Rj2035

Root fragments and nodules were fixed as mentioned above. After that, they were washed with 0.1 M phosphate buffer three times for 15 min each, once with water for 15 min, and dehydrated for 10 min in 10%, 30%, 50%, 70%, 90%, and 100% (v/v) ethanol, and sequentially embedded in plastic Technovit 7100 (Heraeus Kulzer). Sections of 5 μm were made using a microtome (RJ2035, Leica), stained with 0.05% Toluidine Blue (Sigma), mounted in Euparal (Carl Roth), and analyzed with a Leica AU5500B microscope equipped with a DFC425c camera (Leica). Transgenic pMtHDT::GFP::MtHDT nodules and root segments were sectioned into 60-µm slices by vibratome (VT1000, Leica) and stained with propidium iodide for 2 min. Sections were then washed three times in phosphate buffer containing triton x-100 and mounted on slides with MQ water. All confocal images were acquired using Leica SP8 confocal laser scanning microscope (Leica, Germany). GFP and EdU signals were detected with excitation wavelength and detection windows of 488 and 500–530 nm, propidium iodide was detected with excitation wavelength and detection windows of 543 and 580–640 nm.