Fig. 3
P19 RNA silencing suppressor and CP act synergistically to enhance RYMVMg1ΔP1ΔCP/CterPSA RNA accumulation. N. benthamiana leaves were infiltrated with the RYMVMg1ΔP1ΔCP/CterPSA construct alone (RYMVMg1ΔP1ΔCP/CterPSA : empty vector 1:1 ratio) or with CP and P19 constructs either alone (RYMVMg1ΔP1ΔCP/CterPSA:CP or P19:empty vector 1:0,5:0,5 ratio) or together (RYMVMg1ΔP1ΔCP/CterPSA:CP:P19 1:0,5:0,5 ratio). Infiltrated leaves were harvested at 5 dpi. Three independent experiments were carried out. A- Semi-quantitative RT-PCR was performed after reverse transcription of RNA using with a mixture of oligodT and CterPSA specific primers. PCR amplifications was performed with CP and P19 specific primers. NbEF1-α expression was used as an internal control. The results shown here are representative of the three experiments. B- Total soluble proteins were prepared from the same N. benthamiana samples. Equal amounts of proteins (10 µg) were separated by SDS-PAGE and analyzed by Western immunoblotting using anti-CP antibody. The size of the CP signals is indicated in kDa. The results shown are representative of the three experiments. C- RYMVMg1ΔP1ΔCP/CterPSA RNA accumulation was evaluated by q-RTPCR for cDNA samples prepared in A. CterPSA-specific primers were used. Normalization was performed using GAPDH as a housekeeping internal control. Expressions were evaluated relative to the “RYMVMg1ΔP1ΔCP/CterPSA alone” Condition. The three independent experiments are represented by different colors