title: [Prokaryotic expression and purification of human GST-Cdc25C fusion protein and preliminary detection of its function].
abstract: OBJECTIVE
  To clone prokaryotic expression vector of Cdc25C,  purify the fusion protein of GST-Cdc25C,  and identify its function preliminarily.
  
  
  METHODS
  Human Cdc25C coding region was amplified from human mammary cDNA library by PCR,  and cloned into the prokaryotic expression vector pGEX-KG. The fusion protein GST-Cdc25C was expressed in E.coli Rossate and purified by GST-Sepharose 4B beads. The function of purified GST-Cdc25C was identified by GST pull-down assay.
  
  
  RESULTS
  The GST-Cdc25C recombinant plasmid was successfully obtained by double  digestion identification. The inserted fragment was confirmed correctly  by sequencing. SDS-PAGE and Western blot analysis showed that the fusion protein was expressed. The fusion protein of about M(r); 80 000 was successfully induced,  and identified by SDS-PAGE and Western blot analysis. GST pull-down assay showed that GST-Cdc25C could interact with Chk2 which verified its known function.
  
  
  CONCLUSIONS
  Cdc25C was successfully cloned and purified.
authors.affiliation: Department of Oncology, Chinese PLA Postgraduate Medical School, Beijing, China.
authors.forename: Zhong-yi
authors.initials: ZY
authors.lastname: Fan
authors.forename: Xiao-jie
authors.initials: XJ
authors.lastname: Xu
authors.forename: Jia
authors.initials: J
authors.lastname: Cao
authors.forename: Lei
authors.initials: L
authors.lastname: Kang
authors.forename: Bai-yu
authors.initials: BY
authors.lastname: Han
authors.forename: Kai
authors.initials: K
authors.lastname: Jiang
authors.forename: Qi-nong
authors.initials: QN
authors.lastname: Ye
authors.forename: Nan
authors.initials: N
authors.lastname: Du
mesh_terms: D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics*; D015870:Gene Expression; D006801:Humans; D010957:Plasmids / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D020687:cdc25 Phosphatases / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D011993:Recombinant Fusion Proteins; C517009:CDC25C protein, human; D020687:cdc25 Phosphatases