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目的:利用Bac-to-Bac杆状病毒表达系统对Ⅱ型单纯疱疹病毒(HSV-2)糖蛋白D(gD2)进行表达,纯化后评价其免疫效果,以探索HSV-2重组亚单位疫苗的研制。方法:提取HSV-2DNA作为模板,用聚合酶链式反应(PCR)扩增gD2基因,克隆至pFastBac HTA载体中,利用载体中的His基因标记gD2,通过Bac-to-Bac杆状病毒表达系统表达His-gD2融合蛋白,表达产物进行SDS-PAGE和Western-blot检验,用镍柱进行纯化,免疫小鼠评价其免疫原性。结果:HSV-2gD2在昆虫细胞Sf9中得到特异性表达,纯化后的重组蛋白诱导小鼠体内产生高水平的IgG抗体及中和抗体。结论:gD2基因在Bac-to-Bac杆状病毒表达系统中获得表达,表达产物表现出良好的免疫原性及中和活性,为发展HSV-2亚单位疫苗奠定了基础。
OBJECTIVE: To express glycoprotein D (gD2) of herpes simplex virus type 2 (HSV-2) by Bac-to-Bac baculovirus expression system and evaluate its immunogenicity after purification to explore the feasibility of HSV-2 recombinant subunit vaccine Development. Methods: The gD2 gene was amplified by polymerase chain reaction (PCR) and cloned into pFastBac HTA vector. The HisD gene was labeled with gD2 and the Bac-to-Bac baculovirus expression system The His-gD2 fusion protein was expressed. The expressed product was analyzed by SDS-PAGE and Western-blot, and purified by nickel column. Immunized mice were used to evaluate the immunogenicity. Results: HSV-2gD2 was specifically expressed in insect cells Sf9. The purified recombinant protein induced high levels of IgG and neutralizing antibodies in mice. CONCLUSION: The gD2 gene is expressed in Bac-to-Bac baculovirus expression system. The expressed product shows good immunogenicity and neutralizing activity, which lays the foundation for the development of HSV-2 subunit vaccine.