Sequence-Based Marker Assisted Selection in Wheat

Wheat improvement has traditionally been conducted by relying on artificial crossing of suitable parental lines followed by selection of the best genetic combinations. At the same time wheat genetic resources have been characterized and exploited with the aim of continuously improving target traits. Over this solid framework, innovations from emerging research disciplines have been progressively added over time: cytogenetics, quantitative genetics, chromosome engineering, mutagenesis, molecular biology and, most recently, comparative, structural, and functional genomics with all the related -omics platforms. Nowadays, the integration of these disciplines coupled with their spectacular technical advances made possible by the sequencing of the entire wheat genome, has ushered us in a new breeding paradigm on how to best leverage the functional variability of genetic stocks and germplasm collections. Molecular techniques first impacted wheat genetics and breeding in the 1980s with the development of restriction fragment length polymorphism (RFLP)-based approaches. Since then, steady progress in sequence-based, marker-assisted selection now allows for an unprecedently accurate ‘breeding by design’ of wheat, progressing further up to the pangenome-based level. This chapter provides an overview of the technologies of the ‘circular genomics era’ which allow breeders to better characterize and more effectively leverage the huge and largely untapped natural variability present in the Triticeae gene pool, particularly at the tetraploid level, and its closest diploid and polyploid ancestors and relatives

Genome-wide association study for resistance in bread wheat (Triticum aestivum L.) to stripe rust (Puccinia striiformis f. sp. tritici) races in Argentina

Background: Wheat stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), is one of the most devastating diseases of the wheat crop. It causes significant reductions in both grain yield and grain quality. In recent years, new and more virulent races have overcome many of the known resistance genes in Argentinian germplasm. In order to identify loci conferring resistance to the local races of Pst for effective utilization in future breeding programs, a genome-wide association study (GWAS) was performed using a collection of 245 bread wheat lines genotyped with 90 K SNPs. Results: To search for adult plant resistance (APR) the panel was evaluated for disease severity (DS) and area under disease progress curve (AUDPC) in field trials during two years under natural infection conditions. To look for seedling or all-stage resistance (ASR) the panel was evaluated to determine infection type (IT) under greenhouse conditions against two prevalent races in Argentina. The phenotypic data showed that the panel possessed enough genetic variability for searching for sources of resistance to Pst. Significant correlations between years were observed for Pst response in the field and high heritability values were found for DS (H2 = 0.89) and AUDPC (H2 = 0.93). Based on GWAS, eight markers associated with Pst resistance (FDR < 0.01) were identified, of these, five were associated with ASR (on chromosomes 1B, 2A, 3A and 5B) and three with APR (on chromosomes 3B and 7A). These markers explained between 2% and 32.62% of the phenotypic variation. Five of the markers corresponded with previously reported Yr genes/QTL, while the other three (QYr.Bce.1B.sd.1, QYr.Bce.3A.sd and QYr.Bce.3B.APR.2) might be novel resistance loci. Conclusion: Our results revealed high genetic variation for resistance to Argentinian stripe rust races in the germplasm used here. It constitutes a very promising step towards the improvement of Pst resistance of bread wheat in Argentina. Also, the identification of new resistance loci would represent a substantial advance for diversifying the current set of resistance genes and to advance in the improvement of the durable resistance to the disease.EEA BalcarceFil: Franco, María Fiorella. Universidad Nacional de Mar del Plata. Facultad de Ciencias Agrarias; Argentina.Fil: Franco, María Fiorella. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina.Fil: Franco, María Fiorella. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina.Fil: Polacco, A. N. Universidad Nacional de Mar del Plata. Facultad de Ciencias Agrarias; Argentina.Fil: Campos, P. E. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Bordenave; Argentina.Fil: Pontaroli, Ana Clara. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Balcarce; Argentina.Fil: Pontaroli, Ana Clara. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina.Fil: Vanzetti, Leonardo Sebastián. Instituto Nacional de Tecnología Agropecuaria (INTA). Estación Experimental Agropecuaria Marcos Juárez; Argentina

