The Cre protein from bacteriophage P1 promotes site-specific DNA recombination in mammalian cells. This study demonstrates that the Cre protein, which is a 38-kDa protein, can cause DNA synapsis and recombination in a mammalian cell line. A stable mouse cell line was established that expresses the Cre protein under the control of the metallothionein I gene promoter, which is inducible by cadmium ions. DNA recombination was monitored using substrates containing two directly repeated lox sites. One substrate was a circular plasmid with two lox sites flanking a marker gene, and another was a pseudorabies virus containing a lox insertion. Site-specific recombination in vivo was dependent on the presence of the Cre protein and occurred specifically at the 34-base-pair lox sites. These results show that a prokaryotic protein can mediate controlled site-specific DNA synapsis and recombination in mammalian cells, suggesting that Cre-mediated recombination may be a useful tool for understanding and modulating genome rearrangements in eukaryotes. The Cre-lox site-specific recombination system of bacteriophage P1 is simple and well characterized. A single 38-kDa protein, Cre, is necessary and sufficient to catalyze recombination between two lox sites, each 34 bp in length. Recombination can occur between directly repeated sites on the same molecule to excise the intervening DNA segment or between inverted lox sites to give inversion of the intervening DNA segment. Both intra- and intermolecular recombination is catalyzed by Cre with either supercoiled or linear DNA. The Cre-lox system was shown to function efficiently in yeast and now is demonstrated to function in mammalian cells. The study used a mouse cell line, C-127, transformed with vectors containing the metallothionein I gene promoter to establish cell lines 55 and 3C-5. The Cre protein was induced by cadmium ions, and its expression was confirmed by immunoblot analysis. Recombination was assayed using plasmid pRH43 containing two directly repeated lox sites. Southern blot analysis showed that the Cre protein could excise the kanr gene from pRH43 as a 1.9-kb circular DNA molecule containing a single Bgl II site. The results demonstrated that the Cre recombinase is expressed in 55 cells and can perform precise lox-specific recombination on pRH43 to excise a 1.9-kb circular molecule containing the kanr gene. The study also used a herpesvirus vector, PRV42::pBS64, which carries a lox insertion. Precise recombination at the lox sites resulted in the reconstruction of the gIII gene, producing a glycoprotein recognized by a specific antibody. Recombinant viruses formed black plaques whenThe Cre protein from bacteriophage P1 promotes site-specific DNA recombination in mammalian cells. This study demonstrates that the Cre protein, which is a 38-kDa protein, can cause DNA synapsis and recombination in a mammalian cell line. A stable mouse cell line was established that expresses the Cre protein under the control of the metallothionein I gene promoter, which is inducible by cadmium ions. DNA recombination was monitored using substrates containing two directly repeated lox sites. One substrate was a circular plasmid with two lox sites flanking a marker gene, and another was a pseudorabies virus containing a lox insertion. Site-specific recombination in vivo was dependent on the presence of the Cre protein and occurred specifically at the 34-base-pair lox sites. These results show that a prokaryotic protein can mediate controlled site-specific DNA synapsis and recombination in mammalian cells, suggesting that Cre-mediated recombination may be a useful tool for understanding and modulating genome rearrangements in eukaryotes. The Cre-lox site-specific recombination system of bacteriophage P1 is simple and well characterized. A single 38-kDa protein, Cre, is necessary and sufficient to catalyze recombination between two lox sites, each 34 bp in length. Recombination can occur between directly repeated sites on the same molecule to excise the intervening DNA segment or between inverted lox sites to give inversion of the intervening DNA segment. Both intra- and intermolecular recombination is catalyzed by Cre with either supercoiled or linear DNA. The Cre-lox system was shown to function efficiently in yeast and now is demonstrated to function in mammalian cells. The study used a mouse cell line, C-127, transformed with vectors containing the metallothionein I gene promoter to establish cell lines 55 and 3C-5. The Cre protein was induced by cadmium ions, and its expression was confirmed by immunoblot analysis. Recombination was assayed using plasmid pRH43 containing two directly repeated lox sites. Southern blot analysis showed that the Cre protein could excise the kanr gene from pRH43 as a 1.9-kb circular DNA molecule containing a single Bgl II site. The results demonstrated that the Cre recombinase is expressed in 55 cells and can perform precise lox-specific recombination on pRH43 to excise a 1.9-kb circular molecule containing the kanr gene. The study also used a herpesvirus vector, PRV42::pBS64, which carries a lox insertion. Precise recombination at the lox sites resulted in the reconstruction of the gIII gene, producing a glycoprotein recognized by a specific antibody. Recombinant viruses formed black plaques when