Capillary electrophoresis of DNA: Disentangling nucleic acid dynamics in ultradilute polymer solutions.

Abstract

Conformational dynamics of ds-DNA under electrophoretic conditions are observed in solutions of hydroxyethyl cellulose (HEC) both above and below the entanglement limit, C$\sp*$. Both DC and field inversion protocols are investigated. A localized region of entanglement between DNA and polymer is shown to govern the dynamics of DNA separation in both ultradilute and entangled polymer solutions. Although this localized region is relatively stationary in entangled polymer solutions, it migrates in ultradilute polymer solutions with nearly the same velocity as the DNA molecule. This localized entanglement causes extension of a DNA molecule into a U or V shape, which collapses back to a wormlike coil conformation as the DNA molecule moves around the entanglement. The degree of DNA extension is found to be dependent on both the applied field and the polymer concentration. The use of ultradilute polymer solutions for the separation of supercoiled DNA is also examined. Contrary to behavior in gels, supercoiled DNA of less than 16 kbp has a lower mobility than both linear DNA and open circular DNA of equal size. As in gels, bands for supercoiled DNA are wider than bands for linear DNA. These differences in migration and band width allow for rapid discrimination between supercoiled, open circular, and linear DNA within a sample. The differing dependence of mobility on chain length can be explained by assuming that supercoiled DNA migrates as an elastic rod, while linear DNA migrates as a wormlike chain.