159 .
TREPONEMA MEDIUM
What is this page?
Equipment needed:
Autoclave
Centrifuge
Filter
Gassing station
Hungate tubes
Final gas composition:
100 % CO2
read more
Final pH:
6.7 - 7.0
Select a strain (optional):
ml
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
Clarified rumen fluid | 300.00 | ml | ||
|
|
CaCl2 x 2 H2O | 0.12 | g | ||
|
|
MgSO4 x 7 H2O | 0.18 | g | ||
|
|
KH2PO4 | 0.45 | g | ||
|
|
K2HPO4 | 0.45 | g | ||
|
|
NaCl | 0.90 | g | ||
|
|
(NH4)2SO4 | 0.90 | g | ||
|
|
Sodium resazurin
(0.1% w/v) |
0.50 | ml | ||
|
|
L-Cysteine HCl x H2O | 1.00 | g | ||
|
|
Na2CO3 | 1.50 | g | ||
|
|
D-Glucose | 2.00 | g | ||
|
|
Distilled water | 700.00 | ml | ||
| 1 Dissolve ingredients (except cysteine, carbonate and glucose) and sparge medium with 100% CO2 gas for 30 - 45 min to make it anoxic. Add cysteine and adjust pH with KOH to 6.5. Distribute medium under 100% CO2 gas atmosphere into anoxic Hungate-type tubes or serum vials and autoclave. Therafter, add glucose from an anoxic stock solution prepared under 100% N2 gas atmosphere and sterilized by filtration and carbonate from a sterile anoxic stock solution prepared under 80% N2 and 20% CO2 gas atmosphere. Final pH of the medium should be 6.7 - 7.0. | |||||
Clarified rumen fluid #
1000
| 1 Rumen fluid from cow or sheep (obtained from fistulated animals or abattoir refuse) is filtered through muslin, autoclaved at 121°C for 15 min and then centrifuged at 27,000 g for 20 min. The supernatant is made anoxic by sparging with 100% N2 gas for 15 min, dispensed under same gas atmosphere into anoxic serum vials to 30% of volume and then stored frozen at -20°C. |