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Main sol. 142

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Internal Solution-ID: S246

Equipment needed: Autoclave Hungate tubes Filter Gassing station
Compound Amount Unit Conc. [g/L] Conc. [mM]
NaCl 25.00 g 24.876 425.661
(NH4)2SO4 1.00 g 0.995 7.53
MgSO4 x 7 H2O 1.50 g 1.493 6.056
CaCl2 x 2 H2O 0.42 g 0.418 2.843
Trace element solution (Vishniac & Santer, 1957) 0.20 ml - -
Bromothymol blue
(0.1% w/v) 		4.00 ml 0.004 0.006
K2HPO4 0.50 g 0.498 2.856
Na2S2O3 x 5 H2O 5.00 g 4.975 20.046
Seven vitamins solution 1.00 ml - -
Distilled water 1000.00 ml - -
1 Dissolve ingredients (except hydrogenphosphate, thiosulfate and vitamins), adjust pH to 7.2 and autoclave. K2HPO4 and Na2S2O3 are autoclaved separately each in 10% of the final volume. Filter sterilize the vitamins solution. Adjust pH of the complete medium to 7.2 with sterile 0.4% (w/v) Na2CO3 solution. Acidification of the medium during growth causes the pH indicator bromothymol blue to turn from blue to yellow.
2 Note: Growth of most Thiomicrospira strains is more reliable if the medium is prepared under a 80% N2 and 20% CO2 gas atmosphere to make it anoxic and then filled under air atmosphere into Hungate-type tubes (5 ml per vial). The pH is adjusted with a sterile stock solution of Na2CO3 (5% w/v) after autoclaving.
142 THIOMICROSPIRA PELOPHILA MEDIUM