Modification of the electrokinetic properties of reversible electrophoresis gels for the separation and preparation of DNA. Addition of linear polymers.
The effect of adding linear polymers to a novel reversible electrophoretic was measured. Reversible gels are formed using the polyanionic carbohydrate polymer, gellan gum. Gellan gum forms strong stable gels in the presence of divalent cations or diamines. The gels are reversible (return to solution) by changing the ionic environment or pH. Gellan gum is an anionic polymer, and the electrophoresis gels have considerable electroosmotic flow (EOF) toward the negative electrode. We measured the EOF in gellan gum electrophoresis gels as a function of gel concentration, buffer composition, and linear polymer additive. The linear polymers used in this study were polyethylene oxide and hydroxyethyl cellulose. Both polymers reduced EOF in the gels, in a manner dependent on molecular weight. Polymers with high molecular weight were more effective at reducing EOF. The addition of polymers increased the resolution of low molecular weight DNA. Native gellan gum resolved DNA from approx 50,000 to 1000 bp. Addition of the polymers resolved DNA down to approx 50 bp, in some instances. The influence of the polymers on circular plasmid DNA was also investigated. Addition of high molecular weight polyethylene oxide reduced the electrophoretic mobility of the nicked circular form compared to the supercoiled form.
Cole
K D
KD
Biotechnology Division, National Institute of Standards and Technology, Gaithersburg, MD 20899, USA. Kenneth.Cole@nist.gov
Tellez
C M
CM
Nguyen
R B
RB
D004247:DNA / Q000737:chemistry / Q000302:isolation & purification*; D004586:Electrophoresis / Q000379:methods*; D008970:Molecular Weight; D011108:Polymers; D011135:Polysaccharides, Bacterial
D011108:Polymers; D011135:Polysaccharides, Bacterial; C048288:gellan gum; D004247:DNA