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Main sol. 1526e

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Internal Solution-ID: S3163

Equipment needed: Autoclave Hungate tubes Filter Gassing station
Compound Amount Unit Conc. [g/L] Conc. [mM]
NaCl 60.0 g 59.347 1015.52
MgCl2 x 6 H2O 6.0 g 5.935 29.192
KCl 1.5 g 1.484 19.902
Na2SO4 1.0 g 0.989 6.964
NH4Cl 1.0 g 0.989 18.492
CaCl2 x 2 H2O 0.4 g 0.396 2.691
K2HPO4 0.4 g 0.396 2.272
Modified Wolin's mineral solution 10.0 ml - -
Sodium resazurin
(0.1% w/v) 		0.5 ml 4.9e-4 0.002
Sulfur
(powder) 		10.0 g 9.891 308.425
Na2CO3 1.5 g 1.484 13.999
D-Glucose 1.0 g 0.989 5.49
Yeast extract 1.0 g 0.989 -
Wolin's vitamin solution (10x) 1.0 ml - -
L-Cysteine HCl x H2O 0.3 g 0.297 1.689
DL-Dithiothreitol
(DTT) 		0.3 g 0.297 1.923
Distilled water 1000.0 ml - -
1 Dissolve ingredients except sulfur, carbonate, glucose, yeast extract, vitamins, cysteine and DTT, then sparge medium with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Dispense medium under the same gas atmosphere into anoxic Hungate-type tubes or serum vials containing already the appropriate amount of sulfur and autoclave at 110°C for 20 min. Add glucose, yeast extract, vitamins, cysteine and DTT from sterile anoxic stock solutions prepared under 100% N2 gas (vitamins and DTT are sterilized by filtration) and carbonate from a sterile anoxic stock solution prepared under 80% N2 and 20% CO2 gas mixture. Prior to use adjust pH of complete medium to 7.3 - 7.5, if necessary.
1526e LIMIHALOGLOBUS MEDIUM