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Team:TU Delft/protocols/buffers stock solutions
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(New page: ==Buffers and Stock solutions== '''Luria-bertani (LB)''' For 1 liter dissolve in H2O: {| style="color:black; background-color:white;" cellpadding="5" cellspacing="0" border="1" |Compoun...) |
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Autoclave on a liquid cycle. Once broth reaches room temperature, store in cold room. | Autoclave on a liquid cycle. Once broth reaches room temperature, store in cold room. | ||
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Once broth reaches room temperature, add to the broth in aseptic conditions the following amount of each solution: | Once broth reaches room temperature, add to the broth in aseptic conditions the following amount of each solution: | ||
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| - | + | [[*]] For M9-modified medium no glucose solution is added, instead 10 mL of filtered hydrocarbon/ hydrocarbon mixture are added per liter of medium. Be careful when adding the hydrocarbon/hydrocarbon mixture. | |
NOTE: Autoclave the solutions 1 M MgSO4, 1M CaCl2, 20% glucose (w/v) (EXCEPT thiamine-HCl 1M) on a liquid cycle. Filter the solution Thiamine-HCl 1M using a 0.22 μm membrane, store the solution in a cold room when is not required. | NOTE: Autoclave the solutions 1 M MgSO4, 1M CaCl2, 20% glucose (w/v) (EXCEPT thiamine-HCl 1M) on a liquid cycle. Filter the solution Thiamine-HCl 1M using a 0.22 μm membrane, store the solution in a cold room when is not required. | ||
Revision as of 17:56, 9 July 2010
Buffers and Stock solutions
Luria-bertani (LB)
For 1 liter dissolve in H2O:
| Compound/Ingredient | Amount required (1L of medium) |
| Tryptone | 10 g |
| Yeast extract | 5 g |
| NaCl | 10 g |
| Agar+ | 15 g |
* Add agar in case a solid medium is required.
Autoclave on a liquid cycle. Once broth reaches room temperature, store in cold room.
M9
For 1 liter dissolve in H2O:
| Compound/Ingredient | Amount required (1L of medium) |
| Na2HPO4 | 6 g |
| KH2PO4 | 3 g |
| NaCl | 0.5 g |
| (NH4)Cl | 1 g |
| Agar* | 15 g |
* Add agar in case a solid medium is required.
Once broth reaches room temperature, add to the broth in aseptic conditions the following amount of each solution:
| Solution | Amount(1L of medium) |
| 1 M MgSO4 | 2 mL |
| 1 M CaCl2 | 0.1 mL |
| 20% Glucose (w/v)* | 10 mL |
| 1M Thiamine-HCl | 1 mL |
* For M9-modified medium no glucose solution is added, instead 10 mL of filtered hydrocarbon/ hydrocarbon mixture are added per liter of medium. Be careful when adding the hydrocarbon/hydrocarbon mixture.
NOTE: Autoclave the solutions 1 M MgSO4, 1M CaCl2, 20% glucose (w/v) (EXCEPT thiamine-HCl 1M) on a liquid cycle. Filter the solution Thiamine-HCl 1M using a 0.22 μm membrane, store the solution in a cold room when is not required.
Store in cold room if the medium is not used.
Antibiotics
| Antibiotic | Final concentration in the medium |
| Ampicillin | 100 μg/mL |
| Kanamycin | 50 μg/mL |
| Chlorampenicol | 35 μg/mL |
| Tetracycline | 15 μg/mL |
All stock solutions must be kept at -20 ºC and used under aseptic conditions.
TBE Buffer
For 1 liter, dissolve in 950 mL H2O:
| Compound/Ingredient | Amount required (1L of medium) |
| Tris | 54 g |
| Boric Acid | 27.5 g |
| 0.5 M EDTA | 4.65 g or 20 mL |
Adjust to pH 8.0
