Genetic study for seed germination and shattering in Euphorbia lagascae in response to different seed treatments.

Euphorbia lagascae Spreng is a promising emerging oilseed crop, with its seed oil accounting for approximately 50% of the seed weight. Euphorbia oil contains a significant amount of vernolic acid, comprising two-thirds of its composition, which boasts various industrial applications, including acting as a stabilizer-plasticizer and natural dye. However, this species was known to have a high degree of seed-shattering and a low germination rate, which act as two important barriers to large-scale production and exploitation. Therefore, the objective of this study is to determine the genetic control of seed germination and seed-shattering traits in order to develop a reliable pipeline that would be applicable for industries and breeders to select superior E. lagascae lines and design a robust breeding scheme in a short time at reduced labor costs. For this objective, five different wild-type genotypes of E. lagascae that demonstrated high germination potential were crossed with an ethyl methanesulfonate (EMS) mutant genotype that produces non-shattering capsules. The F2 populations from two successful crosses (A and B) were separated into three different treated groups for seed germination evaluation and to study the segregation of 200 individuals per F2 population. The three treatments were: light, gibberellic acid (GA3), and control treatment. Consequently, plants treated with approximately 250 µmol/m2/s of light showed significant improvement in germination up to 75% in cross A and 82.4 % in cross B compared with the control plants and the group treated with 0.05% GA3. According to the chi-square test results, the inheritance pattern of seed germination in response to light treatment follows a 3:1 segregation ratio between germinated and non-germinated seeds, indicating a dominant gene action in the F2 generation. The same conclusion was followed for the shattering trait in the group treated with light, which was also simply inherited as a 3:1 ratio for shattering vs. non-shattering capsules. Our results emphasize the importance and significance of light treatment in producing uniform populations through acceptable germination and shattering resistance of the mutant genotypes of E. lagascae. This is the first report of light treatment that significantly improved seed germination of E. lagascae, which may enhance efforts in the development of this new industrial crop as a feedstock for vernolic acid production.

Improvement of germination rate and hybridization to facilitate breeding of an industrial oil crop, Euphorbia lagascae Spreng.

BACKGROUND: The potential of plant-based sources of vernolic acid to provide agricultural producers with a market diversification opportunity and industrial manufacturers with a renewable, environmentally friendly chemical feedstock is immense. The herbaceous wild spurge or caper spurge (Euphorbia lagascae Spreng) is the most promising source of vernolic acid, containing an average oil content of 50%, of which around 60% is vernolic acid. Its seed yield ranges between 500 and 2000 kg ha-1, and a theoretical yield of 180 kg ha-1 of pure vernolic acid is possible. The objective of this research was to characterize the flower and whole plant morphology so to allow for the development of a method to efficiently hybridize E. lagasce plants for breeding purposes. RESULTS: In this study, we have characterized the flower and whole plant morphology in detail, thereby, developing an efficient method for hybridization of E. lagasce to allow for its breeding and improvement as a novel oil crop. Such method was not described previously in the literature making it difficult to breed this crop. We believe that the method will be of great value to plant breeders working on optimizing the crop, particularly in terms of the development of non-shattering cultivars with enhanced germination potential. CONCLUSIONS: The successful development of this crop through plant breeding could provide substantial economic benefits to farmers by offering them a new industrial oilseed crop. This research could prove invaluable in unlocking the potential of E. lagasce, and in turn, the potential of vernolic acid as a renewable, environmentally friendly source of chemical feedstock.

Unveiling the Mysteries of Non-Mendelian Heredity in Plant Breeding.

Mendelian heredity is the cornerstone of plant breeding and has been used to develop new varieties of plants since the 19th century. However, there are several breeding cases, such as cytoplasmic inheritance, methylation, epigenetics, hybrid vigor, and loss of heterozygosity (LOH), where Mendelian heredity is not applicable, known as non-Mendelian heredity. This type of inheritance can be influenced by several factors besides the genetic architecture of the plant and its breeding potential. Therefore, exploring various non-Mendelian heredity mechanisms, their prevalence in plants, and the implications for plant breeding is of paramount importance to accelerate the pace of crop improvement. In this review, we examine the current understanding of non-Mendelian heredity in plants, including the mechanisms, inheritance patterns, and applications in plant breeding, provide an overview of the various forms of non-Mendelian inheritance (including epigenetic inheritance, cytoplasmic inheritance, hybrid vigor, and LOH), explore insight into the implications of non-Mendelian heredity in plant breeding, and the potential it holds for future research.

