Rapid purification of double-stranded DNA by triple-helix-mediated affinity capture.

A simple and rapid method for the preparation of highly pure plasmid DNA has been developed. The DNA is directly captured from bacterial cell lysates by formation of a triple-helical structure between the plasmid dsDNA and a 20-base biotinylated oligonucleotide attached to streptavidin-coated magnetic beads and then eluted from the beads in pH 9 buffer solution. No phenol extraction, ethanol precipitation, RNase digestion, or CsCl gradient centrifugation is required. A general purpose cloning vector, pHJ19, was constructed for this application from pUC19 DNA by insertion of a 40-base sequence suitable for triple-helix formation. The approach was also found suitable for the purification of lambda bacteriophage DNA.