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Cassettes for the PCR‐mediated construction of regulatable alleles in Candida albicans
Author(s) -
GeramiNejad Maryam,
Hausauer Danielle,
McClellan Mark,
Berman Judith,
Gale Cheryl
Publication year - 2004
Publication title -
yeast
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.923
H-Index - 102
eISSN - 1097-0061
pISSN - 0749-503X
DOI - 10.1002/yea.1080
Subject(s) - biology , ura3 , green fluorescent protein , gene , plasmid , genetics , mutant , selectable marker , candida albicans , promoter , gene expression
The recent availability of genome sequence information for the opportunistic pathogen Candida albicans has greatly facilitated the ability to perform genetic manipulations in this organism. Two important molecular tools for studying gene function are regulatable promoters for generating conditional mutants and fluorescent proteins for determining the subcellular localization of fusion gene products. We describe a set of plasmids containing promoter–GFP cassettes (P MET3 – GFP , P GAL1 – GFP , and P PCK1 – GFP ), linked to a selectable nutritional marker gene ( URA3 ). PCR‐mediated gene modification generates gene‐specific promoter, or gene‐specific promoter– GFP , fusions at the 5′‐end of the gene of interest. One set of primers can be used to generate three strains expressing a native protein of interest, or an amino‐terminal GFP‐tagged version, from three different regulatable promoters. Thus, these promoter cassette plasmids facilitate construction of conditional mutant strains, overexpression alleles and/or inducible amino‐terminal GFP fusion proteins. Copyright © 2004 John Wiley & Sons, Ltd.