| | MgSO4·7H2O | 0.5 | g |
| | CaCl2·2H2O | 0.1 | g |
| | NH4Cl | 0.1 | g |
| | KH2PO4 | 0.1 | g |
| | KCl | 0.1 | g |
| | NaNO3 | 0.85 | g |
| | Sodium acetate | 0.16 | g |
| | Trace element mixture* | 1 | ml |
| | Selenite-tungstate solution** | 1 | ml |
| | Bicarbonate solution*** | 30 | ml |
| | Vitamin solution**** | 1 | ml |
| | Thiosulfate solution**** | 20 | ml |
| | Distilled water | 930 | ml |
| | pH 7.0-7.2 | |
|
| | | Mix ingredients except for bicarbonate, vitamin and thiosulfate solutions. Dispense the medium into suitable culture vessels under a stream of N2/CO2 (80/20) gas, seal with butyl rubber stoppers, and autoclave at 121°C for 20 min. Aseptically and anaerobically add other ingredients using sterile syringes. |
| | | | | | | |
| | | *Trace element mixture |
| | | | HCl (25%; 7.7 M) | 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. |
| | | | | | | |
| | | ***Bicarbonate solution (1.0 M) |
| | | | NaHCO3 | 8.4 | g |
| | | | Distilled water | 100 | ml |
|
| | | Saturated with CO2 gas. Tightly sealed in bottles filled with CO2 gas and then sterilized by autoclaving. |
| | | | | | | |
| | | ****Vitamine solution |
| | | | p-Aminobenzoic acid | 4 | mg |
| | | | Biotin | 1 | mg |
| | | | Nicotinic acid | 10 | mg |
| | | | Ca-pantothenate | 5 | mg |
| | | | Pyridoxine-HCl | 15 | mg |
| | | | Thiamine-HCl | 10 | mg |
| | | | Cyanocobalamin | 5 | mg |
| | | | Sodium phosphate buffer (10 mM; pH7.1) | 100 | ml |
|
| | | Sterilized by filtration. |
| | | | | | | |
| | | ****Thiosulfate solution (1.0 M) |
| | | | Na2S2O3·5H2O | 24.8 | g |
| | | | Distilled water | 100 | ml |
|
| | | Sterilized by filtration and stored under N2 gas. |