One-step Targeted Gene Deletion in Candida Albicans Haploids

Nat Protoc. 2014 Feb;9(2):464-73. doi: 10.1038/nprot.2014.029. Epub 2014 Jan 30.


The recent discovery of haploids in Candida albicans and the construction of tool strains carrying multiple auxotrophic markers have enabled, for the first time, performing one-step gene deletions in this fungal human pathogen. This breakthrough promises to greatly facilitate the molecular and genetic study of C. albicans biology and pathogenicity. However, the construction of gene-deletion mutants in C. albicans haploids involves many technical difficulties, particularly low transformation efficiency and autodiploidization. Here we describe a highly effective protocol for designing and performing one-step gene deletion in C. albicans haploids, which takes ∼11 d to complete (not including plasmid construction, which may take ∼2 weeks). A gene deletion cassette is constructed on a plasmid and subsequently released for transformation by lithium acetate incubation or electroporation. Desired gene-deletion mutants are identified and their ploidy is assessed simultaneously by colony PCR before final confirmation by flow cytometry.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acetates
  • Candida albicans / genetics*
  • Electroporation
  • Flow Cytometry
  • Gene Deletion*
  • Genetic Engineering / methods*
  • Haploidy*
  • Plasmids / genetics
  • Polymerase Chain Reaction


  • Acetates
  • lithium acetate