多种布尼亚目动物多分体负链RNA病毒已经成功建立了微型基因组反向遗传学体系，但是在植物中至今还没有任何多分体负链RNA病毒反向遗传学体系的成功报道。番茄斑萎病毒（TSWV）是在世界范围内农业生产上危害极为严重的一种植物多分体负链RNA病毒，本项目拟在课题组已有TSWV前期研究工作基础之上，全面建立TSWV三条基因组正、负义RNA微型复制子反向遗传学体系，并以此为基础，利用病毒学、分子生物学、生物化学和细胞生物学等多学科的手段深入解析TSWV复制调控机制，明确病毒复制的顺式作用元件和反式作用因子，鉴定复制相关的寄主因子并探明其调控复制的机制，解析TSWV复制工厂及装配机制。本项目的开展不仅可为TSWV为模式的植物多分体负链RNA病毒基因功能研究提供全新的反向遗传学研究体系，同时也为进一步深入了解TSWV的复制调控分子机制提供基础理论知识，为有效防控该病毒病害提供重要理论科学依据。

The minigenome reverse genetics system of a variety of animal multi-segmented negative-strand RNA viruses of Bunyavirale has successfully established, but there have not been any reports of reverse genetics of the plant multi-segmented negative-strand RNA virus in planta. Tomato spotted wilt virus (TSWV) is the most serious plant multi-segmented negative-stranded RNA virus in the world. Based on the our preliminary studies of TSWV, we aim to establish the minireplicon based reverse genetics system for all three genome RNA segments of TSWV in positive and negative strand. Based on the minigenome reverse genetics system, we will integrate the multidisciplinary approaches of plant virology, molecular biology, biochemistry and cell biology to understand the regulatory mechanism of TSWV replication, to characterize the cis-acting elements and transacting factors of viral replication, to identify the host factors involved in viral replication and to explore the mechanism of their regulation of viral replication, to analyze the viral replication factory and to understand how the replication factory was assembled. The implement of this project will not only provide an important reverse genetics platform for the gene function analysis of plant multi-segmented negative-strand RNA viruses, but also will further understand the molecular mechanism of TSWV replication, hence providing an important scientific basis for the high efficient control of the TSWV diseases.