博文
小麦 PAG 2018 摘要 （三）
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61、Introgression of a Newly Discovered Xanthophyll Acyl Transferase into Durum Wheat
The University of Adelaide
The carotenoid lutein is the main yellow pigment in the grain of both bread wheat (AABBDD) and durum wheat (AABB). Lutein confers health benefits and is important in determining the colour of wheat-based food products. During postharvest storage of bread wheat grain, lutein can be converted into lutein esters, which are more stable than free lutein. This esterification does not occur in durum wheat.
Recently, we mapped the esterification trait to a locus on chromosome 7D. We have now confirmed that a gene at that locus encodes a xanthophyll acyl transferase (XAT). This is the first such enzyme to be functionally confirmed in any plant species.
Given the importance of stable yellow pigment for pasta and other durum-based products, we wanted to transfer the Xat1 gene into durum wheat. We crossed a 7D(7A) disomic chromosome substitution line of Langdon durum with DBA-Aurora, an Australian durum cultivar. This provided opportunities for 7D-7A pairing and recombination in the F1 generation. Using marker assays designed for 7D-7A SNPs, we screened F2 progeny for evidence of spontaneous 7D-7A recombination. Plants with recombinant chromosomes were discovered, including one with only a short distal 7D segment that includes the *Xat1 *gene. Homozygous recombinant progeny were developed and are being backcrossed to DBA-Aurora to develop a high-performing durum with improved lutein stability. This work demonstrates that with the use of appropriate crosses and markers, plants with intergenomic recombinations can be discovered for use in breeding.
62、Evaluation of Winter-Survival of Winter Wheat By Drone-Generated Multi-Spectral Imagery Identified Two Quantitative Trait Loci on Chromosome 5A
University of Guelph
The harsh and unpredictable winters in the high latitude regions of the northern hemisphere often leads to high risk of winterkill for winter wheat (Triticum aestivum *L.). One of the key traits that influence winter-survival is the timing of the transition from vegetative to reproductive stage, as wheat loses cold tolerance after the transition. The goal of this research is to investigate the genetic basis of winter-survival in Canadian winter wheat and to identify the combination of key candidate gene alleles that is optimal for high-latitude winter wheat. The Canadian Winter Wheat Diversity Panel (CWWDP), which includes 450 winter wheat genotypes with various levels of winter-hardiness, was phenotyped under field conditions in Ontario, Canada. Normalized difference vegetation index (NDVI) was extracted from multi-spectral imagery captured by unmanned aerial vehicle (UAV) as a measure of winter-survival. The diversity panel was genotyped with the 90K Illumina SNP chip and additionally for allelic variation at the candidate gene loci that have demonstrated significant effect on flowering time and cold tolerance. This included the major vernalization gene loci on group 5 chromosomes (VRN-A1, VRN-B1, and *Vrn-D1), C-Repeat Binding Factor (CBF)-12 *and *-15 on chromosome 5A and photoperiod response loci on chromosome 5D (PPD-D1) and 5A (PPD-A1). Genome-wide association studies identified two major quantitative trait loci on chromosome 5A, which correspond to the frost resistance loci Fr-A1 *and *Fr-A2. This result is consistent with previous reports on the role of chromosome 5A in winter-survival and showcased the potential of UAV-based phenotyping in genetics research.
63、Completion of the ‘Jagger’ Winter Wheat Genome Leads to Identification of Aegilops Ventricosa 2NS Translocation and Impact in Kansas Wheat Breeding
Kansas State University
Since its release 23 years ago, the winter wheat cultivar Jagger has had a huge impact on wheat production in the US and around the globe and became a parental germplasm in many of the current cultivars of the central U.S. Importantly, Jagger also possesses the Aegilops ventricosa 2NS translocation fragment, which is associated with disease resistance against multiple wheat pathogens including the devastating wheat blast fungus. Here we present the first de novo assembly and anchoring of the Jagger genome based on the NRGene DeNovoMAGIC 3.0 pipeline and chromosome conformation capture technology (Hi-C). We were able to successfully anchor ~3000 scaffolds with a cumulative length of 14.2 Gb and an N50 of 10.5 Mb to build 21 draft pseudomolecules. Overall, these draft pseudomolecules showed high collinearity with the reference Chinese Spring genome (IWGSC RefSeq v1). The Jagger genome will be a powerful tool in the analysis of the wheat pan genome and the dissection of important agronomic traits. To illustrate its utility, we delineated the Aegilops**ventricosa 2NS translocation fragment in Jagger, consisting of a ~30 Mb fragment identified through alignment of Jagger chromosome 2A to ‘Chinese Spring’ chromosome 2A. We also developed a pipeline for identification of 2NS translocations in wheat breeding lines using genotyping by sequencing (GBS) data and reference genomes. Our results suggest that the 2NS translocation segment is widely present in KS breeding lines, and is potentially providing a yield benefit over the span of 11 years of the breeding program.
64、A Megabase-Scale Comparative Analysis of Wheat Chromosome 2D from Two Wheat Cultivars Unravels Molecular Mechanisms of Genome Evolution
King Abdullah University of Science and Technology (KAUST)
Recent improvements in DNA sequencing technologies and assembly algorithms have paved the way to generated high-quality genome assemblies of the large and complex genomes of wheat and its wild relatives. These genome assemblies form the basis to study the evolutionary dynamics of wheat genomes on a megabase-size scale.
