The Animal Core Laboratory (Core C) will provide five services for the projects in the grant: 1) Establishment and maintenance of breeding colonies of mutant mice, mainly immunodeficient, for the projects, 2) Genotyping of mice for either breeding or experimentation, 3) Provision of pups for gene therapy experiments, 4) Establishment of strains of mice with congenic markers to facilitate monitoring of cells following administration of transduced cells, 5) Assistance with health monitoring and care of immunodeficient mice. The Core will be housed within the Animal Care Facility at Childrens Hospital Los Angeles and run under the direction of Dr. Kenneth Weinberg. Dr. Weinberg has ten years experience in the breeding, maintenance, genotyping and phenotyping of murine models of immune deficiency. He has established five new strains of mutant mice in his laboratory by breeding of established naturally occurring mutants, transgenics, and knockouts. He has extensive knowledge in the phenotyping of immunodeficient mice by flow cytometry, analyses of immunoreceptor gene rearrangements, functional assays, and immunohistology. 1. Establishment and maintenance of breeding colonies of mutant mice. Types of mice. The Core will establish breeding colonies of the mutant mice needed for individual projects. There are two general types of immunodeficient mice to be used in the projects: 1) mice which are to be engrafted with human progenitors for analyses of differentiation of transduced human cells; 2) mice with specific gene knockouts as models of human disease for testing of gene therapy. Group 1 are either Non- Obese Diabetic/severe combined immune deficiency (NOD/scid) mice or NOD/scid mice that are also deficient for 132-microglobulin (NOD/scid/132M). Group 2 are deficient for either 13-globin(Project 2), BTK-tec (Project 3), IL-7Rc_ or 7c (Project 4), or ADA (Project 5). For Project 3, BTK/tec mice will be bred at the University of Washington. Breeding of normal mice will be outside the scope of the Core's activities - it is more economical to purchase such mice from outside breeders, e.g., Jackson or Charles River Laboratories. Projections for usage of the Project 1 (Crooks) Project 2 (Malik) Project 3 (Rawlings) Project 4 (Weinberg) Project 5 (Kohn) Core for breeding mice are shown in the Immunodeficient Mice for Hu/Mouse Chimeras (Mice/year) 50 (NOD/scid/132M) 60 (NOD/scid) table below Mutant Mice as Disease Models (Mice/year) 80 (Thalassemia intermedia, heterozygous breeding and pups) none 150 (IL-7R_ or ?c +/- Kit w41) 60 (ADA) Breeding. The colonies to be established will be derived from either mice given to the Animal Core Laboratory by the Project investigators. Breeding will be performed with standard IACUC-approved protocols. Male-female pairs will be in individual cages and monitored. Males will be removed from cages with pregnant mice, prior to birth. After weaning, the gender of the pups will be determined by examination, and males and females transferred to separate cages. For breedings of heterozygotes, DNA will be extracted from tail cuttings and used for either PCR or Southern blot based assays. The Core will develop protocols with Dr. Malik to determine the number of heterozygotes in Project 2 to be kept for breeding or used in experiments. We expect 2-3 litters per female mouse, and frequently only one litter. Therefore, the number of breeding mice per year is estimated at 2-3 times the number of mice that will be breeding at any one time. Strategies to increase fertility and health. A general problem with immunodeficient mice has been decreased fertility. Mice which are immunodeficient have a higher risk of infections, which appear to interfere with overall health and fertility, even when the animals survive the infection. Although prophylactic and therapeutic administration of antibiotics is often used to control infections in breeding mice, our experience has been that antibiotic administration frequently alters the flora in the colony and development of resistant infections. The best way that we have found to increase the health of immunodeficient breeders is to make them immunocompetent by