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PMID: 11004171 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

A single-transformation gene function test in diploid Candida albicans.

Journal of bacteriology ·Vol. 182 ·No. 20 ·2000-10-00 ·Pages 5730-6

Enloe B, Diamond A, Mitchell AP

Abstract

The fungal pathogen Candida albicans is naturally diploid, and current gene disruption strategies require two successive transformations. We describe here a genetic construct (UAU1) for which two copies may be selected. Insertion of UAU1 into one genomic site, after a single transformation, allows selection for segregants with two copies of the insertion. Major classes of segregants are those carrying homozygous insertion mutations and allelic triplications, which have two insertion alleles and a wild-type allele. Thus nonessential and essential genes may be distinguished rapidly through PCR tests for homozygosis and triplication. We find that homozygous mutations may be isolated at three nonessential loci (ADE2, RIM20, and YGR189), while only allelic triplications were found at two essential loci (SNF1 and CDC28). We have unexpectedly isolated homozygous mutants with mutations at CDC25; they are viable but defective in filamentation on serum-containing medium. The UAU1 cassette is thus useful to assess rapidly the essentiality of C. albicans genes.

MeSH Terms
Base Sequence Candida albicans/genetics,growth & development DNA Primers Diploidy Gene Deletion Genome, Fungal Homozygote Molecular Sequence Data Mutagenesis, Insertional Plasmids Polymerase Chain Reaction Recombination, Genetic Transformation, Genetic
Chemicals
DNA Primers
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Enloe B
Department of Microbiology, Columbia University, New York, New York 10032, USA.
Diamond A
Mitchell A P
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Article Info
Journal
Journal of bacteriology
Abbr.
J Bacteriol
ISSN
0021-9193
DOI
10.1128/JB.182.20.5730-5736.2000
Published
2000-10-00
Pages
5730-6
Language
English
Region
United States
NLM ID
2985120R
PMCID
PMC94694
Subset
IM
Grants
NIAID NIH HHS · P01 AI037194 · United States
NIAID NIH HHS · P01 AI37194 · United States
External Links
PubMed source DOI: 10.1128/JB.182.20.5730-5736.2000 PMC Full Text View journal details
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