246. VULCANITHERMUS MEDIUM

NH₄Cl 0.33 g

MgCl₂ × 6 H₂O 0.33 g

CaCl₂ × 2 H₂O 0.33 g

KCl 0.33 g

KNO₃ 0.33 g

NaCl 25.0 g

PIPES 3.6 g

Yeast extract 0.5 g

Tryptone 1.0 g

Sucrose 1.0 g

Vitamin solution (see below) 1.0 ml

Trace elements solution (see below) 1.0 ml

Distilled water 1000.0 ml

Vitamin solution:

Biotin 2.0 mg

Folic acid 2.0 mg

Pyridoxine 10.0 mg

Riboflavin 5.0 mg

Pantotenoic acid 5.0 mg

p-Aminobenzoic acid 5.0 mg

Thiamine-HCl 5.0 mg

Nicotinic acid 5.0 mg

Vitamin B₁₂ 0.1 mg

Lipoic acid 5.0 mg

Distilled water 1000.0 ml

Trace element solution:

Nitrilotriacetic acid 12.8 g

FeSO₄ × 7 H₂O 0.1 g

MnCl₂ × 4 H₂O 0.1 g

CoCl₂ × 2 H₂O 0.17 g

CaCl₂ × 2 H₂O 0.1 g

ZnCl₂ 0.1 g

CuCl₂ × 2 H₂O 0.02 g

H₃BO₃ 0.01 g

Na₂MoO₄ × 2 H₂O 0.01 g

NaCl 1.0 g

Na₂SeO₄ 0.017 g

Distilled water 1000.0 ml

pH 6.8

Prepare the medium anaerobically, under N₂, omitting the CaCl₂ × 2 H₂O, MgCl₂ × 6 H₂O, KNO₃, tryptone, yeast extract, vitamins and sucrose. Dispense the medium into vessels suitable for anaerobic growth (Hungate tubes or serum bottles) under an atmosphere of N₂ and autoclave at 121°C 15 min. To the sterile, cooled medium add, from sterile stock solutions the CaCl₂ × 2 H₂O, MgCl₂ × 6 H₂O, KNO₃, tryptone, yeast extract, vitamins and sucrose. The CaCl₂ × 2 H₂O, MgCl₂ × 6 H₂O, KNO₃, tryptone, and yeast extract, stock solutions should be autoclaved at 121°C 15 min, sucrose at 111°C 30 min, while the vitamin solution is sterile filtered.

 

Updated 1.02.2024