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>A Novel Ultrasensitive ECL Sensor for DNA Detection Based on Nicking Endonuclease-Assisted Target Recycling Amplification, Rolling Circle Amplification and Hemin/G-Quadruplex
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A Novel Ultrasensitive ECL Sensor for DNA Detection Based on Nicking Endonuclease-Assisted Target Recycling Amplification, Rolling Circle Amplification and Hemin/G-Quadruplex
In this study, we describe a novel universal and highly sensitive strategy for the electrochemiluminescent (ECL) detection of sequence specific DNA at the aM level based on Nt.BbvCI (a nicking endonuclease)-assisted target recycling amplification (TRA), rolling circle amplification (RCA) and hemin/G-quadruplex. The target DNAs can hybridize with self-assembled capture probes and assistant probes to form “Y” junction structures on the electrode surface, thus triggering the execution of a TRA reaction with the aid of Nt.BbvCI. Then, the RCA reaction and the addition of hemin result in the production of numerous hemin/G-quadruplex, which consume the dissolved oxygen in the detection buffer and result in a significant ECL quenching effect toward the O2/S2O82− system. The proposed strategy combines the amplification ability of TRA, RCA and the inherent high sensitivity of the ECL technique, thus enabling low aM (3.8 aM) detection for sequence-specific DNA and a wide linear range from 10.0 aM to 1.0 pM. At the same time, this novel strategy shows high selectivity against single-base mismatch sequences, which makes our novel universal and highly sensitive method a powerful addition to specific DNA sequence detection.
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机译:在这项研究中,我们描述了一种基于Nt.BbvCI(切口内切核酸酶)辅助靶标循环扩增(TRA),滚环扩增的电化学发光(ECL)在aM水平上检测序列特异性DNA的新型通用且高度灵敏的策略(RCA)和血红素/ G四联体。靶DNA可以与自组装的捕获探针和辅助探针杂交以在电极表面上形成“ Y”连接结构,从而在Nt.BbvCI的帮助下触发TRA反应的执行。然后,RCA反应和血红素的添加导致产生大量的血红素/ G-四链体,这些消耗了检测缓冲液中的溶解氧,并对O 2 sub>产生了显着的ECL猝灭作用/ S 2 sub> O 8 sub> 2- sup>系统。所提出的策略结合了TRA,RCA的扩增能力和ECL技术固有的高灵敏度,从而实现了针对序列特异性DNA的低aM(3.8 aM)检测以及从10.0 aM到1.0 pM的宽线性范围。同时,这种新颖的策略显示出对单碱基错配序列的高度选择性,这使我们新颖且通用的高灵敏度方法成为特定DNA序列检测的有力补充。
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