Team:TU Dresden/Notebook/Protocols/CellsForElectroporation


HiYield Presto Mini Plasmid Kit

  1. The day before start a 1 mL LB culture with the respective antibiotic when necessary and incubate at 30 °C (e.g. with pSC101BADaβγA) or 37 °C with 950 rpm o/n .
  2. Place a bottle of sterile water at least for 2 hours on ice.
  3. Start a 1.4 mL LB culture with the respective antibiotic, when necessary with 30 μL of the o/n culture and incubate at 30 °C (e.g. with pSC101BADaβγA) or 37 °C with 950 for 2 hours.
  4. Put the cells on ice.
  5. Keep the cells on ice whenever possible.
  6. Spin the cells down at 8,000 rpm for 0.5 minutes in a cooling centrifuge at 2 °C.
  7. Discard the supernatant.
  8. Resuspend the pellet in 1 mL of ice cold water with a pipette.
  9. Spin the cells down at 8,400 rpm for 0.5 minutes at 2 °C.
  10. Discard the supernatant.
  11. Resuspend the pellet in 1 mL of ice cold water with a pipette.
  12. Spin the cells down at 8,800 rpm for 0.5 minutes at 2 °C.
  13. Discard as much supernatant as possible.
  14. The remaining solution should be about 30 μL, resuspend the cells in it.
  15. Use the cells immediately.