301. METHYLOFERULA STELLATA MEDIUM

KH₂PO₄ 100.0 mg

(NH₄)₂SO₄ 100.0 mg

MgSO₄ × 7 H₂O 50.0 mg

CaCl₂ × 2 H₂O 10.0 mg

NaCl 20.0 mg

FeEDTA solution 3.0 ml

Trace elements solution (see below) 1.0 ml

Methanol 10.0 ml

Distilled water 1000.0 ml

Trace elements solution:

EDTA 5.0 g

CuCl₂ × 2 H₂O 0.1 g

FeSO₄ × 7 H₂O 2.0 g

ZnSO₄ × 7 H₂O 0.1 g

NiCl₂ × 6 H₂O 0.02 g

CoCl₂ × 6 H₂O 0.2 g

Na₂MoO₄ 0.03 g

Distilled water 1000.0 ml

FeEDTA solution:

FeSO₂ × 7 H₂O 1.54 g

Na₂EDTA 2.06 g

Distilled water 1000.0 ml

Final pH 5:0-5.8.

The medium is fairly weakly buffered so the pH should be checked before and after autoclaving at 121°C for 15 min. Methanol is filter sterilized. The pH should be adjusted with H₃PO₄ (sterilised if added to sterile medium). The strain should be grown with shaking.