TRANSGENIC MOUSE CORE

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The goal of the Transgenic Mouse Core is to provide the investigators of each project with assistance in the design and production of genetically modified mice. To achieve this goal, the core will perform the following functions: Specific Aim 1. Design and construct transgenes and develop genotyping assays. Core personnel will consult with the project leaders regarding the goal for each proposed transgenic model. Appropriate DNA constructs will then be designed using optimal promoter and coding sequences. The required genomic or cDNA clones for the gene of interest will be acquired and cloned downstream from the desired promoter. Transgene DNA will be purified for microinjection, and genotyping assays will be designed and tested. Specific Aim 2. Generate transgenic founders via microinjection of transgene DNA. Fertilized eggs from the C57BL/6 strain of mice will be used for pro-nuclear injection of transgene construct DNA. Potentially transgenic offspring will be identified using genotyping assays developed in Specific Aim 1. Founder mice will be characterized for the ability to transmit the transgene to offspring. The number of independent chromosomal integration sites in each founder will be determined. Germline-competent mice, each with a transgene integrated at a single genomic locus, will be delivered to project leaders. Specific Aim 3. Assist in the characterization of transgenic mice. Quantitative reverse-transcriptase polymerase chain reaction (RT-PCR) assays will be designed to specifically detect transgene mRNAs and then used to determine the tissue distribution of transgene mRNA expression. If transgene expression is low, core personnel will assist the Project Leader in identifying reasons for low expression and optimizing transgene design to overcome the problem. Transgenic mice are increasingly used to address questions in aging and skeletal research. Experiments in transgenic mice allow investigators to determine whether phenomena observed in cell lines and primary cell cultures also occur in vivo. In addition, transgenic mice can be used to test hypotheses that cannot be convincingly addressed in any in vitro system. Thus timely and efficient production of transgenic mice is essential to the overall goal of the Program which is to improve the understanding of the pathophysiology of the bone fragility syndrome of osteoporosis and thereby rationalize and optimize its treatment.