Figure 1. A S. oneidensis MR-1 mutant fitness compendium.

(A) Parallel analysis of MR-1 mutant pools using TagModules. Only the uptags are used to interrogate strain abundance in the upPool, while the downtags are used exclusively to measure strain abundance in the dnPool. The Affymetrix TAG4 microarray illustrated here contains the complement sequences to both the uptags and downtags, therefore the abundance of all strains across both pools is assayed in a single hybridization. In the simple example diagrammed here, strain 3 in the upPool and strain 5 in the dnPool have fitness defects. We hybridize the tags both before (start) and after growth in selective media (condition). We calculate the fitness of a strain as the normalized log₂ ratio of tag intensity of the condition relative to start. (B) Heatmap of the entire fitness dataset. Both genes and experiments were ordered by hierarchical clustering with Euclidean distance as the metric. A subset of the fitness heatmap for mutants in the general secretory pathway is expanded (bottom). (C) Fitness values for dnPool strains (x-axis) and upPool strains (y-axis) on DL-lactate defined media. “Same insertion” indicates identical mutant strains that are represented in both pools; “Other insertion, same gene” indicates independent transposon insertions in the same gene. The dashed line shows x = y. (D) Comparison of gene fitness values for pairs of genes predicted to be in the same operon [50]. The data plotted reflects a single fitness experiment in DL-lactate minimal media, but the color-coding is derived from the entire fitness compendium: points in red are uncorrelated (r<0.3) across 195 fitness experiments. (E) Quality metrics for each of the 195 pool fitness experiments. r(Same) is the fitness correlation of identical mutant strains contained in both the upPool and dnPool (see red triangles in panel C). r(Operon) is the fitness correlation of adjacent genes predicted to the in the same operon (see panel D). (F) Comparison of DL-lactate minimal media pool fitness values (y-axis) and individual strain growth rates (x-axis) for 48 transposon mutants. Individual strain growth rates represent the average of at least three independent experiments. The vertical dotted line represents the growth rate of wild-type MR-1. The horizontal dotted line represents a pool fitness value for a neutral insertion. Some strains had long lag phases and a growth rate could not be calculated (green plus symbols).