Figure 3. Chronology of ibpA Heat-Shock Promoter Evolution in Sublines of the 5A Line.

Line 5A was initiated from a single female in June 1999. In September 2000, it was transfected with a secondary symbiont to generate 5A⁺R^LONG. Subsequently the 5A line acquired the first mutation to the short promoter allele, which became fixed. An identical mutation rose to near fixation in the 5A⁺R^LONG culture in October 2005; both alleles were isolated from that colony to yield 5A⁺R^SHORT and 5A⁺R^LONG. Line 5A⁺R^SHORT lost its Candidatus Serratia symbiotica to generate line 5A⁻R^SHORT, and then in November 2005, 5A⁺R^LONG was cured of Candidatus Serratia symbiotica to generate line 5A⁻R^LONG. Thus the mutation occurred and rose to fixation or near fixation twice in sublines of the 5A laboratory line (Figure 1B).