Compound | Amount | Unit | |||
---|---|---|---|---|---|
|
NaCl | 19.000 | g | ||
|
MgCl2 x 6 H2O | 9.000 | g | ||
|
MgSO4 x 7 H2O | 0.150 | g | ||
|
CaCl2 x 2 H2O | 0.300 | g | ||
|
KCl | 0.500 | g | ||
|
KH2PO4 | 0.420 | g | ||
|
(NH4)2SO4 | 0.100 | g | ||
|
NaBr | 0.050 | g | ||
|
SrCl2 x 6 H2O | 0.020 | g | ||
|
Yeast extract
(BD-Difco) |
0.100 | g | ||
|
NaHCO3 | 2.500 | g | ||
|
Na2SeO4 | 0.020 | g | ||
|
Poorly crystalline iron(III) oxide | 100.000 | mM | ||
|
Trace vitamins | 10.000 | ml | ||
|
Trace mineral solution | 10.000 | ml | ||
|
L-Cysteine HCl x H2O | 0.044 | g | ||
|
FeCl2 x 4 H2O | 0.260 | g | ||
1 Add components except L-cysteine x HCl x H2O and FeCl2 x 4H2O to distilled water, bring volume to 1.0 L and adjust pH to 7.0. Distribute the medium into appropriate culture vessels (e.g., 20 ml of the medium in 120 ml serum bottles) under a H2-CO2 (4:1, v/v) gas mixture, sparge with the same gas mixture for 10 - 15 min, seal with butyl rubber stoppers and autoclave. Separately autoclave concentrated (100- or 200-fold) L-cysteine x HCl x H2O and FeCl2 x 4H2O solutions under a N2 gas atmosphere. Prior to inoculation, add the sterile L-cysteine x HCl x H2O and FeCl2 x 4H2O solutions. Pressurize the inoculated vessels to 100 kPa H2-CO2 (4:1, v/v) and cultivate without shaking. |
1 Dissolve 108 g FeCl3 in 1.0 L distilled water. Adjust pH to 7.0 with 10 N NaOH. Allow the suspension to sit for 30 min, then readjust pH to 7.0. Centrifuge the suspension at 5,000 x g for 20 min and pour off supernatant. Wash the precipitate with distilled water by resuspending in distilled water followed by the centrifugation. Repeat the washing at least 5 times, then, resuspend in 1.0 L distilled water. Use the Fe(III) suspension as 0.67 M solution. More precisely, check the concentration of Cl- to be less than 1 mM and determine the concentration of Fe(III) by using hydroxylamine as described by Lovley and Phillips (Appl. Environ. Microbiol. 54, 751-757, 1986). |
1 Use Trace minerals of Medium No. 151 supplemented with 0.025 g (final) Na2MoO4 x 2H2O, 0.024 g NiCl2 x 6H2O and 0.025 g Na2WO4 x 2H2O (per liter). | |||||
2 Comment: For strain JCM 12378, inoculate with a 10% inoculum. |
Compound | Amount | Unit | |
---|---|---|---|
|
Biotin | 2.0 | mg |
|
Folic acid | 2.0 | mg |
|
Pyridoxine hydrochloride | 10.0 | mg |
|
Thiamine HCl | 5.0 | mg |
|
Riboflavin | 5.0 | mg |
|
Nicotinic acid | 5.0 | mg |
|
Calcium pantothenate | 5.0 | mg |
|
Vitamin B12 | 0.1 | mg |
|
p-Aminobenzoic acid | 5.0 | mg |
|
Lipoic acid | 5.0 | mg |
|
Distilled water | 1000.0 | ml |