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Medium No.	1020
Medium	Methanofollis ethanolicus medium
Composition
Ethanol 0.46 ml   Yeast extract 0.1 g   KH2PO4 0.14 g   MgCl2·6H2O 0.2 g   CaCl2·2H2O 0.15 g   NH4Cl 0.54 g   Sodium acetate 0.82 g   Vitamin solution* 2 ml   Trace element solution** 2 ml   Se/W solution*** 1 ml   Resazurin solution**** 1 ml   NaHCO3 2.5 g   Na2S·9H2O 0.3 g   Cysteine-HCl 0.3 g   Distilled water 1 L     For vial cultures, dissolve all ingredients except for ethanol, vitamin solution, Na2S·9H2O and cysteine-HCl, flush N2/CO2 (80/20) and autoclave. Concentrated Na2S·9H2O and cysteine-HCl solutions are sterilized separately by autoclaving under a N2 atmosphere in tightly closed vessels. Add filter-sterilized 0.8 M ethanol and vitamin solutions, and autoclaved Na2S·9H2O and cysteine-HCl solutions . Inoculum size 10-20%.                   *Vitamin solution       Biotin 0.5 mg       Folic acid 0.9 mg       Pyridoxine-HCl 0.4 mg       Thiamine-HCl 0.7 mg       Riboflavin 0.8 mg       Nicotinic acid 0.2 mg       Ca-pantothenate 0.5 mg       Vitamin B12 2.7 mg       p-Aminobenzoic acid 0.3 mg       Lipoic acid 0.4 mg       Distilled water 1 L                   **Trace element solution       Nitrilotriacetic acid (NTA) 12.8 g       FeCl3·6H2O 1.35 g       MnCl2·4H2O 0.1 g       CoCl2·6H2O 24 mg       CaCl2·2H2O 0.1 g       ZnCl2 0.1 g       CuCl2·2H2O 25 mg       H3BO3 10 mg       Na2MoO4·2H2O 24 mg       NiCl2·6H2O 0.12 g       Distilled water 1 L     First dissolve NTA in 200 ml of water and adjust the pH to 6.5 with KOH.                   ***Se/W solution       Na2SeO3 4 mg       Na2WO4 4 mg       Distilled water 1 L                   ****Resazurin solution       Resazurin 1 g       Distilled water 1 L
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