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目的利用Bac-to-Bac杆状病毒表达系统对单纯疱疹病毒II型(HSV-2)糖蛋白G(gG-2)免疫优势片段gG321-580进行表达,纯化并评价其反应原性,以探索HSV-2免疫诊断试剂的研制。方法提取HSV-2DNA作为模板,PCR扩增gG321-580基因,克隆至pFastBac HTC载体中,通过Bac-to-Bac杆状病毒表达系统表达His-gG321-580融合蛋白,SDS-PAGE和Western-blotting鉴定表达产物,NTA-Ni 2+纯化后间接ELISA评价其反应原性。结果 HSV-2gG321-580在昆虫细胞Sf9中得到特异性表达,纯化后的重组gG321-580蛋白对HSV-2阳性血清抗体有较好的抗原性和特异性。结论 gG321-580蛋白在Bac-to-Bac杆状病毒表达系统中成功表达,表达产物表现出良好的反应原性,为发展HSV-2免疫诊断试剂奠定了基础。
Objective To express, purify and evaluate the immunogenicity of gG321-580 of HSV-2 glycoprotein G (gG-2) by Bac-to-Bac Baculovirus Expression System Development of HSV-2 Immunodiagnostic Reagents. Methods HSV-2 DNA was extracted as a template and gG321-580 gene was amplified by PCR and cloned into pFastBac HTC vector. His-gG321-580 fusion protein was expressed by Bac-to-Bac baculovirus expression system. SDS-PAGE and Western-blotting The expression products were identified, and their reactionogenicities were evaluated by NTA-Ni 2+ purified indirect ELISA. Results HSV-2gG321-580 was specifically expressed in insect cells Sf9. The purified recombinant gG321-580 protein had good antigenicity and specificity for HSV-2 positive serum antibody. Conclusion The recombinant gG321-580 protein was successfully expressed in Bac-to-Bac baculovirus expression system. The expressed product showed good reactivity and laid the foundation for the development of HSV-2 immunodiagnostic reagent.