Insertion of a foreign gene into the beta-casein locus by cre-mediated site-specific recombination

The expression of foreign genes in transgenic animals is generally unpredic table as transgenes are integrated at random after pro-nuclear injection in to fertilized oocytes. In many cases, transgene expression is inhibited by neighbouring chromatin structures or by the repeated nature of the multiple transgene copies present at the integration site. A strategy involving hom ologous and site-specific recombination has been devised by which single co pies of a foreign gene can be inserted specifically into the locus of a hig hly expressed gene. As a first step, a loxP recombination target site is in troduced by homologous recombination into a predetermined gene locus such t hat the loxP sequence is placed next to the promoter region and replaces th e translational initiation signal. In a subsequent site-specific recombinat ion reaction, a gene of interest can be integrated into the pre-existing lo xP site. This biphasic recombination strategy was used to integrate a lucif erase reporter gene into the locus of the murine beta-casein gene in embryo nic stem cells. (C) 1999 Elsevier Science B.V. All rights reserved.