Fig 1.

Schematic overview of the modular plasmid system and quantitative promoter measurements. (A) Schematic representation of the modular golden gate assembly plasmid parts with flanking BsaI cut sites (dashed lines). Overlapping 4 bp overhangs are color coordinated. The modular broad-host-range (BHR) backbone (pBTK402) contains inverted BsaI cut sites and an RFP dropout. (B) Golden Gate Assembly is performed in a one-tube reaction by digesting the backbone and insert part plasmids with BsaI and ligating with T4 ligase. (C) A modular stage-1 plasmid is complete when all overlapping inserts are successfully assembled in order. (D and E) Luciferase assays of P. luteoviolacea strains expressing plasmids with different promoters during exponential, stationary, or biofilm growth driving a Nanoluciferase (NLuc) gene where (D) shows CP25-NLuc-T7, PA3-NLuc-T7, Ptac-NLuc-T7 and (E) compares native MACs macS and macB promoters. Luminescence, as relative luminescence units (RLUs), is normalized to optical density at 600 nm (OD₆₀₀) and plotted on a log base 10 scale. The dashed line indicates P. luteoviolacea cells expressing a non-luminescent plasmid as represented by the dotted line (Y = 524 RLU/OD₆₀₀). Plotted is the mean of three biological replicates. Error bars indicate standard deviations.