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Main sol. 817

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Internal Solution-ID: S1665

Equipment needed: Filter Autoclave Hungate tubes Gassing station
Compound Amount Unit Conc. [g/L] Conc. [mM]
MgCl2 x 6 H2O 2.2 g 2.167 10.661
NaCl 0.9 g 0.887 15.173
KCl 17.0 mg 0.017 0.225
NH4Cl 12.0 mg 0.012 0.221
CaCl2 x 2 H2O
(0.1% w/v) 		7.0 ml 0.007 0.047
K2HPO4 x 3 H2O
(0.1% w/v) 		7.0 ml 0.007 0.04
FeCl3
(0.01% w/v in 0.2 N HCl) 		0.5 ml 4.9e-5 3.0e-4
Yeast extract
(BD Bacto) 		1.0 g 0.985 -
Peptone
(BD Bacto) 		1.0 g 0.985 -
Sodium resazurin
(0.1% w/v) 		0.5 ml 4.9e-4 0.002
Na2CO3 0.5 g 0.493 4.648
Wolin's vitamin solution (10x) 1.0 ml - -
Na2S x 9 H2O 0.5 g 0.493 2.051
Distilled water 1000.0 ml - -
1 Dissolve ingredients except carbonate, vitamins and sulfide. Sparge medium with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic, then dispense under same gas atmosphere into anoxic Hungate-type tubes or serum vials to 30% of their volume and autoclave. Add vitamins and sulfide from sterile anoxic stock solutions prepared under 100% N2 gas and carbonate from a sterile anoxic stock solution prepared under 80% N2 and 20% CO2 gas atmosphere. Vitamins should be sterilized by filtration. The pH of the complete medium should be adjusted to 6.5.
2 After inoculation pressurize tubes or bottles (heavy walled) to 2 bar overpressure with sterile 80% N2 and 20% CO2 gas mixture.
817 THERMOSPHAERA MEDIUM