摘要
以Ⅱ亚单纯疱疹毒-2333株(HSV-2)的基因组DNA为模板扩增编码HSV-2糖蛋白D(glyco-protein D)基因,与载体pFastBacⅠ连接,转化E.coilDH5α感受态细胞,经PCR及测序鉴定正确。将pFastBacⅠ-gD重组质粒转座至DH10Bac获得重组穿梭质粒,转染Sf9昆虫细胞,获得重组杆状病毒。重组杆状病毒感染Sf9细胞表达重组蛋白,经对表达条件进行优化,SDS-PAGE和Western blot鉴定,成功表达HSV-2 gD蛋白,为下一步的工作奠定了基础。
HSV-2 DNA was extracted and used as template in polymerase chain reactions to amplify gD gene. The PCR product was cloned into the donor plasmid pFastBac I . After the transformation and selection, the recombinant plasmid was sequenced. Then, the pFastBac I -gD was introduced into the competent cells E. coli DH10Bac, which containing a shuttel vector, Bacmid. The recombinant bacmid DNA was isolated and transfected into the insect cells Sf9 to produce the first generation recombinant virus. The in- sect Sf9 cells were infected with the recombinant virus to express the target protein. The expressed protein was detected by SDS-PAGE and Western blot. The gD gene was successfully expressed by Bac-to-Bac Baculovirus Expression System. This work was basic for the future.
出处
《动物医学进展》
CSCD
北大核心
2010年第6期58-61,共4页
Progress In Veterinary Medicine
