【摘要】 通过寡核苷酸合成仪,用化学方法合成相当于hcGβ链C末端36肽基因的片段,成功地克隆到噬菌体载体λgt11中,并在宿主菌株E. coli y₁₀₈₉中得到表达。含λgt11 hCG重组体的E. coli y₁₀₈₉所合成的含人类绒毛膜促性腺激素β链c末36肽的杂交蛋白（下称:β—galactosidase—hCG）,通过硫酸铵沉淀、亲和层析和制备性聚丙烯酰胺凝胶电泳,可得电泳纯单一色带的β—gaIactosidase—hCG。蛋白质转印（Western Blot）技术证实β—galactasidase—hCG具有hCGβ链c末端36肽的免疫特性。对流免疫电泳和免疫双扩散试验结果表明β—galactosidase—hCG具免疫原性。从一立升培养液的菌体中可分离纯化约3.8毫克的β—galaetosidase—hCG。更多还原

【Abstract】 The present investigation has been earried out to study hCG B-subunit C-terminal containing hybrind protein synthesized by λgt11 hCG Y1089 through genetic engineering technology. A 36 peptide gene containing h CG B-subunit C-terminal has been artificially synthesized by Oligonucleotide Synthesizer, the double stranded DNA was cloned into an expression vetor λgt11 and the 36 peptide gene fusion product was successfully expressed in host strain E. coli Y1089. h CG B-subunit C-terminal 36 peptide containing hybrid protein (B-galactosidase-h CG) expressed in λgt11 hCG Y1089 was precipitated in the ammonium sulfate solution of 50% saturation, affinity isolated by anti-B-galactosidase sepharose 4B column and finally purified with preparative polyacrylamide gel electrophoresis. Comparative analysis of different preparations revealed that the eluate of affinity column gave a mjor band and two minor bands, and the eluate of preparative polyacrylamide gel electrophoresis only gave one band while the solution of ammonium sulfate precipitated gave more than 30 bands in polyacrylamide gel electrophoresis, it indicated that anti-B-galactosidase affinity chromatography and preparative polyacrylamide gel eleetrophoresis can be used for isolating and purifing B-galactosidase-h CG. The results of western blot technique demonstrated that B-galactosidase-hCG contains immunological property of hCG B-subunit C-terminal. Preliminary studies of antigenicitY of this B-galactosidase-hCG through counter current immunoelectrophoresis and double immunodiffusion techniques have clearly demonstrated that this Purified B-galactosidase-hCG can be crossly immunopreciPitated with rabbit anti-B-galactosidase-hCG serum, it has been concluded that B-galactosidase-hCG possesses the antigenieitY, but the biological activity of this this hybrid protein to neutralizing hCG has to be determined by in vitro or in vivo system.更多还原