Capillary electrophoresis (CE) with a replaceable linear polyacrylamide (LPA) sieving matrix was used to examine sequence-dependent migration of double-stranded DNA fragments. It has been found that DNA conformational effects were significant under high electric field separations, especially using high resolution matrices. Compared to linear DNA-ladder standards, both anomalously slow and rapid DNA fragments were observed, with the degree of anomalous migration depending on the electric field strength, polymer concentration, column temperature, and background electrolyte (denaturants, sodium and magnesium ions, DNA-intercalating dyes). By selecting a combination of electrophoretic conditions (e.g. 3% T LPA, elevated capillary temperature, lower electric field strength and addition of DNA intercalating dyes), molecular weight dependent separations were closely restored.