Genome-wide analysis of cold imbibition stress in soybean, Glycine max.

In Canada, the length of the frost-free season necessitates planting crops as early as possible to ensure that the plants have enough time to reach full maturity before they are harvested. Early planting carries inherent risks of cold water imbibition (specifically less than 4°C) affecting seed germination. A marker dataset developed for a previously identified Canadian soybean GWAS panel was leveraged to investigate the effect of cold water imbibition on germination. Seed from a panel of 137 soybean elite cultivars, grown in the field at Ottawa, ON, over three years, were placed on filter paper in petri dishes and allowed to imbibe water for 16 hours at either 4°C or 20°C prior to being transferred to a constant 20°C. Observations on seed germination, defined as the presence of a 1 cm radicle, were done from day two to seven. A three-parameter exponential rise to a maximum equation (3PERM) was fitted to estimate germination, time to the one-half maximum germination, and germination uniformity for each cultivar. Genotype-by-sequencing was used to identify SNPs in 137 soybean lines, and using genome-wide association studies (GWAS - rMVP R package, with GLM, MLM, and FarmCPU as methods), haplotype block analysis, and assumed linkage blocks of ±100 kbp, a threshold for significance was established using the qvalue package in R, and five significant SNPs were identified on chromosomes 1, 3, 4, 6, and 13 for maximum germination after cold water imbibition. Percent of phenotypic variance explained (PVE) and allele substitution effect (ASE) eliminated two of the five candidate SNPs, leaving three QTL regions on chromosomes 3, 6, and 13 (Chr3-3419152, Chr6-5098454, and Chr13-29649544). Based on the gene ontology (GO) enrichment analysis, 14 candidate genes whose function is predicted to include germination and cold tolerance related pathways were identified as candidate genes. The identified QTLs can be used to select future soybean cultivars tolerant to cold water imbibition and mitigate risks associated with early soybean planting.

Application of SVR-Mediated GWAS for Identification of Durable Genetic Regions Associated with Soybean Seed Quality Traits.

Soybean (Glycine max L.) is an important food-grade strategic crop worldwide because of its high seed protein and oil contents. Due to the negative correlation between seed protein and oil percentage, there is a dire need to detect reliable quantitative trait loci (QTL) underlying these traits in order to be used in marker-assisted selection (MAS) programs. Genome-wide association study (GWAS) is one of the most common genetic approaches that is regularly used for detecting QTL associated with quantitative traits. However, the current approaches are mainly focused on estimating the main effects of QTL, and, therefore, a substantial statistical improvement in GWAS is required to detect associated QTL considering their interactions with other QTL as well. This study aimed to compare the support vector regression (SVR) algorithm as a common machine learning method to fixed and random model circulating probability unification (FarmCPU), a common conventional GWAS method in detecting relevant QTL associated with soybean seed quality traits such as protein, oil, and 100-seed weight using 227 soybean genotypes. The results showed a significant negative correlation between soybean seed protein and oil concentrations, with heritability values of 0.69 and 0.67, respectively. In addition, SVR-mediated GWAS was able to identify more relevant QTL underlying the target traits than the FarmCPU method. Our findings demonstrate the potential use of machine learning algorithms in GWAS to detect durable QTL associated with soybean seed quality traits suitable for genomic-based breeding approaches. This study provides new insights into improving the accuracy and efficiency of GWAS and highlights the significance of using advanced computational methods in crop breeding research.

Functional characterization of Cinnamate 4-hydroxylase gene family in soybean (Glycine max).

