摘要
目的:在毕赤酵母中融合表达人骨保护素(OPG)片段OPG179与人血清白蛋白(HSA)。方法:通过RT-PCR扩增获得OPG179基因,构建表达质粒pHILD2-rhOPG179-HSA,转化至毕赤酵母菌中进行表达、纯化,对分泌表达产物进行SDS-PAGE及Western印迹检测。结果:酶切鉴定与测序结果显示重组表达质粒pHILD2-rhOPG179-HSA正确;经Western印迹检测,融合蛋白rhOPG179-HSA的相对分子质量约为97×103,符合预期值;rhHSA-OPG179在毕赤酵母中的表达量约为80 mg/L。结论:表达并纯化制备了重组骨保护素片段与人血清白蛋白的融合蛋白,为后续的生物学活性研究奠定了一定的基础。
Objective：To express in Pichia pastoris and purify recombinant human osteoprogerin（OPG） fragment OPG^179 and human serum albumin（HSA） fusion protein.Methods：The gene of OPG^179 was amplified by RT-PCR.The plasmid of pHILD2-rhOPG^179-HSA was constructed and then transformed to Pichia pastoris.The rhOPG^179-HSA fusion protein was expressed,purified,and analyzed with SDS-PAGE and Western blot.Results：The expression plasmid of pHILD2-rhOPG^179-HSA was proved correct after the plasmid was identified and sequenced.The molecular mass of fusion protein was about 97 kD.The rhOPG^179-HSA fusion protein was expressed and purified successfully from Pichia pastoris with relative higher-efficient expression level（about 80 mg/L）.Conclusion：The secreted recombinant fusion protein rhOPG^179-HSA has been successfully expressed and purified,and supply for a foundation for its biological activity analysis
出处
《生物技术通讯》
CAS
2011年第1期15-18,23,共5页
Letters in Biotechnology
基金
总装后司令部及卫生局项目(07ZH10)
306医院院级课题(07QN10)
关键词
人骨保护素
人血清白蛋白
融合蛋白
毕赤酵母
表达
纯化
recombinant human osteoprogerin
human serum albumin
fusion protein
Pichia pastoris
expression
purification
