gp350胞外段基因表达载体的构建与表达

Construction and expression of gp350 eukaryotic expression vector

目的构建gp350与C3d融合基因表达载体,并在体外进行表达和鉴定。方法以pGEX-gp350为模板,经PCR获得长846bp的gp350胞外段(25-870),将该扩增片段插入pSG.SS.C3d3.YL载体,构建pSG.SS.gp350.C3d3.YL表达质粒。经限制性内切酶鉴定和DNA序列测定证实后,将该质粒转染Hela细胞,检测其体外表达。结果免疫细胞化学分析结果表明转染细胞有目的分子的表达。结论构建的重组载体可在真核细胞内正确表达,这为基于gp350的鼻咽癌预防性疫苗下一步的动物实验奠定了基础。

Objective To construct the eukaryotic expression vector for gp350 and C3d fusion gene and to express it in vitro. Methods gp350 was amplified by PCR using pGEX-gp350 as a template,and the PCR product was inserted into eukaryotic expression vector pSG. SS. C3d3. YL. The recombinant plasmid was transfected into Hela cells and the expressed product was detected by immunocytoehemistry. Results Immunocytochemistry demonstrated that gp350 existed in the cytoplasm of Hela cells transfected by recombinand plasmid. Conclusion The constructed DNA vaccine can be expressed in vitro,which may pave a way for further studies in animals about gp350-based nasopharyngeal carcinoma vaccine.