# J1101: HALOLACTIBACILLUS 9G MEDIUM


## Main sol. J1101

10	g	NaCl	
0.2	g	KCl	
0.2	g	K2HPO4	
0.1	g	MgCl2 x 6 H2O	
1	g	NH4Cl	
0.1	g	Yeast extract	
2	ml	**Trace vitamins**	
1	ml	**Trace minerals solution**	
0.5	mg	Resazurin	
680	ml	Distilled water	

1. Mix components thoroughly and autoclave under a N2 atmosphere in a bottle capped with a butyl rubber stopper. After cooling, add the following solution to the medium: 

300	ml	**Alkaline solution**	
20	ml	Glucose (autoclaved, 1.0 M)	

2. Anaerobically and aseptically distribute the medium into culture vessels (e.g., 10 ml medium in Hungate tubes) under a N2 gas atmosphere and seal with butyl rubber stoppers. Prior to inoculation, add per liter the following solution (autoclaved and stored under a N2 gas atmosphere): 

8	ml	Na2S x 9 H2O (5%)	

3. Adjust pH to 9.0, if necessary.


## Alkaline solution

2.76	g	Na2CO3	
24	g	NaHCO3	
300	ml	Distilled water	

1. Mix components thoroughly and filter-sterilize.


## Trace vitamins

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 pantothenate	
0.1	mg	Vitamin B12	
5	mg	p-Aminobenzoic acid	
5	mg	Lipoic acid	
1000	ml	Distilled water	


## Trace minerals solution

1.5	g	Nitrilotriacetic acid	
0.2	g	Fe(NH4)2(SO4)2 x 6 H2O	
0.1	g	Na2SeO3	
0.1	g	Na2MoO4 x 2 H2O	
1	g	Na2WO4 x 2 H2O	
0.1	g	MnSO4 x n H2O	
0.1	g	ZnSO4 x 7 H2O	
0.025	g	NiCl2 x 6 H2O	
0.01	g	CuSO4 x 5 H2O	
1000	ml	Distilled water	

1. Dissolve nitrilotriacetic acid and adjust pH to 6.5 with KOH. Then proceed to add minerals.
2. Adjust final pH to 7.0.