Compound | Amount | Unit | |||
---|---|---|---|---|---|
|
Tryptone
(BD Bacto) |
1.00 | g | ||
|
Peptone
(meat) |
1.00 | g | ||
|
Yeast extract
(BD Bacto) |
1.00 | g | ||
|
K2HPO4 | 1.60 | g | ||
|
NaH2PO4 x 2 H2O | 1.00 | g | ||
|
NH4Cl | 0.50 | g | ||
|
MgSO4 x 6 H2O | 0.16 | g | ||
|
Trace element solution SL-11 | 1.00 | ml | ||
|
Sodium resazurin
(0.1% w/v) |
0.50 | ml | ||
|
CaCl2 x 2 H2O | 0.06 | g | ||
|
NaHCO3 | 1.00 | g | ||
|
D-Glucose | 5.00 | g | ||
|
Wolin's vitamin solution (10x) | 1.00 | ml | ||
|
L-Cysteine HCl x H2O | 0.30 | g | ||
|
Na2S x 9 H2O | 0.30 | g | ||
|
Distilled water | 1000.00 | ml | ||
1 Dissolve ingredients (except calcium chloride, bicarbonate, glucose, vitamins, cysteine and sulfide), adjust pH to 7.0 and sparge medium with 100% N2 gas for 30 - 45 min to make it anoxic. Dispense medium under same gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. After sterilization add calcium chloride, glucose, cysteine and sulfide from sterile anoxic stock solutions autoclaved under 100% N2 gas atmosphere. Add bicarbonate from a sterile anoxic stock solution prepared under 80% N2 and 20% CO2 gas mixture. Vitamins are prepared under 100% N2 gas and sterilized by filtration. The pH of the complete medium should be at 7.0. | |||||
2 Note: Supplementation of medium with 10.00 g/l MOPS buffer (pH 6.9 - 7.0; 10% (w/v) anoxic stock solution) may enhance the buffer capacity of the medium. |
Compound | Amount | Unit | |||
---|---|---|---|---|---|
|
Na2-EDTA x 2 H2O | 5.2 | g | ||
|
FeCl2 x 4 H2O | 1.5 | g | ||
|
ZnCl2 | 70.0 | mg | ||
|
MnCl2 x 4 H2O | 100.0 | mg | ||
|
H3BO3 | 6.0 | mg | ||
|
CoCl2 x 6 H2O | 190.0 | mg | ||
|
CuCl2 x 2 H2O | 2.0 | mg | ||
|
NiCl2 x 6 H2O | 24.0 | mg | ||
|
Na2MoO4 x 2 H2O | 36.0 | mg | ||
|
Distilled water | 1000.0 | ml | ||
1 Dissolve EDTA in 800 ml distilled water, adjust pH to 7 using 2 N NaOH and add ferrous chloride. After ferrous chloride has dissolved add other compounds. Finally adjust pH to 6.0 and bring volume to 1000 ml. |
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 |