Here, we provide a comparative analysis of two high-quality assemblies of the 729-megabase-sized chromosomes 2D of wheat landrace Chinese Spring (IWGSC RefSeq v1.0) and the modern Swiss spring wheat line ‘CH Campala Lr22a’. In general, there was a high degree of sequence conservation along the chromosome. Analysis of large insertions and deletions (InDels) revealed four large InDels of a total size of 2.2 Mb. The molecular signatures at their breakpoints enabled to identify unequal crossing over and double-strand break repair as the molecular causes of these InDels. In addition, the gene content of the two chromosomes were compared. This comparison revealed that 99% of the genes were present and collinear in both the cultivars. The fraction of unique single-copy genes observed was 0.44% for Chinese Spring and 0.71% for ‘CH Campala Lr22a’. Hence, our analysis provides evidence that the number of genotype-specific genes is considerably smaller than previously estimated.
65 Identification of Quantitative Trait Loci (QTL) Associated with Fusarium Head Blight and Septoria Resistance in a Maxine/ Redeemer Winter Wheat Population
University of Guelph
Fusarium head blight (FHB) and Septoria tritici blotch (STB) are important wheat diseases in North America. The objective of this study was to map loci associated with FHB traits, STB and plant height in a Maxine/Redeemer winter wheat population. Evaluation of FHB and STB resistance was performed using spray inoculation of a mixture of F. graminearum *and Septoria tritici blotch *isolates, respectively and under natural infections in replicated trials across three environments in Ontario, Canada. FHB disease incidence and severity were recorded and FHB index was calculated. For both diseases, the population showed a continuous distribution pattern and transgressive segregation of progeny. DArT markers were used to generate a genetic map and quantitative trait loci (QTL) analysis were performed by evaluating 105 doubled-haploid lines. FHB resistance QTL were identified on chromosome 2A, 4A, 6A, 3B, 4B, 2D and 3D, while QTL for STB were identified on chromosome 4B and 7A. Plant height QTL were identified on chromosome 4A, 6A, 4B and 2D. QTL identified in this study will be used in winter wheat breeding programs using marker assisted selection (MAS).
66 Activation of Seminal Root Primordia during Wheat Domestication Reveals Underlying Mechanisms of Plant Resilience
Hebrew University of Jerusalem
Seminal roots constitute the early root system of major crops of the Poaceae family. Although variation in seminal root number was described in several crops, mechanisms through which seminal root number (SRN) are controlled and in turn contribute to environmental adaptation are poorly understood. Here, we show that SRN increased upon wheat domestication due to the activation of root primordia which are suppressed in wild wheat, a trait controlled by factors expressed in the germinating embryo. We used variation in seminal root number (SRN) between wild and domesticated wheat to investigate its bearing on water uptake and seedling resilience. The persistence of wild roots at their primordial state promoted seedling recovery from episodic water-stress through re-activation of root primordia following rehydration. In spite of their lower root number, wild seedlings transpired more than domesticated seedlings. Additionally, transpiration rate was associated with higher shoot:root ratio in wild wheat, indicating contrasting strategies of resource allocation between wild and domesticated wheat. Our findings suggest that under well-watered conditions, lower root number enables direction of resources to aboveground without limiting water uptake. Furthermore, the maintenance of roots at their primordial state and their re-activation following rehydration maybe regarded as seedling protective mechanism against episodic water-stress. The results underscore SRN as an adaptive trait that was reshaped upon domestication. Identification of factors associated with the plasticity of the SRN phenotype expands our understanding on the evolutionary dynamics of wheat and may serve to optimize root number in future breeding efforts.
67 IWGSC Phase II: What's Next for the IWGSC
IWGSC
In 2017, the International Wheat Genome Sequencing Consortium (IWGSC) achieved the first high quality, annotated reference sequence of bread wheat, IWGSC RefSeq v1.0. In phase II, the consortium will continue to deliver valuable tools and resources for wheat scientists and breeders through the functional annotation of the reference sequence and the generation of a pan genome that represents the breadth of diversity for bread wheat. An overview of the future activities of the IWGSC will be presented.
68 Genome-Wide Association Mapping for Seedling Heat Tolerance in Winter Wheat
Noble Research Institute, LLC
Heat stress at seedling stage is one of the most common issues of winter wheat in a dual-purpose management system in the southern Plains. However, the genetic mechanism underlying seedling heat tolerance in wheat is not well studied. To dissect the genetic basis of this trait, we conducted a genome-wide association mapping study (GWAS) using 200 hard red winter wheat lines from the Triticeae Coordinated Agricultural Project (TCAP), genotyped using the wheat iselect 90K SNP genotyping array. The plants were initially planted under optimal temperature in growth chambers. At three leaf stage, plants were subjected to two temperature regimes, high temperature (40/35oC day/night) as heat stress treatment, and optimal growth temperature (25/20oC day/night) as control for 14 days. Data were collected on leaf chlorophyll content (LCC), shoot length (SL), number of leaves (LN), and percent of seedling recovery (PSR) under optimal growth temperature following the heat stress treatment. GWAS was performed using mixed linear model (MLM). Significant marker-trait associations were found on all the traits under both optimal and heat-stressed growth conditions. In addition, heat stress responding marker-trait associations were also detected. Once validated, these SNPs will be used in marker-assisted selection of seedling heat tolerance in wheat.
