Comprehensive Interpretation of Gel Electrophoresis Data

Abstract

From temperature analysis of polyacrylamide gel electrophoresis data for rigid-rod DNA analytes, it is proposed that an entropic force term is responsible for the discrepancy between Ogston-Morris-Rodbard-Chrambach model predictions and experimental results. This entropic force originates from reduction of the orientational freedom of anisotropic analytes in small pores of polyacrylamide gels. Time-dependent fluorescence anisotropy decay measurements confirm that, even in the absence of an external field, orientation of anisotropic analytes is restricted in polyacrylamide gels. A new comprehensive model is proposed that takes this effect into consideration. Predictions based on this model are found to compare favorably with experimental data for linear and three-arm asymmetrically branched rigid-rod DNA analytes covering a broad range of molecular aspect ratios and sizes. A new length scale is also proposed for describing the effect of analyte topology on electrophoretic mobility. This length scale reduces to the analyte radius of gyration in the limiting cases of spherically symmetric and linear rigid-rod species. Based on these results, a general approach is proposed for interpreting gel electrophoresis data of charged analytes possessing simple and complex topologies.