Core B is designed provide transgenic animals required by the four projects of the program, including mice that express key molecules/receptors and mice that have defective genes. Core B will breed and screen transgenic animals and backcross them as necessary to render them suitable for the experiments detailed in the individual projects of the program. Because this process is very labor intensive, a centralized facility specialized ot perform this task makes a great deal of sense. Core B will also oversee the collection and preparation of reagents required as key components by at least two or more individual projects. Such reagents would include Ab to potential co-stimulatory molecules and cytokines, antibodies to adhesion molecules, APC cell l lines expressing particular co-stimulatory molecules, cell lines for cytokine bioassays. The Core will obtain the various Tg or KO mice needed for the individual projects, quarantine them if necessary, develop standardized procedures for screening for the Tg/KO gene expression in each case, and will breed and backcross the mice to C57BL/6 and B10.BR inbred strains. The goals are to have healthy, genetically homogenous mice for the investigators in the project. Investigators will be able to obtain these mice for experiments but will maintain the animals separately during the course of the sometimes lengthy adoptive transfer experiments. Many of the experiments conducted so far on the Tg and KO mice have been done with original B6 x SJL founder mice or with mice backcrossed only for one or two generations. In such cases background genes from the non B6 parent may influence experimental results. Use of non-bred mice can also lead to complications due to mismatches in major or minor histocompatibility differences and also to the presence of genes other than the gene of interest due to linkage on the same chromosome. Furthermore, the facility will allow us to generate new strains of mice by inbreeding. For instance TCR Tg mice will be generated with the SCID mutation so that contributions from expression of endogenous alpha chains can be ruled out. Also double KO mice or TCR Tg/KO mice can be created which will allow a more definitive analysis. At the moment we have the following mouse strains to contribute to the core; TCR Tg mice (specific for pigeon cytochrome C (PCC) plus I-Ek (designated AND) backcrossed 5 times to B6 and B10 BR (currently being bred onto a RAG-2 KO background); TCR Tg mice specific for HY plus Dd (already on a SCID background); TCR Tg mice specific for influenza hemagglutinin (HA) peptide plus IAd (CD4 selecting) and TCR Tg specific for HA plus Dd (CD8 selecting); beta2M KO/KO mice; CD4 KO/KO mice backcrossed 8X to B6; IL-2 KO/KO backcrossed 3X to B6; IFNgammaKO/KO backcrossed 3x to B6. IL-4 KO/KO backcrossed 4x to B10.BR; and RAG-2 KO/kO. We also have mum KO/KO mice and mice constitutively expressing high levels of bcl2. Dr. Klinman will provide the Sma58 m ice, which express I=-Ek only on germinal center B cells. Dr. Bradley has already made arrangements to obtain L-, E- and P- selectin Ko mice.
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