739 .
DESULFONEMA ISHIMOTOI MEDIUM
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Equipment needed:
Autoclave
Filter
Gassing station
Hungate tubes
Final pH:
7.0
Select a strain (optional):
ml
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
Solution A | 952 | ml | ||
|
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Solution B | 10 | ml | ||
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Solution C | 1 | ml | ||
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Solution D | 10 | ml | ||
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Solution E | 20 | ml | ||
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Solution F | 10 | ml | ||
| 1 Sparge solution A with 80% N2 and 20% CO2 gas mixture for 30 - 45 min to make it anoxic and reach a pH of around 6, then distribute under the same gas atmosphere into anoxic Hungate-type tubes and autoclave. Solutions B and C are prepared under 100% N2 gas atmosphere and sterilized by filtration. Solutions D and F are autoclaved separately under 100% N2 gas. Solution E is prepared under 80% N2 and 20% CO2 gas gas atmosphere. To complete the medium appropriate amounts of solutions B to F are added to the sterile solution A in the sequence as indicated. Final pH of the medium should be at 7.0. After combining solutions the medium should equilibrate overnight and a white precipitate should be apparent. | |||||
| 2 Note: Addition of 10 - 20 mg sodium dithionite per liter (e.g. from 5% (w/v) solution freshly prepared under N2 and filter-sterilized) just before inoculation may stimulate growth at the beginning. For transfers use 5 - 10% (v/v) inoculum. | |||||
Solution A #
952
| Compound | Amount | Unit | |
|---|---|---|---|
|
|
NaCl | 25.00 | g |
|
|
MgCl2 x 6 H2O | 5.60 | g |
|
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MgSO4 x 7 H2O | 6.80 | g |
|
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CaCl2 x 2 H2O | 1.40 | g |
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KCl | 0.72 | g |
|
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KH2PO4 | 0.14 | g |
|
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NH4Cl | 0.25 | g |
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Trace element solution SL-10 | 1.00 | ml |
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Selenite-tungstate solution | 1.00 | ml |
|
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Sodium resazurin
(0.1% w/v) |
0.50 | ml |
|
|
Distilled water | 950.00 | ml |
Solution B #
10
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
Na-acetate x 3 H2O | 2.50 | g | ||
|
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Disodium succinate | 0.10 | g | ||
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Isobutyric acid | 0.19 | ml | ||
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Distilled water | 10.00 | ml | ||
| 1 Adjust to pH 7.0 with 1 N NaOH. | |||||
Trace element solution SL-10 #
1000
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
HCl
(25%) |
10.0 | ml | ||
|
|
FeCl2 x 4 H2O | 1.5 | g | ||
|
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ZnCl2 | 70.0 | mg | ||
|
|
MnCl2 x 4 H2O | 100.0 | mg | ||
|
|
H3BO3 | 6.0 | mg | ||
|
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CoCl2 x 6 H2O | 190.0 | mg | ||
|
|
CuCl2 x 2 H2O | 2.0 | mg | ||
|
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NiCl2 x 6 H2O | 24.0 | mg | ||
|
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Na2MoO4 x 2 H2O | 36.0 | mg | ||
|
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Distilled water | 990.0 | ml | ||
| 1 First dissolve FeCl2 in the HCl, then dilute in water, add and dissolve the other salts. Finally make up to 1000.00 ml. | |||||
Selenite-tungstate solution #
1000
| Compound | Amount | Unit | |
|---|---|---|---|
|
|
NaOH | 0.5 | g |
|
|
Na2SeO3 x 5 H2O | 3.0 | mg |
|
|
Na2WO4 x 2 H2O | 4.0 | mg |
|
|
Distilled water | 1000.0 | ml |
Wolin's vitamin solution (10x) #
1000
| Compound | Amount | Unit | |
|---|---|---|---|
|
|
Biotin | 20 | mg |
|
|
Folic acid | 20 | mg |
|
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Pyridoxine hydrochloride | 100 | mg |
|
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Thiamine HCl | 50 | mg |
|
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Riboflavin | 50 | mg |
|
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Nicotinic acid | 50 | mg |
|
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Calcium D-(+)-pantothenate | 50 | mg |
|
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Vitamin B12 | 1 | mg |
|
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p-Aminobenzoic acid | 50 | mg |
|
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(DL)-alpha-Lipoic acid | 50 | mg |
|
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Distilled water | 1000 | ml |