A target-triggered dual amplification strategy for sensitive detection of microRNA
A target-triggered dual amplification strategy for sensitive detection of microRNA
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一种用于灵敏检测 microRNA 的靶标触发双扩增策略
DOI：
10.1016/j.bios.2016.04.053
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发表时间：
2016-09-15
影响因子：
12.6
通讯作者：
Zheng, Lei
中科院分区：
文献类型：
--
作者：
Lv, Weifeng;Zhao, Jiamin;Zheng, Lei
microRNA (miRNA) 表达的准确定量分析对于生物医学研究和临床治疗诊断至关重要。在这项研究中，我们报告了一种基于双链体特异性核酸酶（DSN）辅助双信号放大策略的用于灵敏检测 miRNA 的新型传感器。设计并合成了由miRNA识别DNA序列和Taqman探针杂交RNA序列（2'-O-甲基RNA）组成的嵌合探针（DNA/2-OMe-RNA）。在第一轮扩增中，目标 miRNA 的一个分子可以触发嵌合探针的循环切割，通过 DSN 产生 2'-O-甲基 RNA。 2'-O-甲基 RNA 分子随后可以与 Taqman 探针杂交并启动第二轮循环扩增，以产生 DSN 可检测到的荧光。所提出的策略在区分同源 miRNA 方面表现出高度特异性，并且双重信号转导过程能够检测低至 7.3 fM 的 miRNA 浓度。我们进一步将该测定应用于癌细胞中的 miRNA 定量，以证实其适用性。本研究为miRNA检测提供了一种灵敏、特异、简单的方法，在生物医学研究和临床实验室中具有巨大的进一步应用潜力。 (C) 2016 Elsevier B.V. 保留所有权利。
The accurate and quantitative analysis of microRNA (miRNA) expression is critical for biomedical research and clinical theranostics. In this study, we report a novel sensor for the sensitive detection of miRNA based on a duplex-specific nuclease (DSN)-assisted dual signal amplification strategy. A chimeric probe (DNA/2-OMe-RNA) that consists of a miRNA recognition DNA sequence and a Taqman probe hybridization RNA sequence (2'-O-methyl RNA) was designed and synthesized. One molecule of target miRNA can trigger cyclical cleavage of the chimeric probes to produce 2'-O-methyl RNA by DSN in the first round of amplification. The 2'-O-methyl RNA molecules can subsequently hybridize with Taqman probes and initiate the second round of cyclical amplification to generate detectable fluorescence by DSN. The proposed strategy exhibits high specificity in discriminating cognate miRNAs, and the dual signal transduction process enables the detection of miRNA concentrations as low as 7.3 fM. We further applied this assay to miRNA quantification in cancer cells to confirm its applicability. The present study provides a sensitive, specific and simple method for miRNA detection and holds great potential for further application in biomedical research and in the clinical laboratory. (C) 2016 Elsevier B.V. All rights reserved.