表面展示猪圆环病毒2型衣壳蛋白的重组杆状病毒的构建与鉴定

›› 2013, Vol. 40 ›› Issue (7): 6-10.

表面展示猪圆环病毒2型衣壳蛋白的重组杆状病毒的构建与鉴定

靳立明, 张春雷, 廖明, 樊惠英

华南农业大学兽医学院, 农业部兽用疫苗创制重点实验室, 广东广州 510642

收稿日期:2012-12-06 出版日期:2013-07-20 发布日期:2013-07-24
通讯作者: 樊惠英 E-mail:fanhy@scau.edu.cn
作者简介:靳立明(1986-),男,河南人,硕士生,研究方向:预防兽医学。
基金资助:
广州市"珠江科技新星专项"项目(2012J2200086);公益性行业项目(201303034) ;863计划项目(2011AA10A209);973计划项目(2011CB504701)。

Construction and Identification of Recombinant Baculovirus Surface Displaying the Cap Protein of Porcine Circovirus Type 2

JIN Li-ming, ZHANG Chun-lei, LIAO Ming, FAN Hui-ying

MOA Key Laboratory for Animal Vaccine Development, College of Veterinary Medicne, South China Agricultural University, Guangzhou 510642, China

Received:2012-12-06 Online:2013-07-20 Published:2013-07-24

摘要/Abstract

摘要： 通过PCR方法扩增猪圆环病毒2型(porcine circovirus 2,PCV2)核定位序列部分缺失的ORF2基因(pdCap),将其克隆至Bac-to-Bac杆状病毒表达系统的pBACsurf-1质粒中,得到重组质粒pBACsurf-1-pdCap。再将含有pdCap和gp64的融合基因克隆到杆状病毒并转移到质粒pFastBac^TM Dual中,得到重组质粒pFastBac-pdCap。然后将该重组质粒转化至DH10Bac感受态细胞中,经蓝白斑筛选和PCR鉴定,得到重组穿梭质粒Bacmid-pdCap。在脂质体介导下转染Sf9昆虫细胞,获得重组杆状病毒BV-pdCap。间接免疫荧光和Western blotting试验结果表明,该重组杆状病毒成功表达了具有良好反应原性的pdCap蛋白。

关键词: 猪圆环病毒2型; pdCap蛋白; 杆状病毒表达系统

Abstract: The Cap gene partially deleted nuclear localization signal sequence(pdCap) of porcine circovirus 2(PCV2)was cloned into pBACsurf-1 between the upstream gp64 signal sequence and downstresm gp64 mature domain to construct the recombinant plasmid pBACsurf-1-pdCap.The fusion gene containing pdCap and gp64 was subcloned into pFastBac^TM Dual to construct the recombinant plasmid pFastBac-pdCap.And then the plasmid was transformed into E.coli DH10Bac competent cells to obtain the recombinant shuttle vector Bacmid-pdCap.Eventually,the recombinant Bacmid-pdCap was transfected into Sf9 cells to produce the recombinant baculovirus BV-pdCap mediating by Lipofectamine^TM2000.Indirect immunofluorescent and Western blotting assay indicated that the recombinant baculovirus BV-pdCap could express pdCap protein with significant antigencity.

Key words: porcine circovirus 2; pdCap protein; baculovirus expression vector system

中图分类号:

Q785

靳立明, 张春雷, 廖明, 樊惠英. 表面展示猪圆环病毒2型衣壳蛋白的重组杆状病毒的构建与鉴定[J]. , 2013, 40(7): 6-10.

JIN Li-ming, ZHANG Chun-lei, LIAO Ming, FAN Hui-ying. Construction and Identification of Recombinant Baculovirus Surface Displaying the Cap Protein of Porcine Circovirus Type 2[J]. , 2013, 40(7): 6-10.

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