Zymoseptoria tritici fungicide resistance in wheat

Magistritöö Põllumajandussaaduste tootmise ja turustamise õppekavalPopulaarse toiduteravilja nisu olulist saagikuse langust põhjustab kõrreliste helelaiksus, mida põhjustab patogeen Zymoseptoria tritici. Fungitsiidide kasutamine aitab pärssida patogeenide elutegevust ja levikut. Peamiselt kasutatakse Z. tritici tõrjeks fungitsiidirühmadest asoole, suktsinaatdehüdrogenaasi inhibiitoreid ja strobiluriine. Töö eesmärgiks oli võrrelda Z. tritici fungitsiidide tundlikkuse arengut 2019. ja 2020. aasta vahel ja määrata 2019. ja 2020. aastatel Eestis levinud Z. tritici populatsioonis fungitsiidiresistentsust põhjustavad märklaudvalkude mutatsioonid. Eesti talinisu põldudelt isoleeriti kahel aastal kokku 283 Z. tritici puhaskultuuri. Isolaatidel määrati fungitsiidide tundlikkus mikrotiiterplaadimeetodil asoolide suhtes ning mutatsioonid märklaudvalkude geenides (CYP51, SdhB, SdhC ja CytB). 2019 ja 2020 aasta võrdluses leiti, et asoolide tundlikkus toimeainete suhtes oli Eesti Z. tritici populatsioonis varieeruv. Epoksikonasooli ja mefentriflukonasooli tundlikkus suurenes, protiokonasool-destio tundlikkus vähenes ning esines resistentsus tebukonasooli suhtes. Tundlikkuse vähenemist saab seostada märklaudvalkudes tekkinud mutatsioonidega. Asoolide märklaudvalgus CYP51-s olid levinud mutatsioonid D134G, V136A/C, G379A, I381V ja S524T. Tõusutrendi näitasid D134G, V136C, A379G ja S524T. V136A ja I381V sagedus langes kahe aasta võrdluses. Domineerivaim mutatsioon oli I381V. Kõige tugevam seos on asoolide resistentsuse tekkega mutatsioonil S524T. SDHI märklaudvalgu SdhC mutatsioonid N33T, N34T ja N86S on Eesti Z. tritici populatsioonis levinud. Esmakordselt avastati Eesti populatsioonis valgus SdhB esinev mutatsioon N225I. Ükski SdhC ja SdhB mutatsioonidest ei langeta märkimisväärselt SDHI-de tundlikkust. Strobiluriinide märklaudvalgus CytB olev mutatsioon G143A, mis tekitab resistentsuse, esineb 44 kuni 49 % isolaatidest, kuid varieerub tugevalt maakonniti. Kõrreliste helelaikuse tõrjeks sobivad Eestis kasutamiseks nii asoolid, SDHI-d kui strobiluriinid, kuid soovitav on omavahel vahetada erinevaid toimeaineid fungitsiidide resistentsuse tekkeriski minimeerimiseks. Tõrjevahendite valikul tuleks lähtuda ka piirkondlikest eripäradest. Töö vajab edasisi uuringuid, et põllumajandustootjad saaksid ennetada kõrreliste helelaiksuse tekitaja fungitsiidiresistentsust ja kasutada toimeaineid, millele haigustekitaja tundlik on.The significant decrease in the yield of the popular food crop wheat is caused by the Septoria tritici blotch, caused by the pathogen Zymoseptoria tritici. The use of fungicides helps to inhibit the activity and spread of pathogens. Azoles, succinate dehydrogenase inhibitors and strobilurins are mainly used to control Z. tritici. The study aimed to compare the development of susceptibility of the Z. tritici Estonian population to fungicides between 2019 and 2020. Another aim was to determine the mutations in target proteins causing fungicide resistance in the population of Z. tritici prevalent in Estonia in 2019 and 2020. In two years, a total of 283 single spore isolates of Z. tritici were isolated from Estonian winter wheat fields. In each isolate, fungicide susceptibility to azoles was determined by the microtiter plate method and sequencing the pointmutations in target protein genes (CYP51, SdhB, SdhC and CytB). In the comparison of 2019 and 2020, it was found that the sensitivity of azoles varied in the Estonian Z. tritici population. The susceptibility of epoxiconazole and mefentrifluconazole increased, the susceptibility of prothioconazole-destio decreased, and the resistance to tebuconazole prevailed. Decreased sensitivity can be associated with mutations in target proteins. Pointmutations D134G, V136A/C, A379G, I381V and S524T were common in the target protein CYP51. D134G, V136C, A379G and S524T showed an upward trend. The frequency of V136A and I381V decreased in the two-year comparison. The predominant mutation in the population was I381V. The strongest association with azole resistance is with S524T mutation. The SdhC mutations N33T, N34T and N86S of the SDHI target protein are common in the Estonian Z. tritici population. For the first time, the N225I mutation in the SdhB protein was detected in the Estonian population. None of the SdhC and SdhB mutations significantly decrease the sensitivity of SDHIs. The G143A mutation in the strobilurin target protein CytB is present in 44 to 49 % of isolates, but varies widely between the counties. Azoles, SDHIs and strobilurins are suitable for use in Estonia to control the Septoria tritici blotch, but it is recommended to rotate different active ingredients to minimize the risk of developing fungicide resistance. The choice of control measures should also take regional specificities into account. Further research is required to enable farmers to prevent fungicide resistance and to use active substances to which the pathogen is sensitive

