2-MINUTE MINIPREP METHOD FOR PLASMID DNA ISOLATION

An extremely rapid method, INSTA-PREPTM, has been developed to prepare plasmid DNA from 1 to 3 mL miniprep Escherichia coli bacterial cultur es. Direct extraction of plasmid DNA from E. coli bacterial cells is a chieved by a two-phase solution consisting of phenol-chloroform-isoamy l alcohol and water or buffer with efficient separation of the phases by centrifugation in the presence of the INSTA-PREP gel barrier materi al. Processing time, from E. coli culture to usable plasmid DNA, is tw o minutes or less per sample. Supercoiled plasmid DNA yields ranged fr om 3 to 10 mu g per mL of culture depending on plasmid copy number. Pl asmid DNAs prepared by INSTA-PREP were analyzed and are suitable for u se in molecular biology procedures including restriction digestion, li gation with T4 DNA ligase, bacterial transformation PCR, cultured cell transfection and T7 DNA polymer ase or thermostable DNA polymerase-me diated dideoxynucleotide sequencing.