# 358a: ACIDIANUS MEDIUM (ANAEROBIC)


## Main sol. 358a

1.3	g	(NH4)2SO4	
0.28	g	KH2PO4	
0.25	g	MgSO4 x 7 H2O	
0.07	g	CaCl2 x 2 H2O	
0.02	g	FeCl3 x 6 H2O	
10	ml	**Allen's trace element solution**	
5	g	Sulfur (powder)	
0.5	g	Yeast extract (OXOID)	
1000	ml	Distilled water	

1. Dissolve ingredients, except sulfur and yeast extract, bring medium to the boil, then cool to room temperature under 80% H2 and 20% CO2 gas mixture and adjust pH to 2.5 using 10 N H2SO4. Dispense medium under same gas atmosphere into anoxic Hungate-type tubes or serum vials (e.g., 20 ml medium in 100 ml serum bottles) containing already the appropriate amount of sulfur. For sterilization sealed bottles with medium are heated in a boiling water bath for 2 - 3 h on each of 3 successive days. Add yeast extract from a sterile anoxic stock solution prepared under 100% N2 gas atmosphere.
2. Pressurize inoculated bottles to 1 bar overpressure with sterile 80% H2 and 20% CO2 gas mixture.
3. Note: Inoculate with 5% (w/v) culture. Incubate without shaking.


## Allen's trace element solution

180	mg	MnCl2 x 4 H2O	
450	mg	Na2B4O7 x 10 H2O	
22	mg	ZnSO4 x 7 H2O	
5	mg	CuCl2 x 2 H2O	
3	mg	Na2MoO4 x 2 H2O	
3	mg	VOSO4 x 2 H2O	
1	mg	CoSO4 x 7 H2O	
1000	ml	Distilled water	

1. Adjust pH of final solution to 2 with 1 N HCl.