Compound | Amount | Unit | |||
---|---|---|---|---|---|
|
Sea Salt
(SIGMA) |
25.00 | g | ||
|
Na2SO4 | 3.50 | g | ||
|
NH4Cl | 0.25 | g | ||
|
KH2PO4 | 0.20 | g | ||
|
Wolfe's mineral elixir | 1.00 | ml | ||
|
Sodium resazurin
(0.1% w/v) |
0.50 | ml | ||
|
Na2CO3 | 1.50 | g | ||
|
Yeast extract | 0.05 | g | ||
|
Na-pyruvate | 1.00 | g | ||
|
Wolin's vitamin solution (10x) | 1.00 | ml | ||
|
n-Decane | 1.00 | ml | ||
|
Na2S x 9 H2O | 0.10 | g | ||
|
L-Cysteine HCl x H2O | 0.10 | g | ||
|
Distilled water | 1000.00 | ml | ||
1 Dissolve ingredients (except carbonate, yeast extract, pyruvate, vitamins, n-decane and reducing agents) and sparge medium with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Dispense medium under same gas atmosphere into anoxic serum vials (e.g., Balch-type tubes or serum bottles) and autoclave. Add yeast extract, pyruvate, vitamins, n-decane and reducing agents 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. Solutions of vitamins and pyruvate should be sterilized by filtration. Adjust pH of complete medium to 7.2 - 7.4. | |||||
2 Note: Use 20% (v/v) as inoculum and incubate vials in an inverted position. |
Compound | Amount | Unit | |||
---|---|---|---|---|---|
|
MgSO4 x 7 H2O | 30.00 | g | ||
|
MnSO4 x H2O | 5.00 | g | ||
|
NaCl | 10.00 | g | ||
|
FeSO4 x 7 H2O | 1.00 | g | ||
|
CoCl2 x 6 H2O | 1.80 | g | ||
|
CaCl2 x 2 H2O | 1.00 | g | ||
|
ZnSO4 x 7 H2O | 1.80 | g | ||
|
CuSO4 x 5 H2O | 0.10 | g | ||
|
AlK(SO4)2 x 12 H2O | 0.18 | g | ||
|
H3BO3 | 0.10 | g | ||
|
Na2MoO4 x 2 H2O | 0.10 | g | ||
|
(NH4)2Ni(SO4)2 x 6 H2O | 2.80 | g | ||
|
Na2WO4 x 2 H2O | 0.10 | g | ||
|
Na2SeO4 | 0.10 | g | ||
|
Distilled water | 1000.00 | ml | ||
1 First adjust pH to 1.0 with diluted H2SO4, then add and dissolve the salts. |
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 |