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Main sol. J243

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Internal Solution-ID: S3923

Equipment needed: Autoclave Anaerobic workbench Filter
Compound Amount Unit Conc. [g/L] Conc. [mM]
KH2PO4 0.30 g 0.294 2.161
NaCl 0.60 g 0.588 10.066
MgCl2 x 6 H2O 0.10 g 0.098 0.482
CaCl2 x 2 H2O 0.08 g 0.078 0.533
NH4Cl 1.00 g 0.98 18.329
Trace element solution 10.00 ml - -
Trace vitamins 10.00 ml - -
Sodium acetate 6.80 g 6.667 81.271
KHCO3 4.00 g 3.922 39.171
L-Cysteine HCl x H2O 0.50 g 0.49 -
Na2S x 9 H2O 0.50 g 0.49 2.041
Resazurin 1.00 mg 9.8e-4 0.004
Distilled water 1000.00 ml - -
1 Mix ingredients, except Trace vitamins, KHCO3, L-cysteine x HCl x H2O and Na2S x 9H2O, and adjust pH to 7.0. Bring to a boil for 5-10 sec, then, cool down under a stream of N2-CO2 (80:20) and add KHCO3. Dispense the medium into suitable culture vessels (e.g., 50 ml of the medium in 120 ml serum bottles) under a stream of N2-CO2 (80:20) and seal with butyl rubber stoppers. After autoclaving, stand the medium overnight. Separately autoclave cysteine x HCl and Na2S x 9H2O as 5% solutions, respectively, under a N2 atmosphere. Filter-sterilize Trace vitamins under a N2 atmosphere. Prior to inoculation, aseptically and anaerobically add Trace vitamins, L-cysteine x HCl x H2O and Na2S x 9H2O solutions. Pressurize the inoculated bottles to 100 kPa N2-CO2 (80:20).
J243 METHANOTHRIX MEDIUM