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Main sol. 135

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Internal Solution-ID: S231

Equipment needed: Hungate tubes Autoclave Filter Gassing station
Compound Amount Unit Conc. [g/L] Conc. [mM]
NH4Cl 1.00 g 0.979 18.311
KH2PO4 0.33 g 0.323 2.375
K2HPO4 0.45 g 0.441 2.53
MgSO4 x 7 H2O 0.10 g 0.098 0.397
Modified Wolin's mineral solution 20.00 ml - -
Yeast extract 2.00 g 1.959 -
Sodium resazurin
(0.1% w/v) 		0.50 ml 4.9e-4 0.002
NaHCO3 10.00 g 9.794 116.589
D-Fructose 10.00 g 9.794 54.365
Wolin's vitamin solution (10x) 1.00 ml - -
L-Cysteine HCl x H2O 0.50 g 0.49 2.788
Na2S x 9 H2O 0.50 g 0.49 2.039
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 7.5 - 8.0 by adding a sterile anoxic stock solution of sodium carbonate (5% w/v) prepared under 80% N2 and 20% CO2 gas mixture 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