Fig. 4

Application of copper-inducible dCasEV2.1 for activating endogenous genes in N. benthamiana. a Schematic representation of the strategy of copper-mediated dCasEV2.1 induction for targeted gene activation in planta. b Activation of the N. benthamiana DFR (NbDFR) and PAL2 (NbPAL2) genes by the dually copper-regulated dCasEV2.1 under minDFR promoter (Cu/Cu dCasEV2.1) and the constitutive dCasEV2.1 under 35S promoter (Cs/Cs dCasEV2.1). Transcripts were quantified by RT-qPCR at 5 days post-infiltration of dCasEV2.1 in combination with a constitutive guide RNA targeting either NbDFR (gNbDFR), NbPAL2 (gNbPAL2), or the S. lycopersicum MTB gene (unspecific gRNA) and a constitutive CUP2:Gal4. Leaves were treated either with 5 mM copper sulfate or water at 3 days post infiltration. Error bars indicate SD (n = 3). The asterisks represent a significant difference between treatments. Statistical analysis was performed using an unpaired two-tailed t-test for two samples comparisons (P-value ≤ 0.05). The figure includes images from Biorender (biorender..com)