490 .
METHANOLOBUS OREGONENSIS MEDIUM
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Equipment needed:
Autoclave
Gassing station
Hungate tubes
Final gas composition:
100 % N2
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Final pH:
8.5
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ml
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
Yeast extract
(OXOID) |
2.0 | g | ||
|
|
Trypticase peptone
(BD BBL) |
2.0 | g | ||
|
|
NaCl | 29.0 | g | ||
|
|
KCl | 1.5 | g | ||
|
|
NH4Cl | 1.0 | g | ||
|
|
K2HPO4 x 3 H2O | 0.4 | g | ||
|
|
MgCl2 x 6 H2O | 1.7 | g | ||
|
|
2-Mercaptoethanesulfonic acid
(coenzyme M) |
0.5 | g | ||
|
|
Modified Wolin's mineral solution | 10.0 | ml | ||
|
|
Sodium resazurin
(0.1% w/v) |
0.5 | ml | ||
|
|
Na2CO3 | 2.5 | g | ||
|
|
Trimethylamine-HCl | 2.0 | g | ||
|
|
Na2S x 9 H2O | 0.3 | g | ||
|
|
Distilled water | 1000.0 | ml | ||
| 1 Dissolve ingredients except carbonate, trimethylamine and sulfide, then sparge medium with 100% N2 gas for 30 - 45 min to make it anoxic. Distribute medium under the same gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. Before use add trimethylamine and sulfide from sterile anoxic stock solutions autoclaved under 100% N2 gas and carbonate from a sterile anoxic stock solution prepared under 80% N2 and 20% CO2 gas mixture. Adjust pH of the complete medium to 8.5 with a sterile anoxic stock solution of Na2CO3 (5% w/v). | |||||
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 | ||
|
|
CoSO4 x 7 H2O | 0.18 | g | ||
|
|
CaCl2 x 2 H2O | 0.10 | g | ||
|
|
ZnSO4 x 7 H2O | 0.18 | g | ||
|
|
CuSO4 x 5 H2O | 0.01 | g | ||
|
|
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. | |||||