D eclipse c1 confocal

For differential interference contrast (DIC) and bright-field microscopy, all samples were cleared, mounted, and photographed as previously described (Candela et al., 1999 (link)). Micrographs of venation patterns, and leaf primordia expressing ATHB8_pro:GUS were taken under bright field with a Nikon D-Eclipse C1 confocal microscope equipped with a Nikon DS-Ri1 camera, using the NIS-Elements AR 3.1 software (Nikon). Diagrams from leaf cells and venation patterns, and morphometric analysis of leaf cells were obtained as previously described (Pérez-Pérez et al., 2011 (link); Mateo-Bonmatí et al., 2018 (link)). For venation pattern morphometry, the phenoVein (http://www.plant-image-analysis.org) (Bühler et al., 2015 (link)) software was used. Leaf lamina circularity was calculated as 4 · π · area/perimeter². Lamina area and perimeter were measured on diagrams from the leaf lamina with the Fiji Wand tool. GUS assays were performed as previously described (Robles et al., 2010 (link)).

Live and fixed fish were imaged using a Nikon D-Eclipse C1 confocal scanning microscope by taking optical sections through the graft at 5-µm intervals according to a previously described protocol (Hall et al., 2009 (link)).