An improved protocol for the preparation of yeast cells for transformation by electroporation. 1998

J R Thompson, and E Register, and J Curotto, and M Kurtz, and R Kelly
Department of Biochemistry, Merck & Co., Rahway, NJ 07065, USA.

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.

UI MeSH Term Description Entries
D008094 Lithium An element in the alkali metals family. It has the atomic symbol Li, atomic number 3, and atomic weight [6.938; 6.997]. Salts of lithium are used in treating BIPOLAR DISORDER. Lithium-7,Lithium 7
D009693 Nucleic Acid Hybridization Widely used technique which exploits the ability of complementary sequences in single-stranded DNAs or RNAs to pair with each other to form a double helix. Hybridization can take place between two complimentary DNA sequences, between a single-stranded DNA and a complementary RNA, or between two RNA sequences. The technique is used to detect and isolate specific sequences, measure homology, or define other characteristics of one or both strands. (Kendrew, Encyclopedia of Molecular Biology, 1994, p503) Genomic Hybridization,Acid Hybridization, Nucleic,Acid Hybridizations, Nucleic,Genomic Hybridizations,Hybridization, Genomic,Hybridization, Nucleic Acid,Hybridizations, Genomic,Hybridizations, Nucleic Acid,Nucleic Acid Hybridizations
D002176 Candida albicans A unicellular budding fungus which is the principal pathogenic species causing CANDIDIASIS (moniliasis). Candida albicans var. stellatoidea,Candida stellatoidea,Dematium albicans,Monilia albicans,Myceloblastanon albicans,Mycotorula albicans,Parasaccharomyces albicans,Procandida albicans,Procandida stellatoidea,Saccharomyces albicans,Syringospora albicans
D004229 Dithiothreitol A reagent commonly used in biochemical studies as a protective agent to prevent the oxidation of SH (thiol) groups and for reducing disulphides to dithiols. Cleland Reagent,Cleland's Reagent,Sputolysin,Clelands Reagent,Reagent, Cleland,Reagent, Cleland's
D012441 Saccharomyces cerevisiae A species of the genus SACCHAROMYCES, family Saccharomycetaceae, order Saccharomycetales, known as "baker's" or "brewer's" yeast. The dried form is used as a dietary supplement. Baker's Yeast,Brewer's Yeast,Candida robusta,S. cerevisiae,Saccharomyces capensis,Saccharomyces italicus,Saccharomyces oviformis,Saccharomyces uvarum var. melibiosus,Yeast, Baker's,Yeast, Brewer's,Baker Yeast,S cerevisiae,Baker's Yeasts,Yeast, Baker
D014170 Transformation, Genetic Change brought about to an organisms genetic composition by unidirectional transfer (TRANSFECTION; TRANSDUCTION, GENETIC; CONJUGATION, GENETIC, etc.) and incorporation of foreign DNA into prokaryotic or eukaryotic cells by recombination of part or all of that DNA into the cell's genome. Genetic Transformation,Genetic Transformations,Transformations, Genetic
D015139 Blotting, Southern A method (first developed by E.M. Southern) for detection of DNA that has been electrophoretically separated and immobilized by blotting on nitrocellulose or other type of paper or nylon membrane followed by hybridization with labeled NUCLEIC ACID PROBES. Southern Blotting,Blot, Southern,Southern Blot
D018274 Electroporation A technique in which electric pulses, in kilovolts per centimeter and of microsecond-to-millisecond duration, cause a loss of the semipermeability of CELL MEMBRANES, thus leading to ion leakage, escape of metabolites, and increased uptake by cells of drugs, molecular probes, and DNA. Depending on the dosage, the formation of openings in the cell membranes caused by the electric pulses may or may not be reversible. Electric Field-Mediated Cell Permeabilization,Irreversible Electroporation,Reversible Electroporation,Electropermeabilisation,Electric Field Mediated Cell Permeabilization,Electroporation, Irreversible,Electroporation, Reversible

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