title: [Establishment of a targeting protein for Xenopus kinesin-like protein 2 C' terminal SBP-3 x Flag tagged HCT 116 colorectal cancer cell model].
abstract: OBJECTIVE
  To develop a targeting protein for Xenopus kinesin-like protein 2 (TPX2) C' terminal SBP-3 x Flag-tagged HCT 116 cell model.
  
  
  METHODS
  Homologous arms were amplified by polymerase chain reaction (PCR), and then the adeno-associated virus (AAV) -targeting vector of TPX2 was constructed. HCT 116 cells were targeted after the viruses were packaged. Positive cell clones with neomycin resistance gene were obtained by G418 and PCR screening. Finally, the neomycin gene cassette was excised after the targeted clones were infected with adenovirus expressing Cre-recombinase, and the TPX2 C' terminal SBP and 3 x Flag endogenous double-tagged HCT 116 cells were obtained by PCR screening.
  
  
  RESULTS
  Two positive cell clones with neomycin resistance gene were obtained by PCR screening. The positive clones with neomycin resistance gene excised were obtained by Cre adenovirus infection, and the knock-in of SBP-3 x Flag gene was verified by Western blot analysis.
  
  
  CONCLUSIONS
  The TPX2 C' terminal SBP-3 x Flag tagged HCT 116 cell model was successfully established.
authors.affiliation: National Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, CAMS and PUMC, Beijing 100005, China.
authors.forename: Ze-Bin
authors.initials: ZB
authors.lastname: Huang
authors.forename: Ze-Yan
authors.initials: ZY
authors.lastname: Zhang
authors.forename: Xiao-Dong
authors.initials: XD
authors.lastname: Zhang
authors.forename: Shi-Ying
authors.initials: SY
authors.lastname: Miao
authors.forename: Lin-Fang
authors.initials: LF
authors.lastname: Wang
authors.forename: Run-Lei
authors.initials: RL
authors.lastname: Du
mesh_terms: D018797:Cell Cycle Proteins / Q000235:genetics*; D015179:Colorectal Neoplasms / Q000235:genetics / Q000473:pathology*; D000229:Dependovirus / Q000235:genetics; D018390:Gene Targeting; D005822:Genetic Vectors; D045325:HCT116 Cells; D006801:Humans; D008869:Microtubule-Associated Proteins / Q000235:genetics*; D009687:Nuclear Proteins / Q000235:genetics*
chemical_list: D018797:Cell Cycle Proteins; D008869:Microtubule-Associated Proteins; D009687:Nuclear Proteins; C122167:TPX2 protein, human