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Main sol. 1665

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Internal Solution-ID: S3456

Equipment needed: Autoclave Hungate tubes Filter Gassing station
Compound Amount Unit Conc. [g/L] Conc. [mM]
NaCl 20.0 g 19.704 337.174
MgCl2 x 6 H2O 6.0 g 5.911 29.077
Na2SO4 3.0 g 2.956 20.809
CaCl2 x 2 H2O 0.2 g 0.197 1.34
KCl 0.5 g 0.493 6.608
NH4Cl 0.3 g 0.296 5.526
Wolfe's mineral elixir 2.0 ml - -
HEPES
(SIGMA) 		2.4 g 2.365 9.923
Na2-fumarate 1.0 g 0.985 6.156
Yeast extract
(OXOID) 		0.2 g 0.197 -
Sodium resazurin
(0.1% w/v) 		0.5 ml 4.9e-4 0.002
K-phosphate buffer
(0.5 M, pH 7.0) 		2.0 ml - 1
NaHCO3 1.0 g 0.985 11.728
FeCl2 x 4 H2O
(0.2% w/v in 0.02 N HCl) 		10.0 ml 0.02 0.099
Wolin's vitamin solution (10x) 1.0 ml - -
L-Cysteine HCl x H2O 0.5 g 0.493 2.805
Distilled water 1000.0 ml - -
1 Dissolve ingredients (except phosphate buffer, bicarbonate, ferrous chloride, vitamins and cysteine), adjust pH to 7.2 with NaOH and sparge medium with 100% N2 gas for 30 - 45 min to make it anoxic. Dispense medium under the same gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. After autoclaving add phosphate buffer, ferrous chloride, vitamins and cysteine from sterile anoxic stock solutions prepared under 100% N2 gas atmosphere and bicarbonate from a sterile stock solution prepared under 80% N2 and 20% CO2 gas mixture. Vitamins should be sterilized by filtration. Adjust pH of the complete medium to 7.2 - 7.5 , if necessary.
1665 DESULFAMPLUS MEDIUM