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Main sol. 843

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Internal Solution-ID: S1727

Equipment needed: Hungate tubes Filter Centrifuge Autoclave Gassing station
Compound Amount Unit Conc. [g/L] Conc. [mM]
NaCl 1.00 g 0.997 17.06
KCl 0.50 g 0.499 6.687
MgCl2 x 6 H2O 0.40 g 0.399 1.962
CaCl2 x 2 H2O 0.10 g 0.1 0.678
NH4Cl 0.30 g 0.299 5.592
KH2PO4 0.20 g 0.199 1.465
Na2SO4 0.15 g 0.15 1.053
Trace element solution SL-10 1.00 ml - -
Selenite-tungstate solution 1.00 ml - -
Trypticase peptone
(BD BBL) 		1.00 g 0.997 -
Sodium resazurin
(0.1% w/v) 		0.50 ml 5.0e-4 0.002
Na2CO3 1.50 g 1.496 14.11
Sugar mix 30.00 ml - -
Yeast autolysate 50.00 ml - -
Seven vitamins solution 1.00 ml - -
Co-factors solution 20.00 ml - -
DL-Dithiothreitol
(DTT) 		0.40 g 0.399 2.585
Distilled water 900.00 ml - -
1 Dissolve ingredients (except carbonate, sugar mix, yeast autolysate, vitamins, co-factors, and DTT), sparge medium with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Distribute under same gas atmosphere into anoxic Hungate-type tubes or serum vials to 30% of their volume (e.g., 4.5 ml per Hungate-type tube) and autoclave. After sterilization add carbonate from a sterile anoxic stock solution prepared under 80% N2 and 20% CO2 gas atmosphere and sugar mix, yeast autolysate, vitamins, co-factors and DTT from sterile anoxic stock solutions prepared under 100% N2 gas atmosphere and sterilized by filtration. Check pH of complete medium and adjust to 7.2 - 7.4, if necessary.
2 Note: Basal medium and yeast autolysate have to be prepared freshly and cannot be stored for longer periods.
843 HELMUTKOENIGIA MEDIUM