Team:Columbia-Cooper-NYC/Gel Electrophoresis Setup

From 2012.igem.org


Gel Electrophoresis Setup and Run Protocol

Note: The protocol is for creating 60ml of 1% agar gel used to run small DNA sequences of several hundred

To run gels for larger sequences (using the corresponding ladder), create a .7% agar gel

  1. Measure .6g of ultra pure agar
  2. Suspend agar in 60ml of TAE buffer
  3. Place cap on bottle containing the solution
  4. Set microwave for 120 seconds, let solution heat in microwave for first 45 seconds
  5. Take out bottle and shake bottle and place back in microwave
  6. Repeat step 5 every 10-15 seconds there after
  7. After microwaving let solution stand to cool in hood Note: Do not let gel solidify
  8. While cooling, rinse the gel mold with deionized water and make sure rubber gaskets are air tight and in place
  9. Place well molds in place
  10. Add .8µl Ethiolium bromide to TAE/agar mixture
  11. Pour the mixture into mold and let it stand until hardens

To run the gels:

  1. Gently remove wells from hardened gel
  2. For every 5µl of DNA add 1µl of 6x loading dye Note: Need to add loading dye to each sample of DNA)
  3. Add 7-8µl of 2 log ladder in first column
  4. Place desired DNA for inspection in all the other columns
  5. Run gel at steady 150V

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