title: Long-term in vivo gene expression in mouse kidney using φC31 integrase and electroporation.
abstract: BACKGROUND
  Achieving long-term gene expression in kidney will be beneficial for gene therapy of renal and congenital diseases, genetic studies constructing animal disease models, and the functional analysis of disease-related genes.
  
  
  OBJECTIVE
  The purpose of this study was to develop an in vivo long-term gene expression system in murine kidney using φC31 integrase.
  
  
  METHODS
  Gene expression in cultured RENCA, TCMK-1, and HEK293 cells was assessed. The long-term in vivo gene expression system in the kidney was achieved by co-transfecting 5 µg of pORF-luc/attB as a donor plasmid and 20 µg of pCMV-luc as a helper plasmid into the right kidney of mice by electroporation. Luciferase expression levels were measured to determine longevity of the expression.
  
  
  RESULTS
  Significantly high luciferase expression levels were observed in cultured RENCA, TCMK-1, and HEK293 cells over 1 month compared with controls (non-integrase system). The luciferase cDNA sequence was integrated at a pseudo attP site termed mpsL1. In vivo luciferase expression levels in the integrase group were sustained and significantly higher than those in the control group over 2 months. Furthermore, φC31 integrase-transfected cells had less genomic DNA damage caused by integrase expression.
  
  
  CONCLUSIONS
  These results demonstrated that the φC31 integrase system could produce long-term (2 months) in vivo gene expression in mouse kidney.
authors: Department of Drug Delivery Research, Graduate School of Pharmaceutical Sciences, Kyoto University , Kyoto , Japan .
authors: Yuki
authors: Y
authors: Otani
authors: Shigeru
authors: S
authors: Kawakami
authors: Hidefumi
authors: H
authors: Mukai
authors: Yuki
authors: Y
authors: Fuchigami
authors: Fumiyoshi
authors: F
authors: Yamashita
authors: Mitsuru
authors: M
authors: Hashida
mesh_terms: D000818:Animals; D002460:Cell Line; D004249:DNA Damage; D018076:DNA, Complementary / Q000235:genetics; D018274:Electroporation / Q000379:methods; D005260:Female; D005786:Gene Expression Regulation / Q000235:genetics*; D015316:Genetic Therapy / Q000379:methods; D057809:HEK293 Cells; D006801:Humans; D019426:Integrases / Q000235:genetics*; D007668:Kidney / Q000378:metabolism*; D008156:Luciferases / Q000235:genetics*; D051379:Mice; D008813:Mice, Inbred ICR; D010957:Plasmids / Q000235:genetics; D017422:Sequence Analysis, DNA; D013997:Time Factors; D014162:Transfection
chemical_list: D018076:DNA, Complementary; D008156:Luciferases; D019426:Integrases
keywords: electroporation; gene therapy; in vivo long-term gene expression; kidney; plasmid DNA; transfection; φC31 integrase