Electrophoretic charge density and persistence length of DNA as measured by fluorescence microscopy

Individual ethidium-stained DNA molecules, embedded in an agarose gel made with electrophoresis buffer (0.05 molar salt), are observed using a fluorescence microscope. In the first experiment, open circular 66 kilobase pair (kbp) plasmids, immobilized by agarose fibers threaded through their centers, display entropic "rubber" elasticity. The charged molecules extend in an electric field of several volts per centimeter and contract to a compact random coil when the field is removed. The extension of the plasmids as a function of field strength is consistent with the freely jointed chain model when the effective electrophoretic charge density is set at 15 e-per persistence length. In a second experiment, stained linear 48.5 kbp DNA molecules are observed as random coils immobilized in agarose. A measure of their size, here named the "maximal-X-extent," is taken for 100 molecules and found to average 1.47 mu. A Monte Carlo computer simulation of random coils (freely jointed chain model) gives the same maximal-X-extent value when the persistence length is set at 0.08 mu.