Supporting Information
Figure S1. Polyploid evolution of wheat. Hexaploid wheat (T. aestivum) was originated by
natural hybridization between tetraploid wheat (T. turgidum) and the diploid wild relative, Ae.
tauschii. In turn, tetraploid wheat arose by hybridization between two wild diploids (T. urartu
and probably Ae. speltoides). Hybridization events are indicated by broken arrows and
domestication by solid arrows. The cultivated diploid wheat T. monococcum ssp. monococcum
was obtained from domestication of T. monococcum ssp. aegilopoides (syn. T. boeoticum).
Diploids with A-, S- and D-genomes are indicated in boxes with red dotted lines. Species and
subspecies used in this study are indicated by purple boxes.
Figure S2. Percent of reads classified as gene in different subspecies. (a) Percent of genes in
amplicon sequencing which include sequences without premature stop codon and frame shift
(the real genes) and those containing mutation only downstream of PIS which we cannot
distinguish between them as mutations are out of the amplicon. (b) Predicted percent of genes in
amplicon based on the results obtained by Sanger sequencing.
Figure S3. Alignment of the C-terminal end motif of α-gliadin. The first two nucleotide
sequences, present in most of the α-gliadins such as Type 1, 2 and 3, were targeted for
amplification and cloning. In Type 6 α-gliadins, the C-terminal end presents three mismatches
compared with the first two. This hinders the annealing of primers and decreases the
amplification efficiency. Amino acid sequences are shown under corresponding nucleotide
sequences. Mismatches are indicated by red. Accession numbers of Type 6 α-gliadins found in
NCBI database are shown.
Figure S4. Phylogenetic network of α-gliadins constructed using complete coding sequence. In
total, 478 sequences of the tribe Triticeae, excluding polyQ regions (Fig. S2), were used. These
included 207 sequences obtained by Sanger method in this study and 271 found in NCBI
database (124 of Triticum/Aegilops and 147 of other Triticeae species). Type 1 α-gliadins are
indicated in blue, Type 2 in red, Type 3 in green, Type 6 in gray and other types in black.
Variants of the canonical forms are indicated by circles. In the case of Type 1, variants of
Subtype 1.1 are indicated by circles and Subtypes 1.2 or 1.3 by squares. Number indicates
percentage of branch support estimated by 1000 bootstrap test.