Figure 7.

Transient expression of constitutively active phosphomimicking forms SlMKK2^DDand SlMKK4^DDin Nicotiana benthamiana resulted in hypersensitive response and accumulation of ROS. A. Expression of SlMKK2^DD and SlMKK4^DD. Gene expression was analyzed by qRT-PCR and relative expression levels were calculated by comparing with the corresponding values at 0 hr (as a control) after infiltration. B. Proteins of SlMKK2^DD and SlMKK4^DD. Leaf samples were harvested 48 hr after infiltration and total soluble protein extracts were prepared. Proteins were separated by SDS–PAGE and analyzed by immunoblotting using a GFP-specific antibody. Total proteins showing equal loading were examined by Coomassie staining. C. HR-like cell death. Photo was taken 48 hr after infiltration. D. Accumulation of H₂O₂. Detection of H₂O₂ was performed by DAB staining at 48 hr after infiltration. Data presented in (A) are the means ± SD from three independent experiments and different letters above the columns indicate significant differences at p < 0.05 level.