FIGURE 8: Fragmentation of bacterial chromosomes under various conditions. (A) Schematic representation of size-dependent migration of linear DNA through PFGs. Note that the intact circular chromosome stays in the wells. Chromosomal fragmentation is quantified as the percentage of the total signal (well+lane, red rectangle) that enters the lane (blue rectangle), that is: [Signal in lanes x 100 / total signal (well+lane)]. HMW, high molecular weight (≥2 mbp); IMW, Intermediate molecular weight (0.2 – 2.0 mbp); LMW, low molecular weight (≤0.2 mbp). (B) Spontaneous and clastogen-induced chromosomal fragmentation in E. coli. Typically, chromosomal fragmentation is detected in the recBC mutant, deficient both in repair of DSBs and in degradation of linear DNA. However, when fragmentation is massive, it can be detected in wild type cells (such as ciprofloxacin- or phleomycin-induced fragmentation in AB1157 in these examples). Spontaneous fragmentation is measured under conditions when the recBC(Ts) mutants carry additional defects in SeqA, DNA polymerase I or DNA ligase [165, 167]. Clastogen-induced fragmentation is observed upon exposure to phleomycin and ciprofloxacin. Ultraviolet light, which affects only one strand of DNA, still causes significant fragmentation in recBC(Ts) background [166]. CZ, compression zone. (C) Pulsed field gel electrophoresis can be used to calculate the density of breaks in the chromosomal DNA if the average size of the resulting chromosomal fragments is measured.

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