Balagurumoorthy, P., Chen, K., Bash, R. C., Adelstein, S. J. and Kassis, A. I. Mechanisms Underlying Production of Double-Strand Breaks in Plasmid DNA after Decay of ¹²⁵I-Hoechst. Radiat. Res. 166, 333–344 (2006).

Previously, the kinetics of strand break production by ¹²⁵I-labeled m-iodo-p-ethoxyHoechst 33342 (¹²⁵IEH) in supercoiled (SC) plasmid DNA had demonstrated that ∼1 DSB is produced per ¹²⁵I decay both in the presence and absence of the hydroxyl radical scavenger DMSO. In these experiments, an ¹²⁵IEH:DNA molar ratio of 42:1 was used. We now hypothesize that this DSB yield (but not the SSB yield) may be an overestimate due to subsequent decays occurring in any of the 41 ¹²⁵IEH molecules still bound to nicked (N) DNA. To test our hypothesis, ¹²⁵IEH was incubated with SC pUC19 plasmids (¹²⁵IEH:DNA ratio of ∼3:1) and the SSB and DSB yields were quantified after the decay of ¹²⁵I. As predicted, the number of DSBs produced per ¹²⁵I decay is one-half that reported previously (∼0.5 compared to ∼1, ± DMSO) whereas the number of SSBs (∼3/¹²⁵I decay) is similar to that obtained previously (∼90% are generated by OH radicals). Direct visualization by atomic force microscopy confirms formation of L and N DNA after ¹²⁵IEH decays in SC DNA and supports the strand break yields reported. These findings indicate that although SSB production is independent of the number of ¹²⁵IEH bound to DNA, the DSB yield can be augmented erroneously by ¹²⁵I decays occurring in N DNA. Further analysis indicates that 17% of SSBs and 100% of DSBs take place within the plasmid molecule in which an ¹²⁵IEH molecule decays, whereas 83% of SSBs are formed in neighboring plasmid DNA molecules.