Medium data
Search for medium no. = [1430].
Unless otherwise stated, sterilize media by autoclaving at 121°C for 15 min.
1430 SEMI-SOLID HETEROTROPHIC GROWTH MEDIUM FOR ALPHAPROTEOBACTERIA
| NaCl | 20.0 | g |
| MgCl2·6H2O | 6.0 | g |
| Na2SO4 | 3.24 | g |
| KCl | 0.5 | g |
| CaCl2·2H2O | 1.0 | g |
| NH4Cl | 0.3 | g |
| NaHCO3 | 1.26 | g |
| HEPES | 2.4 | g |
| Yeast extract (BD-Difco) | 0.2 | g |
| Disodium succinate | 1.0 | g |
| Modified Wolfe’s mineral solution (see Medium No. 915) | 5.0 | ml |
| Resazurin | 0.2 | mg |
| Distilled waterv | 1.0 | L |
| Agarose | 1.6 | g |
Mix components thoroughly, except agarose, and adjust pH to 6.7. Add agarose to the medium and autoclave under a N2 gas atmosphere. After cooling to 50C, add the following solutions to the medium aseptically and anaerobically (autoclaved or *filter-sterilized). Keep the medium temperature at 50 C until the medium is distributed into sterile screw-capped test tubes as described below:
| 0.5 M Potassium phosphate buffer (pH 7.0) | 1.8 | ml |
| Vitamin solution* (see Medium No. 915) | 0.5 | ml |
| 0.01 M FeSO4·7H2O solution (in 0.02 N HCl) | 3.0 | ml |
| 5% L-Cysteine·HCl·H2O solution | 8.0 | ml |
Adjust pH to 7.0 with sterile HCl or NaOH solution. Dispense the medium to sterile screw-capped test tubes (typically 11 ml medium into sterile Hungate tubes) under air and cap. Allow to stand until the medium is solidified and a O2-gradient formed (as evidenced by a pink upper oxic zone and a colorless lower anoxic zone). Inoculate cells just below the pink-colorless interface in the medium tube using a syringe.
Comment: The medium tubes can be stocked under a N2 gas atmosphere. In this case, prior to use, fill the head space with air and allow to stand until the O2 gradient is formed.
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