一种新细胞因子基因真核表达载体的构建和鉴定

Construction of Eukaryotic Expression Plasmid of a Novel Human B Lymphocyte Stimulator

采用PCR方法 ,以人胎盘cDNA文库为模板 ,扩增出人B淋巴细胞刺激因子 (hBLyS) ,经克隆测序及纯化后 ,再以此PCR产物为模板 ,用Nest PCR方法进一步扩增得到人B淋巴细胞刺激因子的胞膜外功能区域(hsBLyS)的DNA片段 ,纯化、克隆测序鉴定后 .扩增并纯化质粒 ,经酶切、纯化后克隆到真核表达质粒pcDNA3.1(+)中 ,构建成真核表达载体pcDNA3.1(+) /hsBLyS .结果表明 :用此方法制备得到的hsBLyS的DNA片段经测序鉴定与文献报道相符 ,构建的真核表达载体经鉴定也达到了预计的结果 .

Human B Lymphocyte Stimulator (hBLyS) was amplified by using PCR methods from cDNA library of human placenta.After purifying and sequencing, the DNA fragment of functional domain of hsBLyS was amplified by using Nest-PCR methods from the PCR product.The eukaryotic expression plasmid pcDNA3.1(+)/hsBLyS was constructed with reconstruction experiment after purifying and identifying of hsDNA fragment.The experimental result shows that the sequence of the PCR product is coincident with the report of Moore et al.