Herein, we have designed a unique oligonucleotide probe named as bumped switchable molecular probe (BS-MB) and employ it to ultrasensitive detection of miRNA-21. The uniqueness is that the bumped portion renders the BS-MB great design flexibility and divides its long stem into two short stems so that it can play the roles of recognition unit, reporting unit, and polymerization primer and template simultaneously. The opening of BS-MB requires a delayed reaction, allowing the system a better-suppressed background without miRNA-21. In contrast, the introduction of miRNA-21 delayed the full opening of BS-MB, inducing a concurrent three-stage amplification for consuming lots of BS-MBs in one amplification event. During the amplification, lots of target analogues of extended miRNA-21 (E-miRNA-21) and cleavaged miRNA-21 (C-miRNA-21) that can be reused as miRNA-21 to cause further fluorescence enhancement are repeatedly generated. The system retains the simplicity (one oligonucleotide probe) and simplifies the detection procedure (one-step detection). Its mix-to-signaling and one-to-many amplification behaviors are superior to conventional MB based amplification, exhibiting a wide linear response and a low detection limit of 8.1 fM. High specificity against even one-base mutation, desirable recovery rates for testing miRNA-spiked serum samples, and well-defined accuracy for classifying clinically related samples are also demonstrated.

在本文中，我们设计了一种独特的寡核苷酸探针，称为碰撞开关分子探针（BS-MB），并将其用于miRNA-21的超灵敏检测。独特之处在于，隆起部分赋予BS-MB极大的设计灵活性，并将其长茎分为两个短茎，因此它可以同时扮演识别单元，报告单元以及聚合引物和模板的角色。BS-MB的打开需要延迟的反应，从而使系统在没有miRNA-21的情况下具有更好的抑制背景。相反，miRNA-21的引入延迟了BS-MB的完全打开，从而在一个扩增事件中诱导了同时进行的三阶段扩增，从而消耗了大量的BS-MB。在放大过程中 重复生成了许多扩展的miRNA-21（E-miRNA-21）和裂解的miRNA-21（C-miRNA-21）靶标类似物，它们可以作为miRNA-21重复使用，从而进一步增强荧光。该系统保留了简单性（一个寡核苷酸探针）并简化了检测程序（一步检测）。它的混合信号传输和一对多扩增行为优于传统的基于MB的扩增，表现出宽线性响应和低至8.1 fM的检测限。还展示了针对一碱基突变的高特异性，测试miRNA加标血清样品的理想回收率以及对临床相关样品进行分类的明确定义的准确性。该系统保留了简单性（一个寡核苷酸探针）并简化了检测程序（一步检测）。它的混合信号传输和一对多扩增行为优于传统的基于MB的扩增，表现出宽线性响应和低至8.1 fM的检测限。还展示了针对一碱基突变的高特异性，用于测试miRNA掺入的血清样品的理想回收率以及用于对临床相关样品进行分类的明确定义的准确性。该系统保留了简单性（一个寡核苷酸探针）并简化了检测程序（一步检测）。它的混合信号传输和一对多扩增行为优于传统的基于MB的扩增，表现出宽线性响应和低至8.1 fM的检测限。还展示了针对一碱基突变的高特异性，用于测试miRNA掺入的血清样品的理想回收率以及用于对临床相关样品进行分类的明确定义的准确性。