A Recyclable Candida Albicans URA3 Cassette for PCR Product-Directed Gene Disruptions

Yeast. 2000 Jan 15;16(1):65-70. doi: 10.1002/(SICI)1097-0061(20000115)16:1<65::AID-YEA508>3.0.CO;2-M.

Abstract

For some time, gene disruptions in Candida albicans have been made with the hisG-URA3-hisG ('Ura-blaster') cassette, which can be re-used in successive transformations of a single strain after homologous excision of URA3. However, the hisG repeats are too large for efficient PCR amplification of the entire cassette, so it cannot be used for PCR product-directed gene disruptions. We describe here a gene disruption cassette, URA3-dpl200, with 200 bp flanking repeats that permit efficient PCR amplification. After transformation and integration to produce both arg5::URA3-dpl200 and rim101::URA3-dpl200 alleles, we find that arg5::dpl200 and rim101::dpl200 segregants, respectively, can be obtained. We have used the cassette to create rim101::dpl200/rim101::URA3-dpl200 mutants exclusively through PCR product-directed disruption.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Candida albicans / genetics*
  • Fungal Proteins / genetics*
  • Polymerase Chain Reaction*
  • Transformation, Genetic*

Substances

  • Fungal Proteins