FLuc and RLuc activity assays and calculation of relative transcriptional activity

For the FLuc/RLuc assay, leaf samples were collected at 4 dpi. One 0.8 cm diameter disc per agroinfiltrated leaf was excised using a cork borer. Leaf discs were frozen in liquid nitrogen and subsequently homogenized. The Dual‐Glo® Luciferase Assay System (Promega) was used for sample analysis. Briefly, 180 μL of Passive Lysis buffer were added to the homogenized sample, vortexed and centrifuged for 15 min at 14 000 g at 4°C. Crude extract (10 μL) was mixed with 40 μL of LARII and FLuc activity was determined using a GloMax®‐Multi Detection System (Promega) with a 2 s delay and a 10 s integration times. After the measurements, 40 μL of Stop&Glo reagent were added per sample and RLuc activity was determined using the same protocol. Sample FLuc/RLuc ratios were calculated as the average value of the three independent agroinfiltrated leaves. Relative transcriptional activities (RTAs) were calculated as previously described (Vazquez‐Vilar et al., ²⁰¹⁷). Briefly, the FLuc/RLuc ratios in each sample normalized with the FLuc/RLuc ratios produced by a PNOS reference (GB1116) assayed in parallel.