Validation of QTL for grain weight using MAS-derived pairs of NILs in bread wheat (Triticum aestivum L.)

Near isogenic lines (NILs) are ideal material for a variety of genetic studies including validation of specific QTL. In the present study, eight pairs of NILs for grain weight were developed, seven in the background of Raj3765, and one in the background of K9107. For this purpose, marker-assisted selection (MAS) was used for the transfer of three grain weight QTL (QGw.ccsu-1A.2, QGw.ccsu-1A.3 and QGw.ccsu-1B.1) that were earlier identified in our laboratory. Two genotypes of each of the eight pairs of NILs, differed for QTL alleles (QTL_Hgw derived from the donor parent and the QTL_Lgw derived from the recipient parent). Each pair of NILs involved a solitary QTL except one NIL, which differed for all the three QTL. The difference in thousand grain weight (TGW) in two NILs of an individual pair ranged from 2.8 to 7.5 g, thus validating the effect of the QTL for TGW, although the quantum of difference did not always match the phenotypic variance of the corresponding QTL. As expected, the NILs which involved all the three QTL had the maximum difference of 7.5 g in TGW, and the NILs which involved QTL, QGw.ccsu-1A.2 had minimum average difference of 2.8 g for TGW. The NILs produced during the present study may be used in future for MAS and for fine mapping of TGW QTL.

     

Fine mapping TaFLW1, a major QTL controlling flag leaf width in bread wheat (Triticum aestivum L.)

Introduction

Flag leaf width (FLW) is directly related to photosynthetic capacity and yield potential in wheat. In a previous study, Qflw.nau-5A controlling FLW was detected on chromosome 5A in the interval possessing Fhb5 for type I Fusarium head blight (FHB) resistance using a recombinant inbred line population derived from Nanda2419 × Wangshuibai.

Materials and methods

Qflw.nau-5A near-isogenic line (NIL) with the background of Mianyang 99-323 and PH691 was developed and evaluated. FLW inheritance was investigated using two F₂ populations developed from crossing the Qflw.nau-5A NILs with their recurrent parents. One hundred ten and 28 recombinants, which included 10 and 5 types of recombinants, were identified from 2816 F₂ plants with Mianyang 99-323 background and 1277 F₂ plants with PH691 background, respectively, and phenotyped in field trials for FLW and type I FHB resistance. Deletion bin mapping was applied to physically map Qflw.nau-5A.

Results and conclusions

The introduction of Wangshuibai Qflw.nau-5A allele reduced the FLW up to 3 mm. In the F₂ populations, Qflw.nau-5A was inherited like a semi-dominant gene, and was therefore designated as TaFLW1. The FLW of the recombinant lines displayed a distinct two-peak distribution. Recombinants with wider leaves commonly have Mianyang 99-323 or PH691 chromatin in the 0.2 cM Xwmc492-Xwmc752 interval that resided in the 5AL12-0.35–0.57 deletion bin, and recombinants with narrow leaves were Wangshuibai genotype in this interval. Phenotypic recombination between FLW and type I FHB resistance was identified, implying TaFLW1 was in close linkage with Fhb5. These results should aid wheat breeders to break the linkage drag through marker-assisted selection and assist in the map-based cloning of TaFLW1.     

Distribution of gluten proteins in bread wheat (Triticum aestivum) grain

Background and Aims

Gluten proteins are the major storage protein fraction in the mature wheat grain. They are restricted to the starchy endosperm, which forms white flour on milling, and interact during grain development to form large polymers which form a continuous proteinaceous network when flour is mixed with water to give dough. This network confers viscosity and elasticity to the dough, enabling the production of leavened products. The starchy endosperm is not a homogeneous tissue and quantitative and qualitative gradients exist for the major components: protein, starch and cell wall polysaccharides. Gradients in protein content and composition are the most evident and are of particular interest because of the major role played by the gluten proteins in determining grain processing quality.

