Abstract
Tetrahymena thermophila is a free-living ciliated protozoan useful in cell biological and molecular genetic studies. Recently DNA-mediated transformation has been developed for this microorganism (1), and used to study the mechanisms of genome rearrangement (2–4) and structure-function relationships of the RNA component of telomerase, an enzyme responsible for synthesis of chromosomal ends (5,6). Tetrahymena is also an interesting eukaryotic model for studying cell biological processes, such as cell motility, exocytosis, and pattern formation. Furthermore, recent evidence showed that Tetrahymena is one of only a few eukaryotic organisms known in which DNA recombination activity of transforming DNA is known to be exclusively homologous (7–10). Thus, gene replacement can be used in Tetrahymena as a tool to address fundamental problems in molecular cell biology.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Tondravi, M. M., and Yao, M.-C. (1986) Transformation of Tetrahymena thermophila by microinjection of ribosomal RNA genes. Proc. Natl. Acad. Sci. USA 83, 4369–4373.
Godiska, R. and Yao, M.-C. (1990) A programmed site-specific DNA rearrangement in Tetrahymena thermophila requires flanking polypurine tracts. Cell 61, 1237–1246.
Yao, M.-C., Yao, C.-H., and Monks, B. (1990) The controlling sequence for site-specific chromosome breakage in Tetrahymena. Cell 63, 763–772.
Yasuda, L. F., and Yao, M.-C. (1991) Short inverted repeats at a free end signal large palindromic DNA formation in Tetrahymena. Cell 67, 505–516.
Yu, G.-L., Bradley, J. D., Attardi, L. D., and Blackburn, E. H. (1990) In vivo alteration of telomere sequences and senescence caused by mutated Tetrahymena telomerase RNAs. Nature 344, 126–132.
Yu, G.-L. and Blackburn, E. H. (1991) Developmentally programmed healing of chromosomes by telomerase in Tetrahymena. Cell 67, 823–832.
Yu, G.-L., Hasson, M., and Blackburn, E. H. (1988) Circular ribosomal DNA plasmids transform Tetrahymena thermophila by homologous recombination with endogenous macronuclear ribosomal DNA. Proc. Natl. Acad. Sci. USA 85, 5151–5155.
Yao, M.-C. and Yao, C.-H. (1991) Transformation of Tetrahymena to cycloheximide resistance with a ribosomal protein gene through sequence replacement. Proc. Natl. Acad. Sci. USA 88, 9493–9497.
Kahn, R. W., Andersen, B. H., and Brunk, C. F. (1993) Transformation of Tetrahymena thermophila by microinjection of a foreign gene. Proc. Natl. Acad. Sci. USA 90, 9295–9299.
Gaertig, J., Thatcher, T. H., Gu, L., and Gorovsky, M. A. (1994) Electroporation-mediated replacement of a positively and negatively selectable β-tubulin gene in Tetrahymena thermophila. Proc. Natl. Acad. Sci. USA 91, 4549–4553.
Gaertig, J., Kiersnowska, M., and Iftode, F. (1988) Induction of cybrid strains of Tetrahymena thermophila by electrofusion. J. Cell Sci. 89, 253–261.
Gaertig, J. and Iftode, F. (1989) Rearrangement of the cytoskeleton and nuclear transfer in Tetrahymena thermophila cells fused by electric field. J. Cell Sci. 93, 691–703.
Brunk, C. F. and Navas, P. (1988) Transformation of Tetrahymena thermophila by electroporation and parameters effecting cell survival. Exp. Cell Res. 174, 525–532.
Orias, E., Larson, D., Hu, Y.-F., Yu, G.-L., Karttunen, J., Lovlie, A., Haller, B., and Blackburn, E. H. (1988) Replacement of the macronuclear ribosomal RNA genes of a mutant Tetrahymena using electroporation. Gene 70, 295–301.
Gaertig, J. and Gorovsky, M. A. (1992) Efficient mass transformation of Tetrahymena thermophila by electroporation of conjugants. Proc. Natl. Acad. Sci. USA 89, 9196–9200.
Martindale, D. W., Allis, C. D., and Bruns, P. J. (1982) Conjugation in Tetrahymena thermophila. A temporal analysis of cytological stages. Exp. Cell Res. 140, 227–236.
Gaertig, J., Gu, L., Hai, B., and Gorovsky, M. A. (1994) Gene replacement and high frequency vector-mediated electrotransformation in Tetrahymena. Nucleic Acids Res. 22, 5391–5398.
Nanney, D. L. (1980) Experimental Ciliatology. John Wiley, New York.
Doerder, F. P., Deak, J. C., and Lief, J. H. (1992) Rate of phenotypic assortment in Tetrahymena thermophila. Dev. Genet. 13, 126–132.
Gorovsky, M. A., Yao, M.-C., Keevert, J. B., and Pleger, G. L. (1975) Isolation of micro-and macronuclei of Tetrahymena pyriformis, in Methods in Cell Biology, vol. 9 (Prescott, D. M., ed.) Academic, New York, pp. 311–327.
Williams, N. E., Wolfe, J., and Bleyman, L. K. (1980) Long term maintenance of Tetrahymena spp. J. Protozool. 27, 327.
Flacks, M. (1979) Axenic storage of small volumes of Tetrahymena cultures under liquid nitrogen: a miniaturized procedure. Cryobiology 16, 287–291.
Bruns, P. J. and Brussard, T. B. (1974) Pair formation in Tetrahymena pyriformis, an inducible developmental system. J. Exp. Zool. 188, 337–344.
Orias, E. and Bruns, P. J. (1975) Induction and isolation of mutants in Tetrahymena, in Methods in Cell Biology (Prescott, D. M., ed.) Academic, New York, pp. 247–282.
Yao, M.-C. and Yao, C.-H. (1989) Accurate processing and amplification of cloned germ line copies of ribosomal DNA injected into developing nuclei of Tetrahymena thermophila. Mol. Cell. Biol. 9, 1092–1099.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 1995 Humana Press Inc.
About this protocol
Cite this protocol
Gaertig, J., Gorovsky, M.A. (1995). Gene Transfer by Electroporation in Tetrahymena . In: Nickoloff, J.A. (eds) Electroporation Protocols for Microorganisms. Methods in Molecular Biology™, vol 47. Humana Press. https://doi.org/10.1385/0-89603-310-4:331
Download citation
DOI: https://doi.org/10.1385/0-89603-310-4:331
Publisher Name: Humana Press
Print ISBN: 978-0-89603-310-8
Online ISBN: 978-1-59259-534-1
eBook Packages: Springer Protocols