# 1043: IGNISPHAERA MEDIUM


## Main sol. 1043

1.3	g	(NH4)2SO4	
74	mg	CaCl2 x 2 H2O	
0.28	g	MgSO4 x 7 H2O	
0.28	g	KH2PO4	
0.1	g	Yeast extract (OXOID)	
2	g	Trypticase peptone (BD BBL)	
10	ml	**Modified Wolin's mineral solution**	
0.5	ml	Sodium resazurin (0.1% w/v)	
0.5	ml	FeCl3 x 6 H2O (0.1% w/v in 0.2 N HCl)	
0.3	g	L-Cysteine HCl x H2O	
2	g	Starch (soluble)	
0.3	g	Na2S x 9 H2O	
1000	ml	Distilled water	

1. Dissolve ingredients (except ferric chloride, cysteine, starch and sulfide), sparge medium with 100% N2 gas for 30 - 45 min to make it anoxic. Add ferric chloride and solid cysteine, then adjust pH to 6.3. Dispense medium under 100% N2 gas atmosphere into anoxic Hungate-type tubes or serum vials (e.g., 20 ml medium in 50 ml bottles) and autoclave. Prior to inoculation add starch and sulfide from sterile anoxic stock solutions prepared under 100% N2 gas. Adjust pH of complete medium to 6.5, if necessary.


## Modified Wolin's mineral solution

1.5	g	Nitrilotriacetic acid	
3	g	MgSO4 x 7 H2O	
0.5	g	MnSO4 x H2O	
1	g	NaCl	
0.1	g	FeSO4 x 7 H2O	
0.18	g	CoSO4 x 7 H2O	
0.1	g	CaCl2 x 2 H2O	
0.18	g	ZnSO4 x 7 H2O	
0.01	g	CuSO4 x 5 H2O	
0.02	g	AlK(SO4)2 x 12 H2O	
0.01	g	H3BO3	
0.01	g	Na2MoO4 x 2 H2O	
0.03	g	NiCl2 x 6 H2O	
0.3	mg	Na2SeO3 x 5 H2O	
0.4	mg	Na2WO4 x 2 H2O	
1000	ml	Distilled water	

1. First dissolve nitrilotriacetic acid and adjust pH to 6.5 with KOH, then add minerals. Adjust final to pH 7.0 with KOH.