Methods

Protein gradients in the starchy endosperm were investigated using antibodies for specific gluten protein types for immunolocalization in developing grains and for western blot analysis of protein extracts from flour fractions obtained by sequential abrasion (pearling) to prepare tissue layers.

Key Results

Differential patterns of distribution were found for the high-molecular-weight subunits of glutenin (HMW-GS) and γ-gliadins when compared with the low-molecular-weight subunits of glutenin (LMW-GS), ω- and α-gliadins. The first two types of gluten protein are more abundant in the inner endosperm layers and the latter more abundant in the subaleurone. Immunolocalization also showed that segregation of gluten proteins occurs both between and within protein bodies during protein deposition and may still be retained in the mature grain.

Conclusions

Quantitative and qualitative gradients in gluten protein composition are established during grain development. These gradients may be due to the origin of subaleurone cells, which unlike other starchy endosperm cells derive from the re-differentiation of aleurone cells, but could also result from the action of specific regulatory signals produced by the maternal tissue on specific domains of the gluten protein gene promoters.     

The 5S DNA units of bread wheat (Triticum aestivum)

A collection of 44 cloned 5S DNA units fromTriticum aestivum cv. Chinese Spring were grouped into 12 sequence-types based on sequence similarity and the respective consensus sequences were then produced. The relationship between these 12 consensus sequences (T. aestivum S 1-S 8 andT. aestivum L 1-L 4), together with two clones sequenced byGerlach andDyer, and the 5S DNA consensus sequences from diploidTriticum spp. were then determined by numerical methods. Both phenetic and cladistic analyses were carried out. The following wheat 5S DNA sequences were found to group with respective sequences from diploidTriticum spp.:T. aestivum S 4, S 6 withT. tauschii S;T. aestivum S 3 withT. monococcum S andT. monococcum S-Rus 7;T. aestivum L 1 andT. aestivum L-G&D withT. speltoides L;T. aestivum L 2, L 3 withT. tauschii L;T. aestivum L 4 withT. monococcum L andT. monococcum L-Rus 12. The analyses suggested that 5 out of the 65S Dna loci present in wheat were identified at the sequence level. The locus that could not be identified in this analysis was the5S Dna-B 1 locus. A group ofT. aestivum sequences (T. aestivum S 1, S 7, S 8, S-G&D) were found to be distinct from the other 5S DNA sequences in the data base. The existence of the distinct group of 5S DNA sequences suggests that there is a gap in our current understanding of wheat evolution with respect to the5S Dna loci.     

Agrobacterium-mediated In-planta transformation of bread wheat (Triticum aestivum L.)

Journal of Plant Biochemistry and Biotechnology - Agrobacterium-mediated in-planta transformation method allows efficient plant transformation without tissue culture. In the present study, a tissue...     

Allelopathy in wheat (Triticum aestivum)

Wheat (Triticum aestivum) allelopathy has potential for the management of weeds, pests and diseases. Both wheat residue allelopathy and wheat seedling allelopathy can be exploited for managing weeds, including resistant biotypes. Wheat varieties differ in allelopathic potential against weeds, indicating that selection of allelopathic varieties might be a useful strategy in integrated weed management. Several categories of allelochemicals for wheat allelopathy have been identified, namely, phenolic acids, hydroxamic acids and short‐chain fatty acids. Wheat allelopathic activity is genetically controlled and a multigenic model has been proposed. Research is underway to identify genetic markers associated with wheat allelopathy. Once allelopathic genes have been located, a breeding programme could be initiated to transfer the genes into modern varieties for weed suppression. The negative impacts of wheat autotoxicity on agricultural production systems have also been identified when wheat straws are retained on the soil surface for conservation farming purposes. A management package to avoid such deleterious effects is discussed. Wheat allelopathy requires further study in order to maximise its allelopathic potential for the control of weeds, pests and diseases, and to minimise its detrimental effects on the growth of wheat and other crops.     

Precise mapping Fhb5, a major QTL conditioning resistance to Fusarium infection in bread wheat (Triticum aestivum L.)

