# 87: CHLOROFLEXUS MEDIUM (modified)


## Main sol. 87

1	g	Yeast extract	
1	g	Glycyl-glycine	
0.1	g	Na2HPO4 x 2 H2O	
0.1	g	MgSO4 x 7 H2O	
0.1	g	KNO3	
0.5	g	NaNO3	
0.1	g	NaCl	
0.05	g	CaCl2 x 2 H2O	
5	ml	Fe(III) citrate (0.1 g in 100 ml H2O)	
1	ml	**Trace element solution SL-6**	
1050	ml	Distilled water	

1. Adjust pH to 8.2.
2. Boil the medium under a stream of nitrogen gas for a few minutes and distribute 90 ml medium into 100 ml screw-capped bottles. Bubble each bottle with nitrogen gas and close immediately with a rubber septum and screw tight. Autoclave at 121°C for 15 min. After autoclaving inject 1.0 ml of neutralized sulfide solution (0.03% end concentration) to each bottle. This medium can be stored for several months. Incubate the culture at 50°C at a light intensity of 300 - 500 lux. For heavy cell suspension supplement periodically with sterile yeast extract solution (0.1% end concentration, for more details see Ref. 3365, 3366).


## Neutralized sulfide solution 3% (w/v)

3	g	Na2S x 9 H2O	
100	ml	Distilled water	

1. The sulfide solution is prepared in a 250 ml screw-capped bottle with a butyl rubber septum and a magnetic stirrer. The solution is bubbled with nitrogen gas, closed and autoclaved for 15 min. at 121°C. After cooling to room temperature the pH is adjusted to about 7.0 by adding of sterile 2 M H2SO4 drop-wise with a syringe without opening the bottle.


## Trace element solution SL-6

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