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Medium Detail Information

Medium No.
Methanol-using methanogen medium
  KH2PO4 0.227 g
  K2HPO4 0.348 g
  MgSO4·7H2O 0.5 g
  NH4Cl 0.5 g
  CaCl2·2H2O 0.25 g
  NaCl 2.25 g
  Bacto Yeast Extract (Difco) 2 g
  Hipolypepton (Nihon seiyaku) 2 g
  Trace elements solution* 10 ml
  Vitamin solution** 10 ml
  Methanol 10 ml
  NaHCO3 0.8 g
  Resazurin 1 mg
  Cystein-HCl 0.5 g
  Na2S·9H2O 0.5 g
  Distilled water 1 L
    Mix ingredients except vitamin solution, NaHCO3, cysteine-HCl and Na2S·9H2O, and adjust pH to 7 with NaOH. Dispense the medium into suitable culture vessels under a stream of N2/CO2 (80/20) and seal with butyl rubber stoppers. Separately autoclave cysteine-HCl and Na2S·9H2O as 5% solutions under a N2 atmosphere. Sterilize vitamin and 8% NaHCO3 solutions by filtration. Prior to inoculation, aseptically and anaerobically add vitamine, NaHCO3, cysteine-HCl and Na2S·9H2O solutions. Adjust pH to 7.0.
    *Trace elements solution
      Nitrilotriacetic acid (NTA) 12.8 g
      FeCl3·6H2O 1.35 g
      MnCl2·4H2O 0.1 g
      CoCl2·6H2O 0.024 g
      CaCl2·2H2O 0.1 g
      ZnCl2 0.1 g
      CuCl2·2H2O 0.025 g
      H3BO3 0.01 g
      Na2MoO4·2H2O 0.024 g
      NaCl 1 g
      NiCl2·6H2O 0.12 g
      Na2SeO4 0.004 g
      Na2WO4 0.004 g
      Distilled water 1 L
    First dissolve nitrilotriacetic acid and adjust pH to 6.5 with NaOH, and then add minerals. Final pH 7.0.
    **Vitamin solution
      Biotin 2 mg
      Folic acid 2 mg
      Pyridoxine-HCl 10 mg
      Thiamine-HCl 5 mg
      Riboflavin 5 mg
      Nicotinic acid 5 mg
      Ca-pantothenate 5 mg
      p-Aminobenzoic acid 1 mg
      Vitamin B12 0.01 mg
      Distilled water 1 L