69 Indo-UK Research Collaboration to Improve Nitrogen Use Efficiency in Wheat
ICAR.NBPGR
Nitrogen is the major agronomic input that determines the performance and productivity of wheat crop in both India and UK. With nitrogen being the major production cost for farmers, it has a huge environmental footprint, in terms of pollution of ground waters and generation of greenhouse gases. To minimize the use of applied Nitrogen fertilizers a cross-Institute pre-breeding programme (INEW virtual centre) under India-UK partnership is being executed to identify sources of traits and developing markers for use in academic research and transfer to commercial breeding programmes and responsibility for delivering improved wheat varieties to Indian farmers. The Virtual Joint Centre is bringing together major wheat researchers from ICAR.IARI in New Delhi, Indian Institute of Wheat and Barley Research, Karnal, Bourlag Institute for South Asia, Punjab, National Bureau of Plant Genetic resources, New Delhi and National Research Centre for Plant Biotechnology, New Delhi,Punjab Agricultural University. UK partner institutions are Rothamsted research, University of Nottingham, University of Bristol, John Inns Centre and National Institute of Agricultural Botany.
The core of the project is precision field trials being conducted in India and UK using germplasm from both countries, in which the fate of nitrogen in the plant will be followed from root uptake to seed maturity, at limiting and adequate levels of fertilisation. Its impact on grain yield and grain quality in these lines will be studied in detail using the platform technologies (Fig.1) providing information on the relationship between performance and phenology. Major mandate includes integrated study of the genetic, biochemical and molecular basis for improved N use efficiency from mechanisms of nitrogen uptake to partitioning in the grain and effects on processing quality. Candidate genes that control key processes limiting N use efficiency will be identified. The study will be supported by genotyping of germplasm and identification of key genes, enzymes involved, their variation employing high density SNP arrays and transcriptome analysis. Molecular markers developed for key traits will be transferred to wheat breeders in UK and India.
70 Genome-Wide Analysis of MIKC-Type MADS-Box Genes in Wheat: A Primer for Crop Improvement
University College Dublin
MIKC-type MADS-box genes encode transcription factors with prominent roles in plant development. They constitute key regulators of flowering time, floral organ identity, seed and fruit development. MADS-box genes have also been the target of domestication processes in numerous plant species. Understanding the function and evolution of MADS-box genes in crops is therefore of considerable interest for future crop improvement programs.
Here, we present a genome-wide analysis of the MIKC-type MADS-box gene family from wheat. We identified more than 200 MIKC-type MADS-box genes, considerably more than in other monocots, partly due to the hexaploid nature of wheat. MIKC* genes as well as representatives from 15 distinct MIKCc subfamilies were identified. Our preliminary analyses indicate that some MADS-box gene subfamilies (e.g. AGL17-like genes) expanded considerably in wheat whereas others (e.g. AGL6-like genes) have relatively few members as compared to other moncots. Using in silico analyses we deduced a relatively strong conservation of expression pattern within each subfamily, indicating functional similarity among closely related homologs.
We will use the identified MADS-box genes to screen for sequence variation among different wheat lines. This will provide a starting point to reveal how allelic variation in MADS-box genes may affect agronomically important traits in wheat.
71 Classification of Wheat Varieties in Satellite Images to Perform GWAS
Dept. of Crop and Soil Science, Washington State University
With dramatic reduction of sequencing cost, field trials have become the major burden of plant breeding to improve grain yield, quality and resistance to biotic and abiotic stress, especially on large scale. Abundant available satellite images have the potential to provide valuable data in regards to infection from disease, responses to drought and heat, as well as grain yield and quality. Most of these are due to the improvement of sensing resolution and shorter time intervals between pictures taken. The Satellite WorlView-2, launched in 2009, provides sensing at resolution within a meter on a near daily base. We review the satellite imagery resources and usages, including disease detection, crop discrimination, drought tolerance, nitrogen efficiency. Our objective was to inspire the joint usage of remote imagery data and genomic data for the genetic improvement in crops.
72 A Transcriptome Analysis of Genes Involved in the Production of Reactive Oxygen Species By P. triticina during Its Infection on Wheat
Agriculture and Agri-Food Canada
Reactive oxygen species (ROS) play an important role during host and pathogen interactions and are often an indication of induced host defense responses. The importance of these radicals for pathogenesis of the obligate biotrophic fungus, Puccinia triticina *(Pt), has not been investigated. In this study, we demonstrate that *Pt *generates ROS, including superoxide, H202 and hydroxyl radicals, during its infection of wheat. Through pharmacological inhibition, we show that ROS are critical for both *Pturediniospore germination and intercellular growth. A Pt *genome-wide screening identified 291 putative genes associated with general redox homeostasis and the search in RNA-seq data sets representing *Pt urediniospore germination, early and late infection stages on susceptible wheat cultivar Thatcher, found 37 genes annotated to encode known products related to oxidative stress responses. We identified two canonical Pt genes encoding NADPH oxidases (PtnoxA and PtnoxB), as well as a regulatory gene, *PtnoxR. *Real time qPCR analysis showed that all three genes were differentially regulated during urediniospore germination and infection on wheat.
