Main sol. 1526d

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
Na₂CO₃	1.0	g	0.989	9.332
Na-acetate	1.6	g	1.583	19.292
Na₂-fumarate	6.4	g	6.33	39.556
Seven vitamins solution	1.0	ml	-	-
Na₂S x 9 H₂O	0.5	g	0.495	2.059
Distilled water	1000.0	ml	-	-
1 Dissolve ingredients except carbonate, acetate, fumarate, vitamins, cysteine and sulfide, 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 and autoclave. Add acetate, fumarate, vitamins, cysteine and sulfide from sterile anoxic stock solutions prepared under 100% N₂ gas (fumarate and vitamins 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 6.8 - 7.0, if necessary.

LIMISALSIVIBRIO (L21 LS) MEDIUM