Ultra-thin-layer agarose gel electrophoresis - I. Effect of the gel concentration and temperature on the separation of DNA fragments

A novel, rapid and efficient separation method is described for the analysi s of double stranded (ds) DNA fragments in the form of horizontal ultra-thi n-layer agarose gel electrophoresis. This separation technique combines the multilane, high-throughput separation format of agarose slab gel electroph oresis with the excellent performance of capillary electrophoresis. The ele ctrophoretic separation of the fluorophore (Cy5)-labeled dsDNA molecules we re imaged in real time by a scanning laser-induced fluorescence/avalanche p hotodiode detection system. Effects of the gel concentration (Ferguson plot ) and separation temperature (Arrhenius plot) on the migration characterist ics of the DNA fragments are discussed. An important genotyping application is also shown by characterizing the polymorphic region (2X or 4x48 base pa ir repeats) of the dopamine D-4 receptor gene (D4DR, exon III region) for t en individuals, using PCR technology with Cy5-labeled primers and ultra-thi n-layer agarose gel electrophoresis. (C) 1999 Elsevier Science B.V. All rig hts reserved.