[Frontiers in Bioscience 11, 1108-1136, January 1, 2006]

Mammalian genome targeting using site-specific recombinases

Angel-Luis García-Otín 1 and Florian Guillou 2

1 Laboratorio de Investigación Molecular, Instituto Aragonés de Ciencias de la Salud (I+CS), Hospital Universitario Miguel Servet, Zaragoza, Spain. 2 Unité de Physiologie de la Reproduction et des Comportements (PRC), Institut National de la Recherche Agronomique, UMR 6175 INRA-CNRS-Université de Tours Haras Nationnaux, Nouzilly, France

TABLE OF CONTENTS

1. Abstract
2. Introduction
3. Site-specific recombinases
3.1. Cre and FLP recombinases
3.2. Recombinases in a mammalian environment
3.3. Further developments
3.4. Novel recombinases
4. Technology to manipulate the mouse genome
4.1. Gene transfer
4.2. Gene targeting
4.3. Nuclear transfer
5. Strategies for conditional gene targeting
5.1. Generation of floxed alleles
5.2. Promoter-specific driven recombinase expression
5.3. Inducible systems
5.4. Virus-mediated delivery to specific cell or tissue types
5.5. Recombinase-activated siRNA expression
6. Approaches to model generation
6.1. Switching on/off genes in a space- and time-controlled fashion
6.2. Chromosomal rearrangement
6.3. Cell lineage studies
6.4. Humanization of mouse genes
6.5. Cassette exchange or integration
7. Summary and perspectives
8. Acknowledgments
9. References

1. ABSTRACT

Nowadays, a wide array of procedures in mouse technology has been made available to researchers in order to establish valuable models for the study of gene function. The efficiency of gene transfer and gene targeting as methods for producing genetic changes in mice, in addition to continuous advances in molecular biology tools, has converted the mouse into the major experimental model for the study of mammalian physiology. In recent years, the emergence of site-specific recombinases as tools to engineer mammalian genomes has opened new avenues into the design of genetically modified mouse models. The original Cre and FLP recombinases have demonstrated their utility in developing conditional gene targeting, and now other analogous recombinases are also ready to be used, in the same way or in combined strategies, to achieve more sophisticated experimental schemes for addressing complex biological questions. The properties of site-specific recombinases in combination with other biotechnological tools (tet on/off system, siRNA mediated gene silencing, fluorescent proteins, et al.) make them useful instruments to induce precise mutations in specific cells or tissues in a time-controlled manner. This ability can be applied in functional genomics in several ways: from conditional and inducible gene targeting to controlled expression of transgenes and recombination-mediated cassette exchange in mouse models for the study of development or disease phenotypes. This review focuses on the use of site-specific recombinases for mouse genome manipulation. A historical perspective of site-specific recombinases is considered and a number of strategies for achieving inducible or conditional genomic manipulations are contemplated in the context of current techniques for producing genetically modified mice. Finally, several model generation approaches from recent examples in the literature are revised.