Optimized chemiluminescent detection of DNA amplified in the exponential phase of PCR.
We describe a procedure for quantitative detection of nucleic acids by coupling PCR with improved chemiluminescent detection techniques. After performing PCR in the exponential phase in the presence of a trace amount of digoxigenin-11-dUTP, the amplified products are transferred to a positively charged nylon membrane. The membrane is cleaned with a modified method involving sodium dodecyl sulfate and ethanol washing steps to ensure low backgrounds in chemiluminescent detection. The membrane is exposed to two stacked x-ray films to increase the dynamic response in a film exposure. This sensitive and quantitative procedure is useful for molecular biology studies.
Mailman Research Center, McLean Hospital and Harvard Medical School, Belmont, MA 02178.
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Berman
D001483:Base Sequence; D004247:DNA / Q000032:analysis*; D008163:Luminescent Measurements; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction
D004247:DNA