# 454: EBA MEDIUM


## Main sol. 454

0.2	g	KH2PO4	
1	g	NaCl	
0.5	g	KCl	
0.5	g	MgSO4 x 7 H2O	
0.15	g	CaCl2 x 2 H2O	
0.25	g	NH4Cl	
1	ml	**Trace element solution SL-10**	
1	mg	Resazurin	
1	ml	Na2SeO3 x 5 H2O (0.26 g/ 1000 ml 10 mM NaOH)	
2	g	Glycine	
950	ml	Distilled water	

1. pH 7.2 to 7.4
2. Before inoculation add per 100 ml of medium:   

0.2	ml	Biotin (10 mg/l, autoclave for 10 minutes under N2)	
3	ml	NaHCO3 (84 g/l, autoclaved under CO2)	
1	ml	Na2S (60 g/l, autoclaved under N2)	

3. Prepare and distribute medium anaerobically by using the Hungate technique.


## 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.