Engineering high-performance hairpin stacking circuits for logic gate operation and highly sensitive biosensing assay of microRNA
Engineering high-performance hairpin stacking circuits for logic gate operation and highly sensitive biosensing assay of microRNA
复制标题
设计用于逻辑门操作和 microRNA 高灵敏度生物传感测定的高性能发夹堆叠电路
作者：
Yueli Xing;Xinmin Li;Taixian Yuan;Wei Cheng;D;an Li;Tianxiao Yu;Xiaojuan Ding;Shijia Ding
最近，基于支点介导的链置换的发夹堆积电路（HSC）已经被工程化以检测核酸和蛋白质。然而，HSC系统中的三个亚稳态发夹结构在没有催化剂的情况下可能发生非特异性反应，限制了其实际应用。本文提出了一种独特的发夹型设计方法，以消除HSC的电路泄漏，并成功地将高性能HSC应用于逻辑门构建和生物传感。我们首先分析了电路泄漏的来源，并基于表面等离子体共振（SPR）技术优化了HSC系统中发夹结构的立足点长度。接下来，引入了在特定结构域中取代两个核苷酸的新策略，称为“环结构域取代”，以消除泄漏。我们还系统地改变了引入的取代基的位置和数量，以探索它们对电路泄漏抑制的潜在贡献。通过这些努力，HSC的电路泄漏被显着降低。最后，通过设计不同的DNA输入链，可以激活逻辑门来实现输出信号。使用miRNA作为模型分析物，该策略可以检测低至pM水平的miRNA，并使电路泄漏最小化。我们相信这些工作表明DNA电路的重大进展。
ecently, hairpin stacking circuits (HSC) based on toehold-mediated strand displacement have been.engineered to detect nucleic acids and proteins. However, the three metastable hairpins in a HSC system.can potentially react non-specifically in the absence of a catalyst, limiting its practical application. Here,.we developed a unique hairpin design guideline to eliminate circuit leakage of HSC, and the high-.performance HSC was successfully implemented on logic gate building and biosensing. We began by.analyzing the sources of circuit leakage and optimizing the toehold lengths of hairpins in the HSC system.based on the surface plasmon resonance (SPR) technique. Next, a novel strategy of substituting two nucleo-.tides in a specific domain, termed ‘loop-domain substitution’, was introduced to eliminate leakages. We also.systematically altered the positions and numbers of the introduced substitutions to probe their potential.contribution to circuit leakage suppression. Through these efforts, the circuit leakage of HSC was significantly.reduced. Finally, by designing different DNA input strands, the logic gates could be activated to achieve the.output signal. Using miRNA as a model analyte, this strategy could detect miRNA down to pM levels with.minimized circuit leakage. We believe these work indicate significant progress in the DNA circuitry.