One-step gene disruption by cotransformation to isolate double auxotrophs in Candida albicans - PubMed (original) (raw)

One-step gene disruption by cotransformation to isolate double auxotrophs in Candida albicans

R Kelly et al. Mol Gen Genet. 1988 Sep.

Abstract

The Candida albicans LEU2 gene was disrupted by substituting lambda DNA for a small deletion within the LEU2 gene. Cotransformation with a selectable URA3 ARS vector was used to introduce a linear fragment containing the disruption into the genome of a C. albicans ura3 deletion mutant. Cotransformants containing the lambda DNA were identified by colony hybridization and the URA3 plasmid was subsequently cured. Leu2 disrupted heterozygotes were detected by Southern hybridization and one disruptant was subsequently treated with UV irradiation. Only one leu2 ura3 mutant (SGY-484) was isolated out of 11,000 mutagenized cells. SGY-484 was transformed to Leu+ with either the C. albicans or Saccharomyces cerevisiae LEU2 gene. Southern hybridization analysis revealed that the mutant is not homozygous for the disruption; the leu2 mutation reverts and is most likely a point mutation. Unexpectedly, an ade2 ura3 mutant was isolated from the same mutagenesis.

PubMed Disclaimer

References

    1. Microbiol Sci. 1988 Feb;5(2):58-63 - PubMed
    1. Mol Cell Biol. 1987 Jan;7(1):199-208 - PubMed
    1. Gene. 1980 Nov;11(3-4):291-8 - PubMed
    1. Mol Gen Genet. 1980;180(1):107-13 - PubMed
    1. J Bacteriol. 1981 Jun;146(3):833-40 - PubMed

MeSH terms

Substances

LinkOut - more resources