Curing Saccharomyces cerevisiae of the 2 micron plasmid by targeted DNA damage.


Journal Article

Powerful mutagenic screens of yeast Saccharomyces cerevisiae have recently been developed which require strains that lack the endogenous 2 micron plasmid (Burns et al., 1994). Here, we describe a simple and reliable method for curing yeast of the highly stable genetic element. The approach employs heterologous expression of a 'step-arrest' mutant of the Flp recombinase. The mutant, Flp H305L (Parsons et al., 1988), forms long-lived covalent protein-DNA complexes exclusively at 2 micron-borne recombinase target sites. In vivo, the complexes serve as sites of targeted DNA damage. Using Southern hybridization and a colony color assay for plasmid loss, we show that expression of the mutant enzyme results in the effective elimination of the 2 micron from cells.

Full Text

Duke Authors

Cited Authors

  • Tsalik, EL; Gartenberg, MR

Published Date

  • June 30, 1998

Published In

Volume / Issue

  • 14 / 9

Start / End Page

  • 847 - 852

PubMed ID

  • 9818722

Pubmed Central ID

  • 9818722

International Standard Serial Number (ISSN)

  • 0749-503X

Digital Object Identifier (DOI)

  • 10.1002/(SICI)1097-0061(19980630)14:9<847::AID-YEA285>3.0.CO;2-9


  • eng

Conference Location

  • England