title: [Construction and identification of gene vector expressing PTEN while simultaneously silencing Livin].
abstract: OBJECTIVE
  To study the effect of simultaneously increasing PTEN gene expression and inhibiting Livin gene expression on the gastric carcinoma cell line (BGC823) and construct a recombinant vector expressing PTEN while simultaneously silencing Livin.
  
  
  METHODS
  The siRNA expression unit against Livin gene (siLivin) was cleaved from pRNAT-U6.1-Livin vector and then inserted into pCL-neo-PTE to construct the recombinant vector pCL-neo-PTEN-siLivin. Then pCL-neo-PTEN-siLivinp, pCL-neo-PTEN, pRNAT-U6.1-Livin, pCL-neo and pRNAT-U6.1 were respectively transfected into the gastric carcinoma cell line (BGC823) with LipofectAMINE(TM) 2000. The mRNA and protein expression level of PTEN and Livin genes in each cell group was detected by fluorescent quantitative RT-PCR and Western blot.
  
  
  RESULTS
  Recombinant vectors of pCL-neo-PTEN, pRNAT-U6.1-Livin and pCL-neo-PTEN-siLivin were constructed successfully. After transfection with pCL-neo-PTEN-siLivin, the mRNA and protein expression level of PTEN (0.897±0.112) rose in BGC823 cells while Livin gene became silenced. And the characterization of regulated cell bioactivity improved. There were significant differences between transfected and control groups (P<0.05). And the inhibiting effect on the proliferation and metastasis of BGC823 cell by increasing PTEN expression and silencing Livin simultaneously was better than that only by regulating PTEN genes or Livin genes alternatively.
  
  
  CONCLUSIONS
  The recombinant vector of expressing PTEN and silencing Livin gene simultaneously is successfully constructed.
authors.affiliation: Department of General Surgery, First Affiliated Hospital, Zhengzhou University, Zhengzhou 450052, China.
authors.forename: Chun-lin
authors.initials: CL
authors.lastname: Zhao
authors.forename: Jia-xiang
authors.initials: JX
authors.lastname: Wang
authors.forename: Xie-fu
authors.initials: XF
authors.lastname: Zhang
authors.forename: Guo-qiang
authors.initials: GQ
authors.lastname: Zhao
authors.forename: Zhi-ju
authors.initials: ZJ
authors.lastname: Wang
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics*; D045744:Cell Line, Tumor; D015870:Gene Expression; D015972:Gene Expression Regulation, Neoplastic; D020868:Gene Silencing; D005822:Genetic Vectors; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins / Q000235:genetics*; D009363:Neoplasm Proteins / Q000235:genetics*; D051059:PTEN Phosphohydrolase / Q000235:genetics*; D034741:RNA, Small Interfering / Q000235:genetics; D014162:Transfection
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C419866:BIRC7 protein, human; D051034:Inhibitor of Apoptosis Proteins; D009363:Neoplasm Proteins; D034741:RNA, Small Interfering; D051059:PTEN Phosphohydrolase; C494929:PTEN protein, human