Fig. 4. Optimizing alternative IPTG-inducible promoter architectures.

a Top: Design for Pmultiple library. Bottom: The average effect of the distal+ site (rows) on fold change given the distal site identity (column). Here, we examine consensus −10/−35 promoters containing O₁ or O_sym in the proximal site. b Top: Design for Pspacer library. Bottom: Comparison of uninduced expression, induced expression, and fold change between variants composed of the same sequence elements in the Pspacer and Pcombo architectures (two-sided Mann–Whitney U tests, n = 305). We examined only active promoters containing a consensus −10 and/or −35 sequence. In each boxplot, the lower, middle, and upper hinges correspond to the first quartile, median, and third quartile, respectively. Whiskers represent 1.5 × IQR from the lower and upper hinges. c Top: Design for Psteric library. Bottom: The fold change of promoters containing O₁ in both the core and proximal sites and a 56 bp interoperator distance. Here, we examine the effect of the −10 element in conjunction with the strongest UP and extended −10 element combinations. N/A indicates data missing from our analysis. d Distributions of uninduced expression, induced expression, and fold change for variants with fold change ≥2 in each library. The dashed line separates active from inactive sequences and is set as the median of the negative controls + 2*median absolute deviation (two-sided Mann–Whitney U tests with Benjamini–Hochberg correction, *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001). The exact P values are: uninduced (Pspacer–Psteric: P = 0.029, Pspacer–Pmultiple: P = 0.040, Pcombo–Psteric: P = 0.045), induced (Pspacer–Psteric: P = 0.002, Pcombo–Psteric: P = 0.013, Pmultiple–Psteric: P = 0.013), fold change (Pmultiple–Psteric: P = 0.0009, Pspacer–Psteric: P = 0.0045, Pcombo–Psteric: P = 0.040). Source data are available in the Source Data file.