Enhanced ethidium fluorescence of large DNA electrophoresed in gels submersed in an immiscible solvent.
Agarose gel electrophoretic separation of a lambda/HindIII DNA marker containing detectable fragments of 23 to 2 kb was carried out in the conventional submarine apparatus and in the horizontal gel slab apparatus of Wieme, using identical samples, agarose, gel width and procedure for ethidium bromide staining. In the Wieme apparatus, the gel on its microscope slide support is immersed in an immiscible solvent such as petroleum ether or silicone oil. Although band resolution and speed of migration are equivalent between gels run in the two systems, the relative fluorescence intensity of the ethidium bromide-stained bands is substantially more responsive to an increase in DNA length in the Wieme gels than in the submarine gels. The predicted relative fluorescence is by an average factor of 0.5 less than that observed after electrophoresis in the Wieme apparatus but is by an average factor of 3.4 more than that observed on bands derived from the submarine technique.
Costello
R
R
National Institutes of Health, Bethesda, MD 20892-0001.
Wheeler
D
D
Orban
L
L
Chrambach
A
A
D004247:DNA / Q000032:analysis*; D004587:Electrophoresis, Agar Gel; D004996:Ethidium; D005453:Fluorescence
D004247:DNA; D004996:Ethidium