Team:TU Delft/protocols/buffers stock solutions
From 2010.igem.org
(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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- | + | [[*]] Add agar in case a solid medium is required. | |
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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- | + | [[*]] 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: | 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. | ||
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- | '''TBE Buffer''' | + | '''10x TBE Buffer''' |
- | For 1 liter, dissolve in | + | For 1 liter, dissolve in 800 mL H2O: |
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- | Adjust to | + | Adjust volume to 1 liter with H2O |
Latest revision as of 18:45, 12 September 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.
10x TBE Buffer
For 1 liter, dissolve in 800 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 volume to 1 liter with H2O