LB Hpt 35S AdhI 755 bp Waxy-a 755 bp OCS GUS RB

1. ATG TAA 93 173 130 138 333 184 170 11 305 269 1024 181 217 73 434 EcoRI AscI AvrII SacI SpeI HindIII a LB Hpt 35S AdhI 755 bp Waxy-a 755 bp OCS GUS RB OsMPK6 cDNA OsMPK6 cDNA Intron I of rice Clone D87R in vector pDS1301 b T-DNA insertion site of mutant 03Z11FX66 Supplemental Fig. 1 Schematic diagrams of the OsMPK6 gene and its transformation construct. RB and LB, right and left T-DNA border; GUS, -glucuronidase gene; Hpt, hygromycin phosphotransferase gene; 35S, cauliflower mosaic virus 35S promoter; OCS, octopine synthase polyadenylation signal. (a) RNAi construct of OsMPK6. (b)OsMPK6 gene structure and the T-DNA insertion site of OsMPK6-knockout mutant. The coding region (black boxes) of OsMPK6 is interrupted by six introns (thick lines). The positions of 5'- and 3'-untranslated regions (hatched boxes), translation start codon (ATG), and translation stop codon (TAA) are also indicated. The numbers indicate the base pairs of each substructure.

2. Nipponbare IR24 IRBB13 IRBB4 Zhenshan 97 Minghui 63 Nipponbare IR24 IRBB13 IRBB4 Zhenshan 97 Minghui 63 Nipponbare IR24 IRBB13 IRBB4 Zhenshan 97 Minghui 63 Nipponbare IR24 IRBB13 IRBB4 Zhenshan 97 Minghui 63 Nipponbare IR24 IRBB13 IRBB4 Zhenshan 97 Minghui 63 DraI BamHI PstI HindIII EcoRV Supplemental Fig. 2 DNA gel blot analysis of OsMPK6-homologous gene in different rice lines. Minghui 63, Zhanshan 97, IRBB4, IRBB13 and IR24 are indica rice lines and Nipponbare is japonica rice line.

3. a OsMPK6-suppressing T0 plants (D87RZ1-) 120 100 Relative expression level 80 60 40 20 0 3 7 10 13 17 18 19 20 28 30 34 35 36 ck b 140 OsMPK6-suppressing T1 family D87RZ1-20 120 100 80 Relative expression level 60 40 20 0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 ck Supplemental Fig. 3 OsMPK6 expression in OsMPK6-suppressing plants. Zhonghua 11 is wild type (ck). The band density of RNA gel blot analysis was quantified using a computer program Multi Gauge (Version 3.0, Fujifilm, Japan). The band density of rRNA was used to standardize the RNA sample. The expression level of each treatment at each time point is relative to that of wild type (Mei C. et al. 2006. Mol. Plant-Microbe Interact. 19:1127-1137). (a) The phosphoimaging quantification of the RNA gel blot analysis of T0 transgenic plants showing in Fig. 2a. D87RZ1-3 is a negative transgenic plants and others are positive transgenic plants. (b) The phosphoimaging quantification of the RNA gel blot analysis T1 trangenic plants showing in Fig. 2b.

4. a D87RZ1- 20 28 34 ck 100 90 80 70 Lesion area (%) 60 50 40 * 30 * * 20 10 28 0 ck 20 34 D87RZ1- b Supplemental Fig. 4 Resistance of homozygote OsMPK6-suppressing lines (D87RZ1-20, -28 and -34) to Xanthomonas oryza pv. oryzae strain PXO61 at 5- to 6-leaf stage. ck, negative transgenic plants with Zhonghua 11 background. (a) leaves of transgenic plants showing enhanced resistance at 21 days after inoculation. (b) OsMPK6-suppressing plants showed enhanced resistance. Bars represent mean  standard deviation. The asterisk indicates that a significant difference (P < 0.01) in lesion area was detected between transgenic plants and control (ck) at 21 days after bacterial inoculation.

5. 0 2 4 0 0 2 2 4 4 ck 20 28 34 ck 20 28 34 ck 20 28 34 OsMPK6 PR5 rRNA ck 20 28 34 ck 20 28 34 ck 20 28 34 PR10/PBZ1 rRNA PAL ck 20 28 34 ck 20 28 34 ck 20 28 34 ck 20 28 34 WRKY03 Cht1 rRNA NH1 Supplemental Fig. 5 Expression of OsMPK6 and other defense-related genes (PR5, PAL, PR10/PBZ1, WRKY03, NH1, and Cht1) in OsMPK6-suppressing T0 plants analyzed by RNA gel blot. Zhonghua 11 is the wild-type plant (ck). Samples were collected from T0 plants D87RZ1-20, D87RZ1-28, and D87RZ1-34 before pathogen (PXO61) inoculation (0 day) and at 2 and 4 days after pathogen inoculation (lesion mimics developed).