73 Detoxification of Mycotoxins as a Source of Resistance to Fusarium Head Blight: From Brachypodium distachyon to Triticum aestivum
Institute of Plant Sciences Paris-Saclay
Fusarium head blight (FHB) caused by fungi of the Fusarium genus is a widespread disease of wheat (Triticum aestivum) and other small-grain cereal crops. The main causal agent of FHB, Fusarium graminearum, can produce mycotoxins mainly belonging to type B trichothecenes, such as deoxynivalenol (DON) that can negatively affect humans, animals and plants. Several quantitative trait loci (QTLs) for resistance to FHB have been identified some of which have been correlated with efficient DON detoxification, mainly through the conjugation of DON into DON-3-O-glucose (D3G), a reaction catalyzed by UDP-glucosyltransferases (UGTs). Nevertheless, only few studies have conducted functional analyses to directly correlate DON glucosylation and resistance in planta and none were performed on wheat UGT gene(s). Our team, using the model cereal species Brachypodium distachyon, has recently demonstrated that the Bradi5g03300 UGT is able to confer tolerance to DON following glucosylation of DON into DON 3-O-glucose and is involved in the early establishment of quantitative resistance to FHB. In the present work, we transferred the functional analyses conducted on the model species Brachypodium distachyon to bread wheat. In a first approach the B. distachyon Bradi5g03300 gene has been introduced through biolistic-mediated transformation in the wheat variety Apogee, susceptible to FHB. The phenotypic analyses conducted on homozygous transgenic wheat constitutively expressing the Bradi5g03300 gene showed that they exhibit higher resistance to FHB as well as increased root tolerance to DON compared to the control line. In parallel, using a synteny approach between B. distachyon and bread wheat genomes we identified a wheat candidate gene orthologous to the B. distachyon Bradi5g03300 gene. This wheat gene after validation through gene expression pattern during wheat infection, was introduced by transformation into B. distachyon to rapidly determine its ability to conjugate DON into D3G *in planta *and its involvement in FHB resistance. In conclusion, this project contributes to increase the knowledge concerning the functional relationship between DON glucosylation and FHB resistance in wheat and provide candidate genes to include in selection processes.
74 Genetic Architecture of Recombination Rate and Its Effects in Allopolyploid Wheat
Kansas State University
Recombination is a natural process that shapes the landscape of natural alleles within a population. Understanding the genetic basis of how variation in recombination rate is maintained and its effects are important to manipulate the recombination process in crops in order to improve them. We dissected naturally occurring variation in recombination rate present in a spring wheat NAM population and found it is mostly defined by rare alleles with small effects that explain up to 48.3% of the variation in our population. Specifically we identified 66 regions within the genome that contribute to the observed natural variation. Our regions are enriched for meiotic functioning genes, and encompass many conserved recombination genes. Further dissection suggests that the genetic architecture of recombination is predominantly additive and controlled by trans-acting features. In addition, we observed evidence of additive genetic factors that contribute to pericentromeric crossover (CO) frequency without affecting the frequency of telomeric COs. We have also observed a negative effect of linkage drag on deleterious mutation load resulting in excess strong-effect mutations in the pericentromeric genomic regions with constrained recombination. This information suggests manipulation of wheat is possible by influencing CO distribution and frequency, thus unlocking the whole genome without the consequences for linkage drag in wheat improvement.
75 Genomic Prediction and Genome-Wide Association Study of Grain Yield, Kernel Weight, and Kernel Number in a Durum Wheat Breeding Population
North Dakota State University
North Dakota leads the United States in acreage and production of durum wheat [Triticum tugidum L. subsp. durum (Desf.)]. Improvements in durum grain yield can result in substantial increases in profit for both farmers and the state. To date, all cultivar yield improvement in the North Dakota State University Durum (NDSU) Wheat Breeding Program has been achieved with phenotypic selection on replicated-plot yield trials in late generations. Applying genomic selection (GS) in earlier generations prior to replicated-plot testing could potentially increase genetic gain if the prediction model has merit. To understand the prospect of GS in the NDSU durum germplasm, unbalanced yield trials including approximately 1,000 breeding lines were used to generate GS models and predict breeding values of lines from the 2015 and 2016 generations. Generally, forward prediction accuracies increased as lines from additional breeding generations were added to the model. Forward prediction accuracies for the 2016 generation were 0.44, 0.42, and 0.35 for grain yield, kernel weight, and kernel number, respectively. Additionally, genome-wide association mapping revealed quantitative trait loci (QTL) for kernel weight. This information can further our understanding of genetic gain plant breeders can expect when applying GS to complex traits in an active wheat breeding program.
76 Application of CRISPR/Cpf1-Based Genome Editing in Polyploid Wheat
Kansas State University, Department of Plant Pathology
CRISPR/Cas9 has been widely applied in many organisms as a powerful genome editing tool. However, its target sites amenable to editing are limited by the 3’-end NGG proto-spacer adjacent motif (PAM). The CRISPR/Cpf1 requires 5’-end TTV or TTTV PAMs providing opportunities for targeting the AT-rich regions. The genome editing ability of FnCpf1 and LbCpf1 were assessed in wheat by combining transient expression in the wheat protoplasts and next generation sequencing (NGS) of the target regions. While no genome editing events were found for FnCpf1, about 1/3 of the designed targets for LbCpf1 showed the evidence of genome editing. The efficiency of editing was up to 10%, comparable to that of SpCas9 in the wheat protoplasts. We are also testing the editing efficiency of the mutated LbCpf1 (G532R/K538V/Y542R, henceforth LbCpf1m), which in mammalian cells was shown can induce mutations with the 5’-end of the TATV PAM. Multiplex gene editing using Cas9 is somewhat limited by the size of the transgenic constructs. The ability of Cpf1 to process its own CRISPR RNA (crRNA) and the shorter length (43 nucleotides) of crRNA make it a promising multiplex genome editing tool. Multiplex gene editing constructs with different numbers of crRNAs under the control of a single promoter were constructed. All multiplex genome editing constructs generated indels/insertions at the targets sites with the efficiency comparable to that of a single crRNA construct. Our results show that the LbCpf1 can further expand the set of tools available for engineering the wheat genome.
