title: Efficient induction of productive Cre-mediated recombination in retinal pigment epithelium.
abstract: OBJECTIVE
  To dissect gene functions in the retinal pigment epithelium (RPE), we previously generated a tetracycline-inducible RPE-specific Cre mouse line. Although this Cre mouse line was useful for several conditional gene targeting studies that were conducted by different laboratories, its potential has not been fully exploited, presumably due to a lack of knowledge or procedure for inducing Cre expression appropriately in this mouse line. The goal of the current study is to establish a procedure that will improve the reproducibility of Cre-mediated recombination in this mouse line.
  
  
  METHODS
  Analysis of Cre expression and function was performed in double transgenic mice derived from inducible RPE-specific Cre and Cre-activatable ROSA26 lacZ reporter mice. A tetracycline derivative, doxycycline, was supplied to mice intravitreally to induce Cre expression. Cre expression and function were examined with reverse transcription-PCR, immunoblotting, immunostaining, and in situ enzymatic assay for β-galactosidase. Retinal integrity was examined with electroretinography and morphometry.
  
  
  RESULTS
  Intravitreal Dox injection elevated Cre expression significantly and resulted in productive Cre-mediated recombination in approximately 60% of the RPE cells in this mouse line with no apparent change in retinal integrity.
  
  
  CONCLUSIONS
  Our results suggest that productive Cre-mediated recombination in this mouse line can be induced efficiently with intravitreal Dox delivery, with no apparent Dox or Cre toxicity. Therefore, our inducible RPE-specific Cre mice are suitable for Cre/lox-based gene activation and inactivation in adult RPE, which is critical to the effectiveness and suitability of this Cre mouse line in long-term studies requiring conditional gene targeting.
authors.affiliation: Department of Ophthalmology, the Second Affiliated Hospital of Nanchang University, Nanchang, China ; Department of Medicine Endocrinology, University of Oklahoma Health Sciences Center, Oklahoma City, OK.
authors.forename: Shuhua
authors.initials: S
authors.lastname: Fu
authors.affiliation: Department of Medicine Endocrinology, University of Oklahoma Health Sciences Center, Oklahoma City, OK.
authors.forename: Meili
authors.initials: M
authors.lastname: Zhu
authors.affiliation: Department of Ophthalmology, the Second Affiliated Hospital of Nanchang University, Nanchang, China.
authors.forename: Changyun
authors.initials: C
authors.lastname: Wang
authors.affiliation: Department of Medicine Endocrinology, University of Oklahoma Health Sciences Center, Oklahoma City, OK ; Department of Cell Biology, University of Oklahoma Health Sciences Center, Oklahoma City, OK ; Harold Hamm Oklahoma Diabetes Center, University of Oklahoma Health Sciences Center, Oklahoma City, OK.
authors.forename: Yun-Zheng
authors.initials: YZ
authors.lastname: Le
mesh_terms: D000818:Animals; D004318:Doxycycline / Q000008:administration & dosage / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects; D019426:Integrases / Q000235:genetics / Q000378:metabolism*; D058449:Intravitreal Injections; D051379:Mice; D011995:Recombination, Genetic / Q000187:drug effects; D055213:Retinal Pigment Epithelium / Q000187:drug effects / Q000378:metabolism*; D001616:beta-Galactosidase / Q000378:metabolism
chemical_list: C045073:Cre recombinase; D019426:Integrases; D001616:beta-Galactosidase; D004318:Doxycycline
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