# 1479: HALODESULFURIARCHAEUM MEDIUM


## Main sol. 1479

240	g	NaCl	
2.5	g	K2HPO4	
0.5	g	(NH4)2SO4	
10	g	HEPES	
1	ml	**Trace element solution**	
2	g	Sulfur powder	
1000	ml	Distilled water	

1. Prepare the liquid medium. Dispense the medium into Hungate tubes or serum bottles under gassing with N2. Steam medium for 3h on each of 3 successive days. Before inoculation add from sterile stock solutions:

3	mM	MgCl2 x 6 H2O	
5	mM	NaHCO3	
50	mM	Sodium formate	
0.2	g/l	Yeast extract	
1	ml	** Vitamin mix (pfennig & Lippert,1966)**	
1	mM	Disodium sulfide (1M filter sterilized stock)	


## Trace element solution

5	g	EDTA	
2	g	FeSO4 x 7 H2O	
0.1	g	ZnSO4	
0.03	g	MnCl2	
0.3	g	H3BO3	
0.2	g	CoCl2 x 6 H2O	
0.01	g	CuCl2 x 5 H2O	
0.02	g	NiCl2 x 6 H2O	
0.02	g	Na2MoO4 x 2 H2O	
1000	ml	Distilled water	

1. Adjust pH to 3.0-4.0 with HCl.
2. Autoclave at 121 °C for 15 min.
3. Note: autoclave in closed bottles


##  Vitamin mix (pfennig & Lippert,1966)

1. 1: acidic solution (HCl, pH3; 100ml) 

5	mg	Thiamin	
5	mg	Calcium pantothenate	

2. 2: basic solution (100ml 0.1N NaOH) 

2	mg	Biotin	
5	mg	PABA (p-aminobenzoic acid)	
5	mg	Nicotinic acid	
10	mg	Pyridoxine	

3. 3: Neutral solution in 100 ml demi water 

2	mg	Folic acid	
5	mg	Riboflavin	

4. 4: neutral solution in 100 ml demi water 

0.5	mg	B12	

5. Sterilize by filtration.
6. Mix 1:1:1:1 just after addition or add separatlely, each 1ml/L.