Properties of electroporation-mediated DNA transfer in Escherichia coli.
Efficient and reproducible DNA-transfection was attained in E. coli, by electroporation. The yield of the transfectants was affected by pretreatment of the recipient cells as well as by the composition of the electroporation medium. Using a single pulse procedure, relationships among the electrical parameters, the transfection efficiency, and the cellular viability were investigated in 10 mM Tris-HCl buffer (pH 7.5) containing 5% sucrose. Certain sodium salts (e.g., citrate, phosphate, and sulfate) were promotive, whereas Mg2+, DEAE-dextran, and polyvinylpyrrolidone were inhibitory to the transfection. Heterologous nucleic acids (native DNA, denatured DNA, and tRNA) exerted only a marginal effect on transfection with a viral replicative-form DNA. The efficiency of DNA transfer was affected by culture conditions, and bacteria grown at a higher temperature were more competent. The electroporation system was more efficient than an improved CaCl2 method, not only in transfection with viral single- and double-stranded DNAs, but also in transformation with plasmid DNAs.
Taketo
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Department of Biochemistry I, Fukui Medical School.
D002122:Calcium Chloride / Q000494:pharmacology; D004269:DNA, Bacterial / Q000235:genetics* / Q000528:radiation effects; D004560:Electricity; D004926:Escherichia coli / Q000378:metabolism*; D007202:Indicators and Reagents; D010957:Plasmids; D013395:Sucrose; D014161:Transduction, Genetic; D014162:Transfection; D014169:Transformation, Bacterial
D004269:DNA, Bacterial; D007202:Indicators and Reagents; D013395:Sucrose; D002122:Calcium Chloride