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目的 用含乙型肝炎病毒 (HBV)前 S区不同大小的c DNA片段构建酵母双杂交诱饵载体 ,并检测其表达产物对酵母细胞有无毒性作用及对报告基因有无激活作用 .方法 PCR扩增含 HBV前 S区不同大小的 c DNA片段 ,分别克隆入 p U C19质粒 ,经测序正确后 ,再分别亚克隆入酵母双杂交诱饵载体 p GBKT7中 .将重组质粒导入酵母菌 AH10 9,检测其表达产物在酵母细胞中对报告基因有无激活作用 .结果 成功获得含 HBV前 S区不同大小的 c DNA片段 ,HBV前 S区不同片段所表达的蛋白对酵母菌 AH10 9无毒性 ,但是对报告基因均有激活作用 .结论 不能利用酵母双杂交 Gal4系统 3来研究与 HBV前 S区相互作用的蛋白 ;证实了 HBV前S区不同片段所表达的蛋白的自激活功能 ,为进一步研究乙型肝炎病毒致病机制打下了基础
Objective To construct yeast two-hybrid bait vector containing different sizes of pre-S region of hepatitis B virus (HBV), and to detect whether the expressed product has toxicity to yeast cells and whether the reporter gene has activation or not. The DNA fragments containing different sizes of pre-HBV S region were cloned into pUC19 vector and subcloned into the yeast two-hybrid bait vector pGBKT7 after correct sequencing.The recombinant plasmids were introduced into yeast AH109 and detected And the expression products of them had no effect on the reporter gene in yeast cells.Results The c DNA fragments containing different sizes of pre-HBV S region were successfully obtained.The proteins expressed in different fragments of pre-HBV region were non-toxic to yeast strain AH109, The reporter gene can activate.Conclusion The yeast two-hybrid Gal4 system 3 can not be used to study the protein interacting with the pre-HBV region, and the self-activation of proteins expressed in different segments of pre-HBV S region was confirmed. Hepatitis virus pathogenesis laid the foundation