Gel electrophoresis

Requirements for each sample:

•   Elution buffer (0.5 M sodium acetate (pH 7) and 1 mM EDTA (pH 8))
•   95% ethanol
•   80% ethanol
•   TE buffer (10x) (pH 8.0)
•   Gel fragment containing the DNA

Procedure:

•   Excise the region of the gel containing the required DNA band using a scalpel.
•   Add elution buffer to the gel slice until the level of the buffer is a few mm above the level of the excised gel band.
•   Heat the solution in a water bath at 65ᵒ C until the agarose completely melts.
•  Fast-freeze by placing in a -80ᵒ C freezer for 10-15 minutes.
•   Immediately thaw the solution by centrifuging for 10 minutes.
•  The supernatant is transferred to a new tube.
•  Add elution buffer again to the pellet and repeat the steps 3-6.
•  Accumulate the supernatants and add an equal volume of 1-butanol.
•  Rock the mixture for 15 minutes to remove the EtBr completely from the gel.
•  Discard the supernatant and repeat the steps 8 and 9 2-3 times.

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