值的无酶靶标循环扩增（EFTRA）。所开发的高效EFTRA（HEEFTRA）被用作可编程DNA信号转换器，由于反应标准自由能（Δ）更负，其转换效率高于具有完美互补性的传统转换器。一旦痕量的输入靶标miRNA与错配的反应物DNA相互作用，大量二茂铁（Fc）标记的输出DNA就可以通过EFTRA进行转换。令人印象深刻的是，Fc标记的输出DNA可以很容易地被电极表面的DNA四面体纳米探针（DTNP）捕获，在pH = 7.0时形成三链形成寡核苷酸（TFO）以产生灵敏的电化学信号，并且DTNP可以在pH = 10.0时再生，由此可以准确获得转换效率（），这有利于选择合适的错配碱基以获得高效EFTRA（HEEFTRA）。作为概念验证，HEEFTRA作为一种改进的DNA信号转换器成功应用于miRNA-21的超灵敏检测，这为设计其他具有优异效率的信号转换器以推动传感分析、临床诊断和其他领域的最终应用提供了动力。
