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Main sol. 854

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Internal Solution-ID: S1745

Equipment needed: Hungate tubes Filter Autoclave Gassing station
Compound Amount Unit Conc. [g/L] Conc. [mM]
NH4Cl 0.33 g 0.326 6.102
CaCl2 x 2 H2O 0.33 g 0.326 2.22
MgCl2 x 6 H2O 0.33 g 0.326 1.606
KCl 0.33 g 0.326 4.378
KH2PO4 0.33 g 0.326 2.399
NaCl 25.00 g 24.728 423.135
MOPS buffer
(SIGMA) 		3.00 g 2.967 14.18
Modified Wolin's mineral solution 10.00 ml - -
Yeast extract 0.10 g 0.099 -
Na-acetate 5.00 g 4.946 60.287
Sodium resazurin
(0.1% w/v) 		0.50 ml 4.9e-4 0.002
Sulfur
(powdered) 		10.00 g 9.891 308.425
Wolin's vitamin solution (10x) 1.00 ml - -
Na2S x 9 H2O 0.50 g 0.495 2.059
Distilled water 1000.00 ml - -
1 Dissolve ingredients (except sulfur, vitamins, and sulfide), adjust pH to 5.5 - 6.0, and sparge the medium with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Dispense the medium under the same gas atmosphere into anoxic Hungate-type tubes or serum vials that contain already an appropriate amount of sulfur powder. Autoclave 20 min at 110°C. Prior to use add vitamins and sulfide from sterile anoxic stock solutions prepared under 100% N2 gas. Vitamins are sterilized by filtration. Adjust the pH of the complete medium to 6.0 - 6.2, if necessary.
854 HIPPEA MEDIUM