We have developed a gold nanoparticle (AuNP)-based CTG repeat probing system displaying high quenching capability and combined it with isothermal amplification for the detection of miRNA 146a. This method of using a AuNP-based CTG repeat probing system with isothermal amplification allowed the highly sensitive (14 aM) and selective detection of miRNA 146a. A AuNP-based CTG repeat probing system having a hairpin structure and a dTF fluorophore exhibited highly efficient quenching because the CTG repeat-based stable hairpin structure imposed a close distance between the AuNP and the dTF residue. A small amount of miRNA 146a induced multiple copies of the CAG repeat sequence during rolling circle amplification; the AuNP-based CTG repeat probing system then bound to the complementary multiple-copy CAG repeat sequence, thereby inducing a structural change from a hairpin to a linear structure with amplified fluorescence. This AuNP-based CTG probing system combined with isothermal amplification could also discriminate target miRNA 146a from one- and two-base-mismatched miRNAs (ORN 1 and ORN 2, respectively). This simple AuNP-based CTG probing system, combined with isothermal amplification to induce a highly sensitive change in fluorescence, allows the detection of miRNA 146a with high sensitivity (14 aM) and selectivity.

中文翻译：

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使用基于金纳米颗粒的 CTG 重复探测系统和等温扩增进行高灵敏度 MicroRNA 146a 检测

我们开发了一种基于金纳米颗粒 (AuNP) 的 CTG 重复探测系统，显示出高淬灭能力，并将其与等温扩增相结合，用于检测 miRNA 146a。这种使用具有等温扩增的基于 AuNP 的 CTG 重复探测系统的方法允许高度灵敏 (14 aM) 和选择性检测 miRNA 146a。具有发夹结构和 dTF 荧光团的基于 AuNP 的 CTG 重复探测系统表现出高效的淬灭，因为基于 CTG 重复的稳定发夹结构在 AuNP 和 dTF 残基之间施加了较近的距离。少量的miRNA 146a在滚环扩增过程中诱导了CAG重复序列的多个拷贝；然后基于 AuNP 的 CTG 重复探测系统与互补的多拷贝 CAG 重复序列结合，从而诱导从发夹结构到具有放大荧光的线性结构的结构变化。这种基于 AuNP 的 CTG 探测系统与等温扩增相结合，还可以将目标 miRNA 146a 与一个和两个碱基错配的 miRNA（分别为 ORN 1 和 ORN 2）区分开来。这种简单的基于 AuNP 的 CTG 探测系统与等温放大相结合以诱导高度灵敏的荧光变化，允许以高灵敏度 (14 aM) 和选择性检测 miRNA 146a。