Rapid DNA sequencing by horizontal ultrathin gel electrophoresis

Abstract

A horizontal polyacrylamide gel electrophoresis apparatus has been developed that decreases the time required to separate the DNA fragments produced in enzymatic sequencing reactions. The configuration of this apparatus and the use of circulating coolant directly under the glass plates result in heat exchange that is approximately nine times more efficient than passive thermal transfer methods commonly used. Bubble-free gels as thin as 25μm can be routinely cast on this device. The application to these ultrathin gels of electric fields up to 250 volts/cm permits the rapid separation of multiple DNA sequencing reactions in parallel. When used in conjunction with ³²P-based autoradiography, the DNA bands appear substantially sharper than those obtained in conventional electrophoresis. This increased sharpness permits shorter autoradiographic exposure times and longer sequence reads.