Project description:Here is reported the first study of transcriptome analyses using the Illumina HiSeq 4000 platform for three kinds of wheat (G represents Strong gluten wheat, Z represents middle gluten wheat,R represents weak gluten wheat). The variation of wheat varieties with different gluten content is mainly shown in the content of gluten, flour is divided into high gluten powder ( > 30%), medium gluten powder (26%-30%) and low gluten powder ( < 20%), according to the wet gluten content. In total, over 102.6 Gb clean reads were produced and 114, 621 unigenes were assembled; more than 59,085 unigenes had at least one significant match to an existing gene model. Differentially expressed gene analysis identified 2339 and 2600 unigenes which were expressed higher or lower among strong gluten, middle gluten and weak gluten wheat. After functional annotation and classification, three dominant pathways including protein isomerase, antioxidase activity and energy metabolism, and 410 unigenes related to gluten strength polymerization of wheat were discovered. In strong-gluten wheat, low molecular weight subunit content is higher than weak-gluten wheat, and the activity of cysteine synthase and isomerase is increased, which may promote the cross-linking of low molecular weight protein to high molecular weight protein. Meanwhile, POD enzyme strengthens gluten network and CAT enzyme affects gluten polymerization, along with higher ATPase activity, which will provides energy for protein polymerization reaction in comparison of strong-gluten wheat and weak-gluten wheat. The accuracy of these RNA-seq data was validated by qRT-PCR analysis. These data will extend our knowledge of quality characteristics of wheat and provide a theoretical foundation for molecular mechanism research of wheat.
Project description:Rye, wheat and barley contain gluten, proteins that trigger immune-mediated inflammation of the small intestine in people with coeliac disease (CD). The only treatment for CD is a lifelong gluten-free diet. To be classified as gluten-free by the World Health Organisation the gluten content must be below 20 mg/kg, but Australia has a more rigorous standard of no detectable gluten and not made from wheat, barley, rye or oats. The purpose of this study was to devise an LC-MS/MS method to detect rye in food. An MS-based assay could overcome some of the limitations of current immunoassays, wherein antibodies often show cross-reactivity and lack specificity due to the diversity of gluten proteins in commercial food and the homology between rye and wheat gluten isoforms. Comprehensive proteomic analysis of 20 rye cultivars originating from 12 countries enabled the identification of a panel of candidate rye-specific peptide markers. The peptide markers were assessed in 16 cereal and pseudo-cereal grains, and in 10 breakfast cereals and 7 snacks foods. Spelt flour was contaminated with rye at a level of 2% and trace levels of rye were found in a breakfast cereal that based on its labelled ingredients should be gluten-free.
Project description:While the antigenic specificity and pathogenic relevance of immunologic reactivity to gluten in celiac disease have been extensively researched, the immune response to non-gluten proteins of wheat has not been characterized. We aimed to investigate the level and molecular specificity of antibody response to wheat non-gluten proteins in celiac disease. Serum samples from patients and controls were screened for IgG and IgA antibody reactivity to a non-gluten protein extract from the wheat cultivar Triticum aestivum 'Butte 86'. Antibodies were further analyzed for reactivity to specific non-gluten proteins by immunoblotting, following two-dimensional gel electrophoretic separation. Immunoreactive molecules were identified by tandem mass spectrometry. Compared with healthy controls, patients exhibited significantly higher levels of antibody reactivity to non-gluten proteins. The main immunoreactive non-gluten antibody target proteins were identified as serpins, purinins, α-amylase/protease inhibitors, globulins, and farinins. Assessment of reactivity towards purified recombinant proteins further confirmed the presence of antibody response to specific antigens. The results demonstrate that, in addition to the well-recognized immune reaction to gluten, celiac disease is associated with a robust humoral response directed at a specific subset of the non-gluten proteins of wheat
Project description:Drought is among the most limiting factors for sustainable agricultural production. Water shortage at the onset of flowering severely affects the quality and quantity of grain yield of bread wheat (Triticum aestivum). Herein, we measured oxidative stress and photosynthesis-related parameters upon applying transient drought on contrasting wheat cultivars at the flowering initiation stage of ontogenesis. The sensitive cultivar showed ineffective water management and a more severe decline of photosynthesis. Apparently, the tolerant genotype used photorespiration to dissipate excessive light energy. The tolerant cultivar sooner induced superoxide dismutase and showed less inhibited photosynthesis. Such protective effect resulted in less affected yield and spectrum of seed proteome. The tolerant cultivar had a more stable gluten profile, which defines bread-making quality, upon drought. Drought caused the accumulation of medically relevant proteins: (i) components of gluten in the sensitive cultivar and (ii) metabolic proteins in the tolerant cultivar. We propose specific proteins as markers of drought tolerance for guiding efficient breeding: thaumatin-like protein, 14-3-3 protein, peroxiredoxins, peroxidase, FBD domain protein, and Ap2/ERF plus B3 domain protein.
