Team:Freiburg/Protocols/Agarose-Gel

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Agarose gels

Protocol for casting and loading of a DNA fractionation gel

material: TAE-Puffer (0.5x), Agarose, microwave
duration: 90 min

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Casting

1. pour 0.7% or 1% agarose solution into the casting chamber depending on the size of the DNA fragments (the higher concentration, the smaller the fragment)
2. add 4 µL of GelRed solution and distribute it by using the gel comb
3. fix comb in the gel chamber
4. let polymerize
5. Fill gel chamber with TAE-Buffer
6. remove comb (beware for cracks)

CAUTION: hot! ⇒ use gloves for handling
CAUTION: GelRed ist ein strong mutagen!!

Gel loading

1. add 4 µL of ladder into one to two gel pockets (depending on the size of the gel)
2. add running buffer to the DNA samples (6x concentrated)
3. load DNA samples in the gel pockets (volume depending on the size of the gel)
4. apply voltage → 120V
5. Running time depends on the desired quality of band splitting

Cast a new gel

1. weight agarose (depending on concentration) and add the corresponding amount of water
2. heat in the microwave until the solution is has cleared up. Pan from time to time to prevent boiling retardation

CAUTION: Beware boiling retardation and NEVER close the cap
CAUTION: hot! ⇒ use gloves for panning

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