Targeted integrative transformation of Candida tropicalis by electroporation.
A method for integrative transformation of the diploid yeast Candida tropicalis by electroporation has been developed. by linearizing the transforming plasmid DNA containing the URA3 gene prior to electroporation of recipient cells, its integration was targeted to a specific locus in the genome, resulting in single or multiple tandem integrations. The optimal electroporation conditions for this yeast were established and include an electric pulse of 2.25 kV/cm for a duration of 50 ms. Using these conditions, Ura+ transformants were readily obtained at a high frequency (45 transformants/micrograms DNA) as the result of targeted integration of the URA3 gene containing plasmid DNA at the chromosomal ura3 locus. The number of transformants resulting from this procedure is comparable to that achieved with a recently reported spheroplast transformation procedure for C. tropicalis; in addition, it offers the advantages of being simple, rapid and reproducible.
Microbial Technology Department, Cognis, Inc., Santa Rosa, CA 95407.
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D002175:Candida / Q000235:genetics*; D002453:Cell Cycle; D002455:Cell Division; D004247:DNA / Q000494:pharmacology; D004553:Electric Conductivity; D004558:Electric Stimulation / Q000379:methods*; D007477:Ions; D009693:Nucleic Acid Hybridization; D014170:Transformation, Genetic / Q000187:drug effects
D007477:Ions; D004247:DNA