Extended Data Fig. 3: Sr43 localizes to the nucleus and plastids.
From: The wheat stem rust resistance gene Sr43 encodes an unusual protein kinase
a, Agrobacterium-mediated transient expression of a Sr43-GFP fusion construct or a GFP construct driven by the cauliflower mosaic virus (CaMV) 35 S promoter in Nicotiana benthamiana. GFP fluorescence in epidermal cells was recorded at 3 days post infiltration by confocal laser scanning microscopy. b, Agrobacterium-mediated transient expression of a the 35 S:Sr43-GFP with a 35 S:PLSP2A-RFP plastid marker construct in N. benthamiana. GFP or RFP fluorescence in epidermal cells was recorded at 3 days post infiltration by confocal laser scanning microscopy. c, transient expression of a ZmUbi:GFP-Sr43 fusion construct with ZmUbi:NLS-mCherry nuclear marker in wheat mesophyll cells (upper panel) and a ZmUbi:GFP as control (lower panel). GFP and mCherry fluorescence were recorded 16–20 hrs post transfection by confocal laser scanning microscopy. A non-transformed cell is indicated with black arrow. The scale bar in all the images is 20 µm. Nuclei (N), cytoplasm (C) and plastids (P) are indicated on the images with white arrows. The sizes of the GFP and Sr43-GFP fusion proteins are estimated at 27 and 126 kDa, respectively. The experiments were repeated in five (Sr43-GFP, panel a), two (GFP, panel b), three (panel b), five (GFP-Sr43) and two (GFP) independent experiments, respectively, with similar results. In the N. benthamiana experiments, we infiltrated two leaves and screened more than 100 nuclei per leaf.
