# 1132: LC 2


## Main sol. 1132

1	g	MgSO4 x 7 H2O	
0.2	g	CaCl2 x 2 H2O	
0.05	g	KNO3	
1	ml	**Trace element solution SL-10**	
1000	ml	H2O	

1. Dispense 5 ml aliquots into Hungate tubes. After autoclaving add the following components per liter: 

2	ml	**Phosphate buffer** (pH 6.8)	
1	ml	HEPES buffer (pH 7.0, 2 M)	
1	ml	Fe(III)NH4-EDTA (4mg/ml)	
0.3	ml	Methanol	

2. Adjust to pH 6.8.
3. Incubate in sealed containers. The strain held in the DSMZ has been cultivated on methanol, but the original publication indicates that the strain will also grow on methane.


## Trace element solution SL-10

10	ml	HCl (25%)	
1.5	g	FeCl2 x 4 H2O	
70	mg	ZnCl2	
100	mg	MnCl2 x 4 H2O	
6	mg	H3BO3	
190	mg	CoCl2 x 6 H2O	
2	mg	CuCl2 x 2 H2O	
24	mg	NiCl2 x 6 H2O	
36	mg	Na2MoO4 x 2 H2O	
990	ml	Distilled water	

1. First dissolve FeCl2 in the HCl, then dilute in water, add and dissolve the other salts. Finally make up to 1000.00 ml.


## Phosphate buffer

3.6	g	Na2HPO4	
1.4	g	KH2PO4	
100	ml	H2O