1057 .
ETHANOLIGENES MEDIUM
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Equipment needed:
Autoclave
Filter
Gassing station
Hungate tubes
Final gas composition:
100 % N2
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Final pH:
6.0
Select a strain (optional):
ml
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
Tryptone
(BD BBL) |
4.0 | g | ||
|
|
Beef extract
(BD BBL) |
2.0 | g | ||
|
|
Yeast extract
(OXOID) |
1.0 | g | ||
|
|
NaCl | 4.0 | g | ||
|
|
K2HPO4 | 1.5 | g | ||
|
|
FeSO4 x 7 H2O | 0.1 | g | ||
|
|
MgCl2 x 6 H2O | 0.2 | g | ||
|
|
Trace element solution | 1.0 | ml | ||
|
|
Sodium resazurin
(0.1% w/v) |
0.5 | ml | ||
|
|
L-Cysteine HCl x H2O | 0.5 | g | ||
|
|
D-Glucose | 10.0 | g | ||
|
|
Wolin's vitamin solution (10x) | 1.0 | ml | ||
|
|
Distilled water | 1000.0 | ml | ||
| 1 Dissolve ingredients (except cysteine, glucose and vitamins), sparge medium with 100% N2 gas for 30 - 45 min to make it anoxic. Add cysteine and adjust pH to 6.0, then dispense medium under 100% N2 gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. Prior to inoculation add glucose and vitamins from sterile anoxic stock solutions prepared under 100% N2 gas. The vitamin solution should be sterilized by filtration. Adjust pH of complete medium to 6.0, if necessary. | |||||
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 | ||
|
|
CoCl2 x 6 H2O | 0.03 | g | ||
|
|
CaCl2 x 2 H2O | 0.10 | g | ||
|
|
ZnCl2 | 0.10 | g | ||
|
|
CuCl2 | 0.02 | g | ||
|
|
H3BO3 | 0.01 | g | ||
|
|
Na2MoO4 x 2 H2O | 0.03 | g | ||
|
|
NiCl2 x 6 H2O | 0.10 | g | ||
|
|
NaCl | 1.00 | g | ||
|
|
Na2SeO3 x 5 H2O | 0.03 | g | ||
|
|
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 |
|
|
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 |