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为探讨乙型肝炎病毒 (HBV)前S1蛋白的功能 ,在真核生物酵母细胞中表达HBV前S1基因。以HBVayw亚型全长质粒pCP10为模板 ,多聚酶链反应 (PCR)扩增HBV前S1基因 ,克隆到pGEM T载体中 ,双酶切后回收与酵母表达质粒pGBKT7连接。将重组载体转化酵母细胞AH10 9,提取酵母蛋白质 ,进行十二烷基磺酸钠 聚丙烯酰胺凝胶电泳 (SDS PAGE)和Western免疫印迹分析。结果成功地构建了HBV前S1基因酵母表达载体 ,Western免疫印迹显示HBV前S1在酵母细胞中表达 ,表达产物在胞内存在 ,分子量 30kD左右。表明HBV前S1蛋白在酵母细胞中表达成功。
To investigate the function of pre-S1 protein of hepatitis B virus, pre-S1 gene of HBV is expressed in eukaryotic yeast cells. The HBV pre-S1 gene was amplified by polymerase chain reaction (PCR) using the full-length plasmid pCP10 of HBVayw subtype as a template and cloned into pGEM T vector. After double enzyme digestion, it was ligated with the yeast expression plasmid pGBKT7. The recombinant vector was transformed into yeast cell AH10 9, and the yeast protein was extracted for sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS PAGE) and Western blot analysis. Results The yeast pre-S1 gene expression vector was successfully constructed. Western blotting showed that HBV pre-S1 was expressed in yeast cells. The expressed product existed in the cytoplasm with a molecular weight of about 30 kD. It showed that HBV pre-S1 protein was successfully expressed in yeast cells.