小鼠金属硫蛋白在聚胞藻中的金属诱导表达与纯化

应用蓝藻类金属硫蛋白基因启动子(smt O-P)的金属诱导性，在单细胞的聚胞藻PCC 6803中表达小鼠金属硫蛋白结构基因(mMT-1 cDNA)。在大肠杆菌HB 101中构建含有smt O-P和mMT1 cDNA的穿梭表达载体pKT-MRE，经质粒转移，链霉素筛选，Southern和Western杂交分析鉴定得稳定的转基因工程藻落。同时，做小批量锌诱导表达，并纯化了外源蛋白，5L培养液含鲜藻重5.0g，得到3.5mg mMT-1；转基因藻在高金属浓度下的耐受性测定表明，外源基因的表达提高了蓝藻对金属离子的抗性，约为野生藻的2倍。

Metal induced Expression of Mouse Metallothionein 1 cDNAin Synechocystis sp.PCC 6803

Cyanobacterium MT like promoter( smt O P) was used in expression of mMT 1 cDNA in Synechocystis sp.PCC 6803 to enhance metalbinding ability and specialty.Shuttle expression vector pKT MRE was constructed in E.coli (HB101),and through triparental conjugated transfer,Streptomycin screening,Southern and Western blotting analysis,transgenic cyanobacterium clone was acquired.The cultivation and Zn 2 inducing of transgenic cyanobacterium were carried on ,and the foreign mMT 1 was purified through the improved gel filtration and ion exchange chromatography.About 3.5mg mMT 1 were obtained from 5g wet cyanobacterial cells in 5L culture scale.Cyanobacterial metal resistance measurement indicated that the expression of foreign mMT 1 enhanced the transgenic cyanobacterial metalresistance to two times than the wild Synechocystis sp.PCC 6803