| NaCl | 20 | g | | K2HPO4 | 0.14 | g | | CaCl2·2H2O | 1 | g | | NaNO3 | 1 | g | | MgSO4·7H2O | 3.4 | g | | MgCl2·6H2O | 4.2 | g | | KCl | 0.33 | g | | Bacto Yeast Extract (Difco) | 0.1 | g | | Sodium pyruvate | 0.2 | g | | Sodium lactate | 0.2 | g | | Sodium acetate | 0.2 | g | | Trace elements solution* | 10 | ml | | Vitamin solution** | 10 | ml | | NaHCO3 | 0.1 | g | | Resazurin | 1 | mg | | Na2S·9H2O | 0.5 | g | | Distilled water | 1 | L | | | | Mix ingredients except vitamin solution, NaHCO3 and Na2S·9H2O, and adjust pH to 7.5 - 8.0 with HCl. Dispense the medium into suitable culture vessels under a stream of H2/CO2 (80/20) and seal with butyl rubber stoppers. Separately autoclave Na2S·9H2O (5% solution) under a N2 atmosphere. Sterilize vitamin and NaHCO3 solutions by filtration. Prior to inoculation, aseptically and anaerobically add vitamin, NaHCO3 and Na2S·9H2O solutions, and pressurize the inoculated vessels to 150 kPa with H2/CO2 (80/20). | | | | | | | | | | *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 | 0.8 | mg | | | | Na2WO4 | 0.8 | mg | | | | KAl(SO4)2·12H2O | 4 | mg | | | | Distilled water | 1 | L | | | | First dissolve nitrilotriacetic acid and adjust pH to 6.5 with NaOH, and then add minerals. Final pH is 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 |
|