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Medium Detail Information

Medium No.	361
Medium
Composition
Sodium acetate 1 g   Sodium succinate 1 g   KH2PO4 0.5 g   K2HPO4 0.6 g   (NH4)2SO4 1 g   MgSO4·7H2O 0.2 g   NaCl 0.2 g   CaCl2·2H2O 0.05 g   Na2S2O3·5H2O 0.1 g   Yeast extract 0.1 g   Vitamin mixture* 1 ml   Trace element solution** 1 ml   Neutralized Na2S·9H2O 0.5 g   Distilled water 1 L   Agar (if needed) 15 g   pH 7.3-7.5       Mix ingredients (except vitamin mixture) and adjust pH to 7.3-7.5. 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. Separately autoclave neutralized sulfide solution*** under a N2 atmosphere. Sterilize vitamin mixture by filtration. Sterile syringes are used to inoculate and remove samples. Prior to inoculation, aseptically and anaerobically add vitamin mixture and neutralized sulfide solution.                   *Vitamin mixture       Thiamine-HCl 50 mg       Niacine 50 mg       p-Aminobenzoic acid 30 mg       Pyridoxal-HCl 10 mg       Biotin 5 mg       Vitamin B12 5 mg       Distilled water 100 ml                   **Trace element solution       2Na-EDTA 1 g       FeCl3·6H2O 2 g       ZnCl2 0.1 g       MnCl2·2H2O 0.1 g       H3BO3 0.1 g       CoCl2·6H2O 0.1 g       Na2MoO4·2H2O 20 mg       CuCl2·2H2O 10 mg       NiCl2·6H2O 10 mg       Na2SeO3 5 mg       Distilled water 1 L                   ***Neutralized sulfide solution       Na2S·9H2O 1.5 g       Distilled water 100 ml     The sulfide solution is prepared in a vessel sealed with a butyl rubber stopper. Autoclave at 121℃ for 15 min. under a N2 atmosphere. After cooling to room temperature the pH is adjusted to about 7.3 by adding of sterile 2N H2SO4 drop-wise with a syringe without opening the bottle. Appearance of yellow colour indicates the pH is about 8. The vessel should be shaken continuously to avoid precipitation of elemental sulfur. The final solution should be transparent yellow.
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