Figure 5.

CRK28 expression is induced in response to flg22, and heighted expression of CRK28 enhances the flg22-triggered ROS burst. A, qPCR analyses of CRK28 expression in response to treatment with flg22. Four-week-old Col-0 and npro:CRK28-FLAG lines 28-1 and 28-2 were treated with flg22 or water, and leaf samples were collected for RNA extraction at the indicated time points. Expression was normalized to Arabidopsis EF-1α. RQ, Relative quantification. Error bars indicate se; n = 3. Statistical differences were detected by Fisher’s lsd, α = 0.05, and different letters indicate statistical significance. The experiment was repeated twice with similar results. B and C, ROS burst in Col-0, 28-1, and 28-2 after treatment with flg22 and chitin, respectively. Gray and blue bars indicate water and flg22/chitin treatment, respectively. ROS was quantified using a luminol-based assay. The graphs depict total relative light units (ΣRLU) detected over a 30-min period. Error bars indicate se; n = 16. Statistical differences after flg22 or chitin treatment were detected by Fisher’s lsd, α = 0.05. These experiments were repeated three times with similar results. D, MAPK3/6 activation assay in Col-0 and 28-2 after flg22 or water treatment. Three-week-old plants were sprayed with flg22 or water, and tissues were collected at the indicated time points and processed. Phosphorylated MAPK3 and MAPK6 were detected with anti-p44/42 MAPK antibody. The bottom two gels show membranes stained with Coomassie Brilliant Blue (CBB) to indicate protein loading. These experiments were repeated three times with similar results.