110a .
CMC MEDIUM (N2/CO2)
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Selected strain: Laedolimicola ammoniilytica DSM 102317
This strain uses the standard medium without modifications.
Growth conditions: anaerobic
37 °C
Equipment needed:
Autoclave
Filter
Gassing station
Hungate tubes
Final pH:
7.0
Select a strain (optional):
ml
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
Casitone | 30.0 | g | ||
|
|
Yeast extract | 5.0 | g | ||
|
|
K2HPO4 | 5.0 | g | ||
|
|
Sodium resazurin
(0.1% w/v) |
0.5 | ml | ||
|
|
L-Cysteine HCl x H2O | 0.5 | g | ||
|
|
Sugar mix | 10.0 | ml | ||
|
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Na2CO3 | 1.5 | g | ||
|
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Meat filtrate | 1000.0 | ml | ||
| 1 Dissolve ingredients, except cysteine, sugar mix, and carbonate, boil medium for 1 min, then cool to room temperature under 80% N2 and 20% CO2 gas atmosphere, add 0.5 g/l L-cysteine-HCl x H2O and dispense under same gas atmosphere into Hungate-type tubes (for strains demanding meat particles put those first into the tube, use 1 part meat particles to 4 or 5 parts fluid). Autoclave at 121°C for 20 min. After autoclaving add the sugar mix from a sterile anoxic stock solution prepared under 100% N2 gas and carbonate from a sterile anoxic stock solution prepared under 80% N2 and 20% CO2 gas mixture. Adjust the pH of the medium to pH 7.0, if necessary. | |||||
Meat filtrate #
1000
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
Ground beef
(fat free) |
500 | g | ||
|
|
NaOH
(1 N) |
25 | ml | ||
|
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Distilled water | 975 | ml | ||
| 1 Use lean beef or horse meat. Remove fat and connective tissue before grinding. Mix meat, water and NaOH, then boil for 15 min with stirring. Cool to room temperature, skim fat off surface, and filter, retaining both meat particles and filtrate. To the filtrate add water to a final volume of 1000.0 ml. | |||||
Sugar mix #
10
| Compound | Amount | Unit | |
|---|---|---|---|
|
|
D-Glucose | 4 | g |
|
|
Cellobiose | 1 | g |
|
|
D-Maltose | 1 | g |
|
|
Starch
(soluble) |
1 | g |
|
|
Distilled water | 10 | ml |