Site-specific cleavage of chromosomes in vitro through Cre-lox recombination.
Site-specific recombination systems are useful tools for chromosome engineering in vivo and site-specific DNA cleavage methods have applications in genome analysis and gene isolation. Here, we report a new method to fragment chromosomes in vitro using the Cre-lox site-specific recombination system. Two lox sites were targeted into the 5.7 Mb chromosomes I of Schizosaccharomyces pombe. In vitro recombination between chromosomal lox sites and exogenously provided lox oligonucleotides 'cleaved' the chromosome at the defined lox sequences. Site-specific cleavage of lox sites in the tobacco genome was also demonstrated. This recombination-based cleavage method provides a novel approach for structural and functional analyses of eukaryotic chromosomes as it allows direct isolation of chromosome regions that correspond to phenotypes revealed through Cre-lox mediated chromosome rearrangements in vivo. Moreover, recombination with end-labeled lox oligonucleotides would permit the specific end-labeling of chromosome segments to facilitate the long range mapping of chromosomes.
Qin
M
M
Plant Gene Expression Center, US Department of Agriculture, Albany, CA 94710, USA.
Lee
E
E
Zankel
T
T
Ow
D W
DW
D001483:Base Sequence; D002875:Chromosomes / Q000235:genetics*; D008969:Molecular Sequence Data; D010957:Plasmids; D011995:Recombination, Genetic; D012568:Schizosaccharomyces / Q000235:genetics*