Fig. 3

Validation of the subcellular localization by covalent interaction in planta between co-expressed organelle-specific SpyTag and eGFP-SpyCatcher. Organelle-specific SpyTag constructs and eGFP-SpyCatcher were transiently co-expressed in leaves of Nicotiana benthamiana plants. CLSM analysis was performed 72 h/96 h after infiltration. Expression of eGFP-SpyCatcher alone revealed cytosolic and nuclear localization (a–d). When co-expressed with organelle-specific SpyTag constructs, eGFP fluorescence is directed towards the targeted organelles. Chloroplasts were targeted by co-expressing Plastid-SpyTag (e–h), and mitochondria by co-expressing Mito-SpyTag (e–h). Autofluorescence of chloroplasts is shown in blue (a–h), the mitochondria marker IVD-mcherry is shown in red (i–l). Scale bars represent 10 µm