title: Rapid in vivo functional analysis of transgenes in mice using whole body imaging of luciferase expression.
abstract: The use of transgenic animals in biomedical research is increasing rapidly and may be the best means of determining gene function. Generating transgenic animals typically requires time-consuming screening processes, and gene function is assessed by an array of difficult phenotypic and biochemical assays performed ex vivo. To address the unmet need in transgenic research for functional assays performed with ease in living animals, we demonstrate here that in vivo detection of luciferase enzyme as a transcriptional reporter facilitates rapid screening for both the presence and function of transgenes in intact living mice. Using this approach we identified three bioluminescent transgenic founders where the transgene consisted of the heme oxygenase promoter fused to the modified coding sequence of the luciferase gene. These founders were identified from 183 pups and confirmed by PCR analysis. Identification of HO-1-luc homozygotes from back- crossed F2 littermates was then accelerated by in vivo imaging. In another transgenic mouse line, where the transgene was comprised of the bone morphogenic-4 (BMP4) promoter fused to the modified luciferase gene, we were able to identify transgenic animals and in each line we were able to visualize patterns of expression in living animals over time. The light production from these transgenic mice indicated that the desired DNA fragment was functional and different expression profiles apparent at different ages and after gene induction.
authors.affiliation: Department of Pediatrics, Stanford University Medical Center, Stanford University, CA 94305-5208, USA.
authors.forename: W
authors.initials: W
authors.lastname: Zhang
authors.forename: J Q
authors.initials: JQ
authors.lastname: Feng
authors.forename: S E
authors.initials: SE
authors.lastname: Harris
authors.forename: P R
authors.initials: PR
authors.lastname: Contag
authors.forename: D K
authors.initials: DK
authors.lastname: Stevenson
authors.forename: C H
authors.initials: CH
authors.lastname: Contag
mesh_terms: D000818:Animals; D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins / Q000235:genetics; D005260:Female; D020869:Gene Expression Profiling / Q000379:methods*; D005818:Genetic Engineering / Q000379:methods; D006419:Heme Oxygenase (Decyclizing) / Q000235:genetics; D006720:Homozygote; D008156:Luciferases / Q000032:analysis* / Q000378:metabolism*; D008163:Luminescent Measurements; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D011401:Promoter Regions, Genetic; D019076:Transgenes
chemical_list: C525810:Bmp4 protein, mouse; D055415:Bone Morphogenetic Protein 4; D019485:Bone Morphogenetic Proteins; D008156:Luciferases; D006419:Heme Oxygenase (Decyclizing)