共表达O型口蹄疫病毒P12A+3C基因的重组杆状病毒的构建

doi:10.7668/hbnxb.2010.01.001

华北农学报 ›› 2010, Vol. 25 ›› Issue (1): 1-5. doi: 10.7668/hbnxb.2010.01.001

所属专题： 生物技术；

• 论文 • 下一篇

共表达O型口蹄疫病毒P12A+3C基因的重组杆状病毒的构建

田飞鹏^1,2, 曹轶梅², 卢曾军², 孙普², 高云英¹, 刘在新²

1. 西北农林科技大学,动物医学院, 陕西杨凌 712100;
2. 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,国家口蹄疫参考实验室,农业部畜禽病毒学重点开放实验室, 甘肃兰州 730046

收稿日期:2009-12-26 出版日期:2010-02-28
通讯作者: 高云英(1954-),女,陕西韩城人,副研究员,硕士,主要从事动物重大疫病防治的研究;刘在新(1964-),男,甘肃景泰人,研究员,博士,主要从事动物病毒分子生物学研究.
作者简介:田飞鹏(1985-),男,新疆伊犁人,在读硕士,主要从事动物病毒的研究.
基金资助:
国家"863"项目(2006AA10A204);"十一五"国家科技支撑计划项目(2006BAD06A03)

Construction of Recombinant Raculovirus Containing the P12A and 3C Genes of Foot and mouth Disease Virus Type O

TIAN Fei-peng1^1,2, 2², CAO Yi-mei2², LU Zeng-jun2², SUN Pu2¹, GAO Yun-ying1²

1. College of Veterinary Medicine, Northwest Agriculture and Forestry University, Yangling 712100, China;
2. Lanzhou Veterinary Research Institute of Chinese Academy of Agricultural Sciences, State Key Laboratory of Veterinary Etiological Biology, National Foot-and-Mouth Disease Reference Laboratory, Key Laboratory of Animal Virology of Ministry of Agriculture, Lanzhou 730046, China

Received:2009-12-26 Published:2010-02-28

摘要/Abstract

摘要： 构建共表达O型口蹄疫病毒(Foot and mouth disease virus,FMDV) 衣壳蛋白前体P12A和蛋白酶3C的重组杆状病毒,为进一步研究FMDV 空衣壳抗原和基因工程亚单位疫苗奠定基础.从质粒T-OP1中扩增出编码O型FMDV衣壳蛋白前体的P12A 基因,并将其插入到杆状病毒转移载体pFastDual-3C的PH启动子之下,构建重组杆状病毒转移载体pD-P12A3C.通过在大肠杆菌内转座重组,获得重组杆粒B-P12A3C,转染Sf9细胞,获得表达O型FMDV衣壳蛋白的重组杆状病毒.重组杆状病毒经增殖并感染Sf9细胞后,通过双抗体夹心ELISA方法及间接免疫荧光来检测目的蛋白的表达.结果表明,表达产物能被O型FMDV阳性血清识别,具有一定的反应原性,表明重组杆状病毒构建成功,该研究为O型FMDV空衣壳的体外组装及基因工程亚单位疫苗的研究提供了前期材料.

关键词: 口蹄疫病毒, 衣壳蛋白, 重组杆状病毒, 构建

Abstract: The capsid protein precursor P12A gene of foot-and-mouth disease virus(FMDV) type O were amplified from the plasmid T-OP1,the P12A were inserted into the baculovirus transfer vector pFast Dual-3C to construct recombinant transfer vector pD-P12A3C,in which the P12A and 3C were under the control of PH promoter and P10 promoter respectively.The recombinant plasmids were transformed into Escherichia coli DH10Bac(Invitrogen) to construct the recombinant bacmid B-P12A3C,and then,transfected into Sf9 cells,the recombinant baculovirus was harvested.After amplied,the recombinant baculovirus were infected into Sf9 cells.The expressed proteins were analyzed by an indirect sandwich-ELISA and by immuno uorescent assay.These results indicated that the expressed proteins were accurately expressed in Sf9 cells,and displayed specificity to FMDV type O antisera and biologic activation.From this study,The recombinant baculovirus containing the the capsid(P1) and 3C protease coding regions of FMDV type O were successfully obtained,thus providing a basis for the research of FMDV type O empty capsid assembly in vitro and empty capsid vaccine.

Key words: Foot-and-mouth disease virus, Capsid protein, Recombinant baculovirus, Construction

中图分类号:

田飞鹏, 曹轶梅, 卢曾军, 孙普, 高云英, 刘在新. 共表达O型口蹄疫病毒P12A+3C基因的重组杆状病毒的构建[J]. 华北农学报, 2010, 25(1): 1-5. doi: 10.7668/hbnxb.2010.01.001.

TIAN Fei-peng1,2,CAO Yi-mei2,LU Zeng-jun2,SUN Pu2,GAO Yun-ying1,LIU Zai-xin2. Construction of Recombinant Raculovirus Containing the P12A and 3C Genes of Foot and mouth Disease Virus Type O[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2010, 25(1): 1-5. doi: 10.7668/hbnxb.2010.01.001.

http://www.hbnxb.net/CN/Y2010/V25/I1/1

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共表达O型口蹄疫病毒P12A+3C基因的重组杆状病毒的构建

Construction of Recombinant Raculovirus Containing the P12A and 3C Genes of Foot and mouth Disease Virus Type O

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