Validation of Double Gradient Denaturing Gradient Gel Electrophoresis through Multigenic Retrospective Analysis

Among established techniques for the identification of either known or new mutations, denaturing gradient gel electrophoresis (DGGE) is one of the most effective. However, conventional DGGE is affected by major drawbacks that limit its routine application: the differ-ent denaturant gradient ranges and migration times required for different DNA fragments. We developed a modified version of DGGE for high-throughput muta-tional analysis, double gradient DGGE (DG-DGGE), by superimposing a porous gradient over the denaturant gradient, which maintains the zone-sharpening effect even during lengthy analyses. Because of this innova-tion, DG-DGGE achieves the double goals of retaining full effectiveness in the detection of mutations while allowing identical run time conditions for all fragment