Fig. 4. Promoter engineering controls the growth and population size in two-member co-cultures.
a, Diagram of two-member cross-feeding co-cultures (em1-em2); BFP-tagged member is overexpressing em1 and auxotrophic to em2, RFP-tagged member is overexpressing em2 and auxotrophic to em1. Five pairs of co-cultures include ade-Lys, Leu-Trp, Val-Lys, Trp-Lys and His-Lys. Ade-Lys co-culture is used as an example here and the other 4 pairs are shown in Supplementary Figs. 20–23. b, The combinations of five promoters with different strengths from strong (1) to weak (5), plus 0 expression (6). The expressions of target genes em1 and em2 were driven by these five promoters, with no expression in the BFP- and RFP-tagged member, respectively. Then, these 6 BFP-tagged members and 6-RFP-tagged members were combined to form 36 pairs of different two-member co-cultures. c, The strain table for the combinations of ade-Lys two-member co-cultures. The strain is named after the abbreviation of the target gene and the promoter strength; for example, BFP-tagged strain ade+lysΔ overexpressing ADE4op under stronger promoter pCCW12 is abbreviated ade#1. d, OD700 at 48 h of monocultures in synthetic minimal medium as the negative controls, and of the positive control (+ve, BY4741-pHLUM). N = 2 biologically independent samples and data are presented as mean ± s.d. e,f, Heat map of OD700 values (e) and RFP-tagged population percentages at 48 h (f) of 36 pairs of ade-Lys two-member co-cultures. The initial ratio was 1:1 and the initial cell density was OD700 0.078 for each member in these co-cultures. X axis from left to right deonotes the promoter strength of Lys+ (LYS21op) from weak (6) to strong (1); y axis from bottom to top denotes the promoter strength of ade+ (ADE4op) from weak (6) to strong (1). g, Time courses of the growth of all co-cultures and each member. N = 2 biologically independent samples and data are presented as mean ± s.d.