MINERAL MEDIUM (NAGEL AND ANDREESEN)

Distilled water 1000.00 ml

Na2HPO4.2H2O 1.45 g

KH2PO4 0.25 g

CaCl2 0.01 g

MgSO4.7H2O 0.50 g

MnSO4 0.01 g

NH4Cl 0.30 g

NaCl 0.05 g

Vitamin Solution (see below) 5.00 ml

Trace element sol. SL-10 (see below) 1.00 ml

Dissolve and autoclave the phosphates separately. Add the vitamin solution and trace element solution after autoclaving. After mixing, pH should be 7.5.

Trace element solution SL-10:

HCl (25%; 7.7 M) 10.00 ml

FeCl2.4H2O 1.50 g

ZnCl2 70.00 mg

MnCl2.4H2O 100.00 mg

H3BO3 6.00 mg

CoCl2.6H2O 190.00 mg

CuCl2.2H2O 2.00 mg

NiCl2.6H2O 24.00 mg

Na2MoO4.2H2O 36.00 mg

Distilled water 990.00 ml

First dissolve FeCl2 in the HCl, then dilute in water, add and dissolve the other salts. Finally make up to 1000.0 ml.

Vitamin solution:

Vitamin B12 50.00 mg

Pantothenic acid 50.00 mg

Riboflavin 50.00 mg

Pyridoxamine-HCl 10.00 mg

Biotin 20.00 mg

Folic acid 20.00 mg

Nicotinic acid 25.00 mg

Nicotine amide 25.00 mg

?-lipoic acid 50.00 mg

p-aminobenzoic acid 50.00 mg

Thiamine-HCl.2H2O 50.00 mg

Distilled water 1000.00 ml

Stir for some hours, filter sterilize the solution.

Rehydrate and cultivate the lyophilized cells in complex medium (e.g. medium 2 or 45) first. After this reactivation, cultivate on mineral medium 95 with the appropriate carbon source.

Final pH is 7.0. Autoclave for 10 minutes at 120°C.