77 Identification of Quantitative Trait Loci (QTL) Associated with Fusarium Head Blight Resistance in a D8006W/ Superior Winter Wheat Population
Department of Plant Science, University of Manitoba
Fusarium head blight (FHB) caused by Fusarium graminearum is a major disease of wheat in North America. FHB infection reduces grain yield, affects end-use quality, and accumulates mycotoxins such as deoxynivalenol (DON) in the grain. The objective of this research was to identify QTL associated with FHB resistance. A doubled haploid soft white winter wheat population consisting of 107 lines from the cross D8006W/Superior was used. Evaluation for FHB reaction was performed using spray inoculation of a macroconidia mixture of four *F. graminearum *isolates representing two chemotypes in replicated field disease nurseries in three locations in Canada in 2016 and 2017. Disease incidence and severity were recorded 21 days post inoculation and FHB index was calculated. Percentage Fusarium damaged kernels and DON content were measured from collected grain samples. Both parental lines showed moderate reaction across all environments for FHB traits. However, the population showed transgressive segregation for FHB reaction with a wide continuous distribution. Genotyping of the population was performed using the 90K Illumina Infinium iSelect single nucleotide polymorphism array and 5194 high quality SNP were selected for analysis. Linkage mapping and QTL analysis is under processing. This experiment will be repeated in field nurseries in 2018. Significant FHB resistance QTL identified from this project will be used in winter wheat breeding programs using marker assisted selection.
78 Widening the Genepool and Identifying Genes Controlling Key Traits in Rice and Wheat
University of Queensland/QAAFI
Food security can be advanced by capturing more useful diversity in crop improvement and by better understanding the molecular basis of key traits that limit the rate of genetic gain in breeding. Recent genome sequencing research has identified wild populations representing significant new diversity in the primary gene pool of rice. This provides new sources of resistance to pests and disease, diversity to allow adaptation to climate change and novel grain qualities with potential consumer appeal and health benefits. Analysis of the transcriptome of the developing grain of diverse wheat germplasm has identified the genetic control of traits such as milling performance, hardness, and end-use quality (bread and chapatti). Genetic improvements can generate more grain per hectare more flour per tonne of wheat and more end-product per tonne of flour. These studies have demonstrated the great diversity of response to heat stress in the wheat gene pool. When combined these developments offer large improvements in food security.
79 A Near-Finished Reference Genome for the Wheat Blast Fungus Provides Insight on Pathogenicity and Adaptation
Kansas State University
University of Florida
Wheat blast, a devastating new fungal disease caused by the haploid fungus Magnaporthe oryzae pathotype Triticum *(MoT), has been spreading in South America since it was first reported in Brazil in 1985. This disease jumped to Bangladesh in 2016 and now poses a major threat to wheat production in South Asia and beyond. To produce a MoT reference genome, the highly aggressive MoT isolate B71, collected in Bolivia in 2012, was sequenced and assembled with long PacBio reads, Illumina sequences, and a novel scaffold technology using Long-Insert-End-Pair (LIEP) sequences, resulting in a near telomere-to-telomere assembly. The availability of the near-finished assembly facilitated the dissection of genomic structural variation (SV) between B71 and a less aggressive MoT strain isolated in Brazil in 1988, as well as among multiple *M. oryzae isolates adapted for infecting wheat or other crop species. A dispensable mini-chromosome in the B71 genome was identified through the SV analysis and verified, while none or two mini-chromosomes were found in two other MoT isolates. Two effector genes first characterized in the rice blast pathogen, M. oryzae pathotype Oryza, occur together in the MoT mini-chromosome, where both genes maintain characteristic in planta specific expression. The results imply the potential role of the mini-chromosome in the pathogenicity and adaptation of MoT.
80 Identification of Consistent Loci for Fusarium Head Blight Resistance in Northern European Spring Wheat through Genome-Wide Association Mapping
Graminor
Fusarium head blight resistance is quantitative, highly complex and divided into several different resistance types. QTL that are effective against several of the resistance types would be a valuable contribution for resistance breeding against this devastating wheat disease. A panel of 299 spring wheat lines with different geographical origin was tested in spawn-inoculated field trials and subjected to visual FHB assessment. In addition, DON level was analysed in the harvested seed. Anther extrusion (AE) was also assessed, in separate field trials. The panel was genotyped with the Affymetrix 35K SNP chip. Eight QTL, significant in three or more testing environments, were detected associated with both FHB and DON. These QTL were detected on chromosomes 1AS, 1AL, 2BL, 3B, 4AL, 5AL, 7AS and 7BS. AE was negatively correlated with FHB and DON, and association mapping could reveal seven AE QTL that coincided with the QTL detected for FHB and DON. The lines tested in the wheat panel harboured from zero to all the detected QTL, and the results show that resistance can be significantly increased by combining several of these resistance alleles. This information enhances the possibility to select crossing parents to obtain varieties more resistant to FHB and DON.
