Dataset Information

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

ABSTRACT: 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 = 1Bx7^OE > 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 1Bx7^OE was two-fold higher than that of 1Bx7 or 1Dx5 and, therefore, translated a significant increase in the protein amount in 1Bx7^OE. 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.

SUBMITTER: Shin D

PROVIDER: S-EPMC7695314 | biostudies-literature | 2020 Nov

REPOSITORIES: biostudies-literature

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Publications

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

Shin Dongjin D Cha Jin-Kyung JK Lee So-Myeong SM Kabange Nkulu Rolly NR Lee Jong-Hee JH

Plants (Basel, Switzerland) 20201109 11

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 w ...[more]

PMID: 33182289

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Project description:BackgroundLow-molecular-weight glutenin subunits (LMW-GS), encoded by Glu-3 complex loci in hexaploid wheat, play important roles in the processing quality of wheat flour. To date, the molecular characteristics and effects on dough quality of individual Glu-3 alleles and their encoding proteins have been poorly studied. We used a Glu-A3 deletion line of the Chinese Spring (CS-n) wheat variety to conduct the first comprehensive study on the molecular characteristics and functional properties of the LMW-GS allele Glu-A3a.ResultsThe Glu-A3a allele at the Glu-A3 locus in CS and its deletion in CS-n were identified and characterized by proteome and molecular marker methods. The deletion of Glu-A3a had no significant influence on plant morphological and yield traits, but significantly reduced the dough strength and breadmaking quality compared to CS. The complete sequence of the Glu-A3a allele was cloned and characterized, which was found to encode a B-subunit with longer repetitive domains and an increased number of α-helices. The Glu-A3a-encoded B-subunit showed a higher expression level and accumulation rate during grain development. These characteristics of the Glu-A3a allele could contribute to achieving superior gluten quality and demonstrate its potential application to wheat quality improvement. Furthermore, an allele-specific polymerase chain reaction (AS-PCR) marker for the Glu-A3a allele was developed and validated using different bread wheat cultivars, including near-isogenic lines (NILs) and recombinant inbred lines (RILs), which could be used as an effective molecular marker for gluten quality improvement through marker-assisted selection.ConclusionsThis work demonstrated that the LMW-GS allele Glu-A3a encodes a specific LMW-i type B-subunit that significantly affects wheat dough strength and breadmaking quality. The Glu-A3a-encoded B-subunit has a long repetitive domain and more α-helix structures as well as a higher expression level and accumulation rate during grain development, which could facilitate the formation of wheat with a stronger dough structure and superior breadmaking quality.

Project description:Wheat (AABBDD, 2n?=?6x?=?42) is a major dietary component for many populations across the world. Bread-making quality of wheat is mainly determined by glutenin subunits, but it remains challenging to elucidate the composition and variation of low-molecular-weight glutenin subunits (LMW-GS) genes, the major components for glutenin subunits in hexaploid wheat. This problem, however, can be greatly simplified by characterizing the LMW-GS genes in Triticum urartu, the A-genome donor of hexaploid wheat. In the present study, we exploited the high-throughput molecular marker system, gene cloning, proteomic methods and molecular evolutionary genetic analysis to reveal the composition, variation, expression and evolution of LMW-GS genes in a T. urartu population from the Fertile Crescent region.Eight LMW-GS genes, including four m-type, one s-type and three i-type, were characterized in the T. urartu population. Six or seven genes, the highest number at the Glu-A3 locus, were detected in each accession. Three i-type genes, each containing more than six allelic variants, were tightly linked because of their co-segregation in every accession. Only 2-3 allelic variants were detected for each m- and s-type gene. The m-type gene, TuA3-385, for which homologs were previously characterized only at Glu-D3 locus in common wheat and Aegilops tauschii, was detected at Glu-A3 locus in T. urartu. TuA3-460 was the first s-type gene identified at Glu-A3 locus. Proteomic analysis showed 1-4 genes, mainly i-type, expressed in individual accessions. About 62% accessions had three active i-type genes, rather than one or two in common wheat. Southeastern Turkey might be the center of origin and diversity for T. urartu due to its abundance of LMW-GS genes/genotypes. Phylogenetic reconstruction demonstrated that the characterized T. urartu might be the direct donor of the Glu-A3 locus in common wheat varieties.Compared with the Glu-A3 locus in common wheat, a large number of highly diverse LMW-GS genes and active genes were characterized in T. urartu, demonstrating that this progenitor might provide valuable genetic resources for LMW-GS genes to improve the quality of common wheat. The phylogenetic analysis provided molecular evidence and confirmed that T. urartu was the A-genome donor of hexaploid wheat.

Project description:Wheat end-use quality mainly derives from two interrelated characteristics: the compositions of gluten proteins and grain hardness. The composition of gluten proteins determines dough rheological properties and thus confers the unique viscoelastic property on dough. One group of gluten proteins, high molecular weight glutenin subunits (HMW-GS), plays an important role in dough functional properties. On the other hand, grain hardness, which influences the milling process of flour, is controlled by Puroindoline a (Pina) and Puroindoline b (Pinb) genes. However, little is known about the combined effects of HMW-GS and PINs on dough functional properties. In this study, we crossed a Pina-expressing transgenic line with a 1Ax1-expressing line of durum wheat and screened out lines coexpressing 1Ax1 and Pina or lines expressing either 1Ax1 or Pina. Dough mixing analysis of these lines demonstrated that expression of 1Ax1 improved both dough strength and over-mixing tolerance, while expression of PINA detrimentally affected the dough resistance to extension. In lines coexpressing 1Ax1 and Pina, faster hydration of flour during mixing was observed possibly due to the lower water absorption and damaged starch caused by PINA expression. In addition, expression of 1Ax1 appeared to compensate the detrimental effect of PINA on dough resistance to extension. Consequently, coexpression of 1Ax1 and PINA in durum wheat had combined effects on dough mixing behaviors with a better dough strength and resistance to extension than those from lines expressing either 1Ax1 or Pina. The results in our study suggest that simultaneous modulation of dough strength and grain hardness in durum wheat could significantly improve its breadmaking quality and may not even impair its pastamaking potential. Therefore, coexpression of 1Ax1 and PINA in durum wheat has useful implications for breeding durum wheat with dual functionality (for pasta and bread) and may improve the economic values of durum wheat.

Dataset Information

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

Publications

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

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