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Main sol. 1529

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Internal Solution-ID: S3169

Equipment needed: Filter Hungate tubes Autoclave Gassing station
Compound Amount Unit Conc. [g/L] Conc. [mM]
Ferric citrate monohydrate 11.20 g 11.178 42.507
NaCl 19.00 g 18.962 324.471
MgCl2 x 6 H2O 9.00 g 8.982 44.181
MgSO4 x 7 H2O 0.15 g 0.15 0.607
CaCl2 x 2 H2O 0.30 g 0.299 2.037
KCl 0.10 g 0.1 1.339
KH2PO4 0.60 g 0.599 4.4
(NH4)2SO4 0.10 g 0.1 0.755
NaBr 0.05 g 0.05 0.485
SrCl2 0.02 g 0.02 0.126
Trace element solution SL-10 1.00 ml - -
NaHCO3 2.50 g 2.495 29.7
Yeast extract 0.10 g 0.1 -
Na-pyruvate 1.00 g 0.998 9.069
FeCl2 x 4 H2O 0.20 g 0.2 1.004
Wolin's vitamin solution (10x) 1.00 ml - -
L-Cysteine HCl x H2O 0.10 g 0.1 0.568
Distilled water 1000.00 ml - -
1 First, dissolve ferric citrate in water by heating, adjust to pH 5.8, then cool to room temperature. Add other compounds (except bicarbonate, yeast extract, pyruvate, vitamins, cysteine, and ferrous chloride) and sparge medium with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Then add cysteine and ferrous chloride and adjust pH to 6.8 - 7.0 with bicarbonate. Dispense medium under 80% N2 and 20% CO2 gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. Before inoculation add yeast extract, pyruvate, and vitamins, from sterile anoxic stock solutions prepared under 100% N2 gas. Stock solutions of pyruvate and vitamins should be sterilized by filtration. Adjust the pH of the complete medium to 6.8 - 7.0.
2 Note: Use 5 - 10% (v/v) inoculum. The color of the medium supernatant changes from orange-brown to yellow-greenish during growth.
1529 GEOGLOBUS AHANGARI MEDIUM