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Medium No.	1190
Medium	MMJYP2 Medium
Composition
Yeast extract 4 g   Bacto Tryptone (Difco) 4 g   Sodium pyruvate 0.2 g   NaHCO3 1 g   Na2S 0.5 g   MJ synthetic seawater for MMJYP2 medium * 1 L   pH 6.5         Mix ingredients except for pyruvate, sulfide and bicarbonate. Dispense the medium into suitable culture vessels under a stream of N2/CO2 (80/20), seal with butyl rubber stoppers, and autoclave at 121°C for 15 min. After cooling to room temperature, aseptically and anaerobically add the filter-sterilized bicarbonate and pyruvate to the medium, and the pH is adjusted to 6.5 by adding of sterile pH-adjusting agent with a syringe without opening the vessel. Prior to inoculation, aseptically add the stelile neutralized sulfide solution, and pressurize the vessels to 350 kPa with N2/CO2.                   *MJ synthetic seawater for MMJYP2 medium       NaCl 25 g       K2HPO4 0.14 g       CaCl2·2H2O 0.7 g       NH4Cl 0.25 g       MgSO4·7H2O 3.4 g       MgCl2·6H2O 4.2 g       KCl 0.5 g       Balch's trace mineral solution** 10 ml       Distilled water 990 ml                   **Balch's trace mineral solution       Nitrilotriacetic acid (NTA) 1.5 g       MgSO4·7H2O 3 g       MnSO4·2H2O 0.5 g       NaCl 1 g       FeSO4·7H2O 0.1 g       CoSO4·7H2O 0.18 g       CaCl2·2H2O 0.1 g       ZnSO4·7H2O 0.18 g       CuSO4·5H2O 0.01 g       KAl(SO4)2·12H2O 0.02 g       H3BO3 0.01 g       Na2MoO4·2H2O 0.01 g       NiCl2·6H2O 0.025 g       Na2SeO3·5H2O 0.3 mg       Distilled water 1 L       First dissolve nitrilotriacetic acid and adjust pH to 6.5 with KOH, then add minerals. Final pH 7.0 (with KOH).
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