81 Harnessing ‘Left behind’ Drought-Adaptive Alleles for Modern Wheat Improvement
R. H. Smith Institute of Plant Science & Genetics in Agriculture, The Hebrew University of Jerusalem
The genetic diversity in wild ancestors of crop plants has been considerably eroded throughout plant domestication, evolution under cultivation and recent plant breeding, thereby making modern crop germplasm vulnerable to various biotic and abiotic stresses. Therefore, an important task of modern breeding is to identify and reintroduce valuable ‘left behind’ alleles into the modern domesticated gene pool. Introduction of such alleles has been mostly employed for improving biotic stress resistances, while abiotic stress adaptations received only minor attention. Water deficit is the major environmental factor limiting crop productivity, hence developing drought-resistant crop cultivars is essential to ensure a sustainable agricultural production under the ongoing climatic change.
Wild emmer wheat (Triticum turgidum ssp. dicoccoides [Körn.] Thell.), the progenitor of the domesticated durum (T. turgidum *spp. durum* (Desf.) MacKey) and bread (T. aestivum L.) wheats, harbors a rich allelic diversity that is valuable for future improvement. Selected drought-adaptive QTL alleles were introgressed from wild emmer wheat into modern durum and bread wheat cultivars and the resultant near isogenic lines (NILs) were tested for their drought responses. NILs introgressed with wild emmer QTL on chromosome 7A, exhibited under water-limited conditions greater grain yield, osmotic adjustment, photosynthetic capacity and root development compared with their recurrent parent. Selected NILs, carrying the 7A QTL in two genetic backgrounds, tested across 3 years under commercial field conditions, exhibited a significant advantage over their parental cultivars, particularly under drought, thus confirming the potential of this left behind QTL allele for improving drought resistance in modern wheat.
82 Development of Genetic and Genomic Resources to Evaluate Wheat Organellar Genome Variants and their Functional Implications
Department of Plant Pathology, University of MinnesotaCereal Disease Laboratory, United States Department of Agriculture-Agricultural Research Service
Organellar genome diversity represents a potential untapped source for improving agronomic traits. Due to the nature of hybridization and domestication events that led to modern cultivated wheat, organellar (mitochondria and chloroplast) genome diversity was dramatically reduced. However, wheat has the largest known collection of alloplasmic (alien cytoplasm) lines. In these lines, the cytoplasms, and therefore the organelles, of wild relatives have been mismatched with the nuclear genome of domesticated wheat through extensive backcrossing. Disruption of native nuclear-cytoplasmic interactions (NCI) impacts a number of agronomic traits including fertility, biomass, grain yield, and stress response. To understand the functional implications of organellar genome variants and genetic mechanisms underlying NCI, we generated organellar genomic resources. We developed a method to couple organellar DNA enrichment from total gDNA utilizing a pull-down approach and ultra-low input library preparations for long-read sequencing. We sequenced 20 organellar-enriched samples with PacBio, including 13 diverse wild species, T. durum, T. aestivum cv. Chinese Spring, and three alloplasmic lines. We also generated Illumina short-read sequences for >75 cultivars, wild species, and alloplasmics. Comparative analyses are investigating organellar diversity across the Triticum-Aegilops complex and how sequences change in the alloplasmic condition. In parallel, we are generating additional genetic and genomic resources in wheat and the model system Brachypodium distachyon for functional genomics studies. These resources will be useful to the broader community for furthering our understanding of the molecular mechanisms involved in NCI and their affects on plant phenotype as well as for breeding efforts to improve agronomic traits.
83 POTAGE: A Visualisation Tool for Speeding up Gene Discovery in Wheat
The University of Adelaide
POPSEQ Ordered Triticum aestivum *Gene Expression (POTAGE) is a web application which accelerates the process of identifying candidate genes for quantitative trait loci (QTL) in hexaploid wheat. This is achieved by leveraging several of the most commonly used data sets in wheat research. These include the Chromosome Survey Sequences, their order along the chromosomes determined by the population sequencing (POPSEQ) approach, the gene predictions and RNA-Seq expression data. POTAGE aggregates those data sets and provides an intuitive interface for biologists to explore the expression of the predicted genes and their functional annotation in a chromosomal context. The interface accelerates some of the laborious and repetitive tasks commonly undertaken in the process of identifying and prioritising genes which may underlie QTL. We illustrate the utility of POTAGE by showing how a short-list of candidate genes can quickly be identified for a QTL linked to pre-harvest sprouting - a major cause of quality and yield loss in wheat production. The candidate genes identified using POTAGE included *TaMKK3, which was recently reported as a causal gene for seed dormancy in wheat, and a mutation in its barley ortholog has been shown to reduce pre-harvest sprouting. In addition to the public version of POTAGE, we have also developed a Docker image for quickly deploying POTAGE locally and work-flows showing how to add your own expression data sets to a local installation. This is of particular relevance to those who work with unpublished data sets or would like to deploy POTAGE on their own hardware.
