Agarose gel electrophoresis

Pouring Agarose Gel

Purpose: To prepare gels for gel electrophoresis.

Materials:

• gel box, gel tray, comb (all in gel room)
• 1% agarose in TAE (premade, premelted bottles available in peltier incubator; next room over on the floor)
• EtBr (in gel room, upper right hand shelf)
• prewarmed glass bottle

Method:

1. Place gel tray in box to pour. The rubber ends of the tray should form a seal with the sides of the box.
2. Place comb of appropriate width, size, and number into niche at end of the tray.
3. Prewarm a small 100ml bottle (preferably with marks on the side indicating volume); setting it in the water bath for a few minutes should do it, or the incubator.
4. Pour 35ml of 1% agarose in 1xTAE for small gels, 70ml for large gels and place into prewarmed bottle. The marks on the side of the bottle should be a sufficient guide.
5. Add 0.2ul of EtBr per 35ml 1% agarose. EtBr is extremely carcinogenic - be careful with it. (Wear gloves; dispose of tips properly).
6. Swirl bottle to mix EtBr. Pour the mixture into the tray.
7. Let agarose set for ~15 minutes to solidify. After it has set a bit (enough so that you can walk with it), you can set it in the cold room to chill further and solidify completely, saving a few minutes.
8. When gel has solidified, remove the comb. Lift the gel box and rotate it 90 degrees, so that the ends of the gel are now exposed to either end of the box.
9. Add 1x TAE so that it fills the wells on either side of the gel, and covers the surface of the gel - the gel should be immersed in 1x TAE. (There are bottles of 1x TAE in the gel room. If they're empty, 1x TAE is above the sink in the main room.)

You're now ready to load DNA samples and run your gel.