title: Separation of topological forms of plasmid DNA by anion-exchange HPLC: shifts in elution order of linear DNA.
abstract: We sought to establish a single anion-exchange HPLC method for the separation of linear, open circular and supercoiled plasmid topoisomers using purified topoisomeric forms of three plasmids (3.0, 5.5 and 7.6 kb). However, finding one condition proved elusive as the topoisomer elution order was determined to depend on salt gradient slope. The observed change in selectivity increased with plasmid size and was most pronounced for the linear form. Indeed, the elution order of the linear 7.6 kb plasmid was reversed relative to the supercoiled form. This observation may have implications for methods used in quality control of plasmid DNA.
authors.affiliation: BioPharmaceutical Product Development-US, GlaxoSmithKline, 5 Moore Drive, Research Triangle Park, NC 27709, USA.
authors.forename: Clara R
authors.initials: CR
authors.lastname: Smith
authors.forename: Randolph B
authors.initials: RB
authors.lastname: DePrince
authors.forename: Jennifer
authors.initials: J
authors.lastname: Dackor
authors.forename: Debra
authors.initials: D
authors.lastname: Weigl
authors.forename: Jack
authors.initials: J
authors.lastname: Griffith
authors.forename: Magnus
authors.initials: M
authors.lastname: Persmark
mesh_terms: D000837:Anion Exchange Resins; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D002852:Chromatography, Ion Exchange / Q000379:methods*; D004247:DNA / Q000302:isolation & purification*; D046529:Microscopy, Electron, Transmission; D010957:Plasmids / Q000302:isolation & purification*
chemical_list: D000837:Anion Exchange Resins; D004247:DNA