Transforming E. coli Cells by Electroporation

This procedure uses the TOP10 Electrocomp™ E. coli cells (but is identical in any other standard electrocomp cell type).

  1. Thaw the electrocompetent cells on ice.
  2. To the electrocompetent cells, add 1-3 μl of DNA (<100 ng of DNA).
  3. Mix by gently flicking the tube containing the electrocompetent cell + DNA mixture. Let the mixture sit on ice for 1-10 minutes. (Erik Quandt says this is not necessary)
  4. Pipette the mixture into a chilled cuvette, making sure that the mixture is at the bottom of the cuvette by gently tapping the cuvette on a flat surface.
    • Be sure to wipe any condensation off the sides of the cuvette before electroporation.
  5. Place the cuvette in the pulser and press the "Pulse" button.
    • For the TOP10 Electrocomp™ E. coli cells, the Ec l setting is fine.
  6. After electroporation, add 500-1000 μl of SOC/LB to the cuvette to recover the cells.
  7. Transfer the mixture to a 1.5 mL microcentrifuge tube.
  8. Incubate for ~30-60 minutes at 37°C or other appropriate temperaturein a shaking incubator.
    • Be sure to place the tube on its side so the transformed cells will grow properly.
  9. Plate the cells (~ 50 μl) on an LB plate containing the appropriate antibiotic.
  10. Incubate overnight at 37°C or other appropriate temperature.

-- Main.SeanLeonard - 15 Dec 2016

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Topic revision: r1 - 15 Dec 2016 - 20:51:53 - Main.SeanLeonard
 
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