乙型肝炎病毒核心抗原诱饵载体的构建及酵母表达

Construction and expression of a novel HBcAg of hepatitis B virus in yeast

目的构建乙型肝炎病毒核心抗原真核表达载体,并在酵母细胞中进行表达。方法以实验室保存质粒为模板,通过聚合酶链式反应(PCR)扩增HBcAg基因,克隆到pGEM-T载体。测序正确后酶切并连接至酵母表达载体pGBKT7中,转化酵母菌AH109,在色氨酸缺陷型培养基(SD/-Trp/Kana)上筛选阳性菌落后大量表达并提取重组蛋白,再进行SDS-PAGE和Western blot分析。结果成功构建酵母表达载体pGBKT7-HBcAg,Western blot结果显示重组蛋白在酵母细胞中正确表达。结论利用真核表达载体在酵母细胞中成功表达HBcAg蛋白,为研究HBV合并代谢性疾病的机制奠定了基础。

Objective To construct the eukaryotic expression vector of HBcAg gene and express HBcAg recombinant protein in yeast. Methods PCR was performed to amplify the gene of HBcAg from the preserved plasmid, and the gene was cloned into pGEM-T vector. After sequencing, the correct DNA frag- ment was cut and inserted into yeast expression plasmid pGBKT7, then transformed into yeast cell AH109 and screened on the SD/-Trp/Kana. Finally the yeast protein was isolated and analyzed with SDS-PAGE and Western blot. Results The eukaryotic expressive vector was constructed successfully and expressed correctly by results of Western blot. Conclusion The successful expression of HBcAg protein in yeast cells provided the foundation for studying the relationship between HBV and metabolism of glucose and lipid.