稳定表达乙型脑炎病毒NS1蛋白细胞系的建立  

Establishment of cell line stably expressing NS1 protein of Japanese encephalitis virus

作　　者：陈振师[1,2] 刘立科[1,2] 汪恩强[1] 李业南[2] 华荣虹[2] 

机构地区：[1]河北农业大学,河北保定071000 [2]中国农业科学院哈尔滨兽医研究所兽医生物技术国家重点实验室,黑龙江哈尔滨150001

出　　处：《中国预防兽医学报》2011年第12期920-923,共4页Chinese Journal of Preventive Veterinary Medicine

基　　金：国家自然科学基金(30700027);公益性行业(农业)科研专项(200803015;201203082)

摘　　要：为建立稳定表达乙型脑炎病毒(JEV)NS1蛋白的真核细胞系,本研究将编码JEV NS1蛋白的人工合成基因克隆到真核表达载体pCAGG-TK-neo中,构建了重组质粒pCAGG-opti-NS1。重组质粒经脂质体转染RK-13细胞,以含G418的选择性培养基选择培养,经细胞克隆纯化,以间接免疫荧光试验(IFA)筛选表达目的基因的细胞。结果表明,转染的RK-13细胞经G418加压及IFA筛选后,获得表达JEV NS1蛋白的阳性RK-13细胞系。经RT-PCR、western blot和IFA鉴定,该细胞系在传代至第15代后仍然可以稳定表达NS1蛋白。本实验获得了能够稳定表达JEV NS1蛋白的细胞系,为进一步开展JEV相关的研究奠定了基础。Japanese encephalitis virus （JEV） is an important mosquito-borne virus for human and animal, to generate cell line stably expressing NSI protein of JEV, RK-13 cells was transfected with the recombinant pCAGG-opti-NS1 plasmid, which was constructed by introducing the synthetic JEV NS1 gene into eukaryotic vector pCAGG-TK-neo, and subsequently selected with G418 and indirect immunofluorescence assay （IFA）. The results showed that the established cell line was able to express NS1 protein stably up to the fifteenth passages identified by RT- PCR, westem blot and IFA. The cell line expressing NS1 protein of JEV would be useful for further study on JEV.

关 键 词：乙型脑炎病毒 NS1蛋白 细胞系 

分 类 号：S852.65[农业科学—基础兽医学]