High-throughput method for isolating plasmid DNA with reduced lipopolysaccharide content

Isolating plasmid DNA from bacteria is a fundamental step in molecular biol ogy. It is often accomplished by an alkaline lysis of bacteria and the subs equent adsorption of nucleic acids to silica oxide in the presence of chaot ropic substances. Here we show that the addition of such chaotropic reagent s is not required for the efficient DNA isolation with silica oxide. This s urprising finding allowed us to purify plasmid DNA with significantly less lipopolysaccharides (LPS), which is otherwise a common bacterial contaminan t of silica oxide-isolated DNA and inhibits subsequent applications. In add ition, we have implemented a precipitation step that altogether leads to a reduction of the LPS content by a:factor of 900 relative to published metho ds. Our novel protocol facilitates an inexpensive high-throughput analysis of pure plasmids in a 96-well format without the addition of hazardous reag ents.