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2000; Murata et?al. 2001; Kwak et?al. 2003; Munemasa et?al. 2007; Islam et?al. 2010). ABA and MeJA lead [Ca2+]cyt elevation to activate S-type anion channels, which causes in plasma membrane depolarization, selleck compound leading activation of outward-rectifying K+ (K+out)channels, resulting in stomatal closure (Allen et?al. 1999). In addition, inward-rectifying K+ (K+in) channels in guard cells have been suggested to provide a pathway for K+ uptake into guard cells during stomatal opening. Suppression of K+in channel is favourable to stomatal closure (Kwak et?al. 2001; Saito et?al. 2008; Siegel et?al. 2009). K+in channels are inhibited by increases in [Ca2+]cyt (Schroeder & Hagiwara 1989; Lemtiri-Chlieh & MacRobbie 1994) but [Ca2+]cyt-independent K+in channel activation has also been PFI-2 datasheet reported (Grabov & Blatt 1997). In this study, to elucidate SA signalling in Arabidopsis, we examined stomatal closure, ROS production, NO production, [Ca2+]cyt oscillations, and K+in channel activity. Arabidopsis (Arabidopsis thaliana) L. ecotype Columbia was used as wild-type plant in this study. Columbia and atrbohD atrbohF mutant plants were grown in growth chambers (22?��C, 80??mol?m?2?s?1 under a 16?h light/8?h dark regime). Arabidopsis genome initiative numbers for AtRBOH genes are AtRBOHD (At5g47910) and AtRBOHF (At1g64060). Stomatal apertures were measured as described previously with modification (Murata et?al. 2001; Munemasa et?al. 2007; Jahan et?al. 2008; Islam et?al. 2009, 2010). Excised rosette leaves of 4�C6 weeks old Arabidopsis plants were blended in water for 30?s in blender and epidermal tissues were collected with 48??m nylon sieves. The epidermal tissues were dipped into stomatal assay solution containing 50?mM KCl, 50??M CaCl2 and 10?mM MES-Tris, pH?6.15, for 2?h in the light (80??mol?m?2?s?1). Then, SA was added to the stomatal assay solution and stomatal MASP1 apertures were measured 2?h after incubation. Twenty stomatal apertures were measured on each epidermal peel. Three replications were maintained for each stomatal assay experiment. Student's t-test was used to determine statistical significance of the data. We regarded difference at the level of P?