Overexpression of BnaA10.WRKY75 Decreases Cadmium and Salt Tolerance via Increasing ROS Accumulation in Arabidopsis and Brassica napus L.

Soil is indispensable for agricultural production but has been seriously polluted by cadmium and salt in recent years. Many crops are suffering from this, including rapeseed, the third largest global oilseed crop. However, genes simultaneously related to both cadmium and salt stress have not been extensively reported yet. In this study, BnaA10.WRKY75 was screened from previous RNA-seq data related to cadmium and salt stress and further analyses including sequence comparison, GUS staining, transformation and qRT-PCR were conducted to confirm its function. GUS staining and qRT-PCR results indicated BnaA10.WRKY75 was induced by CdCl2 and NaCl treatment. Sequence analysis suggested BnaA10.WRKY75 belongs to Group IIc of the WRKY gene family and transient expression assay showed it was a nuclear localized transcription factor. BnaA10.WRKY75-overexpressing Arabidopsis and rapeseed plants accumulated more H2O2 and O2- and were more sensitive to CdCl2 and NaCl treatment compared with untransformed plants, which may be caused by the downregulation of BnaC03.CAT2. Our study reported that BnaA10.WRKY75 increases sensitivity to cadmium and salt stress by disrupting the balance of reactive oxygen species both in Arabidopsis and rapeseed. The results support the further understanding of the mechanisms underlying cadmium and salt tolerance and provide BnaA10.WRKY75 as a valuable gene for rapeseed abiotic stress breeding.

Enhancing climate change resilience in agricultural crops.

Climate change threatens global food and nutritional security through negative effects on crop growth and agricultural productivity. Many countries have adopted ambitious climate change mitigation and adaptation targets that will exacerbate the problem, as they require significant changes in current agri-food systems. In this review, we provide a roadmap for improved crop production that encompasses the effective transfer of current knowledge into plant breeding and crop management strategies that will underpin sustainable agriculture intensification and climate resilience. We identify the main problem areas and highlight outstanding questions and potential solutions that can be applied to mitigate the impacts of climate change on crop growth and productivity. Although translation of scientific advances into crop production lags far behind current scientific knowledge and technology, we consider that a holistic approach, combining disciplines in collaborative efforts, can drive better connections between research, policy, and the needs of society.

In vivo characterisation of field pea stem wall thickness using optical coherence tomography.

BACKGROUND: Modern field pea breeding faces a significant challenge in selecting lines with strong stems that resist lodging. Traditional methods of assessing stem strength involve destructive mechanical tests on mature stems after natural senescence, such as measuring stem flexion, stem buckling or the thickness of dry stems when compressed, but these measurements may not correspond to the strength of stems in the living plant. Optical coherence tomography (OCT) can be used as a noncontact and nondestructive method to measure stem wall thickness in living plants by acquiring two- or three-dimensional images of living plant tissue. RESULTS: In this proof-of-principle study, we demonstrated in vivo characterisation of stem wall thickness using OCT, with the measurement corrected for the refractive index of the stem tissue. This in vivo characterisation was achieved through real-time imaging of stems, with an acquisition rate of 13 milliseconds per two-dimensional, cross-sectional OCT image. We also acquired OCT images of excised stems and compared the accuracy of in vivo OCT measurements of stem wall thickness with ex vivo results for 10 plants each of two field pea cultivars, Dunwa and Kaspa. In vivo OCT measurements of stem wall thickness have an average percent error of - 3.1% when compared with ex vivo measurements. Additionally, we performed in vivo measurements of both stem wall thickness and stem width at various internode positions on the two cultivars. The results revealed that Dunwa had a uniform stem wall thickness across different internode positions, while Kaspa had a significantly negative slope of [Formula: see text]0.0198 mm/node. Both cultivars exhibited an increase in stem width along the internode positions; however, Dunwa had a rate of increase of 0.1844 mm/node, which is three times higher than that of Kaspa. CONCLUSIONS: Our study has demonstrated the efficacy of OCT for accurate measurement of the stem wall thickness of live field pea. Moreover, OCT shows that the trends of stem wall thickness and stem width along the internode positions are different for the two cultivars, Dunwa and Kaspa, potentially hinting at differences in their stem strength. This rapid, in vivo imaging method provides a useful tool for characterising physical traits critical in breeding cultivars that are resistant to lodging.

Genetic and molecular analysis of stem rot (Sclerotinia sclerotiorum) resistance in Brassica napus (canola type).

