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Team:Newcastle University/Protocols
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Newcastle University
GOLD MEDAL WINNER 2008
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Agarose Gel Electrophoresis
- Add 20ml 50 X TAE to 980ml H2O.
- Add 3g agarose powder to 300ml of the TAE-H2O (1 x TAE) solution.
- Microwave full power for 5 minutes or until solution is clear and agarose has dissolved.
- Leave to stand for 5-10 minutes. Put tape around sides of tray.
- Pour solution into tray, add 4ul of ethidium bromide and mix gently using the comb.
- Clean the comb (this prevents capillary action drawing the gel up between the comb teeth and making 'spikes' around the wells) and reinsert near one end of the tray.
- Leave to set 30-40 mins.
- Remove tape and comb, place tray in electrophoresis machine and pour 1 x TAE solution to just cover the gel.
- Combine 1ul of loading buffer for every 5ul sample being loaded and mix by pipetting up and down.
- Load marker and samples and run the gel.
N.B. We have found that a thin gel works best for loading small samples, but a thicker gel may be preferable if lots of sample needs to be loaded (for example if a fragment needs to be cut from the gel).
N.B. We have run gels at 100V for 20mins and 70V for 60mins. The latter setting/time is better for running larger fragments such as whole plasmids as a high voltage can cause shearing of the DNA.
