902 .
CARCBOXYDIBRACHIUM MEDIUM
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Equipment needed:
Autoclave
Filter
Gassing station
Hungate tubes
Final gas composition:
100 % N2
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Final pH:
7.0
Select a strain (optional):
ml
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
NaCl | 20.00 | g | ||
|
|
KCl | 0.70 | g | ||
|
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MgSO4 x 7 H2O | 3.90 | g | ||
|
|
CaCl2 x 2 H2O | 0.40 | g | ||
|
|
NH4Cl | 0.30 | g | ||
|
|
Na2HPO4 | 0.15 | g | ||
|
|
Na2SiO3 | 0.03 | g | ||
|
|
Yeast extract | 0.05 | g | ||
|
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Trace element solution SL-10 | 1.00 | ml | ||
|
|
Sodium resazurin
(0.1% w/v) |
0.50 | ml | ||
|
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NaHCO3 | 0.50 | g | ||
|
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Na-pyruvate | 2.50 | g | ||
|
|
Wolin's vitamin solution (10x) | 1.00 | ml | ||
|
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L-Cysteine HCl x H2O | 0.45 | g | ||
|
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Na2S x 9 H2O | 0.45 | g | ||
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Distilled water | 1000.00 | ml | ||
| 1 Dissolve ingredients except bicarbonate, pyruvate, vitamins and reducing agents, then adjust pH of solution to 7.0 with NaOH. Sparge medium with 100% N2 gas for 30 - 45 min to make it anoxic, then dispense medium under same gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. Add pyruvate, vitamins, cysteine and sulfide from sterile anoxic stock solutions prepared under 100% N2 gas and bicarbonate from a sterile anoxic stock solution prepared under 80% N2 and 20% CO2 gas mixture. Vitamins should be sterilized by filtration. Adjust pH of complete medium to 7.0, if necessary. | |||||
Trace element solution SL-10 #
1000
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
HCl
(25%) |
10.0 | ml | ||
|
|
FeCl2 x 4 H2O | 1.5 | g | ||
|
|
ZnCl2 | 70.0 | mg | ||
|
|
MnCl2 x 4 H2O | 100.0 | mg | ||
|
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H3BO3 | 6.0 | mg | ||
|
|
CoCl2 x 6 H2O | 190.0 | mg | ||
|
|
CuCl2 x 2 H2O | 2.0 | mg | ||
|
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NiCl2 x 6 H2O | 24.0 | mg | ||
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Na2MoO4 x 2 H2O | 36.0 | mg | ||
|
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Distilled water | 990.0 | ml | ||
| 1 First dissolve FeCl2 in the HCl, then dilute in water, add and dissolve the other salts. Finally 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 |
|
|
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 |