3 - Homologous Recombination in Embryonic Stem Cells as a Means to Generate Mice with Defined Mutations

Abstract

This chapter presents experiments that employ a new method to mutate the mouse homeobox Hox-1.1 gene in ES cells. This method does not require selection and should allow the introduction of small mutations without further alteration of the target locus. All mammalian Hox genes are organized in clusters. These clusters are very similar to the Drosophila antennapedia and Bithorax homeobox clusters in their gene organization and pattern of expression. This clustered organization might be important for gene regulation and cis regulatory elements control the expression of several homeobox genes. It has been shown that the overexpression or ectopie expression of homeobox genes interferes with the normal embryonic development. For this reason, it was a matter concern that disruption of the Hox-1.1 gene with a neor gene, which would introduce a strong promoter into the Hox-1 cluster, would lead to the disregulation of other genes in the cluster. This might make a proper analysis of the Hox-1.1− phenotype impossible. Thus, it was decided to disrupt the Hox-1.1 coding region by integrating a 20-basepair (bp) oligonucleotide into the homeobox.