Cinnamate 4-hydroxylase (C4H) is the first key cytochrome P450 monooxygenase (P450) enzyme in the phenylpropanoid pathway. It belongs to the CYP73 family of P450 superfamily, and catalyzes the conversion of trans-cinnamic acid to p-coumaric acid. Since p-coumaric acid serves as the precursor for the synthesis of a wide variety of metabolites involved in plant development and stress resistance, alteration in the expression of soybean C4H genes is expected to affect the downstream metabolite levels, and its ability to respond to stress. In this study, we identified four C4H genes in the soybean genome that are distributed into both class I and class II CYP73 family. GmC4H2, GmC4H14 and GmC4H20 displayed tissue- and developmental stage-specific gene expression patterns with their transcript accumulation at the highest level in root tissues. GmC4H10 appears to be a pseudogene as its transcript was not detected in any soybean tissues. Furthermore, protein homology modelling revealed substrate docking only for GmC4H2, GmC4H14 and GmC4H20. To demonstrate the function of GmC4Hs, we modified a cloning vector for the heterologous expression of P450s in yeast, and used it for microsomal protein production and enzyme assay. Our results confirmed that GmC4H2, GmC4H14 and GmC4H20 contain the ability to hydroxylate trans-cinnamic acid with varying efficiencies.

Genomic selection for agronomic traits in a winter wheat breeding program.

KEY MESSAGE: rAMP-seq based genomic selection for agronomic traits has been shown to be a useful tool for winter wheat breeding programs by increasing the rate of genetic gain. Genomic selection (GS) is an effective strategy to employ in a breeding program that focuses on optimizing quantitative traits, which results in the ability for breeders to select the best genotypes. GS was incorporated into a breeding program to determine the potential for implementation on an annual basis, with emphasis on selecting optimal parents and decreasing the time and costs associated with phenotyping large numbers of genotypes. The design options for applying repeat amplification sequencing (rAMP-seq) in bread wheat were explored, and a low-cost single primer pair strategy was implemented. A total of 1870 winter wheat genotypes were phenotyped and genotyped using rAMP-seq. The optimization of training to testing population size showed that the 70:30 ratio provided the most consistent prediction accuracy. Three GS models were tested, rrBLUP, RKHS and feed-forward neural networks using the University of Guelph Winter Wheat Breeding Program (UGWWBP) and Elite-UGWWBP populations. The models performed equally well for both populations and did not differ in prediction accuracy (r) for most agronomic traits, with the exception of yield, where RKHS performed the best with an r = 0.34 and 0.39 for each population, respectively. The ability to operate a breeding program where multiple selection strategies, including GS, are utilized will lead to higher efficiency in the program and ultimately lead to a higher rate of genetic gain.

Optimizing genomic selection in soybean: An important improvement in agricultural genomics.

Fast-paced yield improvement in strategic crops such as soybean is pivotal for achieving sustainable global food security. Precise genomic selection (GS), as one of the most effective genomic tools for recognizing superior genotypes, can accelerate the efficiency of breeding programs through shortening the breeding cycle, resulting in significant increases in annual yield improvement. In this study, we investigated the possible use of haplotype-based GS to increase the prediction accuracy of soybean yield and its component traits through augmenting the models by using sophisticated machine learning algorithms and optimized genetic information. The results demonstrated up to a 7% increase in the prediction accuracy when using haplotype-based GS over the full single nucleotide polymorphisms-based GS methods. In addition, we discover an auspicious haplotype block on chromosome 19 with significant impacts on yield and its components, which can be used for screening climate-resilient soybean genotypes with improved yield in large breeding populations.

Genetic variation for effects of drought stress on yield formation traits among commercial soybean [Glycine max (L.) Merr.] cultivars adapted to Ontario, Canada.

Drought stress significantly limits soybean [Glycine max (L.) Merr.] yields in Ontario, Canada. Many studies of genetic variation for drought tolerance compare commercial lines with exotic, unadapted germplasm. We hypothesized that even current commercial cultivars adapted to Ontario would differ significantly for traits related to drought tolerance. In a greenhouse experiment, we grew fifteen soybean cultivars in field soil amended with sand in 1-m rooting columns, which allowed for simulation of field-like soil water profiles and rooting depths. Two watering treatments were imposed from the first flower until maturity by daily restoration of soil water to either 100% (control), or 50% (drought stress) of the maximum soil water holding capacity. Throughout the experiment, we measured volumetric soil water content at different depths in the soil profile, but found no evidence at any developmental stage that the cultivars differed for their ability to extract soil water from different depths. Drought stress reduced seed yield by 51% on average. Similar to the effects of drought in the field, pod number was the yield component most affected, with effects on seeds per pod and single-seed weight being comparatively minor. There were significant cultivar × treatment interactions for seed yield, pod number, shoot dry matter, and water use. We identified two drought-sensitive (Saska and OAC Drayton) and three drought-tolerant (OAC Lakeview, OAC Champion, and PRO 2715R) cultivars based on their ratios of seed yield under drought stress to seed yield under control conditions (seed yield ratio, SYR). Regression and principal component analyses revealed that drought-tolerant (high-SYR) cultivars were consistently those that maintained relatively high values for water use, biomass accumulation and pod number under drought stress; high water use efficiency under drought stress was also associated with a high SYR. One of the cultivars, OAC Lakeview, displayed a distinct mode of drought tolerance, maintaining a very high fraction of its control pod number under drought stress. This study helps define the physiological basis of soybean cultivar differences in drought tolerance, and provides direction for soybean breeders to select traits that could improve yield under drought stress.

