| 人神经生长因子基因在大肠杆菌中的表达 |
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| 摘 要： |
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| 关 键 词： | 人基因组DNA 神经生长因子 大肠杆菌 基因表达 基因治疗 |
Expression of human nerve growth factor gene in E. coli] |
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| Xiaoqiang He,Junjie Chen,Ruohan Wang,Leran You,Ji Liu,Jianye Chen,Jia Lin. Expression of human nerve growth factor gene in E. coli][J]. Journal of West China University of Medical Sciences, 2002, 33(1): 94-97 |
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| Authors: | Xiaoqiang He Junjie Chen Ruohan Wang Leran You Ji Liu Jianye Chen Jia Lin |
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| Affiliation: | Institute of Biochemistry and Molecular Biology, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China. |
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| Abstract: | OBJECTIVE: To investigate the structure and function of human nerve growth factor (beta-NGF) and the gene encoding beta-NGF. METHODS: A pair of specific primers (29 mer) for the sequence encoding human beta-NGF was designed and synthesized. A 380 bp fragment was amplified from human blood genomic DNA by polymerase chain reaction, and cloned into pGEM-T Easy vector. The identified insert fragment from the recombinant pGEM-T-NGF was directionally ligated with linearized pGEX-5T with the compatible termini. E. coli JM 109 was transformed with the expression recombinant p5TNGF and induced by IPTG. RESULTS: The cloned DNA fragment was identified as the full-length sequence encoding human beta-NGF by restriction analysis and DNA sequencing. SDS-PAGE and Western blot revealed the cloned NGF gene expressed as a fusion protein (40.5 x 10(3) u) in the cells transformed by p5TNGF. The soluble fusion protein was determined to be 503.2 mg/L, accounting for 6.8% of the total soluble protein (7.4 g/L) of bacterial cells. This fusion protein was found to have antigenic activities of NGF. CONCLUSION: The clone containing the full-length sequence encoding human beta-NGF is obtained and successfully expressed in E. coli to be of use for studying the biological functions of human beta-NGF gene. |
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