Tetrahymena Stock Center

2% proteose peptone
10 μM FeCl₃ or 90 μM sequestrene (Fe-EDTA)
Absence of glucose and yeast extract is reported not to affect doubling time.

See: Asai DL and Forney JD. 2000. Tetrahymena thermophila. methods in cell biology. San Diego: Academic Pres. Chapter 4.

0.25% proteose peptone
0.25% yeast extract
0.5% glucose
33.3 μM FeCl₃

To avoid precipitate formation, the FeCl₃ is first dissolved in one quarter of the final H₂O volume, and the glucose, yeast and PP are added and dissolved next. The remainder of the H₂O is then added, bottled and autoclaved.

See: Cassidy-Hanley D, Bowen J, Lee JH, Cole E, VerPlank LA, Gaertig J, Gorovsky MA, Bruns PJ. 1997. Germline and somatic transformation of mating tetrahymena thermophila by particle bombardment. Genetics 146(1):135-47.

2% proteose peptone
0.1% yeast extract
0.2% glucose
0.003% sequestrene (Fe-EDTA), can be replaced with 33 μM FeCl₃

See: Gorovsky MA, Yao MC, Keevert JB, Pleger GL. 1975. Isolation of micro- and macronuclei of tetrahymena pyriformis. Methods Cell Biol 9(0):311-27.

1% proteose peptone
0.15% yeast extract
0.01 mM FeCl₃

See: Smith DL and Doerder FP. 1992. Multiple effects of mutation on expression of alternative cell surface protein genes in tetrahymena thermophila. Genetics 130(1):97-104.

0.4% proteose peptone
0.2% yeast extract
1.0% glucose

See: Mori K., Yomo T., Kashiwagi A. 2011. Single-cell isolation and cloning of tetrahymena thermophila cells with a fluorescence-activated cell sorter. J.Eukaryotic Microbiol.Journal of Eukaryotic Microbiology 58(1):37-42.

1% proteose peptone
0.15% yeast extract
0.01 mM FeCl₃
0.2 M NaCl

See: Smith DL and Doerder FP. 1992. Multiple effects of mutation on expression of alternative cell surface protein genes in tetrahymena thermophila. Genetics 130(1):97-104.

1. Unless otherwise specified, all stock solutions are made with distilled water, sterilized by filtration and stored at 4°C.
• Amino acids solutions:
  • A-D: 40-fold concentrated.
  • A, C: adjust pH to 7.
  • B: store frozen.
  • D: dissolve Asp and Glu in water with stirring, use 1 N KOH to prevent the pH from dropping below 7, add Ala and Gly, and adjust pH to 7.
• Nucleosides solution: 10-fold concentrated.
• Salts and chelator solution: 100-fold concentrated.
• Vitamins solutions: 100-fold concentrated.
  • A: store frozen.
  • B: dissolve in 1 ml absolute ethanol, dilute in 100 ml H₂O.
  • C: adjust pH to 7.
• Trace metals solution: 100-fold concentrated, adjust pH to approximately 2 with 1 N HCl.
• Glucose solution: 50-fold concentrated.
2. Begin by dissolving Tyr at 60°C to give 0.2 mg/ml in the final medium, cool and proceed to add the remaining solutions. The pH may be adjusted as required. The medium may be sterilized by autoclaving or by filtration; if autoclaving is used, glucose should be added aseptically after cooling.
3. Special purpose modifications:
   • Amino acid solution A contains all the required amino acids; omission of amino acid solutions B-E yields a minimal defined medium.
   • When inocula of less than 2500 cells/ml of final medium will be used, the medium should be supplemented with hemin at a final concentration of 7.5 μM; a stock solution is prepared by dissolving in 0.01 N NaOH and autoclaving.
   • For growing phagocytosis-deficient cells, the final concentrations of FeCl₃, CuSO₄ and folinic acid should be increased to 1mM, 25 μM and 1 μg/ml respectively.

See:

Szablewski L, Andreasen PH, Tiedtke A, Florin-Christensen J, Florin-Christensen M, Rasmussen L. 1991. Tetrahymena thermophila: Growth in synthetic nutrient medium in the presence and absence of glucose. J Eukaryot Microbiol 38(1):62-5.

Christensen ST and Rasmussen L. 1992. Evidence for growth factors which control cell multiplication in tetrahymena thermophila. Acta Protozoologica 31(4):215-219.

Sanford YM and Orias E. 1981. Phenylketonuric tetrahymena: Phenylalanine hydroxylase mutants and other tyrosine auxotrophs. Proc Natl Acad Sci U S A 78(12):7614-8.

Rasmussen L, Christensen ST, Schousboe P, Wheatley DN. 1996. Cell survival and multiplication. the overriding need for signals: From unicellular to multicellular systems. FEMS Microbiol Lett 137(2-3):123-8.

Asai DL and Forney JD. 2000. Tetrahymena thermophila. methods in cell biology. San Diego: Academic Pres. Chapter 4.

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Simon EM and Hwang SW. 1967. Tetrahymena: Effect of freezing and subsequent thawing on breeding performance. Science 155(763):694-6.

Asai DL and Forney JD. 2000. Tetrahymena thermophila. methods in cell biology. San Diego: Academic Pres. Chapter 4.

0.59 g of Na citrate-2H₂O (2 mM)
0.14 g of NaH₂PO₄ • H₂O (1 mM)
0.14 g of Na₂HPO₄ (1 mM)
0.13 g of CaCl₂ (1.5 mM)
To avoid precipitation of the CA phosphate, the CaCl₂ solution is autoclaved separately from the mixture of sodium salts, and the two solutions are mixed aseptically after cooling.

See: Dryl S. 1959. Antigenic transformation in paramecium aurelia after homologous antiserum treatment during autogamy and conjugation. J Protozool 6(25).

10 mM Tris HCl, pH 7.5

See: Bruns PJ and Brussard TB. 1974. Pair formation in tetrahymena pyriformis, an inducible developmental system. J Exp Zool 188(3):337-44.

0.2% NaCl
0.008% KCl
0.012% CaCl₂

See: Sugai T and Hiwatashi K. 1974. Cytologic and autoradiographic studies of the micronucleus at meiotic prophase in tetrahymena pyriformis. J Protozool 21(4):542-8.

Composition per liter of medium:

20g (2%) PP
0.6g (2mM) Na₃ citrate 2H₂O
0.27g (1 mM) FeCl₃
7.2 mg (30 μM) CuSO₄ • 5H₂O
1 mg (1.7 μM) Ca salt (Lederle)

See: Orias E and Rasmussen L. 1976. Dual capacity for nutrient uptake in tetrahymena. IV. growth without food vacuoles and its implications. Exp Cell Res 102(1):127-37.

Use the standard chemically defined medium as shown above. Then increase the concentrations of FeCl₃, CuSO₄ and folinic acid to 1 mM, 25 μM and 1 μg/ml respectively.