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Anweisungen

Anweisung Lösung Medium
Adjust pH to 7.0. For Bacillus strains the addition of 10.0 mg MnSO4 x H2O is recommended for sporulation. 1 1
Incubate microaerobic, for example in a candle jar. 2 10
Before autoclaving, the pH of the Medium is adjusted with NaOH to 7.5-8.0. After autoclaving, the filter-sterilized vitamin solution (0.1% volume), and each of the separately autoclaved, concentrated solutions are added to the medium at a final concentration described below (#14-16). Then a mix gas (80% H2, 20% CO2) is purged for 5 min. Finally, the mix gas is compressed into gas phase (> 80% volume of the tube or bottle) at 3 atm. 3 1000
After autoclaving add: #5-8 150 1078
For 200 ml of medium add: #1-5 143 1076b
Adjust pH to 7.4; autoclave at 121°C for 15 minutes. 143 1076b
This medium should not be exposed to direct sunlight! Add all basal medium ingredients. Bring the final volume of the medium to 1.0 liter. Dispense to appropriate culture containers. Bubble the medium with 80:20 N2:CO2 (final pH sould be 6.8 to 7.0) - approximately 10 ml of media (anaerobic culture tube) should be gassed for 5 min in the aqueous phase (bubbled) and the headspace gassed for one minute prior to sealing the container. Sterilize per usual procedure. Add electron donor (Acetate-final conc. of 10 mM-recipe below) and electron acceptor (Fe(III)NTA-final conc. of 10 mM-recipe below) from sterile, anaerobic stock solutions using a sterile syringe and needle flushed with anaerobic gas. Store out of direct light. 5 1001
Add 13.6 g sodium acetate to ca. 80 ml distilled water. Bring to final volume of 100 ml. Bubble with anaerobic gas (nitrogen) for 45 min. Seal and sterilize. 6 1001
Add 8.2 g NaHCO3 to ca. 70 ml distilled water. Add 12.8 g of Na3Nitrilotriacetic acid (NTA) to ca. 70 ml distilled water. Add 13.5 g FeCl3 x 6 H2O to ca 70 ml distilled water. Adjust pH to 6.5 using 10N NaOH. Bring solution to final volume of 100 ml. Ingredients will go into solution after stirring for ca. 15 min. Bubble with anaerobic gas (N2) for 45 min and then filter sterilize (0.2 μm filter) into a sterile, anaerobic, serum bottle. 7 1001
Prepare the NH4Cl, CaCl2, carbonate, bicarbonate and sulfide as a single sterile stock solution and add to the sterile medium after autoclaving. The pH should be adjust to 9-9.5. 10 1002
Place 1000 ml of anaerobic water (CO2 water) in a flask. Add the following compounds: #2-7 11 1003
Add solutions A, B, and D (see recipes below): 11 1003
pH the media to 6.0 and gas the flask containing the media with CO2 for 20 min at least. Gas first culture tube in rack and place 5 ml of media in previous tube. Place stopper on loaded culture tube, crimp tube cap onto stopper. Autoclave the medium for 20 min., 121°C. Add 1 ml of O2 to each tube before inoculation. After the inoculation the tubes are pressurized with H2 (138 KP). 11 1003
Adjust the pH to 4 with HCl. 12 1003
Adjust the pH of the solution to 3.0 with HCl. 14 1003
Dissolve ingredients (except bicarbonate, yeast extract, vitamins, glucose and reducing agents), then sparge medium with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic. Add solid bicarbonate and adjust pH to 7.0. Dispense medium under 80% N2 and 20% CO2 gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. Add yeast extract, vitamins (sterilized by filtration), glucose, sulfide and cysteine from sterile anoxic stock solutions prepared under 100% N2 gas. Adjust pH of complete medium to 7.0, if necessary. 15 1004
For DSM 16554, DSM 16555 and DSM 17877 replace glucose with 7.20 g/l sucrose and reduce amount of yeast extract to 0.10 g/l. 1004
For DSM 16556 omit glucose and reduce amount of yeast extract to 0.10 g/l. 1004
For DSM 22659 increase amount of glucose to 5.00 g/l. 1004
For DSM 23815 replace glucose with 2.00 g/l cellobiose added after autoclaving from a sterile anoxic stock solution sterilized by filtration. 1004
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