Two methods to detect DNA fragments produced by restriction enzymes.
This report summarizes two methods for detecting limited amounts of DNA from restriction endonuclease digests. The first is a photographic system for visualizing ethidium bromide-stained DNA fragments in agarose gels which can detect as little as 50-100 pg DNA per band. The second technique is direct sulfonation of DNA fragments bound to nylon membranes followed by visualization of the fragments by nonradioactive immunoblot methods. The immunohistochemical staining can detect 10 pg DNA per band. The direct sulfonation technique is not intended to identify specific DNA sequences; DNA-DNA hybridization with sulfonated probes has previously been described (P. Lebacq, D. Squalli, M. Duchenne, P. Poulety, and M. Johannes (1988) J. Biochem. Biophys. Methods 15, 255-266). Direct sulfonation can be used when samples are relatively free of contaminating nucleic acids and is a useful alternative to end-labeling. These highly sensitive techniques may be suitable when the DNA source is of limited quantity or in instances where radiolabeling is not permitted.
Chapman
R W
RW
Chesapeake Bay Institute, Johns Hopkins University, Shady Side, Maryland 20764.
Brown
B L
BL
D000818:Animals; D003386:Brachyura / Q000235:genetics; D004247:DNA / Q000032:analysis*; D004262:DNA Restriction Enzymes; D004272:DNA, Mitochondrial / Q000032:analysis; D004587:Electrophoresis, Agar Gel; D004996:Ethidium; D007150:Immunohistochemistry; D008722:Methods; D010781:Photography; D013194:Staining and Labeling; D013447:Sulfites
D004272:DNA, Mitochondrial; D013447:Sulfites; D004247:DNA; D004262:DNA Restriction Enzymes; D004996:Ethidium; C009279:sodium bisulfite