391 .
METHANOCOCCUS VOLTAE MEDIUM
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Equipment needed:
Autoclave
Gassing station
Hungate tubes
Final pH:
7.2
Select a strain (optional):
ml
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
Mineral solution 2xW | 500.0 | ml | ||
|
|
Modified Wolin's mineral solution | 10.0 | ml | ||
|
|
LIP-solution | 50.0 | ml | ||
|
|
Na-acetate | 1.0 | g | ||
|
|
Sodium resazurin
(0.1% w/v) |
0.5 | ml | ||
|
|
TYC-solution | 50.0 | ml | ||
|
|
Na2CO3 | 1.5 | g | ||
|
|
L-Cysteine HCl x H2O | 0.5 | g | ||
|
|
Na2S x 9 H2O | 0.5 | g | ||
|
|
Distilled water | 400.0 | ml | ||
| 1 Dissolve ingredients except TYC solution, carbonate, cystein and sulfide. Sparge medium with 80% H2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic, then dispense under same gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. Before use add TYC solution, cysteine and sulfide from sterile anoxic stock solutions prepared under 100% N2 gas and carbonate from a sterile anoxic stock solution prepared under 80% N2 and 20% CO2 gas mixture. Adjust pH of complete medium to 7.2, if necessary. | |||||
Mineral solution 2xW #
1000
| Compound | Amount | Unit | |
|---|---|---|---|
|
|
NaCl | 40.00 | g |
|
|
MgCl2 x 6 H2O | 5.60 | g |
|
|
MgSO4 x 7 H2O | 0.70 | g |
|
|
KCl | 0.68 | g |
|
|
NH4Cl | 0.50 | g |
|
|
K2HPO4 | 0.28 | g |
|
|
CaCl2 x 2 H2O | 0.28 | g |
|
|
Distilled water | 1000.00 | ml |
LIP-solution #
1000
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
L-Leucine | 5.0 | g | ||
|
|
L-Isoleucine | 10.0 | g | ||
|
|
Pantothenate | 0.1 | g | ||
|
|
Distilled water | 1000.0 | ml | ||
| 1 May be stored frozen without sterilization. | |||||
TYC-solution #
1000
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
Casamino acids
(BD Bacto) |
100 | g | ||
|
|
Yeast extract
(OXOID) |
50 | g | ||
|
|
L-Tryptophan | 1 | g | ||
|
|
Distilled water | 1000 | ml | ||
| 1 Prepare and autoclave under 100% N2 gas. | |||||
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. | |||||