Present address: Department of Applied Biological Chemistry, Graduate School of of Agricultural Life Sciences, The University of Tokyo, Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan.
Temporal and spatial regulation of DROOPING LEAF gene expression that promotes midrib formation in rice
Article first published online: 29 NOV 2010
© 2010 The Authors. The Plant Journal © 2010 Blackwell Publishing Ltd
The Plant Journal
Volume 65, Issue 1, pages 77–86, January 2011
How to Cite
Ohmori, Y., Toriba, T., Nakamura, H., Ichikawa, H. and Hirano, H.-Y. (2011), Temporal and spatial regulation of DROOPING LEAF gene expression that promotes midrib formation in rice. The Plant Journal, 65: 77–86. doi: 10.1111/j.1365-313X.2010.04404.x
- Issue published online: 22 DEC 2010
- Article first published online: 29 NOV 2010
- Accepted manuscript online: 19 OCT 2010 11:30PM EST
- Received 9 August 2010; revised 28 September 2010; accepted 11 October 2010; published online 29 November 2010.
Figure S1. Alignments of the nucleotide sequences of CNS-C, CNS-D and CNS-E. Bold lines indicate the CNSs: CNS-C (red), CNS-D (blue) and CNS-E (orange). Nucleotides that are conserved among four species are indicated in black, and those conserved among three species are indicated in gray. CNS-E is divided into three subdomains: E1 (solid line), E2 (dotted line) and E3 (wavy line). Numbers indicate the positions (bp) from the translation initiation site of each DL ortholog.
Figure S2. The large deletion in dl-1 and its flower phenotype. (a) The genomic region around the DL locus. The red box indicates the large deletion (-13,590 to -6,335) far upstream region of dl-1, and the blue box indicates DG1. Exons are indicated by black bars (RAP-DB build 4, http://rapdb.dna.affrc.go.jp/). Thick arrows indicate the orientation of each gene. (b) Flower phenotype of dl-1. The palea and lemma have been removed. st, stamen; ca, carpel. Bar = 500 mm.
Figure S3.GUS reporter analysis to examine the effect of the position of intron 2. (a) and (b) Schematic representation of the GUS reporter constructs. (a) Intron 2 of DL is placed at a region upstream of the 35S minimal promoter-GUS fusion. (b) A DNA segment containing a short promoter region and part of exon 1 is fused to the GUS gene (translational fusion) and intron 2 is placed at a region upstream of the DL promoter. Numbers indicate the position from the putative transcription initiation site (+1). CNSs are shown by orange boxes (CNS-C and CNS-E). Arrows indicate the translation start site (ATG). (c) and (d) GUS staining pattern of a transgenic plant (T0 generation) carrying the constructs shown in (a) and (b), respectively. No expression is observed in (c). Weak and expanded GUS staining is observed in P3, but not in P1 and P2 leaf primordia (this pattern is similar to that observed in DG5::GUS plants; see Figure 3 (b)). Outlines of the leaf primordia (P1 to P3) are traced by unbroken lines. Bar = 100 mm.
Figure S4. Schematic representation of the construction processes used to make the DG::GUS constructs (DG1-DG7). (a) Positions of fragments (fA-fH) used for DG::GUS construction. Letters below the vertical dotted lines indicate the endogenous restriction sites used for construction. The white bar and black bars indicate the 3’ UTR and exons of DL, respectively. (b) Flow chart of pGEM-A, pGEM-B, pGEM-C, pTA-D, pTA-G and pTA-H construction. (c) Flow chart of DGfA::GUS construction. (d) Flow chart of DG3::GUS, DG4::GUS, and DG5::GUS construction. (e) Flow chart of DG6::GUS and DG7::GUS construction. (f) Flow chart of DG1::GUS construction. (g) Flow chart of DG2::GUS construction.
Figure S5. Schematic representation of the construction processes used to make the DG::GUS constructs incorporating internal deletions. (a) Positions of the region deleted in each construct. The region removed from fragment fJ in each construct is shown by bilateral arrows labeled K to Q. The white bar and black bars indicate the 3’ UTR and exons of DL, respectively. Numbers below the vertical dotted lines indicate the position (bp) from the putative transcription initiation site (+1). The endogenous restriction site (ApaI) used for construction is indicated. (b) Flow chart of pT7-J construction. (c) Flow chart of DG8::GUS construction. (d) Flow chart of DG7 DFS::GUS construction. (e) Flow chart of DG7DCNS-E::GUS construction. (f) Flow chart of DG7DE1::GUS, DG7DE2::GUS, and DG7DE3::GUS construction.
Figure S6. Flow chart of the construction processes used to make the DG1::DL-VP16 construct.
Table S1. Primers used in this paper.
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