The mouse is the premier venue for modeling human disease because of the unrivalled technology for manipulating its genome. Embryonic stem (ES) cells, pronuclear injection in fertilized oocytes, and transpositional mutagenesis allow for sophisticated manipulation of the murine germ line. Potentially the most powerful and precise strategy for disease modeling is conditional mutagenesis, in which a specific mutation is made in a somatic cell to produce cell type-specific disease states. Conditional mutagenesis relies on the site-specific recombinase (SSR) Cre to mediate a very precise recombination event between its 34-base pair loxP recombination target sites. The loxP sites have to be placed into the mouse genome at a chosen site, usually by gene targeting in ES cells. Upon exposure to Cre, these sites will be precisely recombined without any other recombination event in the rest of the genome. This remarkable precision is what makes conditional mutagenesis such a valuable technique. Among all SSRs, Cre has proven to be the most effective at mediating recombination in large genomes such as the mouse. However, owing to the complexities of alternative splicing, gene product interactions and genetic redundancy, it will be useful to have the means to mutate a given gene in more than one way, or to combine more than one conditional mutation. To achieve this goal, new tools that perform as well as Cre are required.
The authors show that an SSR that is related closely to Cre, termed Dre, has similar efficiencies to Cre when applied in the mouse. Using recombination tests in E. coli, mammalian cells and the mouse, they show that Dre recombinase produced very specific recombination to rox sites, even in the presence of loxP. Thus, there is no crosstalk between Cre and Dre recombination. They also produced ubiquitous mouse lines for the Dre-rox system and propose using this technique to build more complex models by combining it with the existing Cre and FLP methods of genetic manipulation.
These results present a second option for efficient conditional mutagenesis in the mouse. Whereas Cre will remain the main instrument for conditional mutagenesis, the advent of Dre permits the development of more sophisticated strategies of combinatorial site-specific recombination and conditional mutagenesis. The use of the two SSRs will simplify the use of conditional mutagenesis to analyze alternative splice forms. Given that most mammalian genes are alternatively spliced, this application could play a significant role in future mouse modeling.
