Comparison of the separation of large DNA fragments in the presence and absence of electroosmotic flow at high pH.
This paper describes the analysis of large DNA fragments at pH > 10.0 by capillary electrophoresis (CE) in the presence of electroosmotic flow (EOF) using hydroxyethylcellulose (HEC) solution. HEC solution in the anodic reservoir enters the capillaries filled with high-pH buffer by EOF after sample injection. With respect to resolution, sensitivity, and speed, separation conducted under discontinuous conditions (different pH values of HEC solutions and buffer filling the capillary) is appropriate. Using HEC solution at concentrations higher than its entanglement threshold ensures a good separation of large DNA fragments in the presence of EOF at high pH. In addition to pH and HEC, the electrolyte species, dimethylamine, methylamine, and piperidine, play different roles in determining the resolution. The separation of DNA fragments ranging in size from 5 to 40 kilo base pairs was completed in 6 min using 1.5% HEC prepared in 20 mM methylamine-borate, pH 12.0, and the capillary filled with 40 mM dimethylamine-borate, pH 10.0. In comparison, this method allows faster separations of large DNA fragments compared with that conducted in the absence of EOF using dilute HEC solutions.
Chiu
Tai-Chia
TC
Department of Chemistry, National Taiwan University, Section 4, Roosevelt Road, Taipei, Taiwan.
Chang
Huan-Tsung
HT
D002482:Cellulose / Q000031:analogs & derivatives* / Q000737:chemistry; D004247:DNA / Q000302:isolation & purification*; D004563:Electrochemistry / Q000379:methods*; D006863:Hydrogen-Ion Concentration; D009995:Osmosis
D002482:Cellulose; C002283:hydroxyethylcellulose; D004247:DNA