# 319: SPOROHALOBACTER LORTETII MEDIUM


## Main sol. 319

2	g	Casamino acids (BD Bacto)	
2	g	Nutrient broth (BD Difco)	
2	g	Yeast extract	
4	g	L-Glutamic acid	
105	g	NaCl	
0.75	g	KCl	
10	ml	**Modified Wolin's mineral solution**	
2	ml	FeSO4 x 7 H2O (0.1% w/v in 0.1 N H2SO4)	
0.5	ml	Sodium resazurin (0.1% w/v)	
0.5	g	L-Cysteine HCl x H2O	
10	g	MgCl2 x 6 H2O	
3.7	g	CaCl2 x 2 H2O	
1	ml	**Wolin's vitamin solution (10x)**	
1000	ml	Distilled water	

1. Dissolve ingredients (except cysteine, magnesium chloride, calcium chloride and vitamins), bring medium to the boil, then cool to room temperature under 100% N2 gas atmosphere. Thereafter, add and dissolve cysteine, adjust pH to 6.5 and dispense under same gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. After sterilization add magnesium chloride and calcium chloride from sterile anoxic stock solutions autoclaved under 100% N2 gas and vitamins from a filter-sterilized stock solution prepared under N2 gas.
2. Note: For agar medium add 20.00 g/l agar, 5.00 g/l CaCO3, and 2.00 g/l soluble starch.


## Modified Wolin's mineral solution

1.5	g	Nitrilotriacetic acid	
3	g	MgSO4 x 7 H2O	
0.5	g	MnSO4 x H2O	
1	g	NaCl	
0.1	g	FeSO4 x 7 H2O	
0.18	g	CoSO4 x 7 H2O	
0.1	g	CaCl2 x 2 H2O	
0.18	g	ZnSO4 x 7 H2O	
0.01	g	CuSO4 x 5 H2O	
0.02	g	AlK(SO4)2 x 12 H2O	
0.01	g	H3BO3	
0.01	g	Na2MoO4 x 2 H2O	
0.03	g	NiCl2 x 6 H2O	
0.3	mg	Na2SeO3 x 5 H2O	
0.4	mg	Na2WO4 x 2 H2O	
1000	ml	Distilled water	

1. First dissolve nitrilotriacetic acid and adjust pH to 6.5 with KOH, then add minerals. Adjust final to pH 7.0 with KOH.


## Wolin's vitamin solution (10x)

20	mg	Biotin	
20	mg	Folic acid	
100	mg	Pyridoxine hydrochloride	
50	mg	Thiamine HCl	
50	mg	Riboflavin	
50	mg	Nicotinic acid	
50	mg	Calcium D-(+)-pantothenate	
1	mg	Vitamin B12	
50	mg	p-Aminobenzoic acid	
50	mg	(DL)-alpha-Lipoic acid	
1000	ml	Distilled water