# J470: VULCANIBACILLUS MEDIUM


## Main sol. J470

30	g	Sea Salt (Sigma)	
1	g	NaNO3	
2	ml	**Wolfe's mineral elixir**	
0.05	mg	VOSO4 x n H2O	
1	g	Sodium pyruvate	
0.5	mg	Resazurin	
0.1	g	FeSO4 x 7 H2O	
1	g	NaHCO3	
0.5	g	Yeast extract	
20	ml	**Trace vitamins**	
1000	ml	Distilled water	

1. Dissolve ingredients except FeSO4 x 7H2O, NaHCO3, yeast extract and vitamins, and adjust pH to 7.0. Bring to a boil for 3 min, then cool to room temperature under a N2 gas stream. Dispense the medium under the same gas atmosphere into culture vessels, seal with butyl rubber stoppers and autoclave. Add sterile anaerobic stock solutions of FeSO4 x 7H2O (1%, in 0.1 N H2SO4), NaHCO3 (8%, under a N2 x CO2 gas mixture), yeast extract (10%) and vitamins (filter-sterilized) to the medium. Adjust final pH of the medium to 7.0 - 7.2. Add 10 - 20 mg sodium dithionite per liter [e.g., from 5% (w/v) solution, freshly prepared under N2 and filter-sterilized], if the medium is not completely reduced after inoculation.


## Wolfe's mineral elixir

30	g	MgSO4 x 7 H2O	
5	g	MnSO4 x n H2O	
10	g	NaCl	
1	g	FeSO4 x 7 H2O	
1.8	g	CoCl2 x 6 H2O	
1	g	CaCl2 x 2 H2O	
1.8	g	ZnSO4 x 7 H2O	
0.1	g	CuSO4 x 5 H2O	
0.18	g	AlK(SO4)2 x 12 H2O	
0.1	g	H3BO3	
0.1	g	Na2MoO4 x 2 H2O	
2.8	g	(NH4)2Ni(SO4)2 x 6 H2O	
0.1	g	Na2WO4 x 2 H2O	
0.1	g	Na2SeO4	
1000	ml	Distilled water	

1. First adjust pH to 1.0 with diluted H2SO4, then dissolve the salts.


## Trace vitamins

2	mg	Biotin	
2	mg	Folic acid	
10	mg	Pyridoxine hydrochloride	
5	mg	Thiamine HCl	
5	mg	Riboflavin	
5	mg	Nicotinic acid	
5	mg	Calcium pantothenate	
0.1	mg	Vitamin B12	
5	mg	p-Aminobenzoic acid	
5	mg	Lipoic acid	
1000	ml	Distilled water