猪干扰素-γ基因在毕赤酵母中的分泌表达  

Secreted Expression of Porcine Interferon-gamma Gene in Pichia pastoris

作　　者：黄志清[1] 胡宏宇[1] 陈小玲[2] 任立明[1] 林爱星[1] 陈永福[1] 

机构地区：[1]中国农业大学农业生物技术国家重点实验室,生物学院,北京100094 [2]中国农业大学动物科技学院,北京100094

出　　处：《生物工程学报》2005年第5期731-736,共6页Chinese Journal of Biotechnology

基　　金：国家自然科学基金资助项目(No.30370726)。~~

摘　　要：将去除信号肽的猪干扰素γ(PoIFNγ)基因置于酿酒酵母α因子分泌信号的DNA序列后,构建成pPIC9KαPoIFNγ分泌型重组表达载体,电转化导入毕赤酵母GS115中,经G418筛选后获得2株多拷贝插入的重组子。SDSPAGE和Westernblot分析结果表明,所获得的重组子能够分泌表达出17kD和23kD左右的PoIFNγ特异蛋白,其表达量为108mgL,占培养液总蛋白的60%。实验首次在毕赤酵母表达系统中实现了PoIFNγ基因的分泌表达。The porcine interferon-gamma（PoIFN-γ） gene, in which the sequence encoding signal peptide was replaced by that of the a-factor of Saccharomyces cerevisiae, was cloned into Pichia pastoris expression vector pPIC9K. The recombinant plasmid pPIC9K-α-PoIFN-γ was then transformed into Pichia pastoris GS115 cells by electroporation and stable multicopy recombinant Pichia pastoris strains were selected by G418 resistance. Two recombinants of multiple inserts were obtained. SDS-PAGE and Western blot assays of culture broth from a methanol-induced expression strain demonstrated that recombinant PoIFN-γ, 17kD and 23kD proteins, were secreted into the culture medium. Target proteins, 60% of total proteins, were obtained in the culture medium at the concentration of 108mg/L. This is the first secreted expression of porcine interferon-gamma gene in Pichia pastoris .

关 键 词：猪干扰素-γ 毕赤酵母 分泌表达 

分 类 号：Q786[生物学—分子生物学]