# 210: HALOANAEROBIUM MEDIUM


## Main sol. 210

130	g	NaCl	
8.8	g	MgSO4 x 7 H2O	
1	g	KCl	
10	g	Yeast extract	
10	g	Trypticase (BBL)	
10	ml	**Wolin's vitamin solution**	
10	ml	**Trace element solution**	
1000	ml	Distilled water	

1. Prepare the medium anaerobically under 100% N2. After autoclaving add from sterile solutions: 

25	ml	**Thioglycolate-ascorbate solution**	
25	ml	Glucose (10 % w/v)	
10	ml	NaOH (2 N)	

2. Final pH: 9.0.


## Thioglycolate-ascorbate solution

0.5	g	Na-thioglycolate	
0.5	g	Na-ascorbate	
25	ml	Distilled water	

1. Filter sterilize.


## 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	


## Trace element solution

12.8	g	Nitrilotriacetic acid (NTA)	
0.2	g	FeCl2 x 4 H2O	
0.1	g	MnCl2 x 4 H2O	
0.17	g	CoCl2 x 6 H2O	
0.1	g	CaCl2 x 2 H2O	
0.1	g	ZnCl2	
0.02	g	CuCl2	
0.01	g	H3BO3	
0.01	g	Na2MoO4 x 2 H2O	
0.03	g	NiCl2 x 6 H2O	
1	g	NaCl	
0.03	g	Na2SeO3 x 5 H2O	
1000	ml	Distilled water	

1. First dissolve NTA in 200 ml of distilled water and adjust pH to 6.5 with KOH, then dissolve mineral salts. Finally adjust pH to 6.5 with KOH and make up to 1000.00 ml.