The characterization of composite agarose/hydroxyethylcellulose matrices for the separation of DNA fragments using capillary electrophoresis.
Mixtures of the polysaccharide derivatives, 19% hydroxyethylated SeaPrep agarose (SP-AG) and hydroxyethylcellulose (HEC), in aqueous buffer solutions are applied for the first time to the separation of DNA fragments using capillary electrophoresis (CE). These matrices form unique size-sieving networks that allow the separation of a wide size range of DNA fragments in a single analysis. Relative to their homogeneous counterparts, the composite separation matrices provide enhanced selectivity properties of DNA fragments, especially for fragments greater than 1000 base pairs (bp) in length. Additionally, the effects on separation performance of capillary temperature, the incorporation of a DNA intercalator, and applied field strength are demonstrated. Solution viscosity measurements of the homogeneous and composite matrix solutions were made in order to establish the entanglement threshold concentrations for the unique size-sieving solutions. The relatively low solution viscosities of the composite separation matrices allow reproducible replacement of the separation matrix between analyses. The mechanism of separation of DNA fragments for the composite matrices is proposed.
Siles
B A
BA
The College of William and Mary, Department of Chemistry, Williamsburg, VA 23187-8795, USA. BASile@chem1.chem.wm.edu
Anderson
D E
DE
Buchanan
N S
NS
Warder
M F
MF
D002021:Buffers; D002482:Cellulose / Q000031:analogs & derivatives*; D004247:DNA / Q000302:isolation & purification*; D004560:Electricity; D019075:Electrophoresis, Capillary / Q000379:methods*; D007202:Indicators and Reagents; D007364:Intercalating Agents; D008970:Molecular Weight; D011108:Polymers; D012685:Sepharose; D012996:Solutions; D013696:Temperature; D014783:Viscosity
D002021:Buffers; D007202:Indicators and Reagents; D007364:Intercalating Agents; D011108:Polymers; D012996:Solutions; D002482:Cellulose; C002283:hydroxyethylcellulose; D004247:DNA; D012685:Sepharose