Identifying the molecular and genetic basis of resistance to Sclerotinia stem rot (Sclerotinia sclerotiorum) is critical for developing long-term and cost-effective management of this disease in rapeseed/canola (Brassica napus). Current cultural or chemical management options provide, at best, only partial and/or sporadic control. Towards this, a B. napus breeding population (Mystic x Rainbow), including the parents, F1, F2, BC1P1 and BC1P2, was utilized in a field study to determine the inheritance pattern of Sclerotinia stem rot resistance (based on stem lesion length, SLL). Broad sense heritability was 0.58 for SLL and 0.44 for days to flowering (DTF). There was a significant negative correlation between SLL and stem diameter (SD) (r = -0.39) and between SLL and DTF (r = -0.28), suggesting co-selection of SD and DTF traits, along with SLL, should assist in improving overall resistance. Non-additive genetic variance was evident for SLL, DTF, and SD. In a genome wide association study (GWAS), a significant quantitative trait locus (QTL) was identified for SLL. Several putative candidate marker trait associations (MTA) were located within this QTL region. Overall, this study has provided valuable new understanding of inheritance of resistance to S. sclerotiorum, and has identified QTL, MTAs and transgressive segregants with high-level resistances. Together, these will foster more rapid selection for multiple traits associated with Sclerotinia stem rot resistance, by enabling breeders to make critical choices towards selecting/developing cultivars with enhanced resistance to this devastating pathogen.

Accuracy of Selection in Early Generations of Field Pea Breeding Increases by Exploiting the Information Contained in Correlated Traits.

Accuracy of predicted breeding values (PBV) for low heritability traits may be increased in early generations by exploiting the information available in correlated traits. We compared the accuracy of PBV for 10 correlated traits with low to medium narrow-sense heritability (h2) in a genetically diverse field pea (Pisum sativum L.) population after univariate or multivariate linear mixed model (MLMM) analysis with pedigree information. In the contra-season, we crossed and selfed S1 parent plants, and in the main season we evaluated spaced plants of S0 cross progeny and S2+ (S2 or higher) self progeny of parent plants for the 10 traits. Stem strength traits included stem buckling (SB) (h2 = 0.05), compressed stem thickness (CST) (h2 = 0.12), internode length (IL) (h2 = 0.61) and angle of the main stem above horizontal at first flower (EAngle) (h2 = 0.46). Significant genetic correlations of the additive effects occurred between SB and CST (0.61), IL and EAngle (-0.90) and IL and CST (-0.36). The average accuracy of PBVs in S0 progeny increased from 0.799 to 0.841 and in S2+ progeny increased from 0.835 to 0.875 in univariate vs MLMM, respectively. An optimized mating design was constructed with optimal contribution selection based on an index of PBV for the 10 traits, and predicted genetic gain in the next cycle ranged from 1.4% (SB), 5.0% (CST), 10.5% (EAngle) and -10.5% (IL), with low achieved parental coancestry of 0.12. MLMM improved the potential genetic gain in annual cycles of early generation selection in field pea by increasing the accuracy of PBV.

Optimal Contribution Selection Improves the Rate of Genetic Gain in Grain Yield and Yield Stability in Spring Canola in Australia and Canada.

Crop breeding must achieve higher rates of genetic gain in grain yield (GY) and yield stability to meet future food demands in a changing climate. Optimal contributions selection (OCS) based on an index of key economic traits should increase the rate of genetic gain while minimising population inbreeding. Here we apply OCS in a global spring oilseed rape (canola) breeding program during three cycles of S0,1 family selection in 2016, 2018, and 2020, with several field trials per cycle in Australia and Canada. Economic weights in the index promoted high GY, seed oil, protein in meal, and Phoma stem canker (blackleg) disease resistance while maintaining plant height, flowering time, oleic acid, and seed size and decreasing glucosinolate content. After factor analytic modelling of the genotype-by-environment interaction for the additive effects, the linear rate of genetic gain in GY across cycles was 0.059 or 0.087 t ha-1 y-1 (2.9% or 4.3% y-1) based on genotype scores for the first factor (f1) expressed in trait units or average predicted breeding values across environments, respectively. Both GY and yield stability, defined as the root-mean-square deviation from the regression line associated with f1, were predicted to improve in the next cycle with a low achieved mean parental coancestry (0.087). These methods achieved rapid genetic gain in GY and other traits and are predicted to improve yield stability across global spring canola environments.

