# 1599: CONTUBERNALIS MEDIUM


## Main sol. 1599

0.5	g	NH4Cl	
0.2	g	KH2PO4	
0.1	g	MgCl2 x 6 H2O	
0.2	g	KCl	
4.2	g	NaCl	
3	g	Na2SO4	
0.2	g	Yeast extract	
1	ml	**Trace element solution SL-10**	
1	ml	**Selenite-tungstate solution**	
0.5	ml	Sodium resazurin (0.1% w/v)	
3.3	g	NaHCO3	
6.5	g	NaCO3	
3	g	Na-acetate	
0.5	g	Na2S x 9 H2O	
1000	ml	Distilled water	

1. Dissolve ingredients (except carbonates, acetate and sulfide) and sparge medium with 100% N2 gas for 30 - 45 min to make it anoxic. Then, add and dissolve bicarbonate and carbonate, adjust pH to 9.5, dispense under 100% N2 gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. After sterilization add acetate and sulfide from sterile anoxic stock solutions autoclaved under 100% N2 gas atmosphere. Adjust pH of complete medium to 9.5 - 9.7, if necessary.


## Trace element solution SL-10

10	ml	HCl (25%)	
1.5	g	FeCl2 x 4 H2O	
70	mg	ZnCl2	
100	mg	MnCl2 x 4 H2O	
6	mg	H3BO3	
190	mg	CoCl2 x 6 H2O	
2	mg	CuCl2 x 2 H2O	
24	mg	NiCl2 x 6 H2O	
36	mg	Na2MoO4 x 2 H2O	
990	ml	Distilled water	

1. First dissolve FeCl2 in the HCl, then dilute in water, add and dissolve the other salts. Finally make up to 1000.00 ml.


## Selenite-tungstate solution

0.5	g	NaOH	
3	mg	Na2SeO3 x 5 H2O	
4	mg	Na2WO4 x 2 H2O	
1000	ml	Distilled water