SoyMAGIC: An Unprecedented Platform for Genetic Studies and Breeding Activities in Soybean.

Multi-Parent Advanced Generation Inter-Cross (MAGIC) populations are emerging genetic platforms for high-resolution and fine mapping of quantitative traits, such as agronomic and seed composition traits in soybean (Glycine max L.). We have established an eight-parent MAGIC population, comprising 721 recombinant inbred lines (RILs), through conical inter-mating of eight soybean lines. The parental lines were genetically diverse elite cultivars carrying different agronomic and seed composition characteristics, including amino acids and fatty acids, as well as oil and protein concentrations. This study aimed to introduce soybean MAGIC (SoyMAGIC) population as an unprecedented platform for genotypic and phenotypic investigation of agronomic and seed quality traits in soybean. The RILs were evaluated for important seed composition traits using replicated field trials during 2020 and 2021. To measure the seed composition traits, near-infrared reflectance (NIR) was employed. The RILs were genotyped using genotyping-by-sequencing (GBS) method to decipher the genome and discover single-nucleotide polymorphic (SNP) markers among the RILs. A high-density linkage map was constructed through inclusive composite interval mapping (ICIM). The linkage map was 3,770.75 cM in length and contained 12,007 SNP markers. Chromosomes 11 and 18 were recorded as the shortest and longest linkage groups with 71.01 and 341.15 cM in length, respectively. Observed transgressive segregation of the selected traits and higher recombination frequency across the genome confirmed the capability of MAGIC population in reshuffling the diversity in the soybean genome among the RILs. The assessment of haplotype blocks indicated an uneven distribution of the parents' genomes in RILs, suggesting cryptic influence against or in favor of certain parental genomes. The SoyMAGIC population is a recombined genetic material that will accelerate further genomic studies and the development of soybean cultivars with improved seed quality traits through the development and implementation of reliable molecular-based toolkits.

Identification of quantitative trait loci associated with seed quality traits between Canadian and Ukrainian mega-environments using genome-wide association study.

KEY MESSAGE: Identifying QTL associated with soybean seed quality traits from a diverse GWAS panel cultivated in Canadian and Ukrainian mega-environments may facilitate future cultivar development for foreign markets. Understanding the complex genetic basis of seed quality traits for soybean in the mega-environments (MEs) is critical for developing a marker-assisted selection program that will lead to breeding superior cultivars adapted to specific regions. This study aimed to analyze the accumulation of 14 soybean seed quality traits in Canadian ME and two seed quality traits in Ukrainian ME and identify associated ME specific quantitative trait loci (QTLSP) and ME universal QTL (QTLU) for protein and oil using a genome-wide association study (GWAS) panel consisting of 184 soybean genotypes. The panel was planted in three locations in Canada and two locations in Ukraine in 2018 and 2019. Genotype plus genotype-by-environment biplot analysis was conducted to assess the accumulation of individual seed compounds across different locations. The protein accumulation was high in the Canadian ME and low in the Ukrainian ME, whereas the oil concentration showed the opposite trends between the two MEs. No QTLU were identified across the MEs for protein and oil concentrations. In contrast, nine Canadian QTLSP for protein were identified on various chromosomes, which were co-located with QTL controlling other traits identified in the Canadian ME. The lack of common QTLU for protein and oil suggests that it may be necessary to use QTLSP associated with these traits separately for the Canadian and Ukrainian ME. Additional Ukrainian data for seed compounds other than oil and protein are required to identify novel QTLSP and QTLU for such traits for the individual or combined Canadian and Ukrainian MEs.