Quantitative Trait Loci for Heat Stress Tolerance in Brassica rapa L. Are Distributed across the Genome and Occur in Diverse Genetic Groups, Flowering Phenologies and Morphotypes.

Heat stress events during flowering in Brassica crops reduce grain yield and are expected to increase in frequency due to global climate change. We evaluated heat stress tolerance and molecular genetic diversity in a global collection of Brassica rapa accessions, including leafy, rooty and oilseed morphotypes with spring, winter and semi-winter flowering phenology. Tolerance to transient daily heat stress during the early reproductive stage was assessed on 142 lines in a controlled environment. Well-watered plants of each genotype were exposed to the control (25/15 °C day/night temperatures) or heat stress (35/25 °C) treatments for 7 d from the first open flower on the main stem. Bud and leaf temperature depression, leaf conductance and chlorophyll content index were recorded during the temperature treatments. A large genetic variation for heat tolerance and sensitivity was found for above-ground biomass, whole plant seed yield and harvest index and seed yield of five pods on the main stem at maturity. Genetic diversity was assessed on 212 lines with 1602 polymorphic SNP markers with a known location in the B. rapa physical map. Phylogenetic analyses confirmed two major genetic populations: one from East and South Asia and one from Europe. Heat stress-tolerant lines were distributed across diverse geographic origins, morphotypes (leafy, rooty and oilseed) and flowering phenologies (spring, winter and semi-winter types). A genome-wide association analysis of heat stress-related yield traits revealed 57 SNPs distributed across all 10 B. rapa chromosomes, some of which were associated with potential candidate genes for heat stress tolerance.

Quantitative Inheritance of Sclerotinia Stem Rot Resistance in Brassica napus and Relationship to Cotyledon and Leaf Resistances.

Sclerotinia sclerotiorum is a necrotrophic fungus causing devastating stem rot and associated yield losses of canola/rapeseed (Brassica napus) worldwide, including in Australia. Developing host resistance against Sclerotinia stem rot is critical if this disease in canola/rapeseed is to be successfully managed, as cultural or chemical control options provide only partial or sporadic control. Three B. napus breeding populations, C2, C5 and C6, including the parents, F1, F2, BC1P1, and BC2P2, were used in a field study with an objective of exploring the inheritance pattern of disease resistance (based on stem lesion length [SLL]) and the genetic relationships of disease with stem diameter (SD) or days to first flowering (DTF), and to compare these new adult plant stem resistances against S. sclerotiorum with those of seedling (cotyledon and leaf) resistances in earlier studies. Heritability (broad sense) of SLL was 0.57 and 0.73 for population C2 at 3 and 5 weeks postinoculation and 0.21 for population C5 at 5 weeks postinoculation. Additive genetic variance was evident within all 3 populations for DTF but not for SD. Narrow-sense heritability for DTF was 0.48 (C2), 0.42 (C5), and 0.32 (C6). SD, DTF, and SLL were all inherited independently, with no significant genetic covariance between traits in bivariate analysis. Genetic variance for SLL in populations C2 and C5 was entirely nonadditive, and there was significant nonadditive genetic covariance of SLL at 3 and 5 weeks postinoculation. Generation means analysis in population C2 supported the conclusion that complex epistatic interactions controlled SLL. Several C2 and C5 progeny showed high adult plant stem resistance, which may be critical in developing enhanced stem resistance in canola/rapeseed. Although population C6 showed no genetic variation for SLL resistance in this study, it showed significant nonadditive genetic variance at the cotyledon and leaf stages in earlier studies. We conclude that host resistance varies across different plant growth stages, and breeding must be targeted for resistance at each growth stage. In populations C2, C5, and C6, resistance to S. sclerotiorum in stem, leaf, and cotyledon was always controlled by nonadditive effects such as complex epistasis or dominance. Overall, our findings in relation to the quantitative inheritance of Sclerotinia stem rot resistance, together with the new high-level resistances identified, will enable breeders to select/develop genotypes with enhanced resistances to S. sclerotiorum.

A chickpea genetic variation map based on the sequencing of 3,366 genomes.

