# J1090: MMJS MEDIUM 2


## Main sol. J1090

20	g	NaCl	
0.14	g	K2HPO4	
1	g	CaCl2 x 2 H2O	
3.4	g	MgSO4 x 7 H2O	
4.18	g	MgCl2 x 6 H2O	
0.25	g	NH4Cl	
0.33	g	KCl	
0.5	mg	NiCl2 x 6 H2O	
0.5	mg	Na2SeO3 x 5 H2O	
0.1	mg	Na2WO4 x 2 H2O	
10	mg	(NH4)2Ni(SO4)2 x 6 H2O	
10	ml	**Trace minerals**	
1000	ml	Distilled water	

1. Mix components and adjust pH to 5.5. Distribute the medium into culture vessels (e.g., 20 ml in 60 ml-serum bottles) under a stream of N2, seal with butyl rubber stoppers and add Autoclave. After cooling, add per liter the following solutions (filter-sterilized): 

1	ml	**Trace vitamins**	
3.75	ml	NaHCO3 (8%)	
5	ml	Na2S2O3 (1 M)	

2. Replace the gas phase with N2-CO2-O2 (94:5:1, v/v/v, 100 kPa). Check final pH to be around 6.2.


## Trace minerals

1.5	g	Nitrilotriacetic acid	
3	g	MgSO4 x 7 H2O	
0.5	g	MnSO4 x n H2O	
1	g	NaCl	
0.1	g	FeSO4 x 7 H2O	
0.1	g	CoSO4 x 7 H2O	
0.1	g	CaCl2 x 2 H2O	
0.1	g	ZnSO4 x 7 H2O	
0.01	g	CuSO4 x 5 H2O	
0.01	g	AlK(SO4)2	
0.01	g	H3BO3	
0.01	g	Na2MoO4 x 2 H2O	
1000	ml	Distilled water	

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


## 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