Main sol. 311a

Equipment needed: Autoclave Filter Hungate tubes Gassing station

Compound	Amount	Unit	Conc. [g/L]	Conc. [mM]
NH₄Cl	0.50	g	0.498	9.301
MgSO₄ x 7 H₂O	0.50	g	0.498	2.019
CaCl₂ x 2 H₂O	0.25	g	0.249	1.692
NaCl	2.25	g	2.239	38.31
FeSO₄ x 7 H₂O (0.1% w/v in 0.1 N H₂SO₄)	2.00	ml	0.002	0.007
Trace element solution SL-10	1.00	ml	-	-
Selenite-tungstate solution	1.00	ml	-	-
Yeast extract	2.00	g	1.99	-
Casitone	2.00	g	1.99	-
Sodium resazurin (0.1% w/v)	0.50	ml	5.0e-4	0.002
K₂HPO₄	0.35	g	0.348	1.999
KH₂PO₄	0.23	g	0.229	1.682
Na₂CO₃	1.00	g	0.995	9.388
Wolin's vitamin solution (10x)	1.00	ml	-	-
DL-Dithiothreitol	0.15	g	0.149	0.967
Distilled water	1000.00	ml	-	-
1 Dissolve ingredients (except phosphates, carbonate, vitamins, and dithiothreitol) and sparge medium with 80% H₂ and 20% CO₂ gas mixture for 30 - 45 min to make it anoxic. Dispense under the same gas atmosphere in anoxic Hungate-type tubes or serum vials to 30% of their volume and autoclave. Add phosphates to the medium after autoclaving from sterile stock solutions prepared under 100% N₂ gas and carbonate from a sterile stock solution prepared under 80% N₂ and 20% CO₂ gas mixture. Vitamins and dithiothreitol should be prepared under N₂ gas and sterilized by filtration. Adjust the pH of the completed medium to pH 7.0.

311a	SPOROMUSA MEDIUM (H2/CO2)