title: [Expression of human Id-2 gene in Escherichia coli and preparation of the antisera against human Id-2].
abstract: OBJECTIVE
  To express the fusion protein of glutathione S-transferase (GST) and human Id-2 in E. coli and prepare the polyclonal antibodies against Id-2.
  
  
  METHODS
  The coding sequence of Id-2 gene was amplified by RT-PCR from the total RNA of breast cancer tissue. The recombinant plasmid was identified by PCR, restriction endonuclease digestion analysis and sequencing. The fusion protein GST-Id-2 expressed in E. coli following IPTG induction was purified by glutathione-agarose affinity chromatography and used to immunize rabbits to prepare the polyclonal antibodies against GST-Id-2.
  
  
  RESULTS
  PCR, restriction endonuclease digestion and sequence analyses showed that the Id-2 gene had been correctly inserted into pGEX-6P-1 vector, and the GST-Id-2 fusion protein expressed had a relative molecular mass of approximately 40,000 as shown by SDS-PAGE. The polyclonal antibodies obtained from the rabbit sera were found to specifically react with purified Id-2 by Western blotting, ELISA and agar gel immunodiffusion (AGP).
  
  
  CONCLUSIONS
  The prepared polyclonal antibodies against Id-2 allow effective Id-2 detection and facilitate further investigation of the structure and antigen epitope of Id-2.
authors.affiliation: National Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research institute, Chinese Academy of Agricultural Sciences, Harbin, China. teargone2003@yahoo.com
authors.forename: Tie-Gang
authors.initials: TG
authors.lastname: Tong
authors.forename: Yan
authors.initials: Y
authors.lastname: Lin
authors.forename: Dan-Mei
authors.initials: DM
authors.lastname: Mu
authors.forename: Yu
authors.initials: Y
authors.lastname: Bai
authors.forename: Mu-Lei
authors.initials: ML
authors.lastname: Yang
authors.forename: Min
authors.initials: M
authors.lastname: Zheng
authors.forename: Dong-Lai
authors.initials: DL
authors.lastname: Wu
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000276:immunology; D001943:Breast Neoplasms / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005260:Female; D006801:Humans; D007106:Immune Sera / Q000096:biosynthesis; D051797:Inhibitor of Differentiation Protein 2 / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D011817:Rabbits; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics
chemical_list: D000911:Antibodies, Monoclonal; C497450:ID2 protein, human; D007106:Immune Sera; D051797:Inhibitor of Differentiation Protein 2; D011994:Recombinant Proteins