# 1304: SULFURITALEA MEDIUM


## Main sol. 1304

0.2	g	MgCl2 x 6 H2O	
0.1	g	CaCl2 x 2 H2O	
1.7	g	NaNO3	
0.1	g	(NH4)2SO4	
0.1	g	KH2PO4	
0.1	g	KCl	
1	ml	**Trace element solution SL-10**	
1	ml	**Selenite-tungstate solution**	
0.4	g	Na2S2O3 x 5 H2O	
5	g	NaHCO3	
1	ml	**Wolin's vitamin solution**	
1000	ml	Distilled water	

1. Dissolve ingredients except trace elements, thiosulfate, bicarbonate and vitamins, then sparge medium with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to remove dissolved oxygen and to saturate the solution with CO2. Dispense solution under 80% N2 and 20% CO2 gas atmosphere in vials suitable for anaerobic cultures (e.g., Balch-type tubes) to 30% of volume. Close vials with butyl rubber septa and autoclave. Add trace element solution, selenite-tungstate solution and vitamins from sterile anoxic stock solutions prepared under 100% N2 gas and bicarbonate from a sterile anoxic stock solution prepared under 80% N2 and 20% CO2 gas mixture. The vitamin solution should be sterilized by filtration. Adjust pH of the complete medium to 6.7 - 7.0, if necessary.
2. After inoculation, add 1 bar overpressure of sterile 100% H2 gas.
3. Note: Use at least 10% (v/v) as inoculum.


## 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.


## Selenite-tungstate solution

0.5	g	NaOH	
3	mg	Na2SeO3 x 5 H2O	
4	mg	Na2WO4 x 2 H2O	
1000	ml	Distilled water	


## Wolin's vitamin solution

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 D-(+)-pantothenate	
0.1	mg	Vitamin B12	
5	mg	p-Aminobenzoic acid	
5	mg	(DL)-alpha-Lipoic acid	
1000	ml	Distilled water