Team:Valencia/Gel Electrophoresis

From 2012.igem.org



Agarose Gel Electrophoresis


Reagents and Materials

  • 1X TAE
  • Graduated Cylinder
  • 125 mL flask
  • Agarose
  • Gel Pouring Tray
  • Tape
  • Gel rig
  • SYBR Safe


Protocol

  1. Measure out 100mL of buffer.
  2. Transfer buffer to 125 mL flask.
  3. Weigh out enough agarose to make a 1% gel (in our case 1.0 g of agarose was the right amount).
  4. Transfer agarose to 125mL flask.
  5. Melt agarose in microwave until solution is almose boiling, stirring every 15-20 seconds (should be around 2 minutes).
  6. Allow agarose to cool (do not let it cool to the point where it is hard).
  7. Add 4 uL of SYBR Safe to the cooling agarose.
  8. Assemble the gel pouring apparatus by inserting gate into slots.
  9. Allow gel to cool until flask can be handled comfortably.
  10. Place comb in the gel rig.
  11. Pour agarose into gel tray.
  12. Allow to solidify. While the gel is solidifying prepare the samples. Add your sample and 1 uL 10x Loading Dye, 4 uL of DNA and 5 uL of water.
  13. Pour 1X TAE over gel so that gel is covered by a 3-5mm buffer.
  14. Load samples into lane (Don't forget to load a 1kb+ ladder into one of the lanes).
  15. Hook electrodes to gel apparatus.
  16. Run the apparatus at 100V for 30 - 45 minutes (make sure to watch that the dye does not run off the gel
  17. Visualize the gel and record the results.

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