传染性造血器官坏死病病毒糖蛋白杆状病毒表达系统的构建及其抗原性分析

doi:10.16431/j.cnki.1671-7236.2017.03.033

《中国畜牧兽医》 ›› 2017, Vol. 44 ›› Issue (3): 854-860.doi: 10.16431/j.cnki.1671-7236.2017.03.033

传染性造血器官坏死病病毒糖蛋白杆状病毒表达系统的构建及其抗原性分析

谢三磊¹, 温书香¹, 罗琳², 马志宏², 李桂萍¹, 李铁梁², 姜娜², 邢薇², 孙惠玲¹

1. 北京市农林科学院畜牧兽医研究所, 北京畜禽疫病防控技术北京市重点实验室, 北京 100097;
2. 北京市水产科学研究所, 北京 100068

收稿日期:2016-07-18 出版日期:2017-03-20 发布日期:2017-03-21
通讯作者: 孙惠玲 E-mail:sunhuiling01@163.com
作者简介:谢三磊(1987-),男,河南鄢陵人,硕士,研究方向:兽医病理学,E-mail:1327372534@qq.com
基金资助:
北京市科委一般项目（Z141100002314002）；北京市农林科学院科技创新能力建设专项（KJCX20140401）

Construction of Baculovirus Expression System and Antigenicity Analysis of Infectious Hematopoietic Necrosis Virus Glycoprotein

XIE San-lei¹, WEN Shu-xiang¹, LUO Lin², MA Zhi-hong², LI Gui-ping¹, LI Tie-liang², JIANG Na², XING Wei², SUN Hui-ling¹

1. Beijing Key Laboratory for Prevention and Control of Infectious Diseases in Livestock and Poultry, Institute of Animal Husbandry and Veterinary Medicine, Beijing Academy of Agriculture and Forestry Sciences, Beijing 100097, China;
2. Beijing Fisheries Research Institute, Beijing 100068, China

Received:2016-07-18 Online:2017-03-20 Published:2017-03-21

摘要/Abstract

摘要：

为研究传染性造血器官坏死病病毒（infectious haematopoietic necrosis virus，IHNV）主要结构蛋白糖蛋白（G），本研究采用RT-PCR方法从IHNV中提取RNA并进行反转录，经PCR方法扩增获得1 380 bp的G蛋白基因片段，将其克隆到pFB-LIC-Bse杆状病毒载体中，成功构建了重组质粒pFB-LIC-Bse-G，转化到大肠杆菌DH10Bac感受态细胞中，获得了重组杆粒rBacmid-G。将重组杆粒转染至Sf9昆虫细胞，获得了重组杆状病毒。间接免疫荧光（IFA）和Western blotting分析显示，重组G蛋白可与抗组氨酸单抗（Anti-His）、抗IHNV鼠血清发生特异性反应，表明本研究克隆的IHNV G蛋白在真核表达系统中得到正确表达，且该蛋白具有良好的抗原性。本试验结果为研究G蛋白的功能及开发IHNV新型疫苗奠定了基础。

关键词: 传染性造血器官坏死病病毒; 糖蛋白; 重组杆状病毒

Abstract:

To study the major structural protein glycoprotein (G) of infectious hematopoietic necrosis virus (IHNV), glycoprotein gene (1 380 bp) was amplified by RT-PCR from IHNV. In order to construct a recombinant plasmid pFB-LIC-Bse-G, G gene was cloned into the baculovirus vector pFB-LIC-Bse. Then, the constructed plasmid pFB-LIC-Bse-G was transformed into E.coli DH10Bac. The recombinant bacmids rBacmid-G was got, and then it was transfected to insect Sf9 cells,and the recombinant baculovirus that contained G gene was obtained. Western blotting analysis and indirect immunofluorescence assay (IFA) showed that the recombinant G protein could be recognized by histidine monoclonal antibody (Anti-His) and anti-IHNV antibody of mouse. The results indicated IHNV G protein had been expressed correctly in Sf9 cells. The study laid a foundation for further studying the G protein structure, function and immunological characteristics.

Key words: infectious hematopoietic necrosis virus; glycoprotein; recombinant baculovirus

中图分类号:

S941.41

谢三磊, 温书香, 罗琳, 马志宏, 李桂萍, 李铁梁, 姜娜, 邢薇, 孙惠玲. 传染性造血器官坏死病病毒糖蛋白杆状病毒表达系统的构建及其抗原性分析[J]. 《中国畜牧兽医》, 2017, 44(3): 854-860.

XIE San-lei, WEN Shu-xiang, LUO Lin, MA Zhi-hong, LI Gui-ping, LI Tie-liang, JIANG Na, XING Wei, SUN Hui-ling. Construction of Baculovirus Expression System and Antigenicity Analysis of Infectious Hematopoietic Necrosis Virus Glycoprotein[J]. , 2017, 44(3): 854-860.

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传染性造血器官坏死病病毒糖蛋白杆状病毒表达系统的构建及其抗原性分析

Construction of Baculovirus Expression System and Antigenicity Analysis of Infectious Hematopoietic Necrosis Virus Glycoprotein

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