title: [Cloning, expression and purification of silent information regulator 2 from Giardia lamblia].
abstract: OBJECTIVE
  To clone and express silent information regulator 2 (Sir2) gene from Giardia lamblia.
  
  
  METHODS
  The GlSir2 gene was amplified by PCR from genomic DNA of Giardia lamblia (Chinese strain C2 clone). PCR product was cloned into pMD-19T vector and transformed into E coli JM109. The recombinant plasmid was sequenced and then cloned into the pET28b vector. The pET28b-Gl1Sir2 recombinant plasmid was transformed into E. coli BL21(DE3), followed by expression of the protein induced by IPTG. The recombinant protein was analyzed by SDS-PAGE. Inclusion bodies were dissolved with 8 mol/L urea, and the supernatant was collected and applied to Ni2+ affinity chromatography. The purified recombinant protein was renatured by dialysis and verified by Western blotting using anti-His tag antibody.
  
  
  RESULTS
  GlSir2 gene sequence was cloned. The GISir2 open reading frame (1 680 bp) encoded a 559-amino acid protein with Mr 62 800. The recombinant plasmid pET28b-GlSir2 expressed an inclusion body protein of GISir2 after being induced with IPTG. The protein purity reached above 80% after purification. The purified protein was renatured by dialysis. The recombinant GISir2 was recognized by anti-His tag antibody.
  
  
  CONCLUSIONS
  The coding sequence of GLSir2 gene was cloned and expressed in vitro. The recombinant protein was identified by anti-His tag antibody.
authors.affiliation: Medical School, Dalian Uninversity, Dalian 116622, China.
authors.forename: Xia
authors.initials: X
authors.lastname: Zhang
authors.forename: Hong-Mei
authors.initials: HM
authors.lastname: Ju
authors.forename: Yun-Hua
authors.initials: YH
authors.lastname: Wang
authors.forename: Ya-Jie
authors.initials: YJ
authors.lastname: Li
mesh_terms: D001483:Base Sequence; D003001:Cloning, Molecular; D015870:Gene Expression; D005822:Genetic Vectors; D016829:Giardia lamblia / Q000235:genetics* / Q000378:metabolism; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism*; D037761:Sirtuins / Q000235:genetics / Q000378:metabolism*
chemical_list: D015800:Protozoan Proteins; D037761:Sirtuins