Genome-wide identification, characteristics and expression of the prolamin genes in Thinopyrum elongatum

Background Prolamins, unique to Gramineae (grasses), play a key role in the human diet. Thinopyrum elongatum (syn. Agropyron elongatum or Lophopyrum elongatum), a grass of the Triticeae family with a diploid E genome (2n = 2x = 14), is genetically well-characterized, but little is known about its prolamin genes and the relationships with homologous loci in the Triticeae species. Results In this study, a total of 19 α-gliadin, 9 γ-gliadin, 19 ω-gliadin, 2 high-molecular-weight glutenin subunit (HMW-GS), and 5 low-molecular-weight glutenin subunit (LMW-GS) genes were identified in the Th. elongatum genome. Micro-synteny and phylogenetic analysis revealed dynamic changes of prolamin gene regions and genetic affinities among Th. elongatum, Triticum aestivum, T. urartu and Aegilops tauschii. The Th. elongatum genome, like the B subgenome of T. aestivum, only contained celiac disease epitope DQ8-glia-α1/DQ8.5-glia-α1, which provided a theoretical basis for the low gluten toxicity wheat breeding. The transcriptome data of Th. elongatum exhibited differential expression in quantity and pattern in the same subfamily or different subfamilies. Dough rheological properties of T. aestivum-Th. elongatum disomic substitution (DS) line 1E(1D) showed higher peak height values than that of their parents, and DS6E(6D) exhibited fewer α-gliadins, which indicates the potential usage for wheat quality breeding. Conclusions Overall, this study provided a comprehensive overview of the prolamin gene family in Th. elongatum, and suggested a promising use of this species in the generation of improved wheat breeds intended for the human diet.

Molecular Characterization of Novel x-Type HMW Glutenin Subunit 1B × 6.5 in Wheat

A novel high molecular weight glutenin subunit encoded by the Glu-1B locus was identified in the French genotype Bagou, which we named 1B × 6.5. This subunit differed in SDS-PAGE from well-known 1B × 6 and 1B × 7 subunits, which are also encoded at this locus. Subunit 1B × 6.5 has a theoretical molecular weight of 88,322.83 Da, which is more mobile than 1B × 6 subunit, and isoelectric point (pI) of about 8.7, which is lower than that for 1B × 6 subunit. The specific primers were designed to amplify and sequence 2476 bp of the Glu-1B locus from genotype Bagou. A high level of similarity was found between the sequence encoding 1B × 6.5 and other x-type encoding alleles of this locus.

Proteomic Determination of Low-Molecular-Weight Glutenin Subunit Composition in Aroona Near-Isogenic Lines and Standard Wheat Cultivars

The low-molecular weight glutenin subunit (LMW-GS) composition of wheat (Triticum aestivum) flour has important effects on end-use quality. However, assessing the contributions of each LMW-GS to flour quality remains challenging because of the complex LMW-GS composition and allelic variation among wheat cultivars. Therefore, accurate and reliable determination of LMW-GS alleles in germplasm remains an important challenge for wheat breeding. In this study, we used an optimized reversed-phase HPLC method and proteomics approach comprising 2-D gels coupled with liquid chromatography–tandem mass spectrometry (MS/MS) to discriminate individual LMW-GSs corresponding to alleles encoded by the Glu-A3, Glu-B3, and Glu-D3 loci in the ‘Aroona’ cultivar and 12 ‘Aroona’ near-isogenic lines (ARILs), which contain unique LMW-GS alleles in the same genetic background. The LMW-GS separation patterns for ‘Aroona’ and ARILs on chromatograms and 2-D gels were consistent with those from a set of 10 standard wheat cultivars for Glu-3. Furthermore, 12 previously uncharacterized spots in ‘Aroona’ and ARILs were excised from 2-D gels, digested with chymotrypsin, and subjected to MS/MS. We identified their gene haplotypes and created a 2-D gel map of LMW-GS alleles in the germplasm for breeding and screening for desirable LMW-GS alleles for wheat quality improvement.

Characterization of 1Sty13, a novel high-molecular-weight glutenin subunit from Elymus sibiricus L.

