142 .
THIOMICROSPIRA PELOPHILA MEDIUM
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Equipment needed:
Autoclave
Filter
Gassing station
Hungate tubes
Final pH:
7.2
Select a strain (optional):
ml
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
NaCl | 25.00 | g | ||
|
|
(NH4)2SO4 | 1.00 | g | ||
|
|
MgSO4 x 7 H2O | 1.50 | g | ||
|
|
CaCl2 x 2 H2O | 0.42 | g | ||
|
|
Trace element solution (Vishniac & Santer, 1957) | 0.20 | ml | ||
|
|
Bromothymol blue
(0.1% w/v) |
4.00 | ml | ||
|
|
K2HPO4 | 0.50 | g | ||
|
|
Na2S2O3 x 5 H2O | 5.00 | g | ||
|
|
Seven vitamins solution | 1.00 | ml | ||
|
|
Distilled water | 1000.00 | ml | ||
| 1 Dissolve ingredients (except hydrogenphosphate, thiosulfate and vitamins), adjust pH to 7.2 and autoclave. K2HPO4 and Na2S2O3 are autoclaved separately each in 10% of the final volume. Filter sterilize the vitamins solution. Adjust pH of the complete medium to 7.2 with sterile 0.4% (w/v) Na2CO3 solution. Acidification of the medium during growth causes the pH indicator bromothymol blue to turn from blue to yellow. | |||||
| 2 Note: Growth of most Thiomicrospira strains is more reliable if the medium is prepared under a 80% N2 and 20% CO2 gas atmosphere to make it anoxic and then filled under air atmosphere into Hungate-type tubes (5 ml per vial). The pH is adjusted with a sterile stock solution of Na2CO3 (5% w/v) after autoclaving. | |||||
Trace element solution (Vishniac & Santer, 1957) #
1000
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
Na2-EDTA | 50.00 | g | ||
|
|
ZnSO4 x 7 H2O | 22.00 | g | ||
|
|
CaCl2 x 2 H2O | 5.54 | g | ||
|
|
MnCl2 x 4 H2O | 5.06 | g | ||
|
|
FeSO4 x 7 H2O | 5.00 | g | ||
|
|
(NH4)6Mo7O24 x 4 H2O | 1.10 | g | ||
|
|
CuSO4 x 5 H2O | 1.57 | g | ||
|
|
CoCl2 x 6 H2O | 1.61 | g | ||
|
|
Distilled water | 1000.00 | ml | ||
| 1 Dissolve EDTA in distilled water, adjust pH to 7 using 2 N KOH, then add remaining compounds. Adjust pH of final solution to 6.0 with KOH. | |||||
Seven vitamins solution #
1000
| Compound | Amount | Unit | |
|---|---|---|---|
|
|
Vitamin B12 | 100 | mg |
|
|
p-Aminobenzoic acid | 80 | mg |
|
|
D-(+)-biotin | 20 | mg |
|
|
Nicotinic acid | 200 | mg |
|
|
Calcium pantothenate | 100 | mg |
|
|
Pyridoxine hydrochloride | 300 | mg |
|
|
Thiamine-HCl x 2 H2O | 200 | mg |
|
|
Distilled water | 1000 | ml |