This strain uses the standard medium without modifications.
Compound | Amount | Unit | |||
---|---|---|---|---|---|
|
NaCl | 1.00 | g | ||
|
KCl | 0.50 | g | ||
|
MgCl2 x 6 H2O | 0.40 | g | ||
|
CaCl2 x 2 H2O | 0.10 | g | ||
|
NH4Cl | 0.30 | g | ||
|
KH2PO4 | 0.20 | g | ||
|
Na2SO4 | 0.15 | g | ||
|
Trace element solution SL-10 | 1.00 | ml | ||
|
Selenite-tungstate solution | 1.00 | ml | ||
|
Trypticase peptone
(BD BBL) |
1.00 | g | ||
|
Sodium resazurin
(0.1% w/v) |
0.50 | ml | ||
|
Na2CO3 | 1.50 | g | ||
|
Sugar mix | 30.00 | ml | ||
|
Yeast autolysate | 50.00 | ml | ||
|
Seven vitamins solution | 1.00 | ml | ||
|
Co-factors solution | 20.00 | ml | ||
|
DL-Dithiothreitol
(DTT) |
0.40 | g | ||
|
Distilled water | 900.00 | ml | ||
1 Dissolve ingredients (except carbonate, sugar mix, yeast autolysate, vitamins, co-factors, and DTT), sparge medium with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Distribute under same gas atmosphere into anoxic Hungate-type tubes or serum vials to 30% of their volume (e.g., 4.5 ml per Hungate-type tube) and autoclave. After sterilization add carbonate from a sterile anoxic stock solution prepared under 80% N2 and 20% CO2 gas atmosphere and sugar mix, yeast autolysate, vitamins, co-factors and DTT from sterile anoxic stock solutions prepared under 100% N2 gas atmosphere and sterilized by filtration. Check pH of complete medium and adjust to 7.2 - 7.4, if necessary. | |||||
2 Note: Basal medium and yeast autolysate have to be prepared freshly and cannot be stored for longer periods. |
Compound | Amount | Unit | |
---|---|---|---|
|
Cellobiose | 56.7 | g |
|
Sucrose | 56.7 | g |
|
Maltose x H2O | 60.0 | g |
|
Trehalose | 63.3 | g |
|
D-Xylose | 25.0 | g |
|
Distilled water | 1000.0 | ml |
Compound | Amount | Unit | |||
---|---|---|---|---|---|
|
Yeast extract
(dry baker's yeast, SIGMA YSC-1) |
280 | g | ||
|
Distilled water | 1000 | ml | ||
1 Suspend dry baker's yeast (SIGMA YSC-1) in distilled water and allow cells to autolyse for 48 h at 55°C. Thereafter, cell debris are removed by centrifugation (3.700 x g for 30 min) and the amber-colored supernatant is stored in aliquots at -20°C. |
Compound | Amount | Unit | |||
---|---|---|---|---|---|
|
Thiamine pyrophosphate | 2.50 | g | ||
|
Pyridoxal hydrochloride | 0.25 | g | ||
|
Pyridoxal phosphate | 0.25 | g | ||
|
Ca-folinate | 0.05 | g | ||
|
ß-NAD | 0.05 | g | ||
|
Coenzyme A | 0.05 | g | ||
|
FAD | 0.05 | g | ||
|
Haemin
(dissolved separately in 10 mM NaOH) |
65.00 | mg | ||
|
Nicotinamide | 25.00 | mg | ||
|
Folic acid | 2.50 | mg | ||
|
Riboflavin | 0.50 | mg | ||
|
Distilled water | 1000.00 | ml | ||
1 Adjust pH to 7.0 with 1 N NaOH against indicator paper, filter sterilize and store under 100% N2 gas atmosphere. |
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 | ||
|
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 | 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. |
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 |
Compound | Amount | Unit | |
---|---|---|---|
|
Vitamin B12 | 100 | mg |
|
p-Aminobenzoic acid | 80 | mg |
|
D-(+)-biotin | 20 | mg |
|
Nicotinic acid | 200 | mg |
|
Calcium pantothenate | 100 | mg |
|
Pyridoxine hydrochloride | 300 | mg |
|
Thiamine-HCl x 2 H2O | 200 | mg |
|
Distilled water | 1000 | ml |