活体内遗传编码RNA适体生物传感体系的构建及应用

活体荧光成像传感不仅可以反映活体动物内的细胞分布及基因表达，还能动态观察活体细胞内分子生物学过程，是生物传感分析发展的前沿技术。本项目拟围绕生物探针在活体内驻留时间短，难以对复杂生理和病理过程进行长时间荧光成像关键问题。以课题组在活细胞遗传表达点亮型RNA适体生物探针和体外转录RNA适体信号放大的研究积累为基础，利用基因工程DNA重组和无缝克隆技术，构建生物传感功能元件的质粒DNA，通过病毒载体传递进细胞或活体，借助宿主细胞内的遗传机制，获得细胞内能持续表达的荧光生物传感探针。以mRNA检测为例，探索诱导转录信号放大新策略，构建基于CRISPR/dCas9系统遗传表达RNA适体生物传感通路体系，探讨诱导转录RNA适体信号调控途径，发展生物传感分析新原理和新方法，为相关的生命科学、细胞生物学、肿瘤精准诊断和疾病治疗等领域的基础研究提供有效的技术平台。

In vivo fluorescence imaging is a frontier technology and the development trend of biosensor analysis, it can not only reflect the distribution and expression of gene but also dynamically observe the biological process in vivo. This project will focus on the key issue of providing RNA probe with stable expression for long-term fluorescence imaging of complex physiological and pathological processes in vivo. We will construct the biosensor on the basis of light-up RNA aptamers and plasmid expression technologies, the designed plasmid will be transported into cells or living organisms by viral packaging. With the regulation mechanism in host cells, the RNA aptamer probes that could be stably expressed in cells. Taking the detection of mRNA as example, we will explore the RNA aptamer biosensor of genetic expression and new strategies for signal amplification using clustered regularly interspaced short palindromic repeats(CRISPR/dCas9) system, and develop new principles and methods for biosensor analysis. Futhermore, we will explore the regulatory pathway of the RNA aptamer signal by transcriptional activation methods, and expecting to provide an effective technology platform for basic research in related areas including life science, cell biology, prescie diagnostic and treatment of tumour.