Step by Step:
Steps on how to set up the box:
- Obtain, box, tray, comb, electrodes and a voltage source
- Turn tray 90o so the rubber edges are stuck to the sides of the box
- Place comb in proper holder, so the wells are formed at the top of the gel
- Hook up electrodes to the proper outlets on the voltage source
Steps to make an agarose gel:
- Weigh out allotted amount of agarose, 0.5g for a 1% gel and 1.5g for a 3% gel
- Place the agarose in a Erlynmeyer flask and add 50mL of TAE buffer which contains, Tris base, acetic acid and EDTA
- Heat solution in microwave for 30 seconds and then in 15 second intervals until the agarose is fully dissolved and the solution is clear
- Let flask cool to roughly 60 degrees celcius
- Add a dye such as SYBR Safe or Ethidium Bromide for visualization, ensure gloves are worn at all times since the dyes are carcinogenic or mutagenic
- Pour the agarose gel into the prepared tray. If bubbles appear pop them with a pipette tip
- Allow gel to solidify for roughly 20 minutes or until it turn opaque
- Remove the comb and turn the tray so the wells are towards the black electrode
- Fill the box with running buffer
- Fill the wells with the samples, and allow to run anywhere between 30 min to an hour
In the video Ethidium Bromide was used but in our experiment SYBR Safe was used as the dye to visualize the nucleic acid bands.https://www.youtube.com/watch?v=wXiiTW3pflM