构建表达Flag-GPI的重组乙型肝炎病毒载体

胡孔颖1; 谷陈建1; 吴敏乐2; 陶帅1; 刘楠楠1; 刘晶1; 谢幼华1

doi:10.3969/j.issn.1673-6184.2019.04.002

微生物与感染 >

2019 , Vol. 14 >Issue 4: 199 - 208

DOI: https://doi.org/10.3969/j.issn.1673-6184.2019.04.002

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构建表达Flag-GPI的重组乙型肝炎病毒载体

胡孔颖1 ,
谷陈建1 ,
吴敏乐2 ,
陶帅1 ,
刘楠楠1 ,
刘晶1 ,
谢幼华1

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1. 复旦大学上海医学院基础医学院病原生物学系，教育部、卫健委、医科院医学分子病毒学重点实验室，上海 200032; 2. 上海市浦东医院，复旦大学附属浦东医院检验科，上海 201399

收稿日期: 2019-04-23

网络出版日期: 2019-08-25

基金资助

国家科技重大专项(民口)(2018ZX10301-208)

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Construction of Flag-GPI carrying recombinant hepatitis B virus vector

HU Kongying1 ,
GU Chenjian1 ,
WU Minle2 ,
TAO Shuai1 ,
LIU Nannan1 ,
LIU Jing1 ,
XIE Youhua1

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1. Department of Medical Microbiology and Parasitology, Key Laboratory of Medical Molecular Virology (MOE/NHC/CAMS), School of Basic Medical Sciences, Shanghai Medical College, Fudan University, Shanghai 200032, China; 2. Department of Clinical Laboratory, Shanghai Pudong Hospital, Fudan University, Shanghai 201399, China

Received date: 2019-04-23

Online published: 2019-08-25

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摘要

乙型肝炎病毒(hepatitis B virus，HBV)是小型的嗜肝DNA病毒，能够特异性地感染人肝实质细胞。HBV的易感性可以归于多个方面，包括细胞表面的受体以及细胞内的蛋白或其他因子，目前对HBV易感性的了解还有待深入。本课题利用课题组原有的HBV 5c3c载体和糖基磷脂酰肌醇(glycosylphosphatidylinositol，GPI)的特性，构建了重组HBV载体5c3c-CD59-Flag-GPI和5c3cT-Flag-GPI，转染Huh7细胞后可以将Flag标签锚定在细胞膜上，且可利用Flag抗体通过流式细胞术对其进行分选。在回补HBV包膜蛋白的情况下，5c3cT-Flag-GPI能包装形成完整的重组HBV颗粒。本研究为后续优化重组HBV的包装效率，进而为HBV易感性研究提供工具并奠定基础。

关键词： 乙型肝炎病毒; 5c3c; 糖基磷脂酰肌醇; 重组乙型肝炎病毒载体

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胡孔颖1 , 谷陈建1 , 吴敏乐2 , 陶帅1 , 刘楠楠1 , 刘晶1 , 谢幼华1 . 构建表达Flag-GPI的重组乙型肝炎病毒载体[J]. 微生物与感染, 2019 , 14(4) : 199 -208 . DOI: 10.3969/j.issn.1673-6184.2019.04.002

Abstract

Hepatitis B virus (HBV) is a small hepatophilic DNA virus that can specifically infect human hepatic parenchymal cells. The susceptibility of HBV can be attributed to cell surface receptor(s), intracellular proteins and other factors. A simple and clear method to assay the susceptibility of cells for HBV remains a challenge. Here, we constructed the recombinant HBV vectors 5c3c-CD59-Flag-GPI and 5c3cT-Flag-GPI based on the original HBV 5c3c vector and glycosylphosphatidylinositol (GPI) anchor. After transfection into Huh7 cells, Flag tags can be anchored on the cell membrane, and sorted by flow cytometry using Flag antibody. With the replenishment of HBV envelope protein, 5c3cT-Flag-GPI can form complete recombinant HBV particles. This study laid a foundation for optimization of packaging efficiency of recombinant HBV, which would provide a tool for HBV susceptibility research.

Key words： Hepatitis B virus; 5c3c; Glycosylphosphatidylinositol; Recombinant hepatitis B virus vector

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