Location:2009 Annual Report
1a. Objectives (from AD-416)
1) Apply molecular biology and genomics to understand the structure, function and expression of wheat seed protein genes; 2) Contribute to the understanding of the organization of the wheat genome, with a focus on the group 1 chromosomes; and 3) Develop and utilize Brachypodium as a model system for Triticeae research for grain quality and other traits.
1b. Approach (from AD-416)
1) Use combinations of molecular alterations, heterologous expression, and plant transformations in the application of molecular biology and genetic engineering to wheat high-molecular-weight (HMW) glutenin class seed storage protein genes and other seed protein classes to understand more of the molecular bases of wheat quality and utilization. 2) Contribute to the understanding of the organization of the wheat genome through in-depth studies of wheat seed protein genetic loci and participation in national and international wheat genome characterization projects. 3) Employ DNA microarrays to profile gene expression during wheat seed development and changes in storage protein gene activity and wheat quality. 4) Contribute to the development of Brachypodium as a model for small grains research, to include use in studying wheat seed protein gene controls. [Previously was 5325-21430-011-00D (10/03). Replacing 5325-21000-011-00D (04/08).]
3. Progress Report
We have made significant progress in the genetic and molecular analysis of genes that influence economically important wheat traits. The omega gliadins, which comprise about 5-10% of wheat flour protein, have been implicated in wheat flour quality and wheat-related allergies. To further understand the biology of omega gliadins, we analyzed all available omega gliadin sequence information and investigated the details of the omega gliadin coding regions. Sequences derived from the orthologous Gli-3 loci in chromosome 1A, 1B and 1D were compared. The transcriptional activities of the omega gliadin genes were examined and the evidence of cysteine-containing omega gliadins in wheat was assessed. To further analyze genes coding for major seed storage proteins in wheat, five BAC clones containing LMW-glutenin genes derived from a large contig spanning the protein storage protein regions have been sequenced and annotated. Comparative analysis with the orthologous regions from other cereal genomes has been performed. Several marker systems based upon TE sequences have been developed, mostly based upon polymorphism at their sites of insertion. We exploited this property to develop molecular markers for genetic analysis in wheat. We utilized the retrotransposon-based insertion polymorphism to generate repeat DNA junction markers (RJM) to map four randomly chosen BAC clones to wheat chromosome arms. Our work allowed the rapid development of genome-specific PCR-based RJM using BES and random shotgun sequences from the D genome donor Ae. tauschii for use in hexaploid wheat. In a study together with ARS and non-ARS collaborators, the effect of polyploidy in allohexaploid wheat was investigated. Large-scale analysis of nonadditive gene expression using DNA microarrays showed that approximately 16% of genes displayed nonadditive expression in synthetic T. aestivum. A certain fraction of the genes (2.9%) showed overdominance or underdominance. The data demonstrated that allopolyploidization, per se, results in rapid initiation of differential expression of homoeologous loci and nonadditive gene expression in T. aestivum. The on-going project to develop and utilize Brachypodium as a model system for Triticeae research for grain quality and other traits has progressed significantly. We completed the construction of Brachypodium genome physical map and performed genome-wide comparison with wheat and rice. A comprehensive study of syntenic relationship between Brachypodium and wheat was carried out. Seven wheat BAC contigs each spanning a ~ 1Mb region has been sequenced and annotated and detailed comparative analysis with Brachypodium orthologous regions has been performed. Furthermore, the identity of the major seed storage proteins in Brachypodium was determined for further dissection of storage protein biology in grasses.
1. Structure of the omega-gliadin gene family. Wheat is uniquely suited to make dough mixtures that possess viscoelastic properties that allow a wide range of food products including leavened and unleavened breads, biscuits, cookies, cakes, pasta, and as a common ingredient in many processed foods. However, the components of wheat flour that contribute to dough quality are not fully understood. ARS scientists in Albany, CA characterized the molecular structure of the family of genes that encode the omega gliadins, one of the major proteins found in wheat seed. The results will facilitate our understanding and genetic engineering of omega gliadins contribution to the unique physical properties of wheat doughs in order to improve these properties.
2. Rapid development of Polymerase Chain Reaction (PCR)-based genome-specific repetitive DNA junction markers in wheat. Wheat is a major domestic crop and an important dietary grain in the American diet yet the complexity of the wheat genome impedes genetic analysis of useful genes in wheat. ARS scientists in Albany, CA exploited the unique and abundant transposable element (TE) insertion junction regions from a diploid wheat to develop genome-specific markers for use in hexaploid wheat. The results indicated that high percentage of identified markers from the diploid genome can be directly applied to the hexaploid wheat for various wheat mapping projects. This novel approach allows for easy identification of thousands of unbiased molecular makers that will facilitate the characterization and cloning of agronomically important genes for crop improvement.
Stamova, B., Roessner, U., Suren, S., Chingcuanco, D.L., Bacic, A., Beckles, D. 2008. Metabolic profiling of transgenic wheat over-expressing the high-molecular weight Dx5 glutenin subunit. Metabolomics. 5(2):239-252.