Knockin Mice
To avoid the problems of a standard transgenic, many researchers now rely on knockin mice to study the exogenous expression of a protein. A knockin mouse is generated by targeted insertion of the transgene at a selected locus. The insert is flanked by DNA from a non-critical locus, and homologous recombination allows the transgene to be targeted to that specific, non-critical integration site. (See Figure 1) In this way, a researcher has complete control of the genetic environment surrounding the overexpression cassette and it is likely that the DNA did not incorporate itself into multiple locations. Site-specific knockins result in a more consistent level of expression of the transgene from generation to generation because it is known that the overexpression cassette is present as a single copy. Also, because a targeted transgene is not interfering with a critical locus, the researcher can be more certain that any resulting phenotype is due to the exogenous expression of the protein. Although the generation of a knockin mouse does avoid many of the problems of a traditional transgenic mouse, this procedure requires more time to assemble the vector and to identify ES cells that have undergone homologous recombination.