An ethidium bromide-agarose plate assay for the nonradioactive detection of cDNA synthesis.
Ethidium bromide was used to determine the success of cDNA synthesis reactions. Since ethidium bromide in agarose can be used to quantitate RNA and DNA, conditions under which the greater fluorescence of double-stranded DNA (dsDNA) is utilized were devised to assay dsDNA synthesis from mRNA. Ethidium bromide at 5 micrograms/ml in agarose allowed quantitative detection of cDNA in the range of 0.03 to 0.0015 microgram. Sodium dodecyl sulfate had an adverse effect on the measurement of cDNA. Subsequent cDNA analysis by alkaline gel electrophoresis and staining in 5 micrograms/ml ethidium bromide allowed accurate and rapid sizing of cDNA and required only 0.1-0.05 microgram cDNA.
Christen
A A
AA
U.S. Department of Agriculture, Southern Regional Research Center, New Orleans, Louisiana 70179.
Montalbano
B
B
D004247:DNA / Q000032:analysis* / Q000096:biosynthesis; D004591:Electrophoresis, Polyacrylamide Gel; D004996:Ethidium; D007202:Indicators and Reagents; D012685:Sepharose
D007202:Indicators and Reagents; D004247:DNA; D012685:Sepharose; D004996:Ethidium