梅花鹿超氧化歧化酶的克隆与原核表达

Clone and Expression of a Novel Copper/Zinc-Superoxide Dismutase from Cervus nippon Temminck

目的:利用分子生物学的方法克隆梅花鹿超氧化歧化酶(Cu,Zn-SOD)基因,并将其于大肠杆菌(E.coli)中表达。方法:通过设计扩增引物及RT-PCR技术从梅花鹿肝细胞中克隆出Cu,Zn-SOD基因,将其构建于表达质粒pET21a(+)并转入E.coli Rosetta(DE3)。结果:通过IPTG诱导表达,在Rosetta(DE3)中成功表达出具Cu,Zn-SOD活性的梅花鹿超氧化物歧化酶,三角摇瓶发酵水平约257U/ml发酵液;经过纯化后得到比活3092U/mg的梅花鹿超氧化物歧化酶。结论:获得了梅花鹿的Cu/Zn-SOD基因的全序列,并使其得到表达,对研究鹿茸的功效和保护梅花鹿这一物种具有一定意义。

Objective：A gene encoding a novel copper/zinc-superoxide dismutase （ Cu,Zn-SOD） from Cervus nippon Temminck was cloned and expressed in E.coli Rosetta（ DE3） .Method：By using the method of RT-PCR and PCR,the gene was cloned from the liver cells of Cervus nippon Temminck and fused with the vector pET21a（＋） .After the new vector pET21a（＋）-SOD had been transformed into Rosetta （ DE3） ,the recombinant Rosetta（ DE3） was induced by IPTG.Result：The results showed that a copper/zinc-superoxide dismutase from Cervus nippon Temminck could be successfully expressed with activity 257U/ml.Conclusion：The Cu ,Zn-SOD of Cervus nippon Temminck with enzyme activity was successfully expressed in E.coli Rosetta（ DE3） .