# J1079: HALANAEROARCHAEUM MEDIUM


## Main sol. J1079

240	g	NaCl	
2.5	g	K2HPO4	
0.5	g	NH4Cl	
7	g	HEPES	

1. Add components to distilled water and bring volume to 1.0 L. Autoclave and, after cooling, add per liter the following solutions (autoclaved or *filter-sterilized separately): 

1	ml	MgCl2 (1 M)	
1	ml	**Selenite-tungstate solution**	
1	ml	**Trace element solution**	
10	ml	NaHCO3 (*, 1 M)	

2. To complete the medium, aseptically and anaerobically distribute the medium into culture vessels containing steam-sterilized sulfur (see below, approximately 1.0 g/L) under an argon gas stream, and seal with butyl rubber stoppers. Then, add per liter the following solutions (autoclaved or *filter-sterilized): 

10	ml	Sodium pyruvate (*, 2 M)	
0.2	ml	Yeast extract (10%)	
5	ml	Na2S x 9 H2O (5%)	

3. Basal mineral NaCl medium:


## Sterilization of sulfur

1. Steam sulfur for 3 hr, or autoclave at 105C for 1 hr, on each of 3 successive days.


## Trace element solution

5	g	EDTA	
2	g	FeSO4 x 7 H2O	
0.1	g	ZnSO4 x 7 H2O	
0.03	g	MnCl2 x 4 H2O	
0.3	g	H3BO3	
0.2	g	CoCl2 x 6 H2O	
0.01	g	CuCl2 x 2 H2O	
0.02	g	NiCl2 x 6 H2O	
0.03	g	Na2MoO4 x 2 H2O	
1000	ml	Distilled water	

1. Adjust pH to 3.6.


## Selenite-tungstate solution

0.4	g	NaOH	
6	mg	Na2SeO3 x 5 H2O	
8	mg	Na2WO4 x 2 H2O	
1000	ml	Distilled water