Gag/IFNα-2b融合基因在痘苗病毒中共表达研究

Study on coexpression of HIV-1 gag/IFNα-2b fusion gene in vaccina virus

目的 本实验将编码AIDS病毒 (HIV -1)核心蛋白gag与编码干扰素 (IFNα -2b)基因插入以痘苗病毒复制非必需区血凝素 (HA)基因为侧翼 ,牛痘病毒包涵体 (ATI)启动子和串联的p7 5突变型早期启动子为基本构件的痘苗病毒表达载体 (pJ3 8) ,经与野生型痘苗病毒在细胞内同源重组 ,获得具有生物活性的重组痘苗病毒vJ3 8gag/IFNα -2b。方法 经免疫荧光、Dot -ELISA、SDS -PAGE、Western -blot等方法检测表达产物 ,分析免疫小鼠后血清抗体IgGOD4 90 值与T淋巴细胞的变化。结果 vJ3 8gag/IFNα -2b能表达Gag/IFNα -2b融合蛋白 ,分子量约 60kDa。免疫小鼠实验表明血清抗体IgGOD4 90 值、CD+ 4、CD+ 8T淋巴细胞计数的组间比较 (P <0 0 0 1) ,含IFNα -2b与单纯vJ3 8gag组相比 (P >0 0 5) ,但呈渐进性增高的趋势。 结论 重组痘苗病毒表达的gag/IFNα

ObjectivepJ38 was used as expression vector,gag gene enc oding type Ⅰ core protein of AIDS virus(HIV) and encoding interferon α-2b gene of human were inserted into the downstream of the hybrid promotor,where HA gene of the vaccinia virus unnecessary areas was the flank and there were 38p7.5 prom otor of mutant-type vaccinia virus,A-type inclusion late promotor of vaccinia vi rus.A vJ38gag/IFNα-2b was acquired by homologous recombination with the natural vaccinia virus as selection of the HA - plaque for reporter.MethodsThe IFA,Dot-blot,SDS-PAGE,Western-blot and FCM we re used.ResultsThe recombinant viruses could express gag/IFNα-2 b,whose molecular weight was 60?kDa.In the mice immunization experiment,the OD 490 value of the serum antibodies of HIV-1 was different(P<0 001) i n groups,There was not difference between containing IFNα-2b and vJ38Gag.ConclusionGag/IFNα-2b proteins both had good reactinoge nicity and immunogenictiy.