Project description:Dietary gluten proteins (prolamins) from wheat, rye, and barley are the driving forces behind celiac disease, an organ-specific autoimmune disorder that targets both the small intestine and organs outside the gut. In the small intestine, gluten induces inflammation and a typical morphological change of villous atrophy and crypt hyperplasia. Gut lesions improve and heal when gluten is excluded from the diet and the disease relapses when patients consume gluten. Oral immune tolerance towards gluten may be kept for years or decades before breaking tolerance in genetically susceptible individuals. Celiac disease provides a unique opportunity to study autoimmunity and the transition in immune cells as gluten breaks oral tolerance. Seventy-three celiac disease patients on a long-term gluten-free diet ingested a known amount of gluten daily for six weeks. A peripheral blood sample and intestinal biopsies were taken before and six weeks after initiating the gluten challenge. Biopsy results were reported on a continuous numeric scale that measured the villus height to crypt depth ratio to quantify gluten-induced gut mucosal injury. Pooled B and T cells were isolated from whole blood, and RNA was analyzed by DNA microarray looking for changes in peripheral B- and T-cell gene expression that correlated with changes in villus height to crypt depth, as patients maintained or broke oral tolerance in the face of a gluten challenge.
Project description:Amylase/trypsin-inhibitors (ATIs) are putative triggers of nonceliac gluten sensitivity, but contents of ATIs in different wheat species were not available. Therefore, the predominant ATIs 0.19 + 0.53, 0.28, CM2, CM3, and CM16 in eight cultivars each of common wheat, durum wheat, spelt, emmer, and einkorn grown under the same environmental conditions were quantitated by targeted liquid chromatography-tandem mass spectrometry (LC−MS/MS) and stable isotope dilution assays using specific marker peptides as internal standards. The results were compared to a label-free untargeted LC−MS/MS analysis, in which protein concentrations were determined by intensity based absolute quantitation. Both approaches yielded similar results. Spelt and emmer had higher ATI contents than common wheat, with durum wheat in between. Only three of eight einkorn cultivars contained ATIs in very low concentrations. The distribution of ATI types was characteristic for hexaploid, tetraploid, and diploid wheat species and suitable as species-specific fingerprint. The results point to a better tolerability of einkorn for NCGS patients, because of very low total ATI contents.
Project description:The prevalence of hypersensitivities towards wheat has increased in the last decades. Apart from celiac disease these include allergic and other inflammatory reactions summarized under the term non-celiac wheat sensitivity. One suspected trigger is the family of amylase/trypsin-inhibitors (ATIs), non-gluten proteins that are prominent wheat allergens and that activate the toll-like receptor 4 on intestinal immune cells to promote intestinal and extra-intestinal inflammation. We therefore quantified 13 ATIs in 60 German hexaploid winter wheat cultivars originating from 1891 to 2010 and harvested in three years by targeted liquid chromatography-tandem mass spectrometry combined with stable isotope dilution assay using specific marker peptides as internal standards. The total ATI content and that of the two major ATIs 0.19 and CM3 did not change from old cultivars (first registered from 1891–1950) to modern cultivars (1951–2010). There were also no significant changes in ATI distribution.
Project description:Objective: Germination of wheat maximizes phytochemical content and antioxidant activity while altering chemical composition, gluten content, and pasting properties. We previously reported marked changes in gene expression in soybean prior to germination before the radicle had grown from a seed. The present study investigated whether imibibition induces similar changes in wheat. Methods: Changes in gene expression profiles of wheat during short-term imbibition (0, 16, and 24 h) were evaluated by DNA microarray analysis. Gene Ontology (GO) analysis was carried out to categorize the function of genes with altered expression. The expression of genes encoding enzymes associated with starch breakdown was evaluated by quantitative real-time PCR, and changes in enzymatic activity were assessed with functional assays. Pasting properties of flour made from wheat seeds imbibed for different times were examined with a Rapid Visco Analyzer. The protein profile was determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis, and gluten content was quantified. Results: The GO analysis revealed that genes related to cellulose and cell wall synthesis were upregulated by imbibition for 16 h whereas those associated with polysaccharide catabolism and nucleosome assembly were upregulated in the subsequent 8 h. α-Amylase expression was highest after 24-h imbibition, with a corresponding increase in enzymatic activity. The pasting properties of wheat flour decreased when seeds were imbibed for over 16 h. Gluten was not degraded until 48 h imbibition. Conclusion: Short-term imbibition of wheat can produce a new type of starch with improved physical and functional properties that may be more appealing to consumers.
Project description:The goals of this study are to compare transcriptome profiling (RNA-seq) between two wheat cultivars with different antioxidant actvity and to clarify the differences of these two wheat cultivars.
Project description:We report the transcriptome profile of different cultivars of Fusarium graminearum-infected wheat grains, aiming to search for some different expression genes and pathways to reveal the difference between wheat cultivars.