Transformation

Materials
  1. Compotent cells (-80oC)
  2. Plasmid DNA or ligation products (make sure if the antibiotic resistant gene is ampicillin or others)
  3. LB agar plates with antibiotics
  4. LB broth
  5. Sterile glass beads
  6. Ice
  7. Water bath or heating block set at 37oC or 42oC
Procedure
  1. Put competent cells on ice for about 5 min to thaw.
  2. Add Plasmid DNA or ligation products no more than 15ul into the cell suspension and mix gently.
  3. Stay on ice for 30min with occasional swirling.
  4. Heat 2-5 min at 37oC or 0.5 min at 42oC.
  5. Put on ice for 1 min.
  6. Add LB to 500ul. Incubate at 37oC for 1 hr with agitation.
  7. Plate 50ul on ampicillin plates with different dilutions as needed and spread evenly by putting several glass beads on the surface and shaking. Discard beads.
  8. Incubate overnight at 37oC with dish upside down.
  9. Determine transformation efficiency and analyse colonies.

 

Hosted by www.Geocities.ws

1