The full length CrNPF2.9 gene was subcloned into the pSCA-cassette vector as a YFP fusion, and the Arabidopsis AtTPK1 was subcloned into the into the pSCA-cassette vector as a CFP fusion. The vacuolar STR-CFP and the plasma membrane PM-CFP (CD3-1002) markers were described previously 5 (link),16 (link). All vectors used a 35S promoter for expression. Transient transformation of C. roseus cells by particle bombardment and fluorescence imaging were performed following the procedures previously described 5 (link),49 (link). C. roseus cells were bombarded with DNA-coated gold particles (1 μm) and 1,100 psi rupture disc at a stopping-screen-to-target distance of 6 cm, using the Bio-Rad PDS1000/He system and 100 ng of each plasmid per transformation. For the transient transformation of onion cells, internal epidermis of fresh onion purchased from local producer (Les Vergers de la Breteche, St-Paterne Racan), were peeled and placed on solid vitamin-free MS medium and bombarded following the same protocol. Both cell-types were cultivated for 16 h to 38 h before being harvested and observed. The subcellular localization was determined using an Olympus BX-51 epifluorescence microscope equipped with an Olympus DP-71 digital camera and a combination of YFP and CFP filters.