The steps required to delete genes from the mouse genome are illustrated by showing how a cluster of three flavin-containing monooxygenase (Fmo) genes (Fmo1, Fmo2, andFmo4) were deleted from mouse chromosome 1. Such large deletions are accomplished usingloxP/Cre recombinase technology. Genomic clones corresponding to the genes to be deleted are first isolated, and then appropriate genomic fragments are cloned into vectors containing aloxPsite. This produces targeting vectors, which are electroporated into mouse embryonic stem (ES) cells to allow a homologous recombination event to take place between the mouse genomic fragment, present within the vector, and the homologous sequences in the ES cell genome. Screening of ES cells for recombinants in whichloxPsites have been inserted on either side of the gene cluster to be deleted is described. Recombination by Cre recombinase to produce ES cell lines carrying the deletion on chromosome 1 is also described.
