摘要
目的 :构建艾滋病病毒核心蛋白 (gag)与干扰素 (IFNα - 2b)融合基因表达质粒 ,观察其在痘苗病毒中共表达结果 ,研究其意义。方法 :利用基因重组技术 ,将IFNα - 2b基因片段插入到gag基因的nt5 31位点 ,经脂质体转染与血凝素阴性蚀斑筛选 ,挑出重组痘苗病毒。经免疫荧光、Westernblot和Dot-ELISA鉴定表达产物。结果 :间接免疫荧光实验结果显示 ,转染重组质粒的细胞表面有绿色荧光。免疫印迹实验与Dot-ELISA结果均显示重组质粒转染细胞的裂解物中存在表达的gag IFNα - 2b蛋白。结论 :成功地构建了重组真核细胞表达质粒 ,表达的蛋白具有良好的免疫原性与免疫反应性。
Objective:To construction an eukaryotic expression plasmid and express fusion of HIV-1gag/IFNα-2b in BHK21cell.Methods:An eukaryotic expression plasmid pSFJ38gag/IFNα-2b was constructed by inserting IFNα-2b gene into site (nt531) of gag gene and inserting HIV-1gag/IFNα-2b gene into vaccine vector plasmid pSFJ38,and transfected to BHK21 cell in vitro by liposome-mediated method.The expressed product was detected by indirect immunofluorescence assay(IFA),Western blot and Dot-ELISA.Results:Indirect IFA showed green fluorescence on the surface of transfected cell.Both Western blot and Dot-ELISA proved that the lysate of transfected cells contained gag/IFNα-2b protein.Conclusion:An eukaryotic expression plasmid for HIV-1gag/IFNα-2b was successfully constructed,and the expressed product showed good reactinogenicity and immunogenictiy.
出处
《生物技术》
CAS
CSCD
2003年第6期6-8,共3页
Biotechnology
