Site-specific recombination of yeast 2-μm DNA in vitro

Abstract

Most strains of the yeast Saccharomyces cerevisiae harbor several copies of a 2-μm plasmid circle DNA termed '2 micron'. This circular plasmid contains two 599-base-pair precise inverted repeats across which a site-specific inversion event occurs in vivo. This inversion is promoted by a plasmid-encoded function called 'FLP'. We have cloned the FLP gene of 2-μm DNA under control of a strong yeast promoter and transformed yeast cells with a plasmid containing the cloned FLP gene. Cell-free extracts from such a transformant promote highly efficient inversion of 2-μm DNA in vitro. The reaction requires a cation and works efficiently on supercoiled, relaxed circular, or linear DNA. The FLP activity bears certain similarities to the cre protein, a site-specific recombinase encoded by bacteriophage P1.