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Medium No.	394
Medium	Pyrococcus Medium 2
Composition
Salt base solution* 1 L   Trace minerals** 10 ml   Citric acid 5 mg   Bacto Yeast Extract (Difco) 1 g   Bacto Peptone (Difco) 5 g   Sulfur (powder)*** 30 g   Na2S·9H2O 0.5 g   Resazurin 1 mg     Mix ingredients except sulfur and Na2S·9H2O, and adjust pH to 6.5 with H2SO4. Sterilize by filtration under a N2 atmosphere. Prepare Na2S·9H2O as a 5% solution, and autoclave under a N2 atmosphere. Aseptically and anaerobically add sterile sulfur to the medium. Prior to inoculation, add the sterile Na2S·9H2O solution, and readjust pH to 6.5, if necessary.                   *Salt base solution       NaCl 13.85 g       MgSO4·7H2O 3.5 g       MgCl2·6H2O 2.75 g       KCl 0.325 g       NaBr 0.05 g       H3BO3 15 mg       SrCl2·6H2O 7.5 mg       KI 0.05 mg       CaCl2 0.75 g       KH2PO4 0.5 g       (NH4)2Ni(SO4)2·6H2O 2 mg       Distilled water 1 L                   **Trace minerals       Nitrilotriacetic acid 1.5 g       MgSO4·7H2O 3 g       MnSO4·xH2O 0.5 g       NaCl 1 g       FeSO4·7H2O 0.1 g       CoSO4·7H2O 0.1 g       CaCl2·2H2O 0.1 g       ZnSO4·7H2O 0.1 g       CuSO4·5H2O 0.01 g       AlK(SO4)2 0.01 g       H3BO3 0.01 g       Na2MoO4·2H2O 0.01 g       Distilled water 1 L     Dissolve nitrilotriacetic acid and adjust pH to 6.5 with KOH solution. Then proceed to add minerals. Adjust final pH to 7.0.                   ***Sulfur powder     Sulfur is sterilized separately by autoclaving for 60 min at 105℃ on each of 3 successive days.
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