Supplemental Data

Files in this Data Supplement:

• Supplemental Figure 1 - Supplemental Figure 1. Cell cycle activity during intermediate stages of caryopsis development. Regions showing cell cycle activity as visualised by RNA in situ hybridisation with the S-phase marker, histone H4 at intermediate stages 4DAA, 5DAA, 5-6DAA and 6-7DAA (ii). Scale bar = 0.3mm
• Supplemental Figure 2 - Supplemental Figure 2. Transcript labelling in the modified aleurone showing spatial and temporal variations at 3, 6 and 9 DAA as shown by in situ hybridization of transverse sections of the caryopsis probed with digoxigenin-labeled antisense RNA and viewed under bright-field optics. (A) a typical expression pattern in the modified aleurone as revealed by an END1 (ID 702037156) probe. (B) accumulation of ethylene forming enzyme ACC oxidase transcript (ID 702006333). (C) detection of an unknown gene (ID 702038349) transcript in the peripheral cells of the modified aleurone. (D) detection of an unknown gene transcript in the central endosperm before restriction to the modified aleurone at 9DAA. The main tissue types are annotated as follows: cr, crease region; np, nucellar projection; e, endosperm. Scale bar = 0.3mm
• Supplemental Figure 3 - Supplemental Figure 3. Transcript labelling in the nucellus showing spatial and temporal variations at 3, 6 and 9 DAA as shown by in situ hybridization of transverse sections of the caryopsis probed with digoxigenin-labeled antisense RNA and viewed under bright-field optics. (A) A wheat allergen gene (ID 701968577) is also expressed in the tip of the nucellar projection at 6 and 9 DAA but is already detectable in this zone at 3DAA. (B) A gene encoding a hypothetical protein similar to a family of dehydration response proteins in rice and Arabidopsis (ID 702 013671) is expressed in the integuments, nucellar projection and modified aleurone with strongest expression at 3DAA. (C) A gene of unknown function (ID 702040719) shows extremely high expression in the adaxial nucellar projection and in the cells surrounding the vascular tissue. At 3DAA this expression extends to the xylem bundles in the pericarp. The main tissue types are annotated as follows: cr, crease region; np, nucellar projection; e, endosperm. Scale bar = 0.3mm
• Supplemental Figure 4 - Supplemental Figure 4. Transcript labelling in the integuments showing spatial and temporal variations at 3, 6 and 9 DAA as shown by in situ hybridization of transverse sections of the caryopsis probed with digoxigenin-labeled antisense RNA and viewed under bright-field optics. (A) Expression of a proteinase inhibitor gene (ID 702015496) in both integuments and in the xylem vessels of the vascular bundle. (B) A GDSL-lipase gene (ID 701965571) is expressed in the outer integument (C) A serine proteinase gene (ID 702011770) is expressed in the integuments and the tip of the nucellar projection (D) A chalcone synthase gene (702001184) is expressed in the pigment strand. The main tissue types are annotated as follows: cr, crease region; nu, nucellus; in, integuments; oin outer integuments; np, nucellar projection; e, endosperm; p, pericarp; ps, pigment strand; ma, modified aluerone; v, vascular region. Scale bar = 0.3mm
• Supplemental Figure 5 - Supplemental Figure 5. Expression patterns of putative transcriptional regulators in developing caryopsis at 3DAA (A), 6DAA (C and D) and 9DAA (B, E and F). (A and B) Mads-13 orthologue (ID 701995119) in outer integument and vasculature at 3DAA and at 9DAA. (C) A HMG-like gene (ID 701990762) expressed in a stochastic cell-cycle-phase-specific pattern. (D) A gene encoding a zinc-finger-domain protein (ID 701990488) expressed in nucellar projection, integuments and endosperm. (E) The transcript of a gene encoding a leucine-zipper-domain protein (ID 701769401) is highly restricted to the nuclei of the endosperm. (F) An RCD1-like gene (ID 702012812) expressed in the central endosperm. The main tissue types are annotated as follows: cr, crease region; oin, outer integuments; np, nucellar projection; e, endosperm; v, vascular region. Scale bar = 0.3mm
• Supplemental Table 1 - Supplemental Spreadsheet 1. Summary of wheat genes, predicted functions and patterns of expression. Transcript accumulation for each gene was classified according to the pattern (spotty, general proliferation associated, nuclear) or the cell type showing highest level of signal (pericarp, integuments, nucellus, vascular, modified aleurone, aleurone, central endosperm, or general expression in all cell types). The columns are as follows: B and C, plate and well position of the probe; D, clone ID; E, gene annotation; F-I, slide prefix and position number; J, pattern of transcript accumulation (abbreviations are as follows: spot - spotty; gen - general proliferation associated; pc - pericarp; pe - pericarp epidermis); K, predicted function (abbreviations are as follows rep - cell cycle, chr - chromatin binding, rna - RNA-binding, rot - protein synthesis, tf - transcription factor, sig - signalling, deg - degradation, def - defence, cw - cell wall, sto - storage proteins, met - metabolism (enzymes), tran - transport and membrane proteins, cyto - cytoskeleton); L, DNA sequence of each clone.
• Supplemental Table 2 - Supplemental Spreadsheet 2. Arabidopsis seed mutants corresponding to wheat genes. Columns show A, Arabidopsis mutant, mutant defect; B, At gene number; C, At annotation; E, expression pattern in wheat (abbreviations are as follows: spot - spotty; gen - general proliferation associated; pc - pericarp; pe - pericarp epidermis; ma - modified aleurone; np nucellar projection; int - integuments; en - endosperm; cse - central starchy endopserm); F, wheat gene ID with significant homology.