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Medium No.
869
Medium
Medium for strain C/G2
Composition
  KH2PO4 0.2 g
  NH4Cl 0.25 g
  NaCl 20 g
  MgCl2·6H2O 3 g
  KCl 0.5 g
  CaCl2·2H2O 0.15 g
  Trace element mixture* 1 ml
  Selenite-tungstate solution** 1 ml
  Vitamin mixture*** 1 ml
  Vitamin B12 solution**** 1 ml
  Bicarbonate solution (1.0 M)***** 30 ml
  Dithionite (ca.) 20 mg
  Distilled water 1 L
    Mix ingredients except vitamin mixture, vitamin B12 solution, bicarbonate solution and dithionite, and adjust pH to about 7.5. Dispense the medium into suitable culture vessels under a stream of nitrogen gas, seal with butyl rubber stoppers, and autoclave at 121°C for 15 min. Aseptically and anaerobically add vitamin mixture, vitamin B12 solution, bicarbonate solution and dithionite solution using sterile syringes, and the N2/CO2 (80/20) mixed gas is enclosed with the vessels. Adjust pH to 7.2- 7.8 using compact pH meter. An aqueous solution of sodium glutamate (final concentration, 10 mM) and sodium thiosulfate (final concentration, 20 mM) are added before inoculation. Addition of yeast extract (0.5 - 1 g/L) greatly improves the growth of the strain.
             
    *Trace element mixture
      HCl (25%) 12.5 ml
      FeSO4·7H2O 2100 mg
      H3BO3 30 mg
      MnCl2·4H2O 100 mg
      CoCl2·6H2O 190 mg
      NiCl2·6H2O 24 mg
      CuCl2·2H2O 2 mg
      ZnSO4·7H2O 144 mg
      Na2MoO4·2H2O 36 mg
      Distilled water 987 ml
 
    Sterilized by autoclaving.
             
    **Selenite-tungstate solution
      NaOH 0.4 g
      Na2SeO3·5H2O 6 mg
      Na2WO4·2H2O 8 mg
      Distilled water 1 L
 
    Sterilized by autoclaving.
             
    ***Vitamin mixture
      4-Aminobenzoic acid 4 mg
      D(+)-Biotin 1 mg
      Nicotinic acid 10 mg
      Calcium D(+)-panthothenate 5 mg
      Pyridoxine dihydrochloride 15 mg
      Thiamine-HCl 10 mg
      Distilled water 100 ml
 
    Sterilized by filtration.
             
    ****Vitamin B12 solution
      Cyanocobalamin 5 mg
      Distilled water 100 ml
 
    Sterilized by filtration.
             
    *****Bicarbonate solution (1.0 M)
      NaHCO3 84 g
      Distilled water 1 L
 
    Sterilized by filtration.
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