Effect of urea on the polymer buffer solutions used for the electrophoretic separations of nucleic acids.

Abstract

The present study describes the effect of urea on the properties of hydroxyethyl cellulose (HEC) polymer solutions used for the separation of nucleic acids. Solution properties were investigated by viscosity measurements and supplemented by Raman spectroscopy of the solution components. By using viscosimetry, it was possible to identify that borate, urea, and HEC participate in an interaction causing an increase in the viscosity of dilute solutions. In addition, short polymer chains exhibited a 4-fold decrease in the entanglement threshold in the presence of 4 M urea and 0.356 M total borate concentration. These interactions were found to be specific to HEC. Raman spectroscopy monitored serial addition of 8 M urea to HEC sieving solutions combined with factor analysis indicated formation of minor urea species. Lack of change in the Raman spectrum and relative amount of borate suggested that there is no direct interaction between borate and urea. These effects on HEC sieving solution properties lead to the use of low HEC concentrations that are beneficial for the separation of nucleic acids under denaturing conditions.