Heat Shock Transformation of E. coli

This protocol can be used to transform chemically competent (i.e. from CaCl2) with a miniprepped plasmid or a ligation product.

Note: Never vortex competent cells. Mix cells by gentle shaking.

1. Thaw competent cells on ice. These can be prepared using the CaCl2 protocol.
2. Place 20 ul of cells in a pre-chilled Eppendorf tube.
   • For an Intact Vector: Add 0.5 ul or less to the chilled cells
   • For a Ligation Product: Add 2-3 ul to the chilled cells.
3. Mix gently by flicking the tube.
4. Chill on ice for 10 minutes. This step is optional, but can improve yields when transforming a ligation product.
5. Heat shock at 42 °C for 30 seconds.
6. Return to ice for 2 minutes.
7. Add 200 ul LB medium and recover the cells by shaking at 37 °C.
   Another rich medium can substitute for the recovery.
   The recovery time varies with the antibiotic selection.
   Ampicillin: 15-30 minutes
   Kanamycin or Spectinomycin: 30-60 minutes
   Chloramphenicol: 60-120 minutes
8. Plate out the cells on selective LB.
   Use glass beads to spread the cells.
   The volume of cells plated depends on what is being transformed.
   
   • For an Intact Vector: High transformation efficiencies are expected. Plating out 10 ul of recovered cells should produce many colonies.
   • For a Ligation Product: Lower transformation efficiencies are expected. Therefore you can plate the entire 200 ul volume of recovered cells.
   Note: 200 ul is the maximum volume of liquid that an LB plate can absorb.
9. Incubate at 37 °C. Transformants should appear within 12 hrs.