人胶源神经营养因子基因的克隆及在大肠杆菌中的表达

Cloning the Gene Coding for Human Glial Cell Line derived Neurotrophic Factor and Its Expression in Escherichia coli

人胶源神经营养因子基因的克隆及在大肠杆菌中的表达张晓霆马舒宁卫敏李昌本陈素珍赵寿元（复旦大学遗传学研究所遗传工程国家重点实验室上海２００４３３）胶源神经营养因子（Ｇｌｉａｌｃｅｌｌｉｎｅｄｅｒｉｖｅｄｎｅｕｒｏｔｒｏｐｈｉｃｆａｃｔｏｒ，ＧＤＮＦ...

By using PCR method,the hGDNF gene was cloned from Chinese genomic DNA.The hGDNF gene was subcloned into the expression vector pET3a which is under the control of T7 promoter and then transfected into E.coli BL21(DE3).The constructed strain was designated as pET3aTT/BL21(DE3).The sequence data of cloned hGDNF gene showed that it was identical to that of natural one.Four hours after IPTG induction,a 16kDa extra protein which is about 16.5% out of total bacteria proteins in pET3aTT/DE3(BL21) was appeared in the gel of SDS PAGE.The expression products existed in the form of inclusion body.The protein was purified viaion exchange chromatography.The results of amino acid sequencing proved that the sequence of 15 amino acids at the N terminal of the expressed hGDNF was the same as that deduced from DNA sequence.