High-molecular-weight glutenin subunit (HMW-GS) is a key factor affecting dough-processing quality. 1Sty13 is a novel HMW-GS found in the tetraploid species, Elymus sibiricus L. 1Sty13 has faster electrophoretic mobility than the 1Dy12 subunit on sodium dodecyl sulphate (SDS)-polyacrylamide gel electrophoresis. The gene encoding the 1Sty13 subunit was composed of 1803 nucleotide base pairs with an open reading frame that was 599 amino acids in length. Analysis of the predicted amino acid sequence of 1Sty13 indicated that the N-terminal domain was similar to the y-type subunit, whereas the C-terminal domains were similar to the x-type subunit. Five cysteine residues were found in 1Sty13, which is one less than the published HMW-GS in the St genome. The 1Sty13 protein was purified at a scale sufficient for incorporation into flour for the SDS sedimentation test, which indicated that incorporating 1Sty13 improved dough quality.

Genome-wide Identification, Expression, Characteristics of the Prolamin Superfamily in Thinopyrum Elongatum and Its Kneading Performance in Common Wheat

Background: Prolamins, unique to Gramineae (grasses), play a key role in the human diet. Thinopyrum elongatum (also known as tall wheatgrass, rush wheatgrass, or Eurasian quackgrass) of Elytrigia is genetically well-characterized, but little is known about its prolamin genes and the relationships with homologous loci in the Triticum genus.Results: In this study, a total of 19 α-gliadin, 9 γ-gliadin, 19 ω-gliadin, 2 high-molecular-weight glutenin subunit (HMW-GS), and 5 low-molecular-weight glutenin subunit (LMW-GS) genes in the Th. elongatum genome were annotated. The transcriptome data of Th. elongatum exhibited differential expression in quantity and pattern in the same subfamily or different subfamilies. In addition, microsynteny and phylogenetic analysis revealed dynamic changes of prolamin gene region and genetic affinities among Th. elongatum, T. aestivum, T. urartu, and Aegilops tauschii. The E genome, like the B genome, only contained DQ8-glia-α1/DQ8.5-glia-α1, which provided a theoretical basis for the study of celiac disease (CD). Dough rheological properties of T. aestivum-Th. elongatum disomic substitution (DS) lines 1E(1A), 1E(1D), and 3E(3A) showed much higher peak height values than that of their parent.Conclusions: Overall, this study provides a comprehensive overview of the prolamin gene superfamily in Th. elongatum, and suggests a promising use of this species in the generation of improved wheat breeds intended for the human diet.

Rapid and Easy High-Molecular-Weight Glutenin Subunit Identification System by Lab-on-a-Chip in Wheat (Triticum aestivum L.)

Lab-on-a-chip technology is an emerging and convenient system to easily and quickly separate proteins of high molecular weight. The current study established a high-molecular-weight glutenin subunit (HMW-GS) identification system using Lab-on-a-chip for three, six, and three of the allelic variations at the Glu-A1, Glu-B1, and Glu-D1 loci, respectively, which are commonly used in wheat breeding programs. The molecular weight of 1Ax1 and 1Ax2* encoded by Glu-A1 locus were of 200 kDa and 192 kDa and positioned below 1Dx subunits. The HMW-GS encoded by Glu-B1 locus were electrophoresed in the following order below 1Ax1 and 1Ax2*: 1Bx13 ≥ 1Bx7 = 1Bx7OE > 1Bx17 > 1By16 > 1By8 = 1By18 > 1By9. 1Dx2 and Dx5 showed around 4-kDa difference in their molecular weights, with 1Dy10 and 1Dy12 having 11-kDa difference, and were clearly differentiated on Lab-on-a-chip. Additionally, some of the HMW-GS, including 1By8, 1By18, and 1Dy10, having different theoretical molecular weights showed similar electrophoretic mobility patterns on Lab-on-a-chip. The relative protein amount of 1Bx7OE was two-fold higher than that of 1Bx7 or 1Dx5 and, therefore, translated a significant increase in the protein amount in 1Bx7OE. Similarly, the relative protein amounts of 8 & 10 and 10 & 18 were higher than each subunit taken alone. Therefore, this study suggests the established HMW-GS identification system using Lab-on-a-chip as a reliable approach for evaluating HMW-GS for wheat breeding programs.