Extended Data Fig. 5: Genetic construction and phenotypic characterization of diploid and tetraploid clusters.

a, Procedure for engineering isogenic grande diploid and tetraploid clusters, from each of which four independent petite mutants were isolated. Isolating multiple petite mutants is important because petite mutations are not isogenic and may confound ploidy-phenotype map. Grande and petite clusters correspond to mixotrophic and anaerobic conditions, respectively. b-e, Violin plots showing the distributions of cluster radius (b,c, where c is weighted by cluster volume), cell volume (d), and cell aspect ratio (e) in engineered diploid and tetraploid clusters under mixotrophic and anaerobic conditions (on average, n = 922 clusters (b,c) and 2,458 cells (d,e) measured per sample). Four biological replicates were measured for the mixotrophic condition, and the four independent petite mutants (each with one biological replicate) were measured for the anaerobic condition. For b,c, we measured cluster radius at 4 h (exponential phase) and 24 h (stationary phase) after transferring the culture to fresh media, and the 24-hour measurements are used throughout the paper unless otherwise noted. For c, filled circles show biomass-weighted mean cluster radius (the 24-hour values are the same as the values in Fig. 3e). For d,e, boxes, IQR; center lines, median; whiskers, values within 1.5 × IQR of the first and third quartiles. f,g, Comparison of the biomass-weighted mean cluster radius (f) and mean cell aspect ratio (g) of the engineered petite tetraploid clusters (mean of the four independent petite mutants, the same values as those in Fig. 1c,e) to the PA t0 and PA1-5 t50, t100, t150, and t200 populations (data from Bozdag et al.⁸).