Transient Expression of CsGR-RBP3 Fusion Protein

The coding sequence without the stop codon was amplified by PCR. The specific primers (F3 and F4) are listed in Supplementary Table S2. The PCR product was cloned into a transient expression vector (pCAMBIA 2300-GFP) between the Kpn I and Spe I sites. The digested pCAMBIA 2300-GFP fragment and CsGR-RBP3 fragment were linked with T4-ligase (Invitrogen, United States). CsGR-RBP3-GFP was driven by a cauliflower mosaic virus (CaMV) 35S promoter. The fusion constructs and control vectors were electroporated into Agrobacterium tumefaciens strain GV3101 using Gene Pulser Xcell^TM Electroporation Systems (Bio-Rad, United States). Tobacco (Nicotiana benthamiana) leaves were used for subcellular localization assay using the infiltration method. GFP fluorescence was observed using a fluorescence microscope 574 (Zeiss Axioskop 2 Plus) (Ye et al., 2016 (link)).