TAE
From Protists
Contents |
Background
- Tris acetate with EDTA (TAE) Buffer:
- is best for electrophoresis of large (>20kb) DNA
- is best for applications requiring high resolution
- its low ionic strength and low buffering capacity means that recirculation may be needed for extended run times (>4hrs) or in the largest-sized gels (B2/B3)
- TAE is a more expensive and difficult buffer to make (compared to TBE) so we use it for special gels
- Gels with large bands to increase their resolution
- Gels from which we will extract bands of DNA and extract them for further downstream processing
TAE Solutions
50x TAE
- Measure and pour ~700mL dH2O from the FIX graduated cylinder into a FIX large flask
- Graduated cylinder under sink attached to the Nanodrop/Gel counter
- Large flask in cabinets above FIX heating/stir plate
- Add 18.61g Na2EDTA x 2H20 (MW 372.24)
- In chemical shelves alphabetically as "EDTA disodium salt, dihydrate, crystal"
- Add 57.1mL Glacial Acetic Acid (CH3COOH) in hood with blower on (Use Gloves and Goggles!)
- Glacial Acetic Acid is in the Acid Storage container under the hood across from the Nanodrop/Gel Image counter
- Add 242.0g Tris Base (or TRIZMA base)
- Put a FIX stir bar in the large flask and cover lid with aluminum foil so dirt/dust doesn't get in mixture
- Put flask on the stir plate and allow to stir until all Tris Base and Na2EDTA is dissolved
- Pour mixture back into the FIX graduated cylinder and add dH2O up to 1000mL
- Pour mixture back into the FIX large flask and mix well
- Makes a total of 1L of 50x TAE buffer
- Store in 1L media bottles with orange caps under sink attached to the Nanodrop/Gel counter
Dilute the 50x TAE buffer to 1X TAE before using in gels or other molecular work
1X TAE
- Get the 50x TAE Buffer from under the sink by the gel rigs.
- Combine 100 mL of 50x TAE with 4.9 L of distilled H2O in the "1X TAE" carboy.
- Close the top tightly and mix by shaking.
- Makes a total of 5L of 1x TAE Buffer
IF THE 50x TAE SINK SUPPLY RUNS OUT OR LOW, MAKE MORE
