Direct solid phase sequencing of genomic and plasmid DNA using magnetic beads as solid support

Abstract

Approaches to direct solid phase sequencing of genomic and plasmid DNA have been developed using magnetic beads, coated with steptavidin, as solid support. The DNA is immobilized through selective incorporation of biotin into one of the strands. A single stranded template, suitable for sequencing, is obtained through strand-specific elution. Using this concept, in vitro amplified plasmid DNA and chromosomal DNA were sequenced directly from single colonies. The solid approach ensures that the amplification and the sequencing reactions can be performed under optimal conditions. The system was found to be suitable for sequencing using both isotope- and fluorescent-labelled primers.