title: Tissue-Specific Regulation of Oncogene Expression Using Cre-Inducible ROSA26 Knock-In Transgenic Mice.
abstract: Cre-inducible mouse models are often utilized for the spatial and temporal expression of oncogenes. With the wide number of Cre recombinase lines available, inducible transgenesis represents a tractable approach to achieve discrete oncogene expression. Here, we describe a protocol for targeting Cre-inducible genes to the ubiquitously expressed ROSA26 locus. Gene targeting provides several advantages over standard transgenic techniques, including a known site of integration and previously characterized pattern of expression. Historically, an inherent instability of ROSA26 targeting vectors has hampered the efficiency of developing ROSA26 knock-in lines. In this protocol, we provide individual steps for utilizing Gateway recombination for cloning as well as detailed instructions for screening targeted ES cell clones. By following this protocol, one can achieve germline transmission of a ROSA26 knock-in line within several months.
authors.affiliation: Interdepartmental Program in Translational Biology and Molecular Medicine, Baylor College of Medicine, Houston, Texas.|Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, Texas.
authors.forename: Brandi L
authors.initials: BL
authors.lastname: Carofino
authors.affiliation: Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, Texas.|Genetics and Genome Biology, The Hospital for Sick Children, Toronto, Ontario, Canada.
authors.forename: Monica J
authors.initials: MJ
authors.lastname: Justice
mesh_terms: D000818:Animals; D005786:Gene Expression Regulation; D055879:Gene Knock-In Techniques / Q000379:methods*; D018390:Gene Targeting / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics / Q000378:metabolism; D019426:Integrases / Q000378:metabolism*; D051379:Mice / Q000235:genetics* / Q000378:metabolism; D008822:Mice, Transgenic; D015513:Oncogene Proteins / Q000235:genetics* / Q000378:metabolism; D009928:Organ Specificity
chemical_list: D015513:Oncogene Proteins; C045073:Cre recombinase; D019426:Integrases
keywords: Cre-inducible; Gateway; In-Fusion; JM8A3 ES cells; ROSA26; Southern blotting; oncogene