Additional file 6: of Outgrowth of Rice Tillers Requires Availability of Glutamine in the Basal Portions of Shoots

Figure S6. Comparison of the PAL activity between basal portions of shoots and the third-leaf blade. Rice seedlings were grown hydroponically in the presence of 1 mM NH4Cl until the fourth-leaf stage, and the basal portions of shoots (black column) and third-leaf blades (grey column) were harvested. Mean values with SE of five independent samples are shown. Asterisks denote statistically significant differences between each organ (*, P < 0.05 by Student’s t-test). (EPS 1857 kb

Additional file 3: of Outgrowth of Rice Tillers Requires Availability of Glutamine in the Basal Portions of Shoots

Figure S3. Amino acid analysis in the basal portions of shoots supplied with transient NH4+. Seedlings of the wild type (WT: black column) and two lines of as1 mutants (as1-m1 and as1-m2: dark and light grey column, respectively) at the fourth-leaf stage were grown in water for 3 d, then treated with (+NH4+) or without (-N) 1 mM NH4Cl for 24 h. Mean values with SE of four independent samples are shown. One-way ANOVAs followed by Bonferroni tests were used to identify significant differences between the WT and as1 mutants (P < 0.05). (EPS 1925 kb

Additional file 1: of Outgrowth of Rice Tillers Requires Availability of Glutamine in the Basal Portions of Shoots

Figure S1. qPCR analysis of OsGS1;1 and OsGS1;2 in the basal portions of wild-type shoots using MSX. Wild-type rice seedlings were grown hydroponically in the presence of 1 mM NH4Cl until the fourth-leaf stage and then in water for 3 d. After pre-treatment with 1 mM MSX for 2 h, the seedlings were treated for 8 h with 1 mM NH4+ (MSX + NH4+) or 5 mM glutamine (MSX + Gln). The seedlings without MSX pre-treatment were also treated for 8 h with 1 mM NH4+ (+NH4+), 5 mM glutamine (Gln) or without nitrogen nutrients (-N). qPCR analysis of OsGS1;1 (a) and OsGS1;2 (b) were performed in the shoot basal parts. Fold changes of transcript levels in each treatment relative to those in –N were calculated, and mean values with SE of four independent samples are shown. Asterisks denote statistically significant differences between samples treated without 1 mM NH4+ and each treated sample (*, P < 0.05 by Student’s t-test). (EPS 1807 kb

Additional file 2: of Outgrowth of Rice Tillers Requires Availability of Glutamine in the Basal Portions of Shoots

Figure S2. qPCR analysis of OsAS1 and OsAS2 in roots of wild-type rice using MSX. Wild-type rice seedlings were grown hydroponically in the presence of 1 mM NH4Cl until the fourth-leaf stage and then transferred into water for 3 d. After pre-treatment with 1 mM MSX for 2 h, the seedlings were treated for 8 h with 1 mM NH4+ (MSX + NH4+) or 5 mM glutamine (MSX + Gln). The seedlings without MSX pre-treatment were also treated for 8 h with 1 mM NH4+ (+NH4+), 5 mM glutamine (Gln) or without nitrogen nutrients (-N). qPCR analyses of OsAS1 (a), OsAS2 (b), and actin1 transcripts (c) were performed in roots. Fold changes of transcript levels in each treatment relative to those in –N were calculated, and mean values with SE of four independent samples are shown. Asterisks denote statistically significant differences between samples treated without 1 mM NH4+ and each treated sample (*, P < 0.05 by Student’s t-test). (EPS 1890 kb

Additional file 2: Figure S2. of A temporal and spatial contribution of asparaginase to asparagine catabolism during development of rice grains

Quantitative real-time PCR detection of transcripts for OsAS in rice flag leaves (A) and developing spikelets (B). The flag leaves were harvested during the ripening period from 14 to 35 days after flowering, respectively. Quantitative real-time PCR was performed using gene-specific primers for OsAS1, OsAS2 and actin, like described in Table 1. The relative contents of these transcripts were normalized against actin transcript. Means of three to four independent samples and standard error values (n = 3 to 4) are indicated. (PDF 405 kb

Additional file 1: Figure S1. of A temporal and spatial contribution of asparaginase to asparagine catabolism during development of rice grains

Quantitative real-time PCR detection of transcripts for OsASNase1 (filled column) and OsASNase2 (opened column) in roots, leaf blades and leaf sheaths at different leaf positions in rice. Rice plants were grown hydroponically for 18 d in water and each organ was harvested individually; whole roots, 2nd leaf blade, 3rd leaf blade, 3rd leaf sheath, and 4th expanding leaf blade. Quantitative real-time PCR analyses were performed using gene-specific primers for OsASNase1, OsASNase2 and actin, respectively, as described in Table 1. The relative contents of these transcripts were normalized against actin transcript. Means of four independent samples and standard error values (n = 4) are indicated. Significant differences between OsASNase1 and OsASNase2 were identified by Student’s t-test, and are marked with asterisks: *P < 0.05 (PDF 401 kb