Zero hunger and good health could be realized by 2030 through effective conservation, characterization and utilization of germplasm resources1. So far, few chickpea (Cicer arietinum) germplasm accessions have been characterized at the genome sequence level2. Here we present a detailed map of variation in 3,171 cultivated and 195 wild accessions to provide publicly available resources for chickpea genomics research and breeding. We constructed a chickpea pan-genome to describe genomic diversity across cultivated chickpea and its wild progenitor accessions. A divergence tree using genes present in around 80% of individuals in one species allowed us to estimate the divergence of Cicer over the last 21 million years. Our analysis found chromosomal segments and genes that show signatures of selection during domestication, migration and improvement. The chromosomal locations of deleterious mutations responsible for limited genetic diversity and decreased fitness were identified in elite germplasm. We identified superior haplotypes for improvement-related traits in landraces that can be introgressed into elite breeding lines through haplotype-based breeding, and found targets for purging deleterious alleles through genomics-assisted breeding and/or gene editing. Finally, we propose three crop breeding strategies based on genomic prediction to enhance crop productivity for 16 traits while avoiding the erosion of genetic diversity through optimal contribution selection (OCS)-based pre-breeding. The predicted performance for 100-seed weight, an important yield-related trait, increased by up to 23% and 12% with OCS- and haplotype-based genomic approaches, respectively.

Multivariate genomic analysis and optimal contributions selection predicts high genetic gains in cooking time, iron, zinc, and grain yield in common beans in East Africa.

Common bean (Phaseolus vulgaris L.) is important in African diets for protein, iron (Fe), and zinc (Zn), but traditional cultivars have long cooking time (CKT), which increases the time, energy, and health costs of cooking. Genomic selection was used to predict genomic estimated breeding values (GEBV) for grain yield (GY), CKT, Fe, and Zn in an African bean panel of 358 genotypes in a two-stage analysis. In Stage 1, best linear unbiased estimates (BLUE) for each trait were obtained from 898 genotypes across 33 field trials in East Africa. In Stage 2, BLUE in a training population of 141 genotypes were used in a multivariate genomic analysis with genome-wide single nucleotide polymorphism data from the African bean panel. Moderate to high genomic heritability was found for GY (0.45 ± 0.10), CKT (0.50 ± 0.15), Fe (0.57 ± 0.12), and Zn (0.61 ± 0.13). There were significant favorable genetic correlations between Fe and Zn (0.91 ± 0.06), GY and Fe (0.66 ± 0.17), GY and Zn (0.44 ± 0.19), CKT and Fe (-0.57 ± 0.21), and CKT and Zn (-0.67 ± 0.20). Optimal contributions selection (OCS), based on economic index of weighted GEBV for each trait, was used to design crossing within four market groups relevant to East Africa. Progeny were predicted by OCS to increase in mean GY by 12.4%, decrease in mean CKT by 9.3%, and increase in mean Fe and Zn content by 6.9 and 4.6%, respectively, with low achieved coancestry of 0.032. Genomic selection with OCS will accelerate breeding of high-yielding, biofortified, and rapid cooking African common bean cultivars.

Fast-forward breeding for a food-secure world.

Crop production systems need to expand their outputs sustainably to feed a burgeoning human population. Advances in genome sequencing technologies combined with efficient trait mapping procedures accelerate the availability of beneficial alleles for breeding and research. Enhanced interoperability between different omics and phenotyping platforms, leveraged by evolving machine learning tools, will help provide mechanistic explanations for complex plant traits. Targeted and rapid assembly of beneficial alleles using optimized breeding strategies and precise genome editing techniques could deliver ideal crops for the future. Realizing desired productivity gains in the field is imperative for securing an adequate future food supply for 10 billion people.

A Trimethylguanosine Synthase1-like (TGS1) homologue is implicated in vernalisation and flowering time control.

KEY MESSAGE: A plant-specific Trimethylguanosine Synthase1-like homologue was identified as a candidate gene for the efl mutation in narrow-leafed lupin, which alters phenology by reducing vernalisation requirement. The vernalisation pathway is a key component of flowering time control in plants from temperate regions but is not well understood in the legume family. Here we examined vernalisation control in the temperate grain legume species, narrow-leafed lupin (Lupinus angustifolius L.), and discovered a candidate gene for an ethylene imine mutation (efl). The efl mutation changes phenology from late to mid-season flowering and additionally causes transformation from obligate to facultative vernalisation requirement. The efl locus was mapped to pseudochromosome NLL-10 in a recombinant inbred line (RIL) mapping population developed by accelerated single seed descent. Candidate genes were identified in the reference genome, and a diverse panel of narrow-leafed lupins was screened to validate mutations specific to accessions with efl. A non-synonymous SNP mutation within an S-adenosyl-L-methionine-dependent methyltransferase protein domain of a Trimethylguanosine Synthase1-like (TGS1) orthologue was identified as the candidate mutation giving rise to efl. This mutation caused substitution of an amino acid within an established motif at a position that is otherwise highly conserved in several plant families and was perfectly correlated with the efl phenotype in F2 and F6 genetic population and a panel of diverse accessions, including the original efl mutant. Expression of the TGS1 homologue did not differ between wild-type and efl genotypes, supporting altered functional activity of the gene product. This is the first time a TGS1 orthologue has been associated with vernalisation response and flowering time control in any plant species.