Development of new tools and germplasms for improvement of wheat resistance to Fusarium head blight

Doctor of PhilosophyDepartment of AgronomyGuihua BaiXiaomao LinWheat Fusarium head blight (FHB) is a devastating disease of wheat worldwide, which can significantly reduce grain yield and quality. Although the application of fungicides can reduce FHB damage, growing FHB resistant wheat is the most effective and eco-friendly approach to reduce the losses. To develop locally adapted FHB-resistant hard winter wheat germplasm, we transferred three major QTLs: Fhb1, Qfhs.ifa-5A, and Qfhb.rwg-5A.2 into two hard winter wheat cultivars, ‘Everest’ and ‘Overland’, using marker-assisted backcrossing and multiplex restriction amplicon sequencing (MRASeq). Ten ‘Overland’ background lines and nine ‘Everest’ background lines with better FHB resistance, recurrent parent similar agronomic traits were selected. They can be used as FHB resistant bridge parents for hard winter wheat breeding. To identify native FHB resistant sources, a population of 201 U.S. breeding lines and cultivars were genotyped using 90K wheat SNP arrays and phenotyped for the percentage of symptomatic spikelets (PSS), Fusarium damaged kernels (FDK) and deoxynivalenol (DON), a toxin produced by the pathogen. Genome-wide association studies (GWAS) identified significant trait associations with single nucleotide polymorphisms (SNPs) on chromosomes 1A, 1D, 2B, 3A, 3B, 4A, 5B and 5D. These marker-trait associations (MTAs) were significant for at least two of the three traits or a single trait in at least two experiments. To accelerate the evaluation of the FDK, we developed an algorithm that can separate FDK from healthy kernels with an accuracy of 90% based on color differences using image processing and unsupervised machine learning methods. Discovery and creation of the new FHB resistant germplasms and development of the fast FDK phenotyping algorithm will accelerate the improvement of U.S. hard winter wheat cultivars for FHB resistance

