Luciferase reporter assay
One hundred microliters of the Agrobacterium strain GV3101 containing the pGreenII 0800-target site-LUC plasmid were combined with 900 μl of the Agrobacterium strain GV3101 containing either the 35s:pri miR398b or35s:primiR157plasmid. The mixture was incubated (28°C, 200 rpm, shaker) to a final concentration (OD600 = 1.0), and resuspended in infiltration buffer (10 mM MES, pH 5.6, 10 mM MgCl2 and 150 μM acetosyringone) as described previously (Hellens et al., 2005). The same volume of mixed suspensions were injected into leaves of Nicotiana benthamiana by agro-infiltration. After 48 h, LUC luminescence was examined using a cryogenically cooled CCD camera (Lumazome PyLoN 2048B) as described previously (Chen et al., 2008). LUC and RLUC activities were measured as counts per second using a multimode plate reader (PerkinElmer) using the Promega kit (N1630).