Alm:Agarose gel electrophoresis
(Difference between revisions)
Revision as of 15:52, 20 September 2007
How to use gel electrophoresis to separate, measure and visualize DNA pieces.
- 6X loading dye
- TAE + agarose 1%
- 1X TAE (~200ml)
- gel box (casting tray optional) and power supply
- Ethidium Bromide staining solution
- used TAE destaining solution
- UV box/gel imager
- Microwave TAE agarose 1% to melt. Pour 30 ml for a small gel into a small flask/beaker to cool a bit, then pour into the gel box with the correct comb for your size/number of samples.
- Add 6x loading dye to your samples.
- Make sure the red electrode is at the opposite end from the wells. You can check the progress of the DNA with the loading dyes to check the direc
- Set the time and settings on the power supply by pressing Mode to change fields. (typical settings for DNA: 90V constant voltage, 45 minutes)
- Slide on the lid and start the power supply.
Staining and visualization
- Wear blue/purple nitrile gloves when handling the items at the staining station and the gel imager.
- Place the gel in the plastic box.
- Stain for 1 hour in EtBr solution (concentration??).
- Pour the EtBr back into the light-proof bottle.
- Destain with used TAE for 2 minutes.
- Pour the TAE back into the bottle.
- Image the gel on a UV light box or the computerized gel imager in the Polz lab.
- Print out the image for your notebook, annotating the lanes, marker, and bands.
- Note agreement with/variation from expected result.
- Arne Materna
- Sean Clarke