Natural variation of leaf shape-related traits in wild Einkorn wheat Triticum urartu Thum.

Haruhiko Morihiro and Shigeo Takumi

Laboratory of Plant Genetics, Graduate School of Agricultural Science, Kobe University, Nada-ku, Kobe 657-8501, Japan

Corresponding author: Shigeo Takumi

E-mail: takumi@kobe-u.ac.jp

The genus Triticum mainly consists of four distinct groups: einkorn (2n=2x=14, nuclear genome constitution AA), emmer (4x, AABB), timopheevi (4x, AAGG) and common wheat (6x, AABBDD). Three species comprising cultivated T. monococcum and wild T. boeoticum and T. urartu belong to the einkorn wheat group. It has been widely accepted that Triticum monococcum was domesticated from T. boeoticum (Dvorák et al. 1988; Takumi et al. 1993; Heun et al. 1997), and that Triticum urartu was the A-genome donor for the polyploid species of Triticum (Dvorák 1976; Chapman et al. 1976; Nishikawa 1983; Dvorák et al. 1988; Takumi et al. 1993; Dvorák et al. 1993). T. urartu is sporadically distributed in Armenia, southeast Turkey, northeastern Iraq and Lebanon (Tumanjan 1938; Johnson and Dhaliwal 1976). Natural variation in T. urartu was previously studied based on nuclear DNA variations (Castagna et al. 1997; Mizumoto et al. 2003) and on morphological traits (Yamagishi 1987). Our previous study showed that both nuclear and chloroplast genomes of T. urartu were clearly differentiated from those of T. monococcum and T. boeoticum, and that the T. urartu accessions were classified into two major haplogroups based on their chloroplast DNA variations (Mizumoto et al. 2003; Fig. 1A).

Yamagishi (1987) studied morphological traits of T. urartu to distinguish from T. boeoticum, in which twelve quantitative traits including flag leaf length and width and some qualitative traits such as auricle pigmentation were examined. The first two principal components from the 12 quantitative traits effectively distinguished the T. urartu accessions from the T. boeoticum ones. In addition, the awn trait and anther length could be used as useful markers to identify these two species (Tumanjan 1938; Yamagishi 1987). Besides these previous reports, information about morphological characteristics of T. urartu is still limited. Leaves of adult plants of T. urartu are drooping because of the long auricle (Fig. 1B). For leaf shape, glabrous leaf blade, sheath and auricle were reported as one of diagnostic characters of T. urartu (Tumanjan 1938; Yamagishi 1987). Therefore, intraspecific variation of quantitative traits related with leaf shape is needed to be studied in T. urartu. Here, we reported natural variation of eight leaf-related traits using 29 accessions of T. urartu, and discussed relationship between the leaf shape variation and nuclear and chloroplast DNA variations.

In this empirical study, we analyzed natural variation of leaf shape-related traits using 29 accessions of T. urartu, two T. boeoticum accessions and two T. monococcum accessions (Table 1, Fig. 1A). The sample set of T. urartu represented the entire natural habitat range. For each accession, we used seeds propagated from a single plant by selfing. Seeds of the sample accessions were sown in November 2004. Plants were grown in a field of Kobe University, and the accessions were arranged in the field using a randomized design. For each accession, a single healthy plant was chosen for analysis of morphological variation. All morphological traits were measured using the three tillers of each plant that headed earliest, and the trait averages and standard deviations were calculated. In total, eight leaf shape-related traits were studied. For the flag leaf, blade length (FLL), blade width (FLW), auricle length (FAL) and auricle width (FAW) were measured (Fig. 1B). For the first leaf below flag leaf, blade length, blade width, auricle length and auricle width were abbreviated as 1LL, 1LW, 1AL and 1AW, respectively. Trait measurements were done using the first, second, and third columns and spikes, for use as the trait values in subsequent analyses. The morphological trait data were statistically analyzed using JMP software ver. 5.1.2 (SAS Institute).

Large natural variations were found in most examined traits, especially FLL and ILL (Table 2). Leaf blade shape of T. urartu was comparatively narrower than those of other einkorn wheat species. Among the examined traits, the highest correlations were significantly observed between FLL and 1LL, and between FLW and 1LW (Table 3). To study intraspecific differentiation in T. urartu, we conducted PC analysis based on the eight leaf traits. Scatter plots with the first two PC values (PC1 and PC2) of the 29 accessions were well united and continuous (Fig. 2). The first two principal components, PC1 and PC2, captured 72.47% of the total variation (48.03% for PC1, and 24.44% for PC2; Table 4).

