鲤生长激素酵母工程菌的分泌表达和产物纯化  

Secreted expression and purification of common carp GH in yeast

作　　者：李英华[1,2] 白俊杰[1] 简清[1] 叶星[1] 李新辉[1] 罗建仁[1] 梁旭方[2] 

机构地区：[1]中国水产科学研究院,珠江水产研究所,农业部热带亚热带鱼类选育与养殖重点开放实验室,广东广州510380 [2]暨南大学生命科学技术学院,生物工程系,广东广州510632

出　　处：《中国水产科学》2002年第4期289-292,共4页Journal of Fishery Sciences of China

基　　金：农业部"九五"渔业科技重点项目 (95 -B -96-0 2 -0 2 -0 4) ;中国水产科学研究院重点基金项目 (99-0 8-0 2 )

摘　　要：取本室构建的鲤生长激素毕赤酵母工程菌GS115 (pPICZαA -GH) ,甲醇诱导 ,SDS -PAGE检测 ,证明重组鲤生长激素 (RecombinantCarpGrowthHormone ,rcGH)在酵母中得到分泌表达。重组酵母菌生长曲线显示 ,细胞从培养第 8小时进入对数生长期 ,至 32hOD660 升至峰值 11.5 ,此后OD660 缓慢下降。在以 10 0 %甲醛按 0 .5 %添加量诱导至 72h、pH为 6 .0时 ,rcGH表达量最高。凝胶扫描分析 ,分泌表达的rcGH占上清总蛋白的 36 .4 1% ,表达量为 30 0～ 4 0 0mg/L发酵液。经离子交换层析法纯化表达产物 ,纯度可达 95 %。纯化的rcGH注射奥尼罗非鱼 。The recombined carp growth hormone yeast Gs115(pPICZαA-GH)was cultured in shaking flask at 28-30 ℃ and 200-300 r/min for a night and then induced by methanol for 4 d. The carp growth hormone rcGH was expressed in the yeast GS115(pPICZαA-GH). In order to investigate the influence of inducing time duration and pH on rcGH expression, the samples were designed at four inducing durations (24, 48, 72 and 96 h) and four pH levels (4.5, 5.5, 6.0 and 6.5) to be detected. The results show that when induced for 72 h, rcGH expression reaches the highest level and the optimum pH is 6.0. By analysis of Tiger Gel Image Scanner, the secreted rcGH accounts for 36.41% of the total supernatant proteins with the yield of 300-400 mg/L. The purified product rcGH was obtained by use of Q Sepharose anion exchange chromatography and was dissolved in sterile saline and injected into tilapia intraperitoneally at the dose of 0.1 μg/g body weight for one test group and 1 μg/g body weight for another. Six weeks later the later group got higher growth rate and both of the two groups had higher growth rates than the control.

关 键 词：鲤 生长激素 酵母工程菌 分泌表达 产物纯化 

分 类 号：S965.116[农业科学—水产养殖]