GENE TARGETING SHARED RESOURCE

The Gene Targeting Facility was established in 1992 to assist AECC researchers in the generation of animal models for human cancer. The goals of the facility are to provide services that allow researchers to modify the mouse genome through the methods of gene targeting in embryonic stem (ES) cells and to introduce these changes into the mouse germline. The facility provides: (1) Advice in the theoretical and practical aspects of gene targeting vector design including design of conventional knockout targeting vectors, knock-in targeting vectors as well as targeting vectors for temporal and spatial ablation of genes. The facility also advises on the methodologies required for targeting vector construction, including the use of gridded BAG 129 strain genomic libraries for the rapid isolation of genomic fragments, subcloning strategies and also the use of recombination based cloning. The facility maintains and makes available a panel of vectors and resistance cassettes that facilitate targeting vector construction. In addition, the facility established recombination-based cloning technology ("recombineering") using the RecET technology that allows the modification of BAG clones and also permits the more rapid generation of more complex targeting vectors. (2) The facility provides comprehensive ES cell culture service and maintains stocks of mouse embryonic fibroblasts (MEF) for feeder cell generation and ES cell lines with high germline transmission capacity. The facility offers complete ES cell culture service encompassing all the required techniques but will train and provide advice to investigators who wish to participate in parts of this process. The facility provides services for chimera production. This includes ES cell microinjection into blastocysts, blastocyst transfer and chimera production. To support these activities, stocks of ES cell lines from different sources and of different genetic backgrounds (129J/Sv, 129/B6 and B6) are maintained along with core mouse colonies sufficient to generate enough blastocysts for injection, as well as pseudopregnant recipient mice for the blastocyst transfers and vasectomized males to mate with these females. In addition, the facility is intimately involved in the implementation of new technologies and the training of new investigators in the technical and theoretical aspects of these procedures. This facility will be relocated to the new Price Center for Genetic and Translational Medicine in a larger space adjacent to a new Animal Barrier Shared Resource when that facility opens in December 2007.