Tris-Borate-EDTA (TBE) Buffer 5xStock Preparation
Updated 6/12/2015 07:53 PM
Introduction
Materials
- Consumables
- Boric acid
- Tris-base
- EDTA stock (pH 8.0)
- Disposables
- pipette tips
- weigh dishes/paper
- Equipment
- autoclave
- balance
- media bottle
- pipettor
Procedure
Component Composition
Table1
A A | B B | |
1 1 | Final Volume (mL)* | 1000 |
2 2 | EDTA (pH 8.0) Stock Concentration (M)* | 0.5 |
Table2
A A | B B | C C | |
1 1 | Component | Amount | Unit |
2 2 | Tris-base | 54 | g |
3 3 | Boric acid | 27.5 | g |
4 4 | EDTA (pH 8.0) | 20 | mL |
Preparation
- Add ~80% of the final volume of dH2O to a sterile media bottle.
Generally, the capacity of the bottle should be greater than (not equal to) the final volume of media to be prepared in order to prevent the media from boiling over during the autoclave step.
Ensure you have a volumetric flask measuring your desired final volume.
- Adjust the final volume and molar concentration in the table above and add the calculated amounts of tris-base, boric acid, and EDTA (pH 8.0) to the dH2O.
Solution likely will not completely dissolve until all components are present.
- Once completely dissolved, transfer the solution to a sterile volumetric flask and adjust the final volume using dH2O, then return it to the media bottle.
Ensure the solution is well mixed before returning it to the media bottle.
- Apply autoclave tape to the bottle and autoclave on liquids cycle.
CRITICALLiquids cycle.
- Store at room temperature.
Comments
How would one prepare a working buffer solution for the above stock solution you have just explained?
This protocol is for the preparation of Tris-Borate-EDTA (TBE) buffer at a 5x concentration.
The typical concentration of TBE when used as an agarose gel electrophoresis buffer is 0.5x, so this stock is effectively a 10x concentrated solution of standard electrophoresis buffer.
See: Agarose Gel Electrophoresis Sambrook, J. & Russell, D. W. (2001). Molecular Cloning: A Laboratory Manual, 3 edn. Cold Spring Harbor, N.Y.: Cold Spring Harbor Laboratory Press. pp. 5.8