Zhu Wu-Yang, Fu Shi-Hong, Wang Li-Hua, He Ying, Tang Qing, Liang Guo-Dong
Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention (IVDC, China CDC), Beijing, China.
Bing Du Xue Bao. 2009 Mar;25(2):143-7.
To construct vector system of XJ-160 virus, a Sindbis virus isolated in China, recombinant vector pBRepXJ together with its helper plasmid pBR-H were derived from XJ-160 viral infectious clone pBR-XJ160 by overlap-PCR. To quantitatively and qualitatively verify the function of the replicon system, recombinant plasmids pSinRep-EGFP, pBRepXJ-EGFP, pSinRep-R and pBRepXJ-R were constructed by cloning report genes of enhanced green fluorescent protein (EGFP) or Renilla luciferase (R. luc) into pBRepXJ or pSinRep5, a commercial Sindbis vector. And in Vitro-synthesized RNA from expression vectors were electroporated into BHK-21 cells. The results indicated that the replicon vector system was capable of self-replicating in host cell, and the expression efficiency of heterologous genes corresponded with that of the commercial Sindbis vector (pSinRep5). Our study laid the basis for developing alphavirus vector system with Chinese intellectual property.
为构建中国分离的辛德毕斯病毒XJ-160的载体系统，通过重叠PCR从XJ-160病毒感染性克隆pBR-XJ160获得重组载体pBRepXJ及其辅助质粒pBR-H。为了定量和定性验证复制子系统的功能，通过将增强型绿色荧光蛋白（EGFP）或海肾荧光素酶（R. luc）的报告基因克隆到pBRepXJ或商业辛德毕斯载体pSinRep5中，构建了重组质粒pSinRep-EGFP、pBRepXJ-EGFP、pSinRep-R和pBRepXJ-R。将表达载体体外合成的RNA电穿孔导入BHK-21细胞。结果表明，复制子载体系统能够在宿主细胞中自我复制，异源基因的表达效率与商业辛德毕斯载体（pSinRep5）相当。我们的研究为开发具有中国知识产权的甲病毒载体系统奠定了基础。
