Melbourne/Loading a DNA gel

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Revision as of 07:29, 8 July 2007

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• Applications:
  1. Diagnostic
  2. Gel Purification

• Time to complete protocol:
  • Lab time: 15min.
  • Waiting time:30min-2hrs
• Approximate cost of materials: $

Method from primary and secondary reagents

Primary & secondary Reagents Required including controls

• 0.8% agarose gel
• 1x buffer (same as used to prepare the gel, TAE gives sleaner bands and is better for gel purification than TBE)
• DNA ladder with dye added (stock of 1kb ladder diluted 1 in 21 in loading dye is in the -20 freezer in iGEM box 1)
• DNA to be run with loading dye already added.

Method including controls

1. Prepare gel and place in electrophoresis tank with the wells towards the black/negative electrode.
2. Cover with 1x buffer such that there is a 1-2mm layer of buffer covering the gel and the wells are full.
3. In lane one load 20uL of the ladder DNA
4. The amount of DNA loaded varies depending on concentration, for the diagnostic digests load 20uL.
   • Ensure that you write down the loading order.
5. If you have sufficient lanes load another 20uL of the ladder in the final lane for greater ease in interpretation.
6. Place the cover on the electrophoeresis tank. Make sure that you connect black to black and red to red and plug the cables into the power supply

Equipement Required

References

• PhillipDodson 03:48, 1 July 2007 (EDT):Secondary Reagent Template 1/July/2007