High-Speed Large Scale Chromatographic Purification of Plasmid DNA with a Novel Giant-Pore Stationary Phase

Abstract

A rigid spherical giant-pore poly (glycidyl methacrylate-co-ethylene dimethacrylate) matrix has been prepared by radical suspension–polymerization on the basis of a novel porogenic mode using superfine particles of calcium carbonate as a solid porogen. Scanning electron microscopy reveals that the bead has pores as large as 10 μm. The hydrodynamic properties show that this polymeric material has good strength and a low back pressure of 1.0 MPa at a flow velocity of 3,000 cm h⁻¹. After being modified to be an anion-exchange material, high dynamic binding capacity of plasmid DNA of above 1,000 μg plasmid per mL of bed by a column of this material, could be obtained comparing to the 150 μg plasmid per mL of bed with a Q-Sepharose FF column at the same flow rate. Large-scale preparative plasmid separations (2–20 mL) from cell lysate were investigated. A 75% yield and 94.9% purity of SC plasmid DNA were obtained by a 20 mL column of giant-pore beads at a flow rate of 600 cm h⁻¹.