Machine-Learning-Based Genome-Wide Association Studies for Uncovering QTL Underlying Soybean Yield and Its Components.

A genome-wide association study (GWAS) is currently one of the most recommended approaches for discovering marker-trait associations (MTAs) for complex traits in plant species. Insufficient statistical power is a limiting factor, especially in narrow genetic basis species, that conventional GWAS methods are suffering from. Using sophisticated mathematical methods such as machine learning (ML) algorithms may address this issue and advance the implication of this valuable genetic method in applied plant-breeding programs. In this study, we evaluated the potential use of two ML algorithms, support-vector machine (SVR) and random forest (RF), in a GWAS and compared them with two conventional methods of mixed linear models (MLM) and fixed and random model circulating probability unification (FarmCPU), for identifying MTAs for soybean-yield components. In this study, important soybean-yield component traits, including the number of reproductive nodes (RNP), non-reproductive nodes (NRNP), total nodes (NP), and total pods (PP) per plant along with yield and maturity, were assessed using a panel of 227 soybean genotypes evaluated at two locations over two years (four environments). Using the SVR-mediated GWAS method, we were able to discover MTAs colocalized with previously reported quantitative trait loci (QTL) with potential causal effects on the target traits, supported by the functional annotation of candidate gene analyses. This study demonstrated the potential benefit of using sophisticated mathematical approaches, such as SVR, in a GWAS to complement conventional GWAS methods for identifying MTAs that can improve the efficiency of genomic-based soybean-breeding programs.

The SoyaGen Project: Putting Genomics to Work for Soybean Breeders.

The SoyaGen project was a collaborative endeavor involving Canadian soybean researchers and breeders from academia and the private sector as well as international collaborators. Its aims were to develop genomics-derived solutions to real-world challenges faced by breeders. Based on the needs expressed by the stakeholders, the research efforts were focused on maximizing realized yield through optimization of maturity and improved disease resistance. The main deliverables related to molecular breeding in soybean will be reviewed here. These include: (1) SNP datasets capturing the genetic diversity within cultivated soybean (both within a worldwide collection of > 1,000 soybean accessions and a subset of 102 short-season accessions (MG0 and earlier) directly relevant to this group); (2) SNP markers for selecting favorable alleles at key maturity genes as well as loci associated with increased resistance to key pathogens and pests (Phytophthora sojae, Heterodera glycines, Sclerotinia sclerotiorum); (3) diagnostic tools to facilitate the identification and mapping of specific pathotypes of P. sojae; and (4) a genomic prediction approach to identify the most promising combinations of parents. As a result of this fruitful collaboration, breeders have gained new tools and approaches to implement molecular, genomics-informed breeding strategies. We believe these tools and approaches are broadly applicable to soybean breeding efforts around the world.

Comprehensive Analysis of Cytochrome P450 Monooxygenases Reveals Insight Into Their Role in Partial Resistance Against Phytophthora sojae in Soybean.

Cytochrome P450 monooxygenases (P450) participate in the catalytic conversion of biological compounds in a plethora of metabolic pathways, such as the biosynthesis of alkaloids, terpenoids, phenylpropanoids, and hormones in plants. Plants utilize these metabolites for growth and defense against biotic and abiotic stress. In this study, we identified 346 P450 (GmP450) enzymes encoded by 317 genes in soybean where 26 GmP450 genes produced splice variants. The genome-wide comparison of both A-type and non-A-type GmP450s for their motifs composition, gene structure, tissue-specific expression, and their chromosomal distribution were determined. Even though conserved P450 signature motifs were found in all GmP450 families, larger variation within a specific motif was observed in the non-A-type GmP450s as compared with the A-type. Here, we report that the length of variable region between two conserved motifs is exact in the members of the same family in majority of the A-type GmP450. Analyses of the transcriptomic datasets from soybean-Phytophthora sojae interaction studies, quantitative trait loci (QTL) associated with P. sojae resistance, and co-expression analysis identified some GmP450s that may be, in part, play an important role in partial resistance against P. sojae. The findings of our CYPome study provides novel insights into the functions of GmP450s and their involvements in metabolic pathways in soybean. Further experiments will elucidate their roles in general and legume-specific function.

Genome-Wide Association Study of Soybean Germplasm Derived From Canadian × Chinese Crosses to Mine for Novel Alleles to Improve Seed Yield and Seed Quality Traits.

