牛疱疹病毒Ⅰ型gB、gE基因的克隆及其杆状病毒表达载体的构建

Cloning of gB,gE Gene of Bovine Herpesvirus-1 and Construction of its Baculovirus Expression Vector

本试验用PCR方法扩增了牛疱疹病毒Ⅰ型(bovine herpesvirus-1,BHV-1)Bartha Nu/67株gB、gE基因片段,将其克隆到pGEM-T-easy载体。经转化、筛选、鉴定后将重组质粒经BamHⅠ和EcoRⅠ双酶切后,与经相同方法处理的杆状病毒转移载体pFastBacHTb连接,得到了重组质粒pFBHgB、pFBHgE。经酶切和测序鉴定后,将其转化入含穿梭载体Bacmid的感受态细胞DH10Bac,经抗性、蓝白斑筛选和PCR鉴定,得到了含gB、gE基因的重组穿梭载体。

The gB and gE gene of bovine herpesvirus-1 Bartha Nu/67 strain were amplified by PCR. The gB and gE frag- ment were cloned into pGEM-T-easy vector. The gB and gE gene were subcloned into baculovirus transfer vector and the re- combinant Baculovirus vector pFBHgB, pFBHgE were constructed successfully. Then they were transferred into E. coli DH10Bac and cultured in LB plate. The white bacterial colonies were positive recombinant bacmid named as BacmidgB, Bac- midgE. Thus the result mentioned above laid down theoretical and practical foundations for the expression of gB and gE gene in insect cells.