Plasmid DNA Preparation Isolation of plasmid DNA from bacterial cultures may be the most ubiquitous molecular biology technique around. Among other applications, it is essential for analysis of recombinant clones. Typically miniprep procedures are used for rapid screening or rapidly gathering sequence information from cloning experiments. The first step in any plasmid prep is crude lysate preparation, and most kits supply the necessary reagents. However, some companies sell columns or resins separately, allowing users to prepare their own lysis reagents.
Most kits utilize the alkaline lysis method.
At high pH and in the presence of SDS, cells are lysed and both genomic and plasmid DNA are released and denatured. Bacterial cell wall material and most of the cellular proteins are also denatured during alkaline lysis. When the lysate is neutralized with potassium acetate, the SDS precipitates and the potassium salt and most of the plasmid DNA go into solution. Genomic DNA remains associated with the insoluble cellular debris. DNA is often the finishing touch.