title: [Expression of human nerve growth factor gene in E. coli].
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.
  
  
  CONCLUSIONS
  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.
authors.affiliation: Institute of Biochemistry and Molecular Biology, West China School of Preclinical and Forensic Medicine, Sichuan University, Chengdu 610041, China.
authors.forename: Xiaoqiang
authors.initials: X
authors.lastname: He
authors.forename: Junjie
authors.initials: J
authors.lastname: Chen
authors.forename: Ruohan
authors.initials: R
authors.lastname: Wang
authors.forename: Leran
authors.initials: L
authors.lastname: You
authors.forename: Ji
authors.initials: J
authors.lastname: Liu
authors.forename: Jianye
authors.initials: J
authors.lastname: Chen
authors.forename: Jia
authors.initials: J
authors.lastname: Lin
mesh_terms: D001483:Base Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D020932:Nerve Growth Factor / Q000096:biosynthesis* / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D011993:Recombinant Fusion Proteins; D020932:Nerve Growth Factor