5 Minute Transformation of E. coli

1. Prewarm LB+antibiotic plates at 37C.

2. Remove competent cells from –80C freezer and thaw on ice.

3. As soon as cells are thawed, immediately add 1-10ng DNA per 100ul of cells.

4. Incubate on ice for 5 minutes.

5. Spread cells on plates (100ul on one, 20-50ul on another).

6. Incubate at 37C overnight.Competent Cells

We currently have three types of competent cells:

1. DH5? - general use

2. XL10-Gold – large or ligated DNA

3. XL1-Blue – unmethylated DNA; unlimited DNA or subcloning