The use of C0t-1 probe DNA for the detection of low levels of DNA fragmentation.
In many apoptotic systems the final demise of the DNA results in the generation of multinucleosomal-sized DNA fragments, which appear as a DNA ladder after agarose electrophoresis. Extensive DNA fragmentation can be detected by ethidium bromide staining. Visualization of low levels of DNA fragmentation, however, requires both a more sensitive detection system, as well as a method of DNA extraction, that limits the extent of high molecular weight DNA shearing. We have found that the use of C0t-1 repetitive DNA as a probe for Southern analysis of DNA ladders is a sensitive method to detect low levels of DNA fragmentation. We have applied this methodology in the detection of DNA fragmentation in the normal developing C57BL/6 mouse retina at stages in which there is known DNA fragmentation, as well as in human Y-79 retinoblastoma cells grown in culture. We have found that in many instances in which there is no detectable DNA ladder with ethidium bromide staining a very definitive DNA ladder can be visualized via Southern blot analysis with a C0t-1 DNA probe.
Wong
P
P
National Eye Institute, National Institutes of Health, Bethesda, MD 20892, USA.
Smith
S B
SB
Bora
N
N
Gentleman
S
S
D000818:Animals; D017209:Apoptosis; D002478:Cells, Cultured; D004247:DNA / Q000032:analysis*; D015342:DNA Probes; D018076:DNA, Complementary; D004273:DNA, Neoplasm / Q000032:analysis; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D014407:Tumor Cells, Cultured
D015342:DNA Probes; D018076:DNA, Complementary; D004273:DNA, Neoplasm; D004247:DNA