350 .
THERMOCOCCUS MEDIUM (pH 6.0)
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Equipment needed:
Autoclave
Final pH:
6.0
Select a strain (optional):
ml
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
Medium J280 | 1000 | ml | ||
| 1 After reduction of the medium, readjust pH to 6.0 with 1.0 N H2SO4. | |||||
Main sol. J280 #
1020
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
KCl | 0.330 | g | ||
|
|
MgCl2 x 6 H2O | 2.800 | g | ||
|
|
MgSO4 x 7 H2O | 3.400 | g | ||
|
|
NH4Cl | 0.250 | g | ||
|
|
NaCl | 25.000 | g | ||
|
|
K2HPO4 | 0.300 | g | ||
|
|
Yeast extract
(BD-Difco) |
3.000 | g | ||
|
|
Tryptone
(BD-Difco) |
3.000 | g | ||
|
|
FeSO4 x 7 H2O | 0.025 | g | ||
|
|
NaBr | 10.000 | mg | ||
|
|
Trace minerals | 10.000 | ml | ||
|
|
Trace vitamins | 10.000 | ml | ||
|
|
Sulfur
(powder) |
10.000 | g | ||
|
|
Na2S x 9 H2O | 0.500 | g | ||
|
|
Resazurin | 1.000 | mg | ||
|
|
Distilled water | 1000.000 | ml | ||
| 1 Mix ingredients except sulfur and Na2S x 9H2O, adjust pH to 7.0-7.2 and autoclave under a N2 atmosphere. Steam sulfur for 3 hr on each of 3 successive days. Neutralize Na2S x 9H2O as a 5% solution and autoclave under a N2 gas atmosphere. Distribute the medium and sulfur into culture vessels under a N2 stream, and seal with butyl rubber stoppers. Prior to inoculation, add the sterile Na2S x 9H2O solution, and readjust pH to 7.0-7.2, if necessary. | |||||
| 2 Comment: | |||||
| 3 Most of Thermococcus strains will grow in this medium supplemented with 5.0 g/L maltodextrin and omitted sulfur. | |||||
Trace minerals #
1000
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
Nitrilotriacetic acid | 1.50 | g | ||
|
|
MgSO4 x 7 H2O | 3.00 | g | ||
|
|
MnSO4 x n H2O | 0.50 | g | ||
|
|
NaCl | 1.00 | g | ||
|
|
FeSO4 x 7 H2O | 0.10 | g | ||
|
|
CoSO4 x 7 H2O | 0.10 | g | ||
|
|
CaCl2 x 2 H2O | 0.10 | g | ||
|
|
ZnSO4 x 7 H2O | 0.10 | g | ||
|
|
CuSO4 x 5 H2O | 0.01 | g | ||
|
|
AlK(SO4)2 | 0.01 | g | ||
|
|
H3BO3 | 0.01 | g | ||
|
|
Na2MoO4 x 2 H2O | 0.01 | g | ||
|
|
Distilled water | 1000.00 | ml | ||
| 1 Dissolve nitrilotriacetic acid and adjust pH to 6.5 with KOH solution. Then proceed to add minerals. Adjust final pH to 7.0. | |||||
Trace vitamins #
1000
| Compound | Amount | Unit | |
|---|---|---|---|
|
|
Biotin | 2.0 | mg |
|
|
Folic acid | 2.0 | mg |
|
|
Pyridoxine hydrochloride | 10.0 | mg |
|
|
Thiamine HCl | 5.0 | mg |
|
|
Riboflavin | 5.0 | mg |
|
|
Nicotinic acid | 5.0 | mg |
|
|
Calcium pantothenate | 5.0 | mg |
|
|
Vitamin B12 | 0.1 | mg |
|
|
p-Aminobenzoic acid | 5.0 | mg |
|
|
Lipoic acid | 5.0 | mg |
|
|
Distilled water | 1000.0 | ml |