Cloning CsDDI1 into GFP Vector

The full-length CDS without the stop codon of CsDDI1 was amplified by RT-PCR and was ligated into the C terminus of the green fluorescent protein (GFP) of a transient expression vector (pCAMBIA2300-GFP) between Kpn I and Spe I sites, driven by a cauliflower mosaic virus (CaMV) 35S promoter. The specific primers (F3 and F4) are listed in Table S1. The fusion constructs and control vector were electroporated into Agrobacterium tumefaciens strain GV3101 using Gene PulserXcellTM Electroporation Systems (Bio-Rad, USA) and then transformed into tobacco (Nicotiana benthamiana) leaf using the infiltration method. GFP fluorescence in tobacco leaf was observed 48 h after transfection using a fluorescence microscope (Zeiss Axioskop 2 Plus) (Wang et al., 2018a (link)).