Genome-wide association study (GWAS) has emerged in the past decade as a viable tool for identifying beneficial alleles from a genomic diversity panel. In an ongoing effort to improve soybean [Glycine max (L.) Merr.], which is the third largest field crop in Canada, a GWAS was conducted to identify novel alleles underlying seed yield and seed quality and agronomic traits. The genomic panel consisted of 200 genotypes including lines derived from several generations of bi-parental crosses between modern Canadian × Chinese cultivars (CD-CH). The genomic diversity panel was field evaluated at two field locations in Ontario in 2019 and 2020. Genotyping-by-sequencing (GBS) was conducted and yielded almost 32 K high-quality SNPs. GWAS was conducted using Fixed and random model Circulating Probability Unification (FarmCPU) model on the following traits: seed yield, seed protein concentration, seed oil concentration, plant height, 100 seed weight, days to maturity, and lodging score that allowed to identify five QTL regions controlling seed yield and seed oil and protein content. A candidate gene search identified a putative gene for each of the three traits. The results of this GWAS study provide insight into potentially valuable genetic resources residing in Chinese modern cultivars that breeders may use to further improve soybean seed yield and seed quality traits.

Genetic analysis of sucrose concentration in soybean seeds using a historical soybean genomic panel.

KEY MESSAGE: Significant QTL for sucrose concentration have been identified using a historical soybean genomic panel, which could aid in the development of food-grade soybean cultivars. Soybean (Glycine max (L.) Merr) is a crop of global importance for both human and animal consumption, which was domesticated in China more than 6000 years ago. A concern about losing genetic diversity as a result of decades of breeding has been expressed by soybean researchers. In order to develop new cultivars, it is critical for breeders to understand the genetic variability present for traits of interest in their program germplasm. Sucrose concentration is becoming an increasingly important trait for the production of soy-food products. The objective of this study was to use a genome-wide association study (GWAS) to identify putative QTL for sucrose concentration in soybean seed. A GWAS panel consisting of 266 historic and current soybean accessions was genotyped with 76 k genotype-by-sequencing (GBS) SNP data and phenotyped in four field locations in Ontario (Canada) from 2015 to 2017. Seven putative QTL were identified on chromosomes 1, 6, 8, 9, 10, 13 and 14. A key gene related to sucrose synthase (Glyma.06g182700) was found to be associated with the QTL located on chromosome 6. This information will facilitate efforts to increase the available genetic variability for sucrose concentration in soybean breeding programs and develop new and improved high-sucrose soybean cultivars suitable for the soy-food industry.

Genome-Wide Association Studies of Soybean Yield-Related Hyperspectral Reflectance Bands Using Machine Learning-Mediated Data Integration Methods.

In conjunction with big data analysis methods, plant omics technologies have provided scientists with cost-effective and promising tools for discovering genetic architectures of complex agronomic traits using large breeding populations. In recent years, there has been significant progress in plant phenomics and genomics approaches for generating reliable large datasets. However, selecting an appropriate data integration and analysis method to improve the efficiency of phenome-phenome and phenome-genome association studies is still a bottleneck. This study proposes a hyperspectral wide association study (HypWAS) approach as a phenome-phenome association analysis through a hierarchical data integration strategy to estimate the prediction power of hyperspectral reflectance bands in predicting soybean seed yield. Using HypWAS, five important hyperspectral reflectance bands in visible, red-edge, and near-infrared regions were identified significantly associated with seed yield. The phenome-genome association analysis of each tested hyperspectral reflectance band was performed using two conventional genome-wide association studies (GWAS) methods and a machine learning mediated GWAS based on the support vector regression (SVR) method. Using SVR-mediated GWAS, more relevant QTL with the physiological background of the tested hyperspectral reflectance bands were detected, supported by the functional annotation of candidate gene analyses. The results of this study have indicated the advantages of using hierarchical data integration strategy and advanced mathematical methods coupled with phenome-phenome and phenome-genome association analyses for a better understanding of the biology and genetic backgrounds of hyperspectral reflectance bands affecting soybean yield formation. The identified yield-related hyperspectral reflectance bands using HypWAS can be used as indirect selection criteria for selecting superior genotypes with improved yield genetic gains in large breeding populations.