Efficient mass transformation of Tetrahymena thermophila by electroporation of conjugants.
Conjugating cells of the ciliate Tetrahymena thermophila were electroporated in the presence of plasmid DNA containing a paromomycin-resistant ribosomal RNA gene (rDNA). Cells were selected with paromomycin following 12-24 hr of growth on nonselective medium. Resistant cells appeared after 2-3 days. Processing vectors containing the micronuclear rDNA and somatic vectors containing the macronuclear gene transformed the cells, with the former yielding frequencies up to 900 transformants per microgram of plasmid DNA. A ribosomal protein gene (rpL29) conferring cycloheximide resistance also transformed conjugating cells. The transformation efficiency of the plasmid containing only the rpL29 gene was increased by insertion of an rDNA replication origin and by cotransformation and preselection with an rDNA vector. These results indicate that electroporation can be used for the production of large numbers of transformed Tetrahymena.
Gaertig
J
J
Department of Biology, University of Rochester, NY 14627.
Gorovsky
M A
MA
D000818:Animals; D015139:Blotting, Southern; D003227:Conjugation, Genetic; D004275:DNA, Ribosomal / Q000235:genetics* / Q000302:isolation & purification; D004351:Drug Resistance / Q000235:genetics; D004558:Electric Stimulation; D005821:Genetic Techniques; D005822:Genetic Vectors; D010303:Paromomycin / Q000494:pharmacology; D010957:Plasmids; D016808:Tetrahymena thermophila / Q000187:drug effects / Q000235:genetics*; D014162:Transfection; D014170:Transformation, Genetic
D004275:DNA, Ribosomal; D010303:Paromomycin