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Main sol. 135

Internal Solution-ID: S231

Equipment needed: Hungate tubes Autoclave Filter Gassing station
Compound Amount Unit Conc. [g/L] Conc. [mM]
NH4Cl 1.00 g 1 18.695
KH2PO4 0.33 g 0.33 2.425
K2HPO4 0.45 g 0.45 2.584
MgSO4 x 7 H2O 0.10 g 0.1 0.406
Modified Wolin's mineral solution 20.00 ml - -
Yeast extract 2.00 g 2 -
Sodium resazurin
(0.1% w/v)
0.50 ml 5.0e-4 -
NaHCO3 10.00 g 10 119.038
D-Fructose 10.00 g 10 55.506
Wolin's vitamin solution 10.00 ml - -
L-Cysteine HCl x H2O 0.50 g 0.5 2.847
Na2S x 9 H2O 0.50 g 0.5 2.082
Distilled water 1000.00 ml - -
1 Dissolve ingredients except bicarbonate, fructose, vitamins, cysteine and sulfide, bring to the boil and cool to room temperature under 80% N2 and 20% CO2 gas mixture. Add bicarbonate (solid) and equilibrate the medium with the gas until a pH of around 7.4 is reached. Then distribute under the same gas atmosphere in anoxic Hungate-type tubes or serum vials and autoclave. Before use adjust the pH to 8.0 - 8.2 by adding a sterile anoxic stock solution of sodium carbonate (5% w/v) prepared under 80% N2 and 20% CO2 gas mixture (c. 0.25 ml per 10 ml medium) and add fructose, vitamins (sterilized by filtration), cysteine and sulfide from anoxic sterile stock solutions prepared under 100% N2.
2 Note: For autotrophic growth D-fructose is omitted and a gas atmosphere of 80% H2 and 20% CO2 is used.
135 ACETOBACTERIUM MEDIUM