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Nucleic Acids Research, 1995, Vol. 23, No. 13 2413-2420
© 1995

CHEMISTRY

Double bands in DNA gel elctrophoresis caused by bis-intercalating dyes

Christina Carisson, Mats Johnson and Björn Åkerman*

Department of Physical Chemistry, Calmers University of Technology S-412 96 Göteborg, Sweden

*To whom corespondence should be addressed

Received April 10, 1995. Accepted May 26, 1995.

Many bis-intercalating dyes used for fluorescence detection of DNA In electrophoresis have been reported to give band-splitting and band-broadening, which results In poor resolution and a decreased detection sensitivity. We have studied the dimerlc dyeYOYO-1, and to some extent also TOTO-1 and EthD-1,and found that in complex with DNA these dyes give rise to two components with different electrophoretic mobilities. Electrophoresis experiments and spectroscoplc measurements on the two components show hat they differ In that the DNA molecules have different amounts of dye bound. Our results exclude that the extra bands are caused by intermolecular crosslinking. Incubation of the samples for Increasing times before electrophoresis makes the bands move closer nd closer to each other as the dye molecules become more homogeneously distributed among the DNA molecules. Finally, the two bands merge into one at an intermediate position. This equilibration rocess is xtremely slow at room temperature (days), and is therefore not a practical method to eliminate bandsplittingin routine analysis. However, we find that if the temperature is raised to 50°C, the dye-DNA complexes equilibrate completely In only 2 h.


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