Bacteria containing the plasmid of interest are first cultured, then a sample is centrifuged in order to concentrate cellular material into a pellet in centrifuge tube and then resuspended in buffer P1.

Separately, a Buffer P2 -strong alkaline solution then added. The resulting mixture is left on ice for a few minutes. During this time, the detergent disrupts the cells membranes and allows the alkali into contact with both the chromosomal and plasmid DNA, which it denatures.

Finally, Buffer P3 is added. This acidifies the solution and allows the renaturing of plasmid DNA, but not chromosomal DNA, which is precipitated out of solution. A final centrifugation is carried out, and this time the pellet contains only debris and can be discarded. The plasmid-containing supernatant is carefully removed, Wash with ethanol and eluted for further analysis, such as gel electrophoresis, PCR, Sequencing etc.