The Rapid Boiling Method for Small-Scale Preparation of Plasmid DNA

Abstract

The classic alternative to the alkaline lysis method for plasmid DNA preparation (1; see Chapters 31 and 33) is that of Holmes and Quigley (2), and is commonly known as the rapid boiling method. This method is based on exactly the same principles as the alkaline lysis method. The cells are partially lysed, allowing plasmid molecules to escape, while most of the genomic DNA is trapped in the cell debris, which is then spun out. The remaining genomic DNA is removed by a denaturing step, in this case, high temperature rather than high pH, followed by rapid reannealing. As a consequence of their supercoiling, plasmid molecules are able to reassociate rapidly in these conditions, whereas genomic DNA remains denatured and is lost on the subsequent ethanol- precipitation step.