基于智能核酸分子组装的固相纳米孔均相检测新原理

纳米孔测量是现代分子识别和检测领域最具研究和推广价值的单分子分析方法之一。然而，其真正意义的应用推广仍然受到诸多因素的影响和制约。如，受限于孔径小且单一等问题，分辨率最高的蛋白质孔几乎无法检测半径在5nm以上的待测物。而在稳定性和孔径选择上更具优势的固相纳米孔（玻璃、氮化硅等），却往往因尺寸过宽、制作/修饰工艺过于复杂而造成分辨率、重现性或特异性严重不足。为加快突破这一目前限制固相纳米孔分析应用的首要障碍，本项目提出以智能核酸纳米组装为"增敏探针”的新型传感原理。筛选出适于固相纳米孔精细表征的一系列具有特异性传导和尺寸放大功能的新型核酸组装反应；进而以其作为寡聚核酸、小分子或金属离子等待测物和纳米孔间的“增敏探针”，在全均相条件下实现复杂和多元样品中小尺寸待测物的“高分辨、高特异和可重现”分析。本项目同时有助于揭示各种核酸组装与纳米孔信号之间的内在联系和规律，推动两个前沿领域的交叉融合。

Nanopore detection is one of most promising single molecular techniques used for molecular recognition and detection. Nevertheless, its real and extensive practical applications have still been seriously restricted by some shortcomings. For instance, due to small and limited pore sizes, the protein pores holding the highest resolution may not detect the targets larger than 5 nm. While, even if there are solid pores (such as glass pore and SiNx pore) that are more stable and can be fabricated into much wider sizes, the complicated fabrication or modification procedures may usually lead to low resolution, selectivity, and reproducibility. This has been one of the biggest challenges that affect the practical applications of solid-state pores. With the purpose to reduce or solve this problem, in this project we propose a novel sensing strategy that uses the smart nucleic acid assembly as a “resolution enhancer” to bridge the small targets and solid-state nanopore. After rationally designing a series of target-induced nucleic acid assemblies, those assembly products or structures that can generate clear and unique nanopore signals will be chosen as “resolution enhancers”. Then, small targets previously blinded by the nanopore (such as oligonucleotides, small molecules, and metal ions) will be specifically transduced to these “resolution enhancers” via corresponding DNA assembly reactions. In this way, the recognition of small targets can be executed in homogeneous solution, completely without covalent bond label or modification. Therefore, the detection is simultaneously sensitive, specific, and easy to handle and repeat, being especially useful in multi-analysis and detection of complex samples. Besides extending and enhancing the application ranges of solid-state nanopore, this project is also helpful in deepening the understanding the internal relationship between the nanopore signals and various nucleic acid assemblies,which will finally advance the interdisciplinary of the two areas.