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DNA extraction using Chelex 100

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New Protocol

We tend to use the protocol made available online here: DNA extraction from buccal cells.

In summary:

  1. Mix a pinch of salt with about 100mL water (about a cappucino cup)
  2. Take a sip and swish it around in your mouth; chewing on your cheeks can improve results
  3. Expel 10mL of the result into a 15mL propylene tube, do this over the sink for better hygiene
  4. Place the tube into the centrifuge with appropriate counterweight and spin for 10min
    • At this point you may want to start setting up the hob plate to get some water boiling
  5. You should see a pellet at the bottom of the tube. These are your cells: decant the supernatant (liquid on top) entirely
  6. Use a clean pasteur pipette to transfer 0.5mL (500μL) of 5% chelex 100 into the tube (try and get 50% beads and 50% clear liquid)
  7. Pipette up and down to mix until there are no visible cell lumps left
  8. Close the tube and place it in a beaker of boiling water for 10min to lyse the cells
    • Try to avoid the tube touching the bottom of the beaker (because it may be hotter than 100°C and damage DNA)
  9. Use a pasteur pipette (can be the same as above) to transfer the contents into a 1.5mL Eppendorf tube
    • It's no problem if you can't get the beads, just make sure you get as much of the liquid as possible
  10. Spin in the small centrifuge with appropriate counterweight at max speed for 30-60s
  11. Use the micropipettor to transfer 200μL of the supernatant to a new tube. Make sure you take nothing from the pellet
    • The pellet at the bottom contains protein and lipids, which you want to avoid. The DNA is in the supernatant
  12. Store at -20°C

Old Protocol

  • Fill half a tube (500ul) with 5% chelex 100 (try and get 50% beads and 50% clear liquid)
  • Scrape off some cheek cells into the end of a pipette tip, add this and some saliva to the tube and mix
  • Cover with a drop of paraffin Incubate at 56°C for 30 mins, then at 94°C for 10 mins
  • Centrifuge briefly and take supernatant
  • Store at -20°C