The mission of the transgenic core is to provide technical expertise, guidance in experimental design, and training opportunities in all aspects of the production of transgenic mice. A core laboratory will prevent the unnecessary and time consuming duplication of effort involved in the development of this technology by multiple investigators. The core will additionally provide better quality control, turn around time, and cost effectiveness than either commercially available services or multiple users within one institution. Projects 1 through 4 will use the core to produce transgenic mice that are essential to addressing a major theme of the Program Project: What are the physiological roles of specific molecular components of the hypothalamic-pituitary-adrenal axis? Project 1 will use transgenic mice that have altered naturietic peptide receptor expression to determine the role of these peptides in the control of ACTH secretion. Project 2 will generate mice with pituitary- specific expression of functional POMC minigenes to genetically """"""""rescue"""""""" POMC gene """"""""knock-out"""""""" mice produce by homologus recombination. Project 3 will explore the role of pituitary derived beta-endorphin in the stress response by blocking the posttranslational processing of POMC at particular dibasic residues with dominant negative or antisense type transgenes. Project 4 will use the technical expertise and facilities of the core to produce mutant mice carrying either null alleles of the neural-specific MSH receptors produced by homologous recombination in embryonic stem cells or constitutively activated receptors encoded by dominant transgenes.
These aims will be accomplished through provision of the following services: 1. Embryo collection, nuclear microinjection, blastocyst injections, and oviduct and uterine transfer of manipulated embryos to generate transgenic mice. 2. Genotyping of transgenic mice by a combination of tail DNA dot-blot, Southern blot, or PCR analysis. 3. Supervision and maintenance of a mouse breeding colony providing inbred FVB mice for embryo donations and including stud and vasectomized males used for the initial production of transgenic embryos, and breeding and pedigree analysis of the transgenic mice. FVB mice from the colony will also be available to all investigators for use as controls in physiology experiments or for tissue harvesting. 4. Assistance in physiology experiments using mice including injections, anesthesia, cannulation, and surgical procedures. 5. Permanent storage of cryopreserved transgenic mouse embryos to protect valuable pedigrees from accidental loss, to minimize the continuous breeding of pedigrees that no longer are immediately needed for experiments, and to provide a potential resource for distribution of selected pedigrees to other qualified investigators outside of the institution.
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