Curing Saccharomyces cerevisiae of the 2 micron plasmid by targeted DNA damage.
Published
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
Language
- eng
Conference Location
- England