Gel electrophoresis of DNA fragments in narrow-bore capillaries

Electrophoresis
P LindbergJ Roeraade

Abstract

In this work, we studied the behavior of electrophoretic columns, having an inner diameter (ID) of 2-10 microm, filled with a cross-linked polyacrylamide gel matrix. The usefulness of these columns for DNA sequencing is discussed. Evaluation of column performance included tests of gel stability and migration time reproducibility. Confocal laser-induced fluorescence (LIF)-detection was employed utilizing a 488 nm argon ion laser for separations of C- and T-terminated DNA Sanger fragments. Reducing the inner diameter of the column from 50 microm to 10 microm resulted in an approximately eightfold increase in lifetime, under conditions in which the columns were subjected to a field strength of 1000 V/cm. The 10 microm ID columns were utilized for separation of Sanger fragments, and adequate detection sensitivity was obtained by stacking of the fragments from a deionized sample solution. A linear algorithm for retention data synchronization between individual electropherograms was employed to provide a route towards a reliable automated base calling protocol.

References

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Citations

Feb 5, 2002·Electrophoresis·G A Griess, P Serwer
Aug 27, 1998·Electrophoresis·G W SlaterG Drouin
Feb 24, 2001·Journal of Chromatography. a·Z K Shihabi
Sep 7, 2002·Journal of Chromatography. a·Benjamin Chu, Dehai Liang
Jan 8, 2000·Journal of Biochemical and Biophysical Methods·V Dolník
Aug 18, 2010·The Analyst·Makedonka D GulcevCharles A Lucy
Mar 7, 2003·Chemphyschem : a European Journal of Chemical Physics and Physical Chemistry·Lori A Woods, Andrew G Ewing
Nov 2, 2007·Chemical Reviews·Karel Kleparník, Petr Bocek

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