title: [Cloning and expression of human interleukin-32 and studies on its bioactivity].
abstract: OBJECTIVE
  To clone human interleukin-32(hIL-32) gene and express it in E.coli efficiently.
  
  
  METHODS
  The gene of hIL-32 was amplified by RT-PCR from human peripheral blood mononuclear cells (PBMC) which stimulated with Con A for 60 h. The PCR product was inserted into the pMD18-T vector. The hIL-32 cDNA confirmed by sequencing was inserted into expression vector pET-30a(+) and expressed in E.coli BL21(DE3) strain. The hIL-32 protein expression was induced by IPTG and assayed by SDS-PAGE and coomassie blue stain. The recombination protein was identified by Western blot and its biological activity was analyzed.
  
  
  RESULTS
  DNA sequencing confirmed that the cloned cDNA was identical to the published sequence of hIL-32 that the nucleotide sequence of this gene was 567 bp. The recombinant plasmid pET30a-hIL32 was transformed into E.coli BL21(DE3) strain for expression. An expected 28 kDa protein of hIL-32  was found mainly in the induced host strains by SDS-PAGE and coomassie blue stain. The 28 kDa protein was recognized by anti-IL-32 antibody in western blot. The purified recombination protein could induce the producing of IL-6 in the human PBMC.
  
  
  CONCLUSIONS
  We have successfully cloned the gene and expressed the protein of hIL-32 and the expressed protein has specific bioactivity.
authors.affiliation: School of Public Health, Taishan Medical University, Taian 271016, China. zhaojutian@163.com
authors.forename: Zhao-ju
authors.initials: ZJ
authors.lastname: Tian
authors.forename: Xin-quan
authors.initials: XQ
authors.lastname: Jiang
authors.forename: Qiang
authors.initials: Q
authors.lastname: Chen
authors.forename: Xi-zhu
authors.initials: XZ
authors.lastname: Xu
authors.forename: Yu-hong
authors.initials: YH
authors.lastname: Ma
authors.forename: Feng-ping
authors.initials: FP
authors.lastname: Jiao
authors.forename: Wen-jing
authors.initials: WJ
authors.lastname: Ye
mesh_terms: D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D006801:Humans; D007378:Interleukins / Q000235:genetics* / Q000302:isolation & purification / Q000494:pharmacology; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000302:isolation & purification / Q000494:pharmacology
chemical_list: C496971:IL32 protein, human; D007378:Interleukins; D011994:Recombinant Proteins