1005 .
THERMODESULFOBIUM MEDIUM
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Equipment needed:
Autoclave
Filter
Gassing station
Serum bottle
Final pH:
5.5 - 6.0
Select a strain (optional):
ml
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
K2HPO4 | 0.78 | g | ||
|
|
KH2PO4 | 0.75 | g | ||
|
|
Na2-EDTA | 0.04 | g | ||
|
|
FeSO4 x 7 H2O | 0.01 | g | ||
|
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MgSO4 x 7 H2O | 0.25 | g | ||
|
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CaCl2 x 2 H2O | 0.03 | g | ||
|
|
Trace element solution | 10.00 | ml | ||
|
|
NaCl | 0.20 | g | ||
|
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NH4Cl | 0.50 | g | ||
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Na2SO4 | 2.80 | g | ||
|
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Na-acetate | 0.15 | g | ||
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Wolin's vitamin solution (10x) | 1.00 | ml | ||
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L-Cysteine HCl x H2O | 0.25 | g | ||
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Distilled water | 1000.00 | ml | ||
| 1 Dissolve ingredients (except vitamins and cysteine), adjust pH to 5.5 with 10 N H2SO4 and sparge medium with 80% H2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Dispense medium under same gas atmosphere into anoxic Hungate-type tubes or serum vials (e.g., 20 ml of medium in 50 ml bottles) and autoclave. Prior to inoculation add vitamins (sterilized by filtration) and cysteine from sterile anoxic stock solutions prepared under 100% N2 gas. Adjust pH of complete medium to 5.5 - 6.0, if necessary. | |||||
| 2 After inoculation pressurize vials to 0.5 bar overpressure with sterile 80% H2 and 20% CO2 gas mixture. | |||||
Trace element solution #
1000
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
Nitrilotriacetic acid
(NTA) |
12.80 | g | ||
|
|
FeCl2 x 4 H2O | 1.00 | g | ||
|
|
MnCl2 x 4 H2O | 0.10 | g | ||
|
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CoCl2 x 6 H2O | 0.03 | g | ||
|
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CaCl2 x 2 H2O | 0.10 | g | ||
|
|
ZnCl2 | 0.10 | g | ||
|
|
CuCl2 | 0.02 | g | ||
|
|
H3BO3 | 0.01 | g | ||
|
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Na2MoO4 x 2 H2O | 0.03 | g | ||
|
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NiCl2 x 6 H2O | 0.10 | g | ||
|
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NaCl | 1.00 | g | ||
|
|
Na2SeO3 x 5 H2O | 0.03 | g | ||
|
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Na2WO4 x 2 H2O | 0.04 | g | ||
|
|
Distilled water | 1000.00 | ml | ||
| 1 First dissolve NTA in 200 ml of distilled water and adjust pH to 6.5 with KOH, then dissolve mineral salts. Finally adjust pH to 6.5 with KOH and make up to 1000.00 ml. | |||||
Wolin's vitamin solution (10x) #
1000
| Compound | Amount | Unit | |
|---|---|---|---|
|
|
Biotin | 20 | mg |
|
|
Folic acid | 20 | mg |
|
|
Pyridoxine hydrochloride | 100 | mg |
|
|
Thiamine HCl | 50 | mg |
|
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Riboflavin | 50 | mg |
|
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Nicotinic acid | 50 | mg |
|
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Calcium D-(+)-pantothenate | 50 | mg |
|
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Vitamin B12 | 1 | mg |
|
|
p-Aminobenzoic acid | 50 | mg |
|
|
(DL)-alpha-Lipoic acid | 50 | mg |
|
|
Distilled water | 1000 | ml |