84 Evaluation of the Effect of an Alien Chromosome Segment Translocated from Aegilops sharonensis on Recombination Frequency in Wheat
Graduate School of Agriculture, Kyoto University
Background: The resolution of genetic map depends on meiotic crossover frequency between homologous chromosomes. Due to a low crossover frequency in the wheat genome (Triticum aestivumL.), a large population is needed to obtain informative recombinants. Moreover, it has been demonstrated that artificial allotriploids in the Brassica species exhibit genome-wide elevation in recombination frequency. However, the effect of alien chromosomes on meiotic recombination still remains poorly understood in crop species. Thus, the purpose of this study is to demonstrate if the alien chromatin (segments) can increase recombination/crossover frequency in wheat.
Methods: We developed an F2 population derived from an accession DT4B-4Ssh. The terminal segment of the chromosome 4Ssh of Aegilops sharonensis was translocated to chromosome 4B of Chinese Spring (CS), which was subsequently crossed with a spelta wheat (T. spelta L. var. duhamelianum, accession KT019-001). The F2 population derived from normal CS and KT019-001 was used as control. Linkage maps were constructed using the same set of 494 markers scattered on A and B genome chromosomes.
Results: The complete linkage map derived from DT4B-4Ssh was ~11% longer than from CS. The average crossover numbers significantly increased for the entire A and B genome respectively. The observed elevation of recombination frequencies was not concentrated in a particular chromosomal region. In conclusion, our results indicated that the chromosome segment introduced from *Ae. sharonensis *increased recombination frequency globally. This suggests that small alien chromatin may affect meiotic recombination in wheat.
85 Reduced Height Semi-Dwarf Alleles Significantly Impact Wheat End Use Quality
Montana State University
The genes responsible for dramatic yield increases during the 1950s and 1960s are mutant forms of the Reduced height (Rht) genes. Since then, two semi-dwarfing alleles of Rht, Rht-B1b and Rht-D1b have been widely incorporated into modern wheat cultivars. Despite being some of the most widely utilized yield increasing genes, few studies have examined their effect on wheat end use quality. For this study we compared near isogenic lines created in a standard height spring wheat variety (Fortuna) varying for the presence of the gibberellin insensitive mutations Rht-B1b, Rht-D1b, *or the gibberellin sensitive dwarfing gene *Rht-8. Our agronomic results agreed with previous findings and we observed a 25% height reduction and 13% yield increase in Rht-B1b/Rht-D1b compared to the tall isoline. Grain protein was decreased (from 15.4 to ~13.6%) as was kernel weight (15%) in the Rht-B1b/Rht-D1b isolines. We also saw a slight decrease in the loaf volume in Rht-B1b/Rht-D1b compared to the tall line. However, we observed statistically significant increases in flour yield (2%), falling number (5%), mixing tolerance (56%), and baking mix time (33%). The fact that flour yield was increased is unexpected since Rht-B1b/Rht-D1b also decrease seed size. For almost all parameters, Rht-8 was intermediate between the tall line and Rht-B1b/Rht-D1b. These findings indicate that although Rht-B1b/Rht-D1b decrease seed size and protein content, they positively impact wheat milling yield and do not negatively impact dough strength.
86 Analysis of Wheat Internode Time Course RNAseq Data to Identify Genes Related to Nutrient Remobilisation Depending on Nitrogen Availability
Rothamsted Research
The massive and polyploid genome of wheat has make it challenging for researchers to leverage RNAseq to identify candidate genes underlying complex phenotypes. Most studies in wheat have focused on high-level characteristics of gene expression because the available references were incomplete and fragmented and over 100,000 transcribed genes are assayed simultaneously. In other organisms, well designed and carefully analysed RNAseq experiments have contributed to gain insights into mechanisms controlling complex traits and we intend to follow a similar approach in wheat genomics now that near-complete references are available.
Here we describe the bioinformatics analysis of a time course RNAseq experiment in which internode samples from hexaploid wheat (T. aestivum) plants treated with two nitrogen levels have been collected at 8 time points between anthesis and senescence. The study aims to elucidate the molecular mechanisms involved in nutrient remobilisation from senescing organs to developing tissues and the role of nitrogen availability in senescence. The chloropyll level has been used as a marker of senescence and the DESeq2 Bioconductor package has been used to identify genes that switch their state at its onset.
We demonstrate that it is possible to reduce the number of candidate genes from thousands to dozens using statistical methods to identify genes that have an expression profile of interest. In this work, 9 genes have been found to be upregulated and 8 downregulated in senescence and their functions seem to be related to processes like chlorophyll binding, protein breakdown and cell signalling.
