| Na2HPO4·2H2O | 0.53 | g | | KH2PO4 | 0.41 | g | | NH4Cl | 0.3 | g | | CaCl2·2H2O | 0.11 | g | | MgCl2·6H2O | 0.1 | g | | NaCl | 0.3 | g | | Vitamin solution* | 10 | ml | | Trace elements solution** | 10 | ml | | Bacto Yeast Extract (Difco) | 0.2 | g | | Methanol | 2 | ml | | Resazurin | 0.5 | mg | | NaHCO3 | 4 | g | | Na2S·9H2O | 0.5 | g | | Distilled water | 1 | L | | | | Mix ingredients except for vitamin, CaCl2·2H2O, MgCl2·6H2O, methanol and Na2S·9H2O, and autoclave under a N2/CO2 atmosphere (80/20). Separately autoclave Na2S·9H2O solution as 5% solution under a N2 atmosphere. Separately autoclave 1.1% CaCl2·2H2O and 1% MgCl2·6H2O solution under a N2/CO2 atmosphere. Prior to inoculation, add the filter-sterile vitamin, mgnesium and calcium solution, filter-sterile methanol and Na2S·9H2O. Final pH is 7.0-7.2. | | | | | | | | | | *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 | | | | | | | | | | **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. |
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