Qfhi.nau-5A is a major quantitative trait locus (QTL) against Fusarium graminearum infection in the resistant wheat germplasm Wangshuibai. Genetic analysis using BC(3)F(2) and BC(4)F(2) populations, derived from selfing two near-isogenic lines (NIL) heterozygous at Qfhi.nau-5A that were developed, respectively, with Mianyang 99-323 and PH691 as the recurrent parent, showed that Qfhi.nau-5A inherited like a single dominant gene. This QTL was thus designated as Fhb5. To fine map it, these two backcross populations and a recombinant inbred line (RIL) population derived from Nanda2419?×?Wangshuibai were screened for recombinants occurring between its two flanking markers Xbarc56 and Xbarc100. Nineteen NIL recombinants were identified from the two backcross populations and nine from the RIL population. In the RIL recombinant selection process, selection against Fhb4 present in the RIL population was incorporated. Genotyping these recombinant lines with ten markers mapping to the Xbarc56-Xbarc100 interval revealed four types of Mianyang 99-323-derived NIL recombinants, three types of PH691-derived NIL recombinants, and four types of RIL recombinants. In different field trials, the percentage of infected spikes of these lines displayed a distinct two-peak distribution. The more resistant class had over 55% less infection than the susceptible class. Common to these resistant genotypes, the 0.3-cM interval flanked by Xgwm304 and Xgwm415 or one of these two loci was derived from Wangshuibai, while none of the susceptible recombinants had Wangshuibai chromatin in this interval. This interval harboring Fhb5 was mapped to the pericentromeric C-5AS3-0.75 bin through deletion bin mapping. The precise localization of Fhb5 will facilitate its utilization in marker-assisted wheat breeding programs.     

Fine mapping Fhb4, a major QTL conditioning resistance to Fusarium infection in bread wheat (Triticum aestivum L.)

Qfhi.nau-4B is a major quantitative trait locus (QTL) against Fusarium graminearum infection identified in the Fusarium head blight-resistant germplasm Wangshuibai. To fine map this QTL, a recombinant inbred line (RIL) population of 530 lines derived from Nanda2419 × Wangshuibai and the BC₃F₂ population derived from the cross of a Qfhi.nau-4B near isogenic line (NIL) with susceptible cultivar Mianyang 99-323 as the recurrent parent were screened for recombinants occurred between microsatellite markers Xbarc20 and Xwmc349 that flank Qfhi.nau-4B. A total of 95 recombinants were obtained, including 45 RIL recombinants obtained through reverse-selection of Qfhi.nau-5A and 50 NIL recombinants from the BC₃F₂ population. Genotyping these recombinant lines with 22 markers mapping to the Xbarc20 and Xwmc349 interval revealed fourteen genotypes of the RIL recombinants as well as of the NIL recombinants. Two-year field evaluation of their resistance to Fusarium infection showed that these lines could be clearly classified into two groups according to percentage of infected spikes. The more resistant class had over 60% less infection than the susceptible class and were common to have Wangshuibai chromatin in the 1.7-cM interval flanked by Xhbg226 and Xgwm149. None of the susceptible recombinants had this Wangshuibai chromatin. Qfhi.nau-4B was thus confined between Xhbg226 and Xgwm149 and named Fhb4. The interval harboring Fhb4 was mapped to 4BL5-0.86–1.00 bin using Chinese Spring deletion lines, a region with about 5.7 times higher recombination rate than the genome average. This study established the basis for map-based cloning of Fhb4.     

Molecular characterization of a gene encoding N-myristoyl transferase (NMT) from Triticum aestivum (bread wheat).