Towards map-based cloning of a Hessian fly resistance gene H34 in wheat

Doctor of PhilosophyDepartment of AgronomyAllan K FritzWheat is a staple food crop worldwide and insect damage is a major constraint for its production. Among the insects, Hessian fly (HF, Mayetiola destructor) is a destructive pest that significantly reduces wheat grain yield. To date, 37 HF resistant genes have been named, but diagnostic markers for these genes are lacking, which hampers their deployment in wheat breeding. HF resistance gene H34 on the short arm of chromosome 6B was one of the genes from a U.S. winter wheat Clark. To finely map H34, a cross was made between two F12 recombinant inbred lines (RIL115-S and RIL118-R) derived from Ning7840 x Clark. RIL118-R carries the resistance allele and RIL118-S carries the susceptibility allele at H34. Screening 286 (RIL115-S x RIL118-R) F3 lines using flanking Kompetitive Allele Specific PCR (KASP) markers identified five heterogenous inbred families (HIFs) segregating at H34. The first round of screening of 159 homozygous recombinant plants from five different HIFs using the KASP markers delimited H34 to a 5.0 Mb interval. Genotype-by-sequencing (GBS) analysis of the four pairs of near-isogenic lines (NILs) from the selected HIFs identified additional SNPs in the H34 region that further narrowed the H34 region to 1.3 Mb after screening 75 additional homozygous recombinant NILs. RNA-sequencing (RNA-seq) of the four pairs of NILs identified three differentially expressed genes (DEGs) in the H34 interval and they were considered as the putative H34 candidate genes for further study. Using the sequences of the DEGs and GBS-SNPs identified in the H34 interval, seven KASP markers were designed and validated to be diagnostic in a US winter wheat panel of 203 lines. These markers can be used in gene pyramiding of H34 with other HF resistance genes using marker-assisted selection (MAS) in the U.S. wheat-breeding programs. Furthermore, studying mechanism of HF resistance in wheat using RNA-seq data revealed that genes encoding defense proteins, stress-regulating transcription factors, and secondary metabolites were strongly up regulated within the first 48 hours of larval feeding, revealing an early defense in resistant wheat plants in response to larval attack. Also, HF feeding on resistant plants triggered the secretions of R-gene receptors by HF to initiate a hypersensitive response (HR) in the plants. This HR response resulted in production of reactive oxygen species (ROS) to up regulate the downstream genes involved in cell wall fortification and activation of different transcription factors (TFs), which prevents HF to access the nutrients in the resistant plants and eventually results in the death of HF larvae. The new knowledge generated in this study will aid in better understanding of HF-resistant mechanisms and developing new crop improvement strategies to increase HF resistance in wheat

Wheat Improvement

This open-access textbook provides a comprehensive, up-to-date guide for students and practitioners wishing to access in a single volume the key disciplines and principles of wheat breeding. Wheat is a cornerstone of food security: it is the most widely grown of any crop and provides 20% of all human calories and protein. The authorship of this book includes world class researchers and breeders whose expertise spans cutting-edge academic science all the way to impacts in farmers’ fields. The book’s themes and authors were selected to provide a didactic work that considers the background to wheat improvement, current mainstream breeding approaches, and translational research and avant garde technologies that enable new breakthroughs in science to impact productivity. While the volume provides an overview for professionals interested in wheat, many of the ideas and methods presented are equally relevant to small grain cereals and crop improvement in general. The book is affordable, and because it is open access, can be readily shared and translated -- in whole or in part -- to university classes, members of breeding teams (from directors to technicians), conference participants, extension agents and farmers. Given the challenges currently faced by academia, industry and national wheat programs to produce higher crop yields --- often with less inputs and under increasingly harsher climates -- this volume is a timely addition to their toolkit

A chromosome-based draft sequence of the hexaploid bread wheat (<em>Triticum aestivum</em>) genome.

An ordered draft sequence of the 17-gigabase hexaploid bread wheat (Triticum aestivum) genome has been produced by sequencing isolated chromosome arms. We have annotated 124,201 gene loci distributed nearly evenly across the homeologous chromosomes and subgenomes. Comparative gene analysis of wheat subgenomes and extant diploid and tetraploid wheat relatives showed that high sequence similarity and structural conservation are retained, with limited gene loss, after polyploidization. However, across the genomes there was evidence of dynamic gene gain, loss, and duplication since the divergence of the wheat lineages. A high degree of transcriptional autonomy and no global dominance was found for the subgenomes. These insights into the genome biology of a polyploid crop provide a springboard for faster gene isolation, rapid genetic marker development, and precise breeding to meet the needs of increasing food demand worldwide