SINDBIS VIRUS DETERMINANTS OF INFECTION IN MOSQUITOS

描述(改编自申请人摘要)：申请人提出 研究辛德比斯病毒与埃及伊蚊的相互作用。为了做到这一点，他们 将使用SIN的分子克隆，这些分子克隆已经被设计成表达 感兴趣的表型特征，并携带方便的标记基因。 他们将检查两个具有感染性的SIN DNA克隆，其中一个感染伊蚊 中肠低效(2J病毒)和其他感染中肠的病毒 并在蚊子中引起传播性感染(MRE1001 病毒)。MRE1001病毒是2J病毒非结构基因的嵌合体 病毒与中肠感染株MRE16的结构基因 紧张。2J株和嵌合MRE1001株都已经 被设计成含有第二个亚基因组RNA，与正常衣壳相比具有3个启动子 他们将首先比较编码GFP(绿色荧光蛋白)的区域 口服和注射给药时这三种病毒的生长曲线 路线，并比较器官取向、外在潜伏期和 病毒的传播潜力。然后，他们将创建更多 MRE1001和2J株结构基因的逐个基因嵌合体 明确中肠趋向性和扩散的遗传决定因素。 最后，他们将引入特定位点的突变和缺失，以进一步 完善病毒肠道复制和传播的遗传决定因素。在……里面 他们提议创造一种新的传染性DNA克隆的相关研究 来自MRE16菌株的SIN。该菌株将与MRE1001菌株在以下方面有所不同 所有的非结构基因和结构基因都将衍生出 来自中肠复制、传播的病毒株。

DESCRIPTION (Adapted from the Applicant's Abstract): The applicants propose to study the interactions of Sindbis virus with Aedes aegypti. To do this they will use molecular clones of SIN that have been engineered to express phenotypic characteristics of interest and which bear convenient marker genes. They will examine two infectious DNA clones of SIN, one of which infects Aedes midguts inefficiently (the 2J virus) and the other which infects midguts efficiently and causes disseminated infections in the mosquito (the MRE1001 virus). The MRE1001 virus is a chimera between he nonstructural genes of the 2J virus and the structural genes from a midgut infecting strain, the MRE16 strain. The 2J strain and the chimeric MRE1001 strains have both been engineered to contain a second sub-genomic RNA 3-prime to the normal capsid region which encodes GFP (green fluorescent protein) They will first compare growth curves of these three viruses when given by the oral or injection routes, and compare the organ tropisms, extrinsic incubation periods, and transmission potentials of the viruses. They will then create additional chimeras, gene by gene, of the structural genes of the MRE1001 and 2J strains to define the genetic determinants of midgut tropism and dissemination. Finally, they will introduce site specific mutations and deletions to further refine the genetic determinants of viral gut replication and dissemination. In a related line of research they propose to create a new infectious DNA clone of SIN from the MRE16 strain. This strain will differ from the MRE1001 strain in that all the non-structural genes as well as the structural genes will derive from a mid-gut replicating, disseminating virus strain.