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Main sol. 233

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Internal Solution-ID: S494

Equipment needed: Autoclave Hungate tubes Filter Gassing station
Compound Amount Unit Conc. [g/L] Conc. [mM]
KCl 0.34 g 0.332 4.458
MgCl2 x 6 H2O 4.00 g 3.91 19.233
MgSO4 x 7 H2O 3.45 g 3.372 13.683
NH4Cl 0.25 g 0.244 4.569
CaCl2 x 2 H2O 0.14 g 0.137 0.931
K2HPO4 0.14 g 0.137 0.786
NaCl 18.00 g 17.595 301.083
Modified Wolin's mineral solution 10.00 ml - -
Fe(NH4)2(SO4)2 x 6 H2O
(0.1% w/v) 		2.00 ml 0.002 -
Sodium resazurin
(0.1% w/v) 		0.50 ml 4.9e-4 0.002
NaHCO3 1.00 g 0.978 11.636
Methanol
(50% v/v) 		10.00 ml 7.742 241.618
Wolin's vitamin solution (10x) 1.00 ml - -
L-Cysteine HCl x H2O 0.50 g 0.489 2.783
Na2S x 9 H2O 0.50 g 0.489 2.035
Distilled water 1000.00 ml - -
1 Dissolve ingredients (except bicarbonate, vitamins, methanol, cysteine and sulfide), sparge medium with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Add and dissolve bicarbonate and adjust pH to 6.5, then distribute under same gas atmosphere into anoxic Hungate-type tubes or serum vials to 30% of their volume and autoclave. After sterilization add methanol (from 50% v/v solution), cysteine and sulfide from sterile anoxic stock solutions autoclaved under 100% N2 gas. Vitamins are prepared under N2 gas atmosphere and sterilized by filtration. Adjust pH of complete medium to 6.5, if necessary.
233 METHANOLOBUS TINDARIUS MEDIUM