336. ALKALIPHILIC SPIROCHAETE MEDIUM

Na₂CO₃ 10.0 g

NaHCO₃ 15.0 g

NaCl 10.0 g

K₂HPO₄ 0.2 g

NH₄Cl 1.0 g

KCl 0.2 g

Na₂S × 9 H₂O 1.0 g

Yeast extract 0.5 g

Sucrose 5.0 g

Vitamin solution (see below) 10.0 ml

Trace element solution (see below) 1.0 ml

Distilled water 1000.0 ml

Trace element solution:

Nitrilotriacetic acid 1.5 g

MgSO₄ × 7 H₂O 3.0 g

MnSO₄ × H₂O 0.5 g

NaCl 1.0 g

FeSO₄ × 7 H₂O 0.1 g

CoSO₄ × 7 H₂O 0.18 g

CaCl₂ × 2 H₂O 0.1 g

ZnSO₄ × 7 H₂O 0.18 g

CuSO₄ × 5 H₂O 0.01 g

KAl(SO₄)₂ × 12 H₂O 0.02 g

H₃BO₃ 0.01 g

Na₂MoO₄ × 2 H₂O 0.01 g

NiCl₂ × 6 H₂O 0.03 g

Na₂SeO₃ × 5 H₂O 0.3 mg

Na₂WO₄ × 2 H₂O 0.4 mg

Distilled water 1000.0 ml

Vitamin solution:

Biotin 2.0 mg

Folic acid 2.0 mg

Pyridoxine-HCl 10.0 mg

Thiamine-HCl × 2 H₂O 5.0 mg

Riboflavin 5.0 mg

Nicotinic acid 5.0 mg

D-Ca-pantothenate 5.0 mg

Vitamin B₁₂ 0.1 mg

p-Aminobenzoic acid 5.0 mg

Lipoic acid 5.0 mg

Distilled water 1000.0 ml

Prepare the medium under N₂, omitting Na₂CO₃, NaHCO₃, Na₂S × 9 H₂O, sucrose and vitamin solution. Boil the medium and cool under nitrogen, add the Na₂CO₃ and NaHCO₃ and adjust the pH to 9.7 with 6N NaOH (about 15 ml). Autoclave at 121°C for 15 min and add the Na₂S × 9 H₂O (neutralized), vitamins and sucrose from sterile stock solutions.

 

Updated 1.02.2024