# 579: GEOBACTER MEDIUM


## Main sol. 579

13.7	g	Ferric citrate monohydrate	
1.5	g	NH4Cl	
0.6	g	NaH2PO4	
0.1	g	KCl	
2.5	g	Na-acetate	
10	ml	**Modified Wolin's mineral solution**	
2.5	g	NaHCO3	
1	ml	**Wolin's vitamin solution (10x)**	
1000	ml	Distilled water	

1. First dissolve ferric citrate in water by heating and adjust to pH 6.0, then add other medium ingredients except bicarbonate and vitamins. Sparge medium with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic, then add and dissolve bicarbonate and adjust pH to 6.8. Thereafter, dispense medium under 80% N2 and 20% CO2 gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. Add vitamins to the sterile medium from an anoxic stock solution prepared under 100% N2 gas and sterilized by filtration. Adjust pH of complete medium to 6.7 - 7.0, if necessary.
2. Note: Use 5 - 10% (v/v) inoculum.


## 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.


## Wolin's vitamin solution (10x)

20	mg	Biotin	
20	mg	Folic acid	
100	mg	Pyridoxine hydrochloride	
50	mg	Thiamine HCl	
50	mg	Riboflavin	
50	mg	Nicotinic acid	
50	mg	Calcium D-(+)-pantothenate	
1	mg	Vitamin B12	
50	mg	p-Aminobenzoic acid	
50	mg	(DL)-alpha-Lipoic acid	
1000	ml	Distilled water