Myristoyl-CoA:protein N-myristoyl transferase (NMT; EC 2.3.1.97) acylates the Gly residue abutting the N-terminal Met with a myristic acid following the removal of the Met residue in certain eukaryotic proteins, and in some cases myristoylation is essential to cell growth and survival. We report the cloning of a full-length cDNA encoding NMT from Triticum aestivum (TaNMT). The cDNA included a predicted open reading frame of 1317 nucleotides, which encoded a predicted protein of 438 amino acids containing all of the residues that are important for NMT activity. The TaNMT amino acid and nucleotide sequences were compared with NMTs from 14 other species encompassing a wide array of taxonomic groups. Among the experimentally validated NMTs, TaNMT was most similar to that of Arabidopsis thaliana. Southern blot analysis of wheat genomic DNA showed that TaNMT is encoded by a single copy gene, with one copy per haploid genome. We expressed TaNMT in Escherichia coli cells and determined that the recombinant protein possessed NMT activity, catalyzing the N-myristoylation of peptides from known or putatively myristoylated proteins from plants and animals without a strong preference for the plant peptides. TaNMT is the second experimentally validated plant NMT sequence and the first from a monocotyledonous species.     

Control of flowering time and spike development in cereals: the earliness per se Eps-1 region in wheat, rice, and Brachypodium

The earliness per se gene Eps-A ^m 1 from diploid wheat Triticum monococcum affects heading time, spike development, and spikelet number. In this study, the Eps1 orthologous regions from rice, Aegilops tauschii, and Brachypodium distachyon were compared as part of current efforts to clone this gene. A single Brachypodium BAC clone spanned the Eps-A ^m 1 region, but a gap was detected in the A. tauschii physical map. Sequencing of the Brachypodium and A. tauschii BAC clones revealed three genes shared by the three species, which showed higher identity between wheat and Brachypodium than between them and rice. However, most of the structural changes were detected in the wheat lineage. These included an inversion encompassing the wg241-VatpC region and the presence of six unique genes. In contrast, only one unique gene (and one pseudogene) was found in Brachypodium and none in rice. Three genes were present in both Brachypodium and wheat but were absent in rice. Two of these genes, Mot1 and FtsH4, were completely linked to the earliness per se phenotype in the T. monococcum high-density genetic map and are candidates for Eps-A ^m 1. Both genes were expressed in apices and developing spikes, as expected for Eps-A ^m 1 candidates. The predicted MOT1 protein showed amino acid differences between the parental T. monococcum lines, but its effect is difficult to predict. Future steps to clone the Eps-A ^m 1 gene include the generation of mot1 and ftsh4 mutants and the completion of the T. monococcum physical map to test for the presence of additional candidate genes.     

Deciphering the genetics of flowering time by an association study on candidate genes in bread wheat (Triticum aestivum L.)

Earliness is very important for the adaptation of wheat to environmental conditions and the achievement of high grain yield. A detailed knowledge of key genetic components of the life cycle would enable an easier control by the breeders. The objective of the study was to investigate the effect of candidate genes on flowering time. Using a collection of hexaploid wheat composed of 235 lines from diverse geographical origins, we conducted an association study for six candidate genes for flowering time and its components (vernalization sensitivity and earliness per se). The effect on the variation of earliness components of polymorphisms within the copies of each gene was tested in ANOVA models accounting for the underlying genetic structure. The collection was structured in five groups that minimized the residual covariance. Vernalization requirement and lateness tend to increase according to the mean latitude of each group. Heading date for an autumnal sowing was mainly determined by the earliness per se. Except for the Constans (CO) gene orthologous of the barley HvCO3, all gene polymorphisms had a significant impact on earliness components. The three traits used to quantify vernalization requirement were primarily associated with polymorphisms at Vrn-1 and then at Vrn-3 and Luminidependens (LD) genes. We found a good correspondence between spring/winter types and genotypes at the three homeologous copies of Vrn-1. Earliness per se was mainly explained by polymorphisms at Vrn-3 and to a lesser extent at Vrn-1, Hd-1 and Gigantea (GI) genes. Vernalization requirement and earliness as a function of geographical origin, as well as the possible role of the breeding practices in the geographical distribution of the alleles and the hypothetical adaptive value of the candidate genes, are discussed.     

QTL mapping of 1000-kernel weight, kernel length, and kernel width in bread wheat (Triticum aestivum L.)