Status and advances in mining for blackleg (Leptosphaeria maculans) quantitative resistance (QR) in oilseed rape (Brassica napus).

KEY MESSAGE: Quantitative resistance (QR) loci discovered through genetic and genomic analyses are abundant in the Brassica napus genome, providing an opportunity for their utilization in enhancing blackleg resistance. Quantitative resistance (QR) has long been utilized to manage blackleg in Brassica napus (canola, oilseed rape), even before major resistance genes (R-genes) were extensively explored in breeding programmes. In contrast to R-gene-mediated qualitative resistance, QR reduces blackleg symptoms rather than completely eliminating the disease. As a polygenic trait, QR is controlled by numerous genes with modest effects, which exerts less pressure on the pathogen to evolve; hence, its effectiveness is more durable compared to R-gene-mediated resistance. Furthermore, combining QR with major R-genes has been shown to enhance resistance against diseases in important crops, including oilseed rape. For these reasons, there has been a renewed interest among breeders in utilizing QR in crop improvement. However, the mechanisms governing QR are largely unknown, limiting its deployment. Advances in genomics are facilitating the dissection of the genetic and molecular underpinnings of QR, resulting in the discovery of several loci and genes that can be potentially deployed to enhance blackleg resistance. Here, we summarize the efforts undertaken to identify blackleg QR loci in oilseed rape using linkage and association analysis. We update the knowledge on the possible mechanisms governing QR and the advances in searching for the underlying genes. Lastly, we lay out strategies to accelerate the genetic improvement of blackleg QR in oilseed rape using improved phenotyping approaches and genomic prediction tools.

Gaining Acceptance of Novel Plant Breeding Technologies.

Ensuring the sustainability of agriculture under climate change has led to a surge in alternative strategies for crop improvement. Advances in integrated crop breeding, social acceptance, and farm-level adoption are crucial to address future challenges to food security. Societal acceptance can be slow when consumers do not see the need for innovation or immediate benefits. We consider how best to address the issue of social licence and harmonised governance for novel gene technologies in plant breeding. In addition, we highlight optimised breeding strategies that will enable long-term genetic gains to be achieved. Promoted by harmonised global policy change, innovative plant breeding can realise high and sustainable productivity together with enhanced nutritional traits.

In silico simulation of future hybrid performance to evaluate heterotic pool formation in a self-pollinating crop.

Hybrid vigour has the potential to substantially increase the yield of self-pollinating crops such as wheat and rice, but future hybrid performance may depend on the initial strategy to form heterotic pools. We used in silico stochastic simulation of future hybrid performance in a self-pollinating crop to evaluate three strategies of forming heterotic pools in the founder population. The model included either 500, 2000 or 8000 quantitative trait nucleotides (QTN) across 10 chromosomes that contributed to a quantitative trait with population mean 100 and variance 10. The average degree of dominance at each QTN was either 0.2, 0.4 or 0.8 with variance 0.2. Three strategies for splitting the founder population into two heterotic pools were compared: (i) random split; (ii) split based on genetic distance according to principal component analysis of SNP genotypes; and (iii) optimized split based on F1 hybrid performance in a diallel cross among the founders. Future hybrid performance was stochastically simulated over 30 cycles of reciprocal recurrent selection based on true genetic values for additive and dominance effects. The three strategies of forming heterotic pools produced similar future hybrid performance, and superior future hybrids to a control population selected on inbred line performance when the number of quantitative trait nucleotides was ≥2000 and/or the average degree of dominance was ≥0.4.

INDEL variation in the regulatory region of the major flowering time gene LanFTc1 is associated with vernalization response and flowering time in narrow-leafed lupin (Lupinus angustifolius L.).