87 Towards Chromatin Immunoprecipitation (ChIP)-DNA Isolation and Epigenome Development in Durum Wheat Tissues
Delaware State University
Chromatin Immunoprecipitation (ChIP) examines the connections between proteins and DNA contained in the genome of an organism by observing histone modifications. Histone modifications incorporate the ability to relax and condense chromatin, which in turn, helps govern gene expression and repression. In acetylation, the chromatin relaxes, which allows transcription factors to reach their target, causing gene expression. In methylation, the chromatin condenses, restricting transcription factors from reaching their target, obstructing gene expression. Tri-methylated lysine four on histone three (H3K4me3) plays a role in activation in gene promoter regions. Histone-DNA interactions might play an important role in fighting drought, adjusting to heat stress, and combating diseases during plant developmental stages. In this work, we will optimize ChIP-DNA extractions in Durum wheat and observe ChIP-DNA isolates in four tissues (leaf, root, head, and two week old seedling) to detect variances in histone-DNA interactions among the group using Illumina sequencing. By optimizing the ChIP-DNA isolation protocol in Durum wheat we may better understand histone modifications and the role they play in fighting abiotic and biotic stresses. Additionally, we will use quantitative PCR to identify chromosome remodeling gene expression differences among tissue types in wheat. Knowing where these particular genes are located in the genome may help us understand their importance. The objective of this study is to serve as a blueprint for examining spatial histone methylation and acetylation in wheat using the ChIP assay.
88 Assessing the Consequences of Key Events of the Hexaploid Wheat Genome Evolution: Structural and Evolutive Analysis of an Ancestral Chromosomes Fusion Point and of a Region Resulting from Ancient and Recent Polyploidizations
French Plant Genomic Center CNRGV - INRA
Studying the bread wheat genome’s structure has been a tremendous challenge for decades. Its complexity (hexaploidy, high rate of repetitive elements) and size make wheat genome both complex to decipher and interesting to investigate as the result of multiple evolutionary forces (ancient and recent polyploidization, chromosomes fusions, deletion or sub-functionalization of homeologous genes). To study these events we first aimed at characterizing a chromosomes fusion locus (CF) and comparing the structure of the copies of a 2 Mb region carried by chromosomes 1A, 1B, 1D, 3A, 3B, 3D. Before the release of a wheat reference genome, our strategy was to identify BACs spanning the regions of interest using available resources (chromosome specific BAC libraries, physical maps, genome zipper, WGP) and to sequence BACs using PacBio technology. Despites all these resources, it remained difficult to sequence the CFs which are highly rearranged regions and the six copies of the region of interest. The release of the IWGSC reference genome followed by its annotation have changed our strategy by giving access to the whole genome sequence. Thanks to this sequence, we have redefined the orthologous relationship between hexaploid wheat genes and rice genes (as rice has conserved the structure of the common grass ancestor). We have identified various rearrangements and precisely outlined the CF regions. Following this analysis at the genome scale, we focused on the rearrangements of the CF regions of chromosomes 1A, 1B, 1D and on the fates of homeologous genes among the six copies of a region of interest.
89 Improved Markers for a Pre-Harvest Sprouting QTL on Wheat Chromosome 3B
Agriculture & Agri-Food Canada
A QTL for sprouting index on chromosome 3B was identified in the AC Domain x RL4452 bi-parental mapping population (Cabral et al. 2014, BMC Plant Biology 14:340). The flanking markers derived from the Illumina iSelect 90K chip define a region of 185 Mbp in the wheat RefSeq 1.0 genome sequence containing over 1180 annotated high confidence genes. Additional markers are needed to narrow down this region for wheat breeding purposes as well as to eventually identify candidate genes. As a first step to identify additional markers SNPs previously validated in the KASP assay were identified by locating QTL flanking markers from the 'AC Domain' x RL4452 3B map on the 'Avalon' x 'Cadenza' map. Validated KASP markers mapped on Avalon x Cadenza in the region between QTL flanking markers were chosen (http://lgcapps.com/assays/wheat/) for genotyping and mapping on the 'AC Domain' x RL4452 population. Subsequently the full genomic region between flanking markers from the IWGSC RefSeq1.0 genome sequence was used to identify SNPs between the parental genotypes in a sequence database derived from exome capture sequencing of Canadian wheat genotypes. 1492 SNPs were identified in 288 annotated genes. The SNPs will be useful to further narrow down the QTL region and provide robust markers for selection of lines with increased tolerance to pre-harvest sprouting.
90 Discovering Superior Alleles Underlying Drought and Heat Tolerance in Common Wheat
Institute of Crop Science, Chinese Academy of Agricultural Sciences
Environmental stresses, drought and heat, especial the stresses in grain filling period are the primary causes of grain yield losses in wheat (Triticum aestivum L.). Abundant wheat germplasm resources with tolerance to drought (DT) and heat (HT) have been identified. They are important gene resources for wheat improvement. However, very little is known about these resources, such as what tolerant-genes these germplasm possess, in which accessions have the superior alleles underlying the tolerances, and how to use these alleles effectively. Our researches showed that some genes involved in the DT and HT, also contributed to the yield-related traits. Here, we dissect the functions of gene Ta**SnRK2 (sucrose non-fermenting1-related protein kinase 2), TaPP2A (protein phosphatase 2A), and TaSPL (squamosa-promoter binding protein, SBP-box). TaSnRK2 and TaPP2A not only responded to abiotic stress, but also significantly associated with 1000-grain weight under terminal drought and heat stress. Two of the SPL *members, *TaSPL20 and TaSPL21 associated with 1000-grain weight and plant height in multi-environment. According to the functional analysis of naturally occurring variants, favorable alleles/haplotypes were identified with the perfect markers. They increased 1000-grain weight, and reduced plant height in well-watered, drought, and heat environments. Our study suggests that during domestication and breeding of wheat in China, superior alleles of each gene were selected and exploited to varying degrees due to their large effects on the yield-related traits.
https://m.sciencenet.cn/blog-1094241-1094517.html
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