DNA sequencing using capillary array electrophoresis

Abstract

A DNA sequencing method is presented that utilizes capillary array electrophoresis, two-color fluorescence detection, and a two-dye labeling protocol. Sanger DNA sequencing fragments are separated on an array of capillaries and detected on-column using a two-color, laser-excited, confocal-fluorescence scanner. The four sets of DNA sequencing fragments are separated in a single capillary and then distinguished by using a binary coding scheme where each fragment set is labeled with a characteristic ratio of two dye-labeled primers. Since only two dye-labeled primers are required, it is possible to select dyes that have identical mobility shifts. It is also shown that the ratio of the signal in the two detection channels provides a reliable identification of the sequencing fragment. DNA sequencing results on a 25-capillary array are presented.