| 稳定表达乙脑病毒结构蛋白的细胞系的建立 |
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| 引用本文： | 刘珊,黄莺,杨鹏,孙志伟,俞炜源.稳定表达乙脑病毒结构蛋白的细胞系的建立[J].病毒学报,2009(4). |
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| 作者姓名： | 刘珊 黄莺 杨鹏 孙志伟 俞炜源 |
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| 作者单位： | 军事医学科学院生物工程研究所蛋白质工程室; |
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| 摘 要： | 以乙型脑炎病毒SA14-14-2疫苗株全长基因组克隆质粒pBR-JTF为模板,通过PCR分别扩增prM-E及C-prM-E基因片段,构建表达乙型脑炎病毒结构蛋白的真核表达质粒pCJE-ME及pCJE-CME。将这2种重组质粒用脂质体法转染BHK-21细胞后,质粒pCJE-ME表现明显的细胞毒性,转染细胞不能存活;质粒pCJE-cME可导致筛选到表达JEV结构蛋白的稳定细胞系,这种稳定表达JEV结构蛋白的细胞系通过PCR扩增细胞系基因组、ELISA、Western Blot、间接免疫荧光等方法得到鉴定。研究结果表明在C蛋白存在下,乙型脑炎病毒C-prM-E蛋白可以在BHK细胞中稳定表达,为研制JEV新型复制子颗粒疫苗提供了便利工具。
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| 关 键 词： | 乙型脑炎病毒 结构蛋白 细胞系 |
Construction of Mammalian Cell Lines Continuously Expressing JEV Structural Proteins |
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| LIU Shan,HUANG Ying,YANG Peng,SUN Zhi-wei,YU Wei-yuan.Construction of Mammalian Cell Lines Continuously Expressing JEV Structural Proteins[J].Chinese Journal of Virology,2009(4). |
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| Authors: | LIU Shan HUANG Ying YANG Peng SUN Zhi-wei YU Wei-yuan |
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| Institution: | Lab for Protein Engineering;Institute of Biotechnology;Academy of Military Medical Sciences;Beijing 100071;China |
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| Abstract: | Based on the infectious clone of JEV vaccine strain SA14-14-2,prM-E genes and C-prM-E genes were cloned into pCDNA3.1 vector.The recombinant plasmid pCJE-ME was transfected into BHK-21 cells,the expressed proteins were toxic to the cell growth and accelerated the cell death.But when transfected with the plasmid pCJE-CME,the cell lines continuously expressing structural proteins could be selected with G418.And the expression products of pCJE-CME vector could be detected by ELISA,Western Blot and IFA assay.It... |
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