| 转录病毒载体介导的稳定表达口蹄疫病毒3D基因的BHK21细胞系的建立 |
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| 引用本文： | 毕研丽,沈小燕,丛国正,刘湘涛,常惠芸,才学鹏.转录病毒载体介导的稳定表达口蹄疫病毒3D基因的BHK21细胞系的建立[J].微生物学报,2008,48(8):1115-1120. |
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| 作者姓名： | 毕研丽 沈小燕 丛国正 刘湘涛 常惠芸 才学鹏 |
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| 作者单位： | 中国农业科学院兰州兽医研究所,家畜疫病病原生物学国家重点实验室,农业部畜禽病毒学重点开放实验室,国家口蹄疫参考实验室,兰州,730046 |
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| 基金项目： | 中国科学院所长基金，国家科技支撑计划 |
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| 摘 要： | 目的]研究口蹄疫病毒RNA聚合酶在BHK-21细胞中的稳定表达状况,为研究RNA聚合酶生物学活性及其基囚工程疫苗研制提供科学依据.方法]从重组质粒pMD18-T-3D扩增口蹄疫病毒3D基因,通过分子克隆技术构建莺组逆转录病毒表达载体pBPSTR1-3D.用pBPSTR1-3D和pVSV-G双质粒瞬时转染GP2-293包装细胞,收获重组逆转录病毒,然后感染BHK-21细胞,嘌呤霉素持续筛选12 d后获得阳性克隆,并用有限稀释法挑选单个阳性细胞克隆.结果]应用PCR、RT-PCR技术可从体外反复传代的阳性细胞中扩增到3D基因,证实目的外源基因能转录并被稳定整合进宿主细胞基因组中.经SDS-PAGE、Western blot、间接免疫荧光检测到在不同代次的阳性细胞中有目的蛋白表达.结论]本试验利用逆转录病毒载体介导的基因转移技术,将外源基因插入到靶细胞的基因组中,构建了稳定表达口蹄疫病毒RNA聚合酶的包装细胞系,为研究3D基因表达及其蛋白定位提供了方便,也为下一步研究RNA聚合酶生物学功能和疫苗研制提供了科学依据.
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| 关 键 词： | 口蹄疫病毒 3D基因 逆转录病毒载体pBPSTR1 BHK-21细胞系 |
| 修稿时间： | 2008/4/20 0:00:00 |
Establishment of BHK-21 cell lines stably expressing FMDV 3Dpol gene by retroviral-mediated gene transfer technique |
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| Yanli Bi,Xiaoyan Shen,Guozheng Cong,Xiangtao Liu,Huiyun Chang and Xuepeng Cai.Establishment of BHK-21 cell lines stably expressing FMDV 3Dpol gene by retroviral-mediated gene transfer technique[J].Acta Microbiologica Sinica,2008,48(8):1115-1120. |
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| Authors: | Yanli Bi Xiaoyan Shen Guozheng Cong Xiangtao Liu Huiyun Chang and Xuepeng Cai |
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| Institution: | State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Animal Virology of Ministry of Agriculture, National FMD Reference Laboratory, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, China;State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Animal Virology of Ministry of Agriculture, National FMD Reference Laboratory, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, China;State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Animal Virology of Ministry of Agriculture, National FMD Reference Laboratory, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, China;State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Animal Virology of Ministry of Agriculture, National FMD Reference Laboratory, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, China;State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Animal Virology of Ministry of Agriculture, National FMD Reference Laboratory, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, China;State Key Laboratory of Veterinary Etiological Biology, Key Laboratory of Animal Virology of Ministry of Agriculture, National FMD Reference Laboratory, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou 730046, China |
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| Abstract: | |
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| Keywords: | foot-and-mouth disease virus 3Dpol gene pBPSTR1 vector BHK-21 cell line |
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