Gel-Capillary Electrophoresis Analysis of Oligonucleotides

Abstract

Gel-capillary electrophoresis is a new alternative to traditional electrophoretic techniques for the analysis of oligonucleotides (1–5). The advantages are dramatically decreased analysis time, excellent resolution, in-capillary detection, reduced sample quantities, and automation. Capillary electrophoresis (CE) is already established as an important analytical tool for other biomolecules, such as proteins, peptides, and high-mol-wt, double-stranded nucleic acids (6,7). Now the CE method has been extended to single-stranded oligonucleotides using special polymeric, gel-filled capillaries. In a gel-matrix capillary, as with polyacrylamide slab gel electrophoresis (PAGE), DNA separates primarily, and predictably, on the ratio of mass-to-charge under the influence of an electric field. The same elution pattern of small oligonucleotides followed by the largest, usually the product, is obtained. Single base resolution can often be attained beyond 100 bases. The analysis, called an electropherogram, is quantitative. It can be displayed, stored, integrated, and printed like HPLC chromatograms. Slab gels (PAGE), on the other hand, are visualized at a single time point in the analysis, do not easily yield to integrative quantitation, and are thus a subjective method of analysis. The resolution of Gel CE surpasses the current traditional techniques, HPLC and PAGE,