Transformation of Tetrahymena thermophila by electroporation and parameters effecting cell survival.
We have successfully transformed Tetrahymena thermophila by electroporation, a process of electrically introducing DNA. The DNA used for transformation contains a mutant ribosomal RNA gene (rDNA) that confers resistance to paromomycin on the transformed cells. This mutant rDNA replicates more rapidly than the endogenous rDNA of the transformed cells so that the mutant rDNA becomes predominant within several generations. This mutant rDNA also carries a restriction polymorphism that readily distinguishes it from the endogenous rDNA of the transformed cells. Substantial nuclease activity is released from the cells during electroporation and must be neutralized in order for transformation to be effective. Cell survival is inversely proportional to the electrical energy dissipated (joules) in the medium. Electroporation is a convenient and effective means of introducing transforming DNA into T. thermophila.
D000818:Animals; D002463:Cell Membrane Permeability; D004275:DNA, Ribosomal / Q000235:genetics*; D004351:Drug Resistance / Q000235:genetics; D004553:Electric Conductivity; D005796:Genes; D005818:Genetic Engineering / Q000379:methods*; D006863:Hydrogen-Ion Concentration; D009154:Mutation; D010303:Paromomycin / Q000494:pharmacology; D012150:Polymorphism, Restriction Fragment Length; D013768:Tetrahymena / Q000187:drug effects / Q000235:genetics* / Q000502:physiology; D014170:Transformation, Genetic
D004275:DNA, Ribosomal; D010303:Paromomycin