# 88c: ZESTOSPHAERA MEDIUM


## Main sol. 88c

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**	
2	g	Na2S2O3 x 5 H2O	
1	g	Yeast extract (OXOID)	
1	g	Tryptone peptone (Bacto BD)	
0.5	g	Casamino acids (Difco BD)	
0.5	g	Na2S x 9 H2O	
1000	ml	Distilled water	

1. Dissolve ingredients (except thiosulfate, yeast extract, Tryptone, Casamino acids and sulfide), sparge medium with 80% N2 and 20% CO2 gas mixture to make it anoxic and dispense medium under same gas atmosphere into serum bottels. After autoclaving add thiosulfate, yeast extract, Tryptone, Casamino acids and sulfide from sterile anoxic stock solutions prepared under 100% N2 gas. Prior to inoculation check pH and adjust to 6.0, if necessary.


## 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.