The Transgenic Shared Resource trains and assists AECCC investigators in the production and characterization of transgenic mice through introduction of DNA sequences into the germline, usually by microinjection into the pronuclei of fertilized oocytes. This facility provides advice on transgene constructs, preparation of DNA in a form suitable for successful injection, micro-injection of oocytes, transfer to pseudo-pregnant recipients, and care of pups to weaning at which time mice are returned to investigators for analysis. To support these services, a core colony of mice is maintained to generate pseudopregnant females, superovulated mice for oocyte retrieval, vasectomized males, etc. The facility provides rigorous testing of pre-implantation culture medium, anesthetics for mice, hormones for super-ovulation, and reagents for DNA purification. In addition, colony rederivation is provided for pathogen-infected mice, in vitro fertilization to rescue colonies that are having difficulty breeding and transient transgene analysis for lethal genes. The facility has extensive microinjection stations maintained and operated by a highly experienced staff with long-standing experience in generating transgenic mice. At least three founders are guaranteed per construct injected although the number is usually greater. There has been essentially a 100% success rate in generating transgenic mice with minimal re-injection necessary. Between 15-20 principal investigators use the facility each year and many have multiple constructs injected. Investigators with little-or-no experience in mouse biology are trained in relevant methodologies as well as more general aspects of mouse genetics, reproduction, husbandry, and mammalian developmental biology. Investigators who plan to use transgenic mice extensively often take the yearly course in """"""""Mouse Developmental Genetics"""""""" sponsored by the facility. This has propagated a large cadre of investigators and trainees at AECCC skilled in transgenic methodologies. Dr. Jeffrey Pollard replaced Dr. Ron DePinho as faculty supervisor in 1998 and has implemented several changes. Stringent quality-control procedures were adapted and all DNA to be injected is now purified by the transgenic facility staff. The FVB strain of mice has become the preferred recipient for transgenes because of its relative ease of injection and the inbred nature of the strain. In addition, the facility has been successful at establishing transgenic mice using BAC DNA. Mice generated in this facility were the first to show that the tetracycline-regulated system is operable in vivo. More recently, the facility has aided in the generation of mice in which tissue regulation is effected through the ecdysone-regulatory system. Advice and assistance on the use of these systems are available to all AECCC.
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