Genetically altered mice are important models to study the mechanism of inherited and acquired diseases. Furthermore, these mice are extremely useful to test the efficacy of new therapies and to validate potential drug targets. Although the generation of mice carrying point mutations or deletions of specific genes is quite well established, it is still a relatively long procedure. Careful planning of the gene targeting is therefore essential. In this chapter we describe the general procedure of generating genetically altered mice and discuss potential problems that could be encountered and their solution.