Kernel size and morphology influence the market value and milling yield of bread wheat (Triticum aestivum L.). The objective of this study was to identify quantitative trait loci (QTLs) controlling kernel traits in hexaploid wheat. We recorded 1000-kernel weight, kernel length, and kernel width for 185 recombinant inbred lines from the cross Rye Selection 111 × Chinese Spring grown in 2 agro-climatic regions in India for many years. Composite interval mapping (CIM) was employed for QTL detection using a linkage map with 169 simple sequence repeat (SSR) markers. For 1000-kernel weight, 10 QTLs were identified on wheat chromosomes 1A, 1D, 2B, 2D, 4B, 5B, and 6B, whereas 6 QTLs for kernel length were detected on 1A, 2B, 2D, 5A, 5B and 5D. Chromosomes 1D, 2B, 2D, 4B, 5B and 5D had 9 QTLs for kernel width. Chromosomal regions with QTLs detected consistently for multiple year-location combinations were identified for each trait. Pleiotropic QTLs were found on chromosomes 2B, 2D, 4B, and 5B. The identified genomic regions controlling wheat kernel size and shape can be targeted during further studies for their genetic dissection.     

The origin of the B-genome of bread wheat (Triticum aestivum L.)

Understanding the origin of cultivated wheats would further their genetic improvement. The hexaploid bread wheat (Triticum aestivum L., AABBDD) is believed to have originated through one or more rare hybridization events between Aegilops tauschii (DD) and the tetraploid T. turgidum (AABB). Progenitor, of the A-genome of the tetraploid and hexaploid wheats has generally been accepted to be T. urartu. In spite of the large number of attempts and published reports about the origin of the B-genome in cultivated wheats, the donor of the B-genome is still relatively unknown and controversial and, hence, remains open. This genome has been found to be closely related to the S-genome of the Sitopsis section (Ae. speltoides, Ae. longissima, Ae. sharonensis, Ae. searsii, and Ae. bicornis) of the genus Aegilops L. Among Sitopsis species, the most positive evidence has been accumulated for Ae. speltoides as the progenitor of the B-genome. Therefore, one or more of the Sitopsis species were proposed frequently as the B-genome donor. Although several reviews have been written on the origin of the genomes of wheat over the years, this paper will attempt for the first time to review the immense literature on the subject, with a particular emphasis on the B-genome which has attracted a huge attention over some 100 years. The ambiguity and conflicting results in most of the methods employed in deducing the precise B-genome donor/s to bread wheat are also discussed.     

Detection and mapping of QTL for earliness components in a bread wheat recombinant inbred lines population

Earliness, an adaptative trait and factor of variation for agronomic characters, is a major trait in plant breeding. Its constituent traits, photoperiod sensitivity (PS), vernalization requirement (VR) and intrinsic earliness (IE), are largely under independent genetic controls. Mapping of major genes and quantitative trait loci (QTL) controlling these components is in progress. Most of the studies focusing on earliness considered it as a whole or through one (or two) of its components. The purpose of this study was to detect and map QTL for the three traits together through an experimental design combining field trials and controlled growth conditions. QTL were mapped in a population of F₇ recombinant inbred lines derived by single-seed descent from a cross between two French varieties, Renan and Récital. A map was previously constructed, based on 194 lines and 254 markers, covering about 77% of the genome. Globally, 13 QTL with a LOD>2.5 were detected, of which four control PS, five control VR and four control IE. Two major photoperiod sensitive QTL, together explaining more than 31% of the phenotypic variation, were mapped on chromosomes 2B and 2D, at the same position as the two major genes Ppd-B1 and Ppd-D1. One major VR QTL explaining (depending on the year) 21.8–39.6% of the phenotypic variation was mapped on 5A. Among the other QTL, two QTL of PS and VR not referenced so far were detected on 5A and 6D, respectively. A VR QTL already detected on 2B in a connected population was confirmed.     

