135 .
ACETOBACTERIUM MEDIUM
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Selected strain: Acetobacterium wieringae DSM 1911
This strain uses the standard medium without modifications.
Growth conditions: anaerobic
30 °C
Equipment needed:
Autoclave
Filter
Gassing station
Hungate tubes
Final pH:
7.5 - 8.0
Select a strain (optional):
ml
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
NH4Cl | 1.00 | g | ||
|
|
KH2PO4 | 0.33 | g | ||
|
|
K2HPO4 | 0.45 | g | ||
|
|
MgSO4 x 7 H2O | 0.10 | g | ||
|
|
Modified Wolin's mineral solution | 20.00 | ml | ||
|
|
Yeast extract | 2.00 | g | ||
|
|
Sodium resazurin
(0.1% w/v) |
0.50 | ml | ||
|
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NaHCO3 | 10.00 | g | ||
|
|
D-Fructose | 10.00 | g | ||
|
|
Wolin's vitamin solution (10x) | 1.00 | ml | ||
|
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L-Cysteine HCl x H2O | 0.50 | g | ||
|
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Na2S x 9 H2O | 0.50 | g | ||
|
|
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. | |||||
Modified Wolin's mineral solution #
1000
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
Nitrilotriacetic acid | 1.50 | g | ||
|
|
MgSO4 x 7 H2O | 3.00 | g | ||
|
|
MnSO4 x H2O | 0.50 | g | ||
|
|
NaCl | 1.00 | g | ||
|
|
FeSO4 x 7 H2O | 0.10 | g | ||
|
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CoSO4 x 7 H2O | 0.18 | g | ||
|
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CaCl2 x 2 H2O | 0.10 | g | ||
|
|
ZnSO4 x 7 H2O | 0.18 | g | ||
|
|
CuSO4 x 5 H2O | 0.01 | g | ||
|
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AlK(SO4)2 x 12 H2O | 0.02 | g | ||
|
|
H3BO3 | 0.01 | g | ||
|
|
Na2MoO4 x 2 H2O | 0.01 | g | ||
|
|
NiCl2 x 6 H2O | 0.03 | g | ||
|
|
Na2SeO3 x 5 H2O | 0.30 | mg | ||
|
|
Na2WO4 x 2 H2O | 0.40 | mg | ||
|
|
Distilled water | 1000.00 | ml | ||
| 1 First dissolve nitrilotriacetic acid and adjust pH to 6.5 with KOH, then add minerals. Adjust final to pH 7.0 with KOH. | |||||
Wolin's vitamin solution (10x) #
1000
| Compound | Amount | Unit | |
|---|---|---|---|
|
|
Biotin | 20 | mg |
|
|
Folic acid | 20 | mg |
|
|
Pyridoxine hydrochloride | 100 | mg |
|
|
Thiamine HCl | 50 | mg |
|
|
Riboflavin | 50 | mg |
|
|
Nicotinic acid | 50 | mg |
|
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Calcium D-(+)-pantothenate | 50 | mg |
|
|
Vitamin B12 | 1 | mg |
|
|
p-Aminobenzoic acid | 50 | mg |
|
|
(DL)-alpha-Lipoic acid | 50 | mg |
|
|
Distilled water | 1000 | ml |