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Main sol. J1067

Internal Solution-ID: S5132

Equipment needed: Filter Autoclave
Compound Amount Unit Conc. [g/L] Conc. [mM]
FeS solution 50.00 ml - -
Modified Wolfe's mineral solution 50.00 ml - -
Agar , Noble
(BD-Difco)
1.50 g 1.333 3.964
1 Top layer:
Modified Wolfe's mineral solution 1000.00 ml - -
Trace minerals 10.00 ml - -
2 After cooling of the top layer solution, add per liter the following solutions (filter-sterilized):
Trace vitamins 10.00 ml - -
NaHCO3
(8%)
5.25 ml - -
3 Bottom layer:
4 Autoclave both layer solutions. Shortly after autoclaving, pipette the bottom layer into each culture vessel (typically 5 ml in 60 ml glass vials, use pipette tips that have been cut to enlarge the opening). The bottom layer is allowed to cool for at least 30 min to ensure its solidification.
5 Add the top layer solution in 5 to 6 times the volume of the bottom layer to the culture vessels containing the bottom layer (e.g., 25 to 30 ml in 60 ml glass vials with 5 ml of the bottom layer).
6 Then, cap the culture vessels with silicone rubber stoppers, and replace the gas phase with a N2-CO2 (4:1, v/v) gas mixture.
7 Comment: The medium can be stored in culture vessels for a certain time by capping with butyl rubber stoppers, instead of silicon rubber stoppers and keeping anoxic in the vessels. Silicone rubber stopper is O2 permeable and creates a mircoaerophilic condition in the culture vessels.
J1067 FRESHWATER IRON-OXIDIZING BACTERIA MEDIUM-II