1104 .
DETHIOBACTER MEDIUM
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Equipment needed:
Filter
Gassing station
Hungate tubes
Water bath
Final pH:
9.5
Select a strain (optional):
ml
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
NaCl | 6.00 | g | ||
|
|
K2HPO4 | 0.50 | g | ||
|
|
NaHCO3 | 8.00 | g | ||
|
|
Na2CO3 | 22.00 | g | ||
|
|
Sulfur
(powdered) |
1.50 | g | ||
|
|
NH4Cl | 0.20 | g | ||
|
|
MgCl2 x 6 H2O | 0.20 | g | ||
|
|
Trace elements solution (Pfennig & Lippert,1966) | 1.00 | ml | ||
|
|
Selenite-tungstate solution | 1.00 | ml | ||
|
|
Yeast extract | 0.05 | g | ||
|
|
Ethanol absolute | 0.60 | ml | ||
|
|
Na-acetate | 0.40 | g | ||
|
|
Na2S2O3 x 5 H2O | 2.50 | g | ||
|
|
Wolin's vitamin solution (10x) | 1.00 | ml | ||
|
|
Na2S x 9 H2O | 0.24 | g | ||
|
|
Distilled water | 1000.00 | ml | ||
| 1 Dissolve sodium chloride and hydrogenphosphate, then sparge solution with 100% N2 gas for 30 -45 min to make it anoxic. Add and dissolve carbonates, then dispense under 100% N2 gas atmosphere into anoxic Hungate-type tubes or serum vials containing already the appropriate amount of sulfur, only to 30% of their volume to allow for a large headspace. Sterilize medium by heating cultivation vessels in a water bath to 90 - 100°C for 1- 2 hours on each of 3 successive days. After sterilization add ammonium chloride, magnesium chloride, trace elements, yeast extract, ethanol, acetate, thiosulfate, vitamins and sulfide from sterile anoxic stock solutions prepared under 100% N2 gas. The stock solutions of thiosulfate and vitamins should be sterilized by filtration. Adjust pH of the complete medium to 9.5. | |||||
| 2 After inoculation add sterile 100% H2 gas to 0.5 bar overpressure. | |||||
Trace elements solution (Pfennig & Lippert,1966) #
1000
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
EDTA | 5.00 | g | ||
|
|
FeSO4 x 7 H2O | 2.20 | g | ||
|
|
ZnSO4 x 7 H2O | 0.10 | g | ||
|
|
MnCl2 x 4 H2O | 0.03 | g | ||
|
|
H3BO3 | 0.03 | g | ||
|
|
CoCl2 x 6 H2O | 0.20 | g | ||
|
|
CuCl2 x 2 H2O | 0.03 | g | ||
|
|
NiCl2 x 6 H2O | 0.03 | g | ||
|
|
Na2MoO4 x 2 H2O | 0.03 | g | ||
|
|
Distilled water | 1000.00 | ml | ||
| 1 pH 3.0-4.0 | |||||
Selenite-tungstate solution #
1000
| Compound | Amount | Unit | |
|---|---|---|---|
|
|
NaOH | 0.5 | g |
|
|
Na2SeO3 x 5 H2O | 3.0 | mg |
|
|
Na2WO4 x 2 H2O | 4.0 | mg |
|
|
Distilled water | 1000.0 | 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 |
|
|
Riboflavin | 50 | mg |
|
|
Nicotinic acid | 50 | mg |
|
|
Calcium D-(+)-pantothenate | 50 | mg |
|
|
Vitamin B12 | 1 | mg |
|
|
p-Aminobenzoic acid | 50 | mg |
|
|
(DL)-alpha-Lipoic acid | 50 | mg |
|
|
Distilled water | 1000 | ml |