The genealogical structure of the leaf shape variations was examined using 29 T. urartu accessions. In a graph of the first two axes from the principal component analysis based on eight leaf traits, the plots of the two haplogroups formed separate clusters (Fig. 2A). The cluster with positive PC2 values mainly contained the HG4 accessions, whereas the HG5 accessions belonged to the other cluster with negative PC2 values. The subgroup diversification of T. urartu was largely caused by FLL and 1LL (Table 4). The principal component analysis of the eight leaf traits indicated that two haplogroups based on the chloroplast DNA variations were diverged in the T. urartu population. Significant difference of the leaf length was observed between the two haplogroups (Table 5). The HG5 accessions with the short leaf length contained accessions mainly collected in Armenia and Lebanon, and the HG4 accessions with the long leaf length included a lot of accessions in Iran and Turkey. Significant difference between the HG4 and HG5 accessions was also found in 1AL and 1AW. The HG5 accessions significantly showed longer and wider auricles of the first leaf below flag leaf than the HG4 ones, indicating the auricle length was not necessarily associated with the leaf length.

Five different clusters could be genealogically divided in the T. urartu accessions revealed by the AFLP analysis of total DNA (Mizumoto et al. 2003). In the graph of the first two axes from the principal component analysis, there was no correlation between the plots and the five clusters (Fig. 2B). However, significant differences were found for the six leaf traits, FLL, FAL, 1LL, 1LW, 1AL and 1AW, among the five groups (Table 6). The differences of FLL and 1LL out of the six leaf traits were corresponding to the haplogroup difference.

In conclusion, analysis of natural variation in leaf shape-related traits showed that the T. urartu accessions could be divided into two major classes; one provided larger leaves and another had smaller leaves. The larger-leaf type accessions belonged to HG4 according to the chloroplast DNA variation analysis in einkorn wheat. Clusters based on the nuclear genome variations, group 1 to 5, also showed significant differences in many leaf shape-related traits. However, the difference patterns did not necessarily reflect the phylogenetic relationship among the five nuclear genome clusters. Because T. urartu is an A-genome donor for common wheat, further genetic studies on the intraspecific variations should be important for wheat breeding.

References

Castagna R, Gnocchi S, Perenzin M, Heun M (1997) Genetic variability of the wild diploid wheat Triticum urartu revealed by RFLP and RAPD markers. Theor Appl Genet 94: 424-430.

Chapman V, Miller TE, Riley R (1976) Equivalence of the A genome of bread wheat and that of T. urartu. Genet Res 27: 69-76.

Dvorák J (1976) The relationship between the genome of Triticum urartu and the A and B genomes of Triticum aestivum. Can J Genet Cytol 18: 371-377.

Dvorák J, McGuire PE, Cassidy B (1988) Apparent sources of the A genomes of wheats inferred from polymorphism in abundance and restriction fragment length of repeated nucleotide sequences. Genome 30: 680-689.

Dvorák J, Terlizzi PD, Zhang HB, Resta P (1993) The evolution of polyploid wheats: identification of the A genome donor species. Genome 36: 21-31.

Heun M, Schäfer-Pregl R, Klawan D, Castagna R, Accerbi M, Borghi B, Salamini F (1997) Site of einkorn wheat domestication identified by DNA fingerprinting. Science 278: 1312-1314.

Johnson BL, Dhaliwal HS (1976) Reproductive isolation of Triticum boeoticum and T. urartu and the origin of the tetraploid wheats. Amer J Bot 63: 1088-1094.

Mizumoto K, Hirosawa S, Nakamura C, Takumi S (2003) Nuclear and chloroplast genome genetic diversity in the wild einkorn wheat, Triticum urartu, revealed by AFLP and SSLP analyses. Hereditas 137: 208-214.

Nishikawa K (1983) Species relationship of wheat and its putative ancestors as viewed from isozyme variation. Proc 6th Int Wheat Genet Symp, Kyoto: 59-63.

Takumi S, Nasuda S, Liu YG, Tsunewaki K (1993) Wheat phylogeny determined by RFLP analysis of nuclear DNA. 1. Einkorn wheat. Jpn J Genet 68: 73-79.

Tumanjan MG (1938) A new species of wild wheat Triticum urartu Thum. Proc Arm Bran Acad Sci USSR, Erevan 2: 210-215.

Yamagishi Y (1987) Phylogenetic differentiation between two species of the wild diploid wheats. Genbunsha Kyoto. pp 170.