利用微流控芯片技术构建多功能纳米颗粒用于细胞内基因表达调控的实时原位动态分析

The study about the mechanism of intracellular gene regulation is a hotspot in the research of cell biology and pathophysiology. It is of great significance to the reveal of pathogenic mechanism of the disease, the search for therapeutic targets and the development of drugs. Conventional research about gene regulation mechanism usually go a route of cell stimulation, cell sacrifice and sample extraction, sample analysis outside the cells. So it is difficult to obtain the in situ, real-time and continuous analysis data of the processes of gene regulation in cells, which is a serious impediment to the integrity and coherence of the mechanism research and to some extent affect the accuracy. The project focus on solving technical difficulties faced by the real-time dynamic analysis of intracellular gene regulation. Based on the controllable synthesis with microfluidics technology, the multi-functional nanoparticles with a cavity structure will be established by the precise control of microfluid and the molecular self-assembling in microfluidic channels. The multi-functional nanoparticles will be used for efficiently delivering and tracing of the gene regulation molecules and probes, in order to achieve the in situ real-time dynamic analysis of intracellular gene regulation. This technology will provide new ideas and methods to the study of gene regulation mechanism, by achieving direct visual observation of intracellular gene regulation, thus greatly improve the integrality and visualization of the mechanism research of gene regulation and bring a new perspective to it.

细胞内基因调控机理研究是细胞生物学、病理生理学等学科的研究热点，对于揭示疾病的致病机理、寻找治疗靶点、开发药物等具有重要意义。常规的基因调控机理研究主要遵循对细胞施加刺激、处死细胞并提取样本、在细胞外进行样本分析的研究路线，因而难以获得细胞内基因调控过程的原位、实时、连续的分析数据，严重妨碍了基因调控机理研究的整体性和连贯性，并在一定程度上影响了研究准确性。本项目针对细胞内基因调控过程实时动态分析的技术难点，拟利用微流控芯片可控合成技术，通过对流体的精确控制和通道内的分子自组装，构建具有空腔结构的复合纳米颗粒，用于基因调控分子和分析探针的高效有序递送和示踪，实现细胞内基因调控过程的原位实时动态化分析。该技术的成功开发可为基因调控机理研究提供新思路、新手段，以直观可视化的观测方式展现细胞内的基因调控过程，从而极大地提高基因调控相关研究的整体性和可视性，为人类的基因调控机理研究带来全新视角。

探究不同调控因子对关键基因表达量的影响，可以为遗传学、发育学、致病机理、药物开发等相关研究提供关键信息。本研究通过构建具有核壳型结构的荧光纳米探针，建立了细胞内关键mRNA分子的实时成像和检测方法，可以实现对细胞的关键基因表达量的特异性分析和原位分析检测，并通过参数优化实现了纳米探针在细胞内的有序解离和分子释放，使得细胞内基因调控因子起效和mRNA检测得以自动分步骤完成。利用该探针探究了两种常见基因调控因子对肿瘤耐药性相关基因表达量的影响。本研究为基因调控机理研究提供了新方法，有助于提高相关研究的整体性和可视性，对人类的基因调控机理研究、基因相关疾病的病理研究和诊断治疗具有一定的理论价值和实际应用价值。