Cassettes for the PCR-mediated construction of regulatable alleles in Candida albicans

Maryam Gerami-Nejad, Danielle Hausauer, Mark McClellan, Judith Berman, Cheryl Gale

Research output: Contribution to journalArticlepeer-review

38 Scopus citations

Abstract

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 (PMET3-GFP, PGALI -GFP, and PPCK1-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.

Original languageEnglish (US)
Pages (from-to)429-436
Number of pages8
JournalYeast
Volume21
Issue number5
DOIs
StatePublished - Apr 15 2004

Keywords

  • Candida albicans
  • Epitope tagging
  • GAL1 promoter
  • Green fluorescent protein
  • MET3 promoter
  • PCK1 promoter
  • Polymerase chain reaction
  • Regulatable expression

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