A novel and versatile nanomachine for ultrasensitive and specific detection of microRNAs based on molecular beacon initiated strand displacement amplification coupled with catalytic hairpin assembly with DNAzyme formation
A novel and versatile nanomachine for ultrasensitive and specific detection of microRNAs based on molecular beacon initiated strand displacement amplification coupled with catalytic hairpin assembly with DNAzyme formation
复制标题
一种新型多功能纳米机器，用于超灵敏和特异性检测 microRNA，基于分子信标引发的链置换扩增，结合催化发夹组装和 DNAzyme 形成
作者：
Yan Yurong;Shen Bo;Wang Hong;Sun Xue;Cheng Wei;Zhao Hua;Ju Huangxian;Ding Shijia
MicroRNA 是一种小调节分子，可用作临床诊断的潜在生物标志物，人们一直致力于开发一种简单、快速、序列选择性的 microRNA 分析方法。在这里，我们报告了一种简单而通用的比色策略，用于基于分子信标引发的链置换扩增（SDA）和催化发夹组装（CHA）以及DNAzyme形成来超灵敏和特异性地测定microRNA。目标 microRNA 的存在会触发链置换扩增，释放切口 DNA 触发器，从而引发 CHA 产生大量 CHA 产物。同时，众多的CHA产品可以与血红素结合形成G-四链体/血红素DNAzyme，一种著名的辣根过氧化物酶（HRP）模拟物，催化比色反应。此外，SDA 混合物的纯化已被开发用于消除基质干扰，以减少非特异性 CHA 产物。在最佳条件下并使用有前途的扩增策略，所建立的比色纳米机器（生物传感器）在5 fM至5 nM的动态响应范围内表现出高灵敏度和选择性，检测限低至1.7 fM（S/N = 3）。此外，还开发了一种多功能比色生物传感器，只需改变分子信标的 miRNA 识别域即可检测不同的 miRNA。因此，这种比色生物传感器可能成为生物医学研究和临床分子诊断的潜在替代工具。
MicroRNAs are small regulatory molecules that can be used as potential biomarkers of clinical diagnosis, and efforts have been directed towards the development of a simple, rapid, and sequence-selective analysis of microRNAs. Here, we report a simple and versatile colorimetric strategy for ultrasensitive and specific determination of microRNAs based on molecular beacon initiated strand displacement amplification (SDA) and catalytic hairpin assembly (CHA) with DNAzyme formation. The presence of target microRNAs triggers strand displacement amplification to release nicking DNA triggers, which initiate CHA to produce large amounts of CHA products. Meanwhile, the numerous CHA products can combine with hemin to form G-quadruplex/hemin DNAzyme, a well-known horseradish peroxidase (HRP) mimic, catalyzing a colorimetric reaction. Moreover, the purification of the SDA mixture has been developed for eliminating matrix interference to decrease nonspecific CHA products. Under the optimal conditions and using the promising amplification strategy, the established colorimetric nanomachine (biosensor) shows high sensitivity and selectivity in a dynamic response range from 5 fM to 5 nM with a detection limit as low as 1.7 fM (S/N = 3). In addition, a versatile colorimetric biosensor has been developed for detection of different miRNAs by only changing the miRNA-recognition domain of molecular beacon. Thus, this colorimetric biosensor may become a potential alternative tool for biomedical research and clinical molecular diagnostics.