乙脑/登革2型嵌合病毒的构建  

Construcion of a Chimeric Japanese Encephalits Virus/Dengue Virus-2

作　　者：韦艳[1] 陆鹏[1] 于建石[1] 李建东[1] 刘琴芝[1] 张全福[1] 李川[1] 苗芳[1] 张硕[1] 杭小同[1] 李德新[1] 

机构地区：[1]中国疾病预防控制中心病毒病预防控制所病毒基因工程国家重点实验室,北京100052

出　　处：《病毒学报》2009年第3期185-189,共5页Chinese Journal of Virology

基　　金：国家自然科学基金(30770087)

摘　　要：在乙脑病毒SA14-14-2株复制子载体pPartial△prM/E中克隆入DV2(Dengue virus serotype 2)的prM/E基因,构建乙脑/登革2型嵌合体克隆。将嵌合体克隆线性化后体外转录,获得的RNA转染BHK-21细胞,5～7d可观察到CPE。收获病毒上清液分别感染BHK-21细胞及C6/36细胞。接种于C6/36细胞中的嵌合病毒可使细胞出现CPE,RT-PCR、间接免疫荧光和Western blot检测显示:获得的嵌合病毒具有预期嵌合性核酸并能表达DV2的包膜蛋白,但不能在BHK-21细胞中传代培养。成功构建的乙脑/登革2型感染性克隆为进一步研究登革病毒疫苗奠定了基础。The prM/E gene of DV2 was cloned into the JEV （SA14-14-2 strain） replicon vector which had been constructed previously, and the resulting recombinant plasmid was named pPartial△prM/E. The constructed chimeric clone was linearized and then was transcripted into RNA in vitro. The produced RNA was transfected into BHK-21 cells. Five to seven days later, CPE could be observed on the transfected BHK-21eells, and then the supernatant containing the chimeric virus was collected. The Supernatant was inoculated to BHK-1 cells and C6/36 cells, respectively. CPE could be observed about 4 days post the infection of C6/36cell with the chimeric virus. The results from RT-PCR, IFA, Western blot showed that the virus contained the chimeric RNA and the envelop protein of DV2. However, the chimeric virus could not be passaged in BHK-21 cell. The successful construction of the infectious clone JE/DEN-2 laid the basis for the further research of the DV vaccine.

关 键 词：乙脑 登革 嵌合病毒 感染性克隆 

分 类 号：R373.31[医药卫生—病原生物学]