Quantitation of ethidium-stained closed circular DNA in agarose gels.
The fluorescence of ethidium bromide (EB) bound to equimolar amounts of supercoiled form I and unstrained linear form III pBR322, SV40 and PM2 DNA in agarose gels has been measured by scanning a photographic negative of the gel with a microdensitometer. For SV40 and PM2 DNA, commonly used staining conditions cause both forms, i.e. linear and supercoiled, to fluoresce to the same extent. This obviates the need to use a correction factor for the fluorescence of form I DNA when measuring the amount of this form relative to the amounts of unstrained forms in agarose gels. In the case of PBR322 DNA, form I was found to fluoresce approximately 20% more than form III DNA.
Shubsda
M F
MF
Department of Chemistry, Syracuse University, New York 13244-4100, USA.
Goodisman
J
J
Dabrowiak
J C
JC
D001435:Bacteriophages / Q000737:chemistry; D004270:DNA, Circular / Q000032:analysis*; D004278:DNA, Superhelical / Q000032:analysis; D004279:DNA, Viral / Q000032:analysis; D003720:Densitometry; D004587:Electrophoresis, Agar Gel / Q000379:methods*; D004996:Ethidium; D005069:Evaluation Studies as Topic; D005456:Fluorescent Dyes; D010957:Plasmids / Q000032:analysis; D013539:Simian virus 40 / Q000737:chemistry
D004270:DNA, Circular; D004278:DNA, Superhelical; D004279:DNA, Viral; D005456:Fluorescent Dyes; D004996:Ethidium