Visualization of DNA in agarose gels as migrating colored bands: applications for preparative gels and educational demonstrations.
Visualization of DNA in electrophoretic gels typically requires UV radiation and the fluorescent dye ethidium bromide. Alternatively, we report here that by inclusion of visible dyes in standard agarose gels, DNA bands are observable in ambient light as they are separating. Such bands can be directly recovered from gels (approximately 50% yield) and used in standard enzymatic reactions (ligation, endonucleolytic cleavage, random labeling, PCR, and cycle-sequencing) without purification. Of 14 common commercially available stains that could visualize fractionating DNA, Nile blue was chosen for more extensive analysis as it gave the sharpest and most persistent bands and is not known to be toxic. Bands containing greater than 40 ng DNA could be detected by direct visual inspection of gels during electrophoresis. Drying the gels increased sensitivity to 4 ng. We describe relevant molecular features of these dyes and detail simple assays that may be employed to find other useful, and perhaps superior, dyes. This method also lends itself to situations in which easy visualization and convenience of DNA electrophoresis are important, such as classroom demonstrations.
Adkins
S
S
Mental Health Research Institute, University of Michigan, 205 Zina Pitcher Place, Ann Arbor, Michigan, 48109-0720, USA.
Burmeister
M
M
D015139:Blotting, Southern; D004247:DNA / Q000302:isolation & purification*; D015247:Deoxyribonuclease HindIII / Q000378:metabolism; D004587:Electrophoresis, Agar Gel / Q000379:methods*; D004996:Ethidium; D005456:Fluorescent Dyes; D006801:Humans; D006863:Hydrogen-Ion Concentration; D010078:Oxazines; D016133:Polymerase Chain Reaction; D011754:Pyronine
D005456:Fluorescent Dyes; D010078:Oxazines; C008619:Nile Blue; D004247:DNA; D015247:Deoxyribonuclease HindIII; D004996:Ethidium; D011754:Pyronine