FIGURE 1.

Experimental and data-analysis workflow. (A) Starting cultures of fluorescence-tagged gene deletion (xΔ_RFP) and wild-type reference (WT_CFP) are grown separately. (B) Saturated cultures are mixed (usually in a 1:2 WT/xΔ ratio for increased dynamic range) and inoculated into SC aging medium in a semi-deep well plate until stationary phase (SP). (C) Competitive-aging cultures are sampled regularly at days t; outgrowth cultures in fresh medium are monitored at T hours with simultaneous measurement of absorbance at 600 nm (OD₆₀₀), and raw RFP and CFP signals (Supplementary Figure S1). Possible differences in growth rate (G) are taken into account. (D) Data analysis uses the change in fluorescent-signal ratio in the outgrowth cultures to estimate the relative survivorship of each mutant (S_w). Data is fitted to a multiple linear-regression model considering the change of ln(RFP/CFP) empirical measurements over time as a function of the starting strains’ proportion (A), the mutant’s relative growth rate (G), systematic batch errors in the measurements (C), and the mutant’s survivorship relative to the wild-type (G), our parameter of interest (Supplementary Figure S2 and Note S1).