Insertional Mutations in Transgenic Mice

In contrast to the rapid rate of progress in the developmental genetics of invertebrate species such as Drosophila melanogaster or the nematode C. elegans, the genetic analysis of mammalian development remains a difficult undertaking. Several approaches are now making it possible to identify, and to analyze at the molecular level, genes that are potentially important in mammalian development. This chapter presents the analysis of mutations caused by the insertion of foreign DNA or viruses into the mouse germ line. It describes the methods to identify insertional mutations causing recessive prenatal lethality, and summarizes their phenotypic effects. The presence of a prenatal recessive lethal mutation in a transgenic mouse line can be discerned by the failure to obtain homozygous animals among the progeny of an intercross. In cases in which host DNA sequences flanking a transgene insertion have already been cloned, a flanking probe may be used to identify homozygotes, based on Southern blot analysis. In other cases, the identification of homozygous transgenic mice has usually relied on the quantitation of the transgene copy number: homozygous mice carry twice the number of copies as do heterozygous mice. The relative transgene copy number can be estimated either by Southern blot or by dot blot hybridization, in each case comparing the signal obtained with a probe for the transgene to the signal obtained with a probe for an endogenous gene.