Physiological traits associated with heat tolerance in bread wheat (Triticum aestivum L.)

Field experiments for evaluating heat tolerance-related physiological traits were conducted for two consecutive years using a mapping population of recombinant inbred lines (RILs) from the cross RAJ4014/WH730. Chlorophyll content (Chl) and chlorophyll fluorescence (CFL) were recorded under timely sown (TS) and late sown (LS) conditions. Late sowing exposes the terminal stage of plants to high temperature stress. Pooled analysis showed that CFL and Chl differed significantly under TS and LS conditions. The mean value of CFL (Fv/Fm) and Chl under both timely and late sown conditions were used as physiological traits for association with markers. Regression analysis revealed significant association of microsatellite markers viz., Xpsp3094 and Xgwm131 with coefficients of determination (R²) values for CFL (Fv/Fm) and Chl as 12 and 8 %, respectively. The correlation between thousand grain weight (TGW) with Chl and CFL were 14 and 7 % and correlation between grain wt./spike with Chl and CFL were 15 and 8 %, respectively. The genotypes showing tolerance to terminal heat stress as manifested by low heat susceptibility index (HSI = 0.43) for thousand grain weight, were also found having very low Chl, HSI (−0.52). These results suggest that these physiological traits may be used as a secondary character for screening heat-tolerant genotypes.     

Development and genetic mapping of sequence-tagged microsatellites (STMs) in bread wheat (Triticum aestivum L.)

The density of SSRs on the published genetic map of bread wheat (Triticum aestivum L.) has steadily increased over the last few years. This has improved the efficiency of marker-assisted breeding and certain types of genetic research by providing more choice in the quality of SSRs and a greater chance of finding polymorphic markers in any cross for a chromosomal region of interest. Increased SSR density on the published wheat genetic map will further enhance breeding and research efforts. Here, sequence-tagged microsatellite profiling (STMP) is demonstrated as a rapid technique for the economical development of anonymous genomic SSRs to increase marker density on the wheat genetic map. A total of 684 polymorphic sequence-tagged microsatellites (STMs) were developed, and 380 were genetically mapped in three mapping populations, with 296 being mapped in the International Triticeae Mapping Initiative W7984 × Opata85 recombinant inbred cross. Across the three populations, a total of 479 STM loci were mapped. Several technological advantages of STMs over conventional SSRs were also observed. These include reduced marker deployment costs for fluorescent-based SSR analysis, and increased genotyping throughput by more efficient electrophoretic separation of STMs and a high amenability to multiplex PCR.Electronic Supplementary Material Supplementary material is available in the online version of this article at and is accessible for authorized users.     

An intervarietal genetic linkage map of Indian bread wheat (Triticum aestivum L.) and QTL maps for some metric traits

Bread wheat (Triticum aestivum L.) exhibits very narrow genetic diversity and hence there is high relatedness among cultivated varieties. However, a population generated from an intervarietal cross, with the parents differing in a large number of traits, could lead to the generation of QTL maps which will be useful in practice. In this report a genetic linkage map of wheat is constructed using a cross between two Indian bread wheat varieties: Sonalika and Kalyansona. The linkage map consisted of 236 markers and spanned a distance of 3639 cM, with 1211.2 cM for the A genome, 1669.2 cM for the B genome, 192.4 cM for the D genome and 566.2 cM for unassigned groups. Linkage analysis defined 37 linkage groups of which 24 were assigned to 17 chromosomes. The genetic map was used to identify QTLs by composite internal mapping (CIM) for three metric traits, viz. culm length (CL), flag leaf length (FLL) and flag leaf breadth (FLB). Of 25 QTLs identified in this study, 15 have not been reported previously. Multitrait CIM (MCIM) analysis was carried out for traits that were significantly correlated such as FLB-FLL and CL-FLB-FLL. Detection of a large number of QTLs for the three traits analysed suggests that in parent cultivars that are not too diverse, the differences at genetic level detected as polymorphisms may be mostly associated with QTLs for the observed differences.