358. THERMOANAEROBACTERIUM MEDIUM

KH₂PO₄ 0.3 g

Na₂HPO₄ × 12 H₂O 5.3 g

NH₄Cl 1.0 g

MgCl₂ × 6 H₂O 0.2 g

Trace elements solution (see below) 10.0 ml

FeSO₄ × 7 H₂O solution (0.1% w/v in 0.1 N H₂SO₄) 1.5 ml

Yeast extract 1.0 g

Na-resazurin solution (0.1% w/v) 0.5 ml

D-Glucose 5.0 g

Vitamin solution (see below) 5.0 ml

Na₂S × 9 H₂O 0.5 g

Distilled water 1000.0 ml

Trace element solution:

Nitrilotriacetic acid (NTA) 12.8 g

FeCl₂ × 4 H₂O 0.2 g

MnCl₂ × 4 H₂O 0.1 g

CoCl₂ × 6 H₂O 0.17 g

CaCl₂ × 2 H₂O 0.1 g

ZnCl₂ 0.1 g

CuCl₂ 0.02 g

H₃BO₃ 0.01 g

Na₂MoO₄ × 2 H₂O 0.01 g

NiCl₂ × 6 H₂O 0.03 g

NaCl 1.0 g

Na₂SeO₃ × 5 H₂O 0.03 g

First dissolve NTA in 200 ml of distilled water and adjust pH to 6.5 with KOH, then dissolve mineral salts. Finally adjust pH to 6.5 with KOH and make up to 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

Dissolve ingredients (except glucose, vitamins and sulfide), adjust pH to 6.0, sparge medium with 100% N₂ gas for 30 – 45 min to make it anoxic. Dispense under same gas atmosphere into Hungate-type tubes or serum vials and autoclave. After sterilization add glucose and sulfide from anoxic stock solutions autoclaved under 100% N₂ gas and vitamins from a filter-sterilized anoxic stock solution prepared under 100% N₂. Adjust pH of complete medium to 6.0 – 6.5, if necessary.

 

Updated 11.08.2021