Compound | Amount | Unit | |||
---|---|---|---|---|---|
|
Solution A | 46.0 | ml | ||
|
Solution B | 2.6 | ml | ||
|
Solution C | 5.0 | ml | ||
|
Solution D | 45.0 | ml | ||
|
Solution E | 2.6 | ml | ||
|
Solution F | 0.1 | ml | ||
|
Solution G | 1.0 | ml | ||
1 Adjust pH to 6.0. | |||||
2 Mix solution D, C and E in the cotton-stoppered Erlenmeyer flask. Bubble with CO2 in an ice bath under sterile conditions (5-10 min). | |||||
3 Fill 50 ml of the mixed solution (C,D,E) into each 100 ml bottle of solution A (for a total volume of approx. 96 ml). | |||||
4 Before using, add 2,6 ml solution B (Na2S x 9 H2O), 0.1 ml solution F (Vitamin B12) and 1 ml of solution G (2,5% Ammonium-acetate and Magnesium-acetate) to each 100ml bottle. | |||||
5 Adjust the pH with filter-sterilised 1M Na2CO3 to 7.1-7.3. | |||||
6 Fill in sterile, N2 gassed screw-cap tubes under N2 gas. | |||||
7 For the growth of DSM 11081 feed the culture with neutralized sulfide feeding solution regularly (approx. every 3 days). 0,5-1 ml/ 100 mL |
Compound | Amount | Unit | |||
---|---|---|---|---|---|
|
CaCl2 x 2 H2O | 0.25 | g | ||
|
Distilled water | 460.00 | ml | ||
1 Fill 10x 46ml in 100 ml screw-cap bottles. Bubble with N2/CO2 and autoclave at 121°C for 15 min. |
Compound | Amount | Unit | |||
---|---|---|---|---|---|
|
Na2S x 9 H2O | 2 | g | ||
|
Distilled water | 135 | ml | ||
1 Prepare in a screw-cap bottle, bubble with N2 to replace air, close tightly and autoclave. |
Compound | Amount | Unit | |||
---|---|---|---|---|---|
|
Resazurin
(0,1%) |
0.5 | ml | ||
|
Distilled water | 450.0 | ml | ||
1 Autoclave in a cotton-stoppered Erlenmeyer flask with an outlet tube for medium, connected to a glass outlet at the bottom of the vessel and has, at the other end, a silicon rubber tube with a pinch cock and a bell for aseptic dispensing of the medium into bottles. Cool to room temperature under an atmosphere of N2/CO2 in an ice bath (10-15 min). |
Compound | Amount | Unit | |||
---|---|---|---|---|---|
|
Ammonium chloride | 0.40 | g | ||
|
MgSO4 x 7 H2O | 0.50 | g | ||
|
KCl | 0.34 | g | ||
|
KH2PO4 | 0.34 | g | ||
|
Trace element solution SL-12 B | 1.00 | ml | ||
|
Distilled water | 25.00 | ml | ||
1 Filter sterilize into sterile, gas-tight, 100 ml screw-cap bottle. |
Compound | Amount | Unit | |||
---|---|---|---|---|---|
|
Ammonium acetate | 2.5 | g | ||
|
Magnesium acetate | 2.5 | g | ||
|
Distilled water | 100.0 | ml | ||
1 Autoclave at 121°C for 15 min. |
Compound | Amount | Unit | |
---|---|---|---|
|
Distilled water | 1000.0 | ml |
|
Na2-EDTA | 3.0 | g |
|
FeSO4 x 7 H2O | 1.1 | g |
|
CoCl2 x 6 H2O | 190.0 | mg |
|
MnCl2 x 2 H2O | 50.0 | mg |
|
ZnCl2 | 42.0 | mg |
|
NiCl2 x 6 H2O | 24.0 | mg |
|
Na2MoO4 x 2 H2O | 18.0 | mg |
|
H3BO3 | 300.0 | mg |
|
CuCl2 x 2 H2O | 2.0 | mg |
Compound | Amount | Unit | |||
---|---|---|---|---|---|
|
Na2S x 9 H2O | 3 | g | ||
|
Distilled water | 100 | ml | ||
1 The sulfide solution is prepared in a 250 ml screw-capped bottle with a butyl rubber septum and a magnetic stirrer. The solution is bubbled with nitrogen gas, closed and autoclaved for 15 min. at 121°C. After cooling to room temperature the pH is adjusted to about 7.0 by adding of sterile 2 M H2SO4 drop-wise with a syringe without opening the bottle. |