Narrow-leafed lupin (Lupinus angustifolius L.) cultivation was transformed by 2 dominant vernalization-insensitive, early flowering time loci known as Ku and Julius (Jul), which allowed expansion into shorter season environments. However, reliance on these loci has limited genetic and phenotypic diversity for environmental adaptation in cultivated lupin. We recently predicted that a 1,423-bp deletion in the cis-regulatory region of LanFTc1, a FLOWERING LOCUS T (FT) homologue, derepressed expression of LanFTc1 and was the underlying cause of the Ku phenotype. Here, we surveyed diverse germplasm for LanFTc1 cis-regulatory variation and identified 2 further deletions of 1,208 and 5,162 bp in the 5' regulatory region, which overlap the 1,423-bp deletion. Additionally, we confirmed that no other polymorphisms were perfectly associated with vernalization responsiveness. Phenotyping and gene expression analyses revealed that Jul accessions possessed the 5,162-bp deletion and that the Jul and Ku deletions were equally capable of removing vernalization requirement and up-regulating gene expression. The 1,208-bp deletion was associated with intermediate phenology, vernalization responsiveness, and gene expression and therefore may be useful for expanding agronomic adaptation of lupin. This insertion/deletion series may also help resolve how the vernalization response is mediated at the molecular level in legumes.

Nondestructive Phenomic Tools for the Prediction of Heat and Drought Tolerance at Anthesis in Brassica Species.

Oilseed Brassica species are vulnerable to heat and drought stress, especially in the early reproductive stage. We evaluated plant imaging of whole plant and flower tissue, leaf stomatal conductance, leaf and bud temperature, photochemical reflectance index, quantum yield of photosynthesis, and leaf gas exchange for their suitability to detect tolerance to heat (H) and/or drought (D) stress treatments in 12 Brassica genotypes (G). A replicated factorial experiment was set up with 7 d of stress treatment from the beginning of anthesis with various levels of three factors H, D, and G. Most phenomics tools detected plant stress as indicated by significant main effects of H, D, and H×D. Whole plant volume was highly correlated with fresh weight changes, suggesting that whole plant imaging may be a useful surrogate for fresh weight in future studies. Vcmax, the maximum carboxylation rate of photosynthesis, increased rapidly on day 1 in H and H+D treatments, and there were significant interactions of G×H and G×D. Vcmax of genotypes on day 1 in H and H+D treatments was positively correlated with their harvested seed yield. Vcmax on day 1 and day 3 were clustered with seed yield in H and H+D treatments as shown in the heatmaps of genotypic correlations. TPU, the rate of triose phosphate use, also showed significant positive genotypic correlations with seed yield in H+D treatments. Flower volume showed significant interactions of G×H and G×D on day 7, and flower volume of genotypes on day 7 in H was positively correlated with their harvested seed yield. There were few interactions of G×H or G×D for leaf stomatal conductance, leaf and bud temperature, photochemical reflectance index, and quantum yield of photosynthesis. Vcmax, TPU, and volume of flowers are potential nondestructive phenomic traits for heat or combined heat and drought stress tolerance screening in Brassica germplasm.

A High-Density Genetic Map of an Allohexaploid Brassica Doubled Haploid Population Reveals Quantitative Trait Loci for Pollen Viability and Fertility.

A doubled haploid (DH) mapping population was obtained from microspore culture of an allohexaploid F1 from the cross between two recently-synthesized allohexaploid Brassica lines. We used single nucleotide polymorphism (SNP) genetic variation based on restriction-site associated DNA (RAD) sequencing to construct a high density genetic linkage map of the population. RAD libraries were constructed from the genomic DNA of both parents and 146 DH progenies. A total of 2.87 G reads with an average sequencing depth of 2.59 × were obtained in the parents and of 1.41 × in the progeny. A total of 290,422 SNPs were identified from clustering of RAD reads, from which we developed 7,950 high quality SNP markers that segregated normally (1:1) in the population. The linkage map contained all 27 chromosomes from the parental A, B and C genomes with a total genetic distance of 5725.19 cM and an average of 0.75 cM between adjacent markers. Genetic distance on non-integrated linkage groups was 1534.23 cM, or 21% of total genetic distance. Out of 146 DH progenies, 91 had a complete set of 27 chromosomes as expected of a hexaploid species, and 21 out of 27 chromosomes showed high collinearity between the physical and linkage maps. The loss of chromosome(s) or chromosome segment(s) in the DH population was associated with a reduction in pollen viability. Twenty-five additive QTL were associated with pollen viability and fertility-related traits (seed number, seed yield, pod length, plant height, 1000-seed weight). In addition, 44 intra-genomic and 18 inter-genomic epistatic QTL pairs were detected for 4 phenotypic traits. This provides confidence that the DH population may be selected for improved pollen viability and fertility in a future allohexaploid Brassica species.