基于液相体系的microRNA电化学检测新方法及其响应机制研究

作为一种重要的转录后基因调节分子，微小核糖核酸（miRNA）表达水平变化与多种疾病密切相关，有望成为一类新型的生物标志物。目前常用的miRNA检测方法多需要复杂操作、昂贵设备，很大程度限制了其临床普及。本项目在前期已合成出具备生物相容性好、导电能力强、超顺磁性强等优点的金磁复合纳米材料的基础上，探讨并验证利用其作为miRNA液相检测微载体的可行性。在此基础上拟通过构建核酸多酶标记体系、制备Nafion-石墨烯修饰丝网印刷工作电极的手段，分别从放大电流信号以及增加电极电子传导能力两方面增加分析的灵敏度。研究将通过多种表征手段阐明其响应机制，探讨并优化分析条件，建立定量分析的数学模型，最后通过检测肿瘤细胞株与临床血清样本miRNA以验证该方法的检测效能，旨在为构建一种高敏、快速、经济、简便的电化学miRNA检测技术提供实验基础与理论依据。

MicroRNAs (miRNAs) are small non-coding RNAs that regulate the activity of specific mRNA targets and play crucial role in a wide variety of biological processes, including gene expression, development, proliferation, differentiation and apoptosis. Implicated in human diseases, miRNA expression signatures will become an emerging class of molecular biomarkers. Though many methods for miRNA analysis have been reported at present, most of them are time-consuming and expensive, which greatly limit their application in clinical laboraory. In this project, we aim to discuss and validate the feasibility of a liquid-phase detection strategy for miRNA using Fe3O4/Au nanoparticle as a class of microcarrier. In our early study, we have demonstrated that the Fe3O4/Au nanoparticle we synthesized shows great advantages of biocompatibility, conductive ability, and strong superparamagnetism. We are going to increase the sensitivity analysis in amplification current signal and electrode electronic conduction respectively by constructing nucleic acid multienzyme marking system and Nafion-graphene modified screen printing working electrode based on previous results. Besides we intend to elucidate the response mechanism by a variety of methods of characterization and establish the mathematical model in quantitative analysis by optimization analysis conditions. Finally we verify the detection efficiency of this method by detecting miRNA expression of tumor cell and clinical serum sample, aiming to construct an economic and simple electrochemical miRNA detection technology with high sensitivity and speed for future clinical application.