COMPARATIVE MEASUREMENTS OF RADIATION-INDUCED DNA DOUBLE-STRAND BREAKS BY GRADED VOLTAGE AND PULSED-FIELD GEL-ELECTROPHORESIS

We compared the ability of graded-voltage gel electrophoresis (GVGE) a nd pulsed-field gel electrophoresis (PFGE), to resolve DNA containing double-strand breaks (dsb) induced in human MS751 cells after exposure to gamma-radiation. For quantitation, prelabelling of the cells with [2-C-14]-thymidine prior to electrophoresis and subsequent scintillati on counting of excited bands was found to give closely similar results to Southern blotting and radiolabelled probe hybridization followed b y phosphorimager quantitation. Compared with PFGE, DNA subjected to GV GE migrated further and generated distinct DNA bands. Dsb were detecte d and quantifiable with GVGE at much lower doses than with PFGE: the r adiation dose limits were approximately 2 and 10 Gy respectively. At a ll doses used, the amounts of dsb detected by GVGE were higher than th ose by PFGE. GVGE coupled with Southern hybridization and phosphorimag er analysis is thus a more sensitive approach to assessing dsb and the ir relationship with radiation sensitivity. The approach is also conve nient when processing large numbers of samples and has the additional advantage of avoiding cell prelabelling such as in the case of cells e xtracted directly from human tumour biopsies and normal tissues.