Molecular Identification And Breeding Utilization Of High Molecular Weight Glutenin Genes In Aegilops Tauschii

The evolution bottleneck of common wheat (Triticum aestivum L.) largely reduced the genetic diversity of common wheat lines, especially in its D genome. Aegilops tauschii Coss., the D-genome progenitor of common wheat, has a wide geographical distribution and a large of genetic variations. High-molecular-weight glutenin subunits (HMW-GS) play an important role in determining bread-making quality of common wheat, which is largely determined by the Glu-D1locus in D-genome. Ae. tauschii has a much higher HMW-GS diversity than common wheat on Glu-D1locus. Identification and utilization of HMW-GS genes in Aegilops tauschii will be useful to improve the quality of common wheat.This study is aimed to report the discovery and characterization of novel x-and y-type HMW-GS alleles from Ae. tauschii, to discuss their phylogenetic relationships with previously identified alleles on the Glu-D1loci of T. aestivum and Ae. tauschii, to introduce novel HMW-GS genes from Ae. tauschii into elite common wheat lines via synthetic hexaploid wheat, and to evaluate the effect of novel HMW-GSs from Ae. tauschii on the quality of flour. The main results are as follows:1. A pair of novel high molecular weight glutenin subunits (HMW-GS)1Dx3.1t and1Dy11*t were revealed and characterized from Aegilops tauschii Coss. subspecies tauschii accession AS60. SDS-PAGE band of1Dx3.1t was between those of1Dx2and1Dx3, while1Dy11*t was between1Dy11and1Dy12. The lengths of1Dx3.1t and1Dy11*t were2,514bp and1,968bp, encoding836and654amino acid residues, respectively. Their authenticity was confirmed by successful expression of the coding regions in Escherichia coli. Network analysis indicated that1Dx3.1t together with other five rare alleles only detected in Asia common wheat populations represented the ancestral sequences in Glu-D1locus. Neighbor-joining tree analysis of previously cloned x-type and y-type alleles in the Glu-D1locus supported the hypothesis that more than one Ae. tauschii genotypes were involved in the origin of hexaploid wheat and that different Ae. tauschii accessions contributed the D genome to common wheat and Ae. cylindrical Host, respectively. An Ae. tauschii accession with1Dx3.1t or a closely related allele probably have involved in the origin of common wheat. Since accession AS60used in this study belonged to typical ssp. tauschii, present results suggested the possibility that ssp. tauschii was involved in the evolution of common wheat.2. A pair of novel high-molecular-weight glutenin subunits (HMW-GS)1Dx5.3t and1Dy12.1**t were revealed and characterized from Ae. tauschii accession PI554324. SDS-PAGE band of10x5.3t was between those of1Bx6and1Bx7, while1Dy12.1**twith slightly faster migration rate than that of1Dyl2. The lengths of1Dx5.3t and1Dy12.1**t were2115bp and1986bp, encoding703and660amino acid residues, respectively. Their authenticity was confirmed by successful expression of the coding regions in Escherichia coli.1Dx5.3t is the shortest of the known Dx-type alleles.1Dy12.1**t is also a special subunit since it has an additional cysteine in the front of the central repetitive domain. This cysteine that is not existed in previously reported Dy-type genes may be useful for improving bread wheat quality. Median-joining Network analysis indicated that1Dyl2.1*may be a key site in the genealogy of the Glu-Dy.3. To reveal the transmission route of Ae. tauschii accessions in Yili River valley region of Xinjiang and the middle reaches of the Yellow River of China, HMW-GS genes of nine Chinese Ae. tauschii accessions from these regions were cloned, sequenced, and compared each other. The results from phylogenetic analysis suggested that at least two Ae. tauschii accessions were transmmited into Xinjiang and the middle reaches of the Yellow River in the history. One possible route is that AS60-like accession in Middle East was eastward spread to Yili River valley region by human activities or natural extension. It was gradually evolved as accession AS71at there. This accession was also eastward spread to the middle reaches of the Yellow River and gradually evolved to AS75or its similar accessions. In addition, another Ae. tauschii accession from the middle east was also eastward spread to Yili River valley region. It was gradually evolved to AS72at there. This accession was also continued to eastward spread to the middle reaches of the Yellow River. The pathway of Chinese Ae. tauschii transmission may be the famous Silk Road. The original accession was gradually evolved to different groups in order to adapt to different habitats.4. Nine synthetic hexaploid wheats were developed by crossing nine T. turgidum lines belonging to four subspecies with Ae. tauschii accession AS60. Their authenticity was confirmed by morphological observation and SDS-PAGE analysis. These synthetic hexaploid wheats showed many undesirable agronomic traits, which can be used as the bridge for the gene transferring of the nine T. turgidum and Ae. tauschii AS60parents. Their agronomic traits can be improved by repeatedly backcrossing with elite common wheat cultivars.5. After backcrossing synthetic hexaploid wheat Syn-SAU-1(AS2255×AS60) with elite common wheat cultivars Chuanmai32, Chuanyu18, and Chuanmai42, and then selfing three times, four common wheat sister lines (23、24、25and26) were developed. The four lines showed desirable agronomy characters and were considered as elite lines, which was confirmed at Wenjiang of Sichuan province in2009-2010and2010-2011crop seasons and at Dujiangyan and Wenjiang of Sichuan provinve in2011-2012crop season and Xining of Qinghai province in2011-2012. The lines23and24have the HMW-GS1Dx3.1t+1Dy11*t, while25and26have1Dx5+1Dy10at Glu-Dl locus. All the four lines had1Ax1at Glu-Al locus and1Bx6+1By8at Glu-B1locus. Based on the analysis on11quality parameters of the four lines, it was indciated that1Dx3.1t+1Dy11*t was slightly worse than1Dx5+1Dy10on the baking quality of wheat flour.