An improved protocol for the preparation of yeast cells for transformation by electroporation.
Pretreatment of yeast cells with lithium acetate (LiAc) and dithiothreitol (DTT) enhances the frequency of transformation by electroporation. The method shows improvements of 6-67-fold in wild-type strains derived from commonly used Saccharomyces cerevisiae genetic backgrounds. In addition, 15-300-fold improvement in transformation frequency was achieved with several mutant strains of S. cerevisiae that transformed poorly by conventional procedures. Both DTT and lithium acetate were necessary for maximal transformation frequencies. Pretreatment with lithium and DTT also resulted in an approximately 3-5-fold increase in the electroporation transformation frequency of the pathogenic fungus Candida albicans.
Thompson
J R
JR
Department of Biochemistry, Merck & Co., Rahway, NJ 07065, USA.
Register
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Curotto
J
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Kurtz
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Kelly
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D015139:Blotting, Southern; D002176:Candida albicans / Q000187:drug effects / Q000235:genetics*; D004229:Dithiothreitol / Q000494:pharmacology; D018274:Electroporation / Q000379:methods*; D008094:Lithium / Q000494:pharmacology; D009693:Nucleic Acid Hybridization; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics*; D014170:Transformation, Genetic
D008094:Lithium; D004229:Dithiothreitol