THE USE OF C(0)T-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 g eneration of multinucleosomal-sized DNA fragments, which appear as a D NA ladder after agarose electrophoresis. Extensive DNA fragmentation c an be detected by ethidium bromide staining. Visualization of low leve ls of DNA fragmentation, however, requires both a more sensitive detec tion system, as well as a method of DNA extraction, that limits the ex tent of high molecular weight DNA shearing. We have found that the use of C(0)t-1 repetitive DNA as a probe for Southern analysis of DNA lad ders 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 ther e 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 ver y definitive DNA ladder can be visualized via Southern blot analysis w ith a C(0)t-1 DNA probe.