基于核酸适配体和自组装DNA纳米结构的双分子荧光互补技术及其生物传感应用研究

Realizing the real-time monitoring of intracellular biological molecules with good biocompatibility, high sensitivity, excellent selectivity is of great significance in biology science research and clinical disease diagnosis. Due to its high signal-to-noise ratio and excellent sensitivity, fluorescent proteins based bimolecular fluorescence complementation (BiFC) probes are widely applied for the study of biological macromolecules interaction, however, realize little application in real-time sensing of the concentration change of biological molecules, especially non-protein targets. To broaden the application of BiFC in biosensors development, this project intends to combine BiFC with nucleic acid aptamer, one kind of ssDNA or ssRNA specificly recognizing a series of targets including small molecules, proteins, cells and even tissues, with high affinity and high specificity. Moreover, self-assembled DNA nanostructures, with the features of good addressable ability, nanoscale spatial resolution and excellent dynamic structure control performance, are utilized for the precise assembly of aptamer and fluorescent protein fragments, as well as the accurate control of key parameters such as the distance and stoichiometric ratio of fluorescent protein fragments. Combining the advantages of aptamer and self-assembled DNA nanostructures, BiFC fluorescent probes would be able to realize their application in the biosensing of intracellular biological molecules with good biocompatibility, high sensitivity and excellent selectivity, to provide a new tool for scientific research and clinical diagnosis.

实现细胞内生物分子的高安全性、高灵敏度、高特异性的实时检测对于生命科学研究和临床疾病诊断具有重要的意义。基于安全无毒的荧光蛋白发展而成的双分子荧光互补技术能够实现荧光信号从无到有的变化，因而具有噪音低、灵敏度高等优点，目前主要用于生物大分子相互作用的研究，但较少应用于生物分子的浓度变化等信息的传感。本项目拟利用靶标范围广、具有高亲和力和高特异性识别能力的核酸适配体作为识别元件来拓宽双分子荧光互补技术在生物传感领域的应用。同时结合自组装DNA纳米结构良好的可寻址性、纳米级的空间分辨率和优异的动态调控性能，实现荧光蛋白和核酸适配体的精确组装，并精准控制荧光蛋白片段的距离、计量比等关键参数，以发展一系列适用于小分子、蛋白等不同类型生物靶标的荧光蛋白探针，从而实现生物分子的高安全性、高灵敏度、高特异性的实时传感，获得生物靶标的浓度、分布等信息，为科学研究和临床诊断提供新的方法。

本项目利用自组装DNA纳米结构良好的可寻址性、纳米级的空间分辨率和优异的动态调控性能，实现荧光分子和功能核酸的精确组装，并精准控制距离、计量比等关键参数，发展一系列不同类型生物靶标的荧光探针，为生物分子的浓度、分布等信息的实时定量研究提供新的工具。在本项目支持下课题组取得了一系列研究成果，发表了第一或通讯作者SCI论文（均标注基金号）11篇（其中影响因子大于10的论文3篇），包括Nature Communications 1篇，Journal of the American Chemical Society 1篇，Chemical Society Reviews 1篇，Analytical Chemistry 2篇。