a, Outline of the evolutionary history of line 17 from its founding by ancestral SM1 SBW25 through ten generations of the life cycle, by which time a mutS mutation had arisen. At the eleventh generation, the mutS (A1489C) mutation in WS22 was reverted to wild type (mutSWT) by in vitro manipulation. The two WS22 lineages (with and without the mutS mutation) were taken through three additional ‘expedited’ life-cycle generations, although with a cycling period of 24 h for mutS (A1489C) and 48 h for mutSWT. Genome sequences were acquired from five different time points (SM1, WS8, WS22, SM27 and WS28, indicated by surrounding boxes). For details of mutations see Supplementary Table 1. b, The mutS-dependent genetic switch. Four independent cultures of the generation-11 WS (WS22), with and without the mutS (A1489C) mutation, were passaged through an additional three ‘expedited’ generations of the life cycle (WS22–WS28). The nucleotide sequence of wspR was determined at each stage. Depicted is the tract of guanine residues beginning at nucleotide 742: WspR is active (and the phenotype is WS) when the tract length is seven residues, but inactive (and the phenotype is SM) when the tract length is eight. Red arrows show slippage events resulting in the gain or loss of a single guanine residue. Transitioning between phases is more reliable (fewer extinction events) and more likely to occur via the tract of guanine residues in the presence of mutS (A1489C). Lines of mutS (A1489C) were passaged on a 24-h cycle; mutSWT lines were passaged on a 48-h cycle (on a 24-h cycle mutSWT lines were extinct before the first generation completed—ancestral lines are incapable of transitioning on a 48-h cycle). Extinction events occurred whenever the extant phase failed to produce the next stage in the life cycle; death of a line allowed birth of an extant lineage (diagonal arrows).