利用杆状病毒实现外源基因在哺乳动物细胞内表达

doi:10.14088/j.cnki.issn0439-8114.2020.23.036

湖北农业科学 ›› 2020, Vol. 59 ›› Issue (23): 159-163.doi: 10.14088/j.cnki.issn0439-8114.2020.23.036

利用杆状病毒实现外源基因在哺乳动物细胞内表达

邹桂莲^1,2, 罗青平¹, 邵华斌¹, 温国元¹, 杨烨², 商雨¹

1.湖北省农业科学院畜牧兽医研究所/农业农村部畜禽细菌病防治制剂创制重点实验室/畜禽病原微生物学湖北重点实验室,武汉 430064;
2.长江大学动物科学学院,湖北荆州 434025

收稿日期:2020-11-20 出版日期:2020-12-10 发布日期:2020-12-30
通讯作者: 商雨(1987-),男,湖北黄梅人,助理研究员,博士,主要从事家禽病毒病治病机制和新型疫苗开发研究,(电子信箱)suppershangyu@126.com
作者简介:邹桂莲(1993-),女,广西富川人,在读硕士研究生,研究方向为耐热疫苗,(电话)15927732410(电子信箱)zguilianlotus@163.com
基金资助:
国家自然科学基金青年基金项目(31800143);现代农业产业技术体系建设专项(CARS-41-G13);湖北省农业科学院青年基金项目(2018NKYJJ08);湖北省农业科技创新中心项目(2019-620-000-001-17)

The expression of exogenous genes in mammalian cells by using baculovirus

ZOU Gui-lian^1,2, LUO Qing-ping¹, SHAO Hua-bin¹, WEN Guo-yuan¹, YANG Ye², SHANG Yu¹

1. Key Laboratory of Prevention and Control Agents for Animal Bacteriosis / Hubei Provincial Key Laboratory of Animal Pathogenic Microbiology /Institute of Animal Husbandry and Veterinary,Hubei Academy of Agricultural Sciences,Wuhan 430064,China;
2. College of Animal Science,Yangtze University,Jingzhou 434025,Hubei,China

Received:2020-11-20 Online:2020-12-10 Published:2020-12-30

摘要/Abstract

摘要： 杆状病毒是一类专职感染无脊椎动物的病毒,目前表达载体和基因转导载体广泛应用于科学研究和生物医药领域。研究在杆状病毒Bac-to-Bac操作系统基础上将杆状病毒载体改造成能在哺乳动物细胞内表达外源基因的载体。首先,将Bac-to-Bac操作系统中pFastBac-Dual质粒中的PH和p10启动子替换成巨细胞病毒(CMV)启动子和真核生物延伸因子(EF1)基因的启动子;然后将egfp基因和mCherry基因分别插入到EFI启动子和CMV启动子下游;通过转座获得表达egfp和mCherry基因的重组杆状病毒基因组,并转染Sf9细胞获得重组杆状病毒。重组杆状病毒可以在昆虫细胞和哺乳动物细胞内产生绿色荧光和红色荧光,表明以CMV启动子和EF1启动子构建的重组杆状病毒既可以在昆虫细胞内表达外源基因,又可以通过转导在哺乳动物细胞内表达外源基因。该载体可以应用于特殊蛋白的真核表达和基因治疗药物的研究开发。

关键词: 杆状病毒, 基因表达, 哺乳动物细胞

Abstract: Baculovirus is a kind of virus that infects invertebrates,and they are widely used in scientific research and biomedicine as expression vector and gene transduction vector.Based on “Bac-to-Bac”operating system of baculovirus,we transformed baculovirus vector into a transduction vector with capable of expressing exogenous genes in mammalian cells in this study.First,the PH and p10 promoters of pFastBac-Dual plasmids in the Bac-to-Bac operating system were replaced with the CMV promoter and EF1 promoter.Then,the egfp gene and mCherry gene were inserted into the downstream of EF1 promoter and CMV promoter respectively.The transposition was performed to obtain the recombinant baculovirus genomeexpressing egfp and mCherry genes,and recombinant baculovirus was obtained by transfect the genome to Sf9 cells.The recombinant baculovirus can generate green and red fluorescence in insect cells and mammalian cells,indicating that the recombinant baculovirus constructed with CMV promoter and EF1 promoter can express exogenous genes in insect cells as well as mammalian cells through transduction.This vector can be used for eukaryotic expression of special proteins and research and development of gene therapy drugs.

Key words: baculovirus, gene expression, mammalian cells

中图分类号:

Q819

邹桂莲, 罗青平, 邵华斌, 温国元, 杨烨, 商雨. 利用杆状病毒实现外源基因在哺乳动物细胞内表达[J]. 湖北农业科学, 2020, 59(23): 159-163.

ZOU Gui-lian, LUO Qing-ping, SHAO Hua-bin, WEN Guo-yuan, YANG Ye, SHANG Yu. The expression of exogenous genes in mammalian cells by using baculovirus[J]. HUBEI AGRICULTURAL SCIENCES, 2020, 59(23): 159-163.

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利用杆状病毒实现外源基因在哺乳动物细胞内表达

The expression of exogenous genes in mammalian cells by using baculovirus

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