1248 .
MINERAL SALTS MEDIUM WITH TCE
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Equipment needed:
Anaerobic workbench
Autoclave
Filter
Final pH:
7.1
Select a strain (optional):
ml
| Compound | Amount | Unit | |||
|---|---|---|---|---|---|
|
|
NaCl | 1.000 | g | ||
|
|
MgCl2 x 6 H2O | 0.500 | g | ||
|
|
KH2PO4 | 0.200 | g | ||
|
|
NH4Cl | 0.300 | g | ||
|
|
KCl | 0.300 | g | ||
|
|
CaCl2 x 2 H2O | 0.015 | g | ||
|
|
FeCl2 solution | 1.000 | ml | ||
|
|
Trace element solution | 1.000 | ml | ||
|
|
Selenite-tungstate solution | 1.000 | ml | ||
|
|
Sodium formate | 0.340 | g | ||
|
|
Sodium acetate | 0.410 | g | ||
|
|
Resazurin | 0.250 | mg | ||
|
|
Distilled water | 1000.000 | ml | ||
| 1 Mix components thoroughly, bring to a boil and cool down under a N2-CO2 (4:1, v/v) gas mixture. Add the following solutions: | |||||
|
|
NaHCO3
(8%) |
30.000 | ml | ||
|
|
L-Cysteine HCl x H2O
(5%) |
6.000 | ml | ||
|
|
Na2S x 9 H2O
(5%) |
10.000 | ml | ||
| 2 Adjust pH of the medium to 7.0-7.2, if necessary. Dispense the medium into culture vessels (e.g., 100 ml in 120 ml-serum bottles) under the same gas mixture and seal with butyl rubber stoppers or Teflon-lined stoppers. Then, anaerobically add per liter the following solution: | |||||
|
|
Tetrachloroethene | 60.000 | μl | ||
| 3 Finally, add per liter the following solution from an anaerobic stock (filter-sterilized): | |||||
|
|
Trace vitamins | 10.000 | ml | ||
| 4 Dissolve tetrachloroethene with ultrasound. Autoclave in a closed basket. Do not remove the culture vessels from autoclave before temperature of the media has reached below 70C. | |||||
FeCl2 solution #
1000
| Compound | Amount | Unit | |
|---|---|---|---|
|
|
HCl
(7.7 M, 25%) |
10.0 | ml |
|
|
FeCl2 x 4 H2O | 1.5 | g |
|
|
Distilled water | 990.0 | ml |
Trace element solution #
1000
| Compound | Amount | Unit | |
|---|---|---|---|
|
|
ZnCl2 | 70 | mg |
|
|
MnCl2 x 4 H2O | 100 | mg |
|
|
H3BO3 | 6 | mg |
|
|
CoCl2 x 6 H2O | 190 | mg |
|
|
CuCl2 x 2 H2O | 2 | mg |
|
|
NiCl2 x 6 H2O | 24 | mg |
|
|
Na2MoO4 x 2 H2O | 36 | mg |
|
|
Distilled water | 1000 | ml |
Selenite-tungstate solution #
1000
| Compound | Amount | Unit | |
|---|---|---|---|
|
|
NaOH | 0.4 | g |
|
|
Na2SeO3 x 5 H2O | 6.0 | mg |
|
|
Na2WO4 x 2 H2O | 8.0 | mg |
|
|
Distilled water | 1000.0 | ml |
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 |