Robert W. Holley Center for Agriculture & Health

A-Maize-ing Maize is one of our oldest and most important crops, having been domesticated approximately 9000 years ago in central Mexico. Schnable et al. (p. 1112 ; see the cover) present the results of sequencing the B73 inbred maize line. The findings elucidate how maize became diploid after an ancestral doubling of its chromosomes and reveals transposable element movement and activity and recombination. Vielle-Calzada et al. (p. 1078 ) have sequenced the Palomero Toluqueño ( Palomero ) landrace, a highland popcorn from Mexico, which, when compared to the B73 line, reveals multiple loci impacted by domestication. Swanson-Wagner et al. (p. 1118 ) exploit possession of the genome to analyze expression differences occurring between lines. The identification of single nucleotide polymorphisms and copy number variations among lines was used by Gore et al. (p. 1115 ) to generate a Haplotype map of maize. While chromosomal diversity in maize is high, it is likely that recombination is the major force affecting the levels of heterozygosity in maize. The availability of the maize genome will help to guide future agricultural and biofuel applications (see the Perspective by Feuillet and Eversole ).

This paper reports the genome sequence of domesticated tomato, a major crop plant, and a draft sequence for its closest wild relative; comparative genomics reveal very little divergence between the two genomes but some important differences with the potato genome, another important food crop in the genus Solanum. Tomato (Solanum lycopersicum) is a major crop plant and a model system for fruit development. Solanum is one of the largest angiosperm genera1 and includes annual and perennial plants from diverse habitats. Here we present a high-quality genome sequence of domesticated tomato, a draft sequence of its closest wild relative, Solanum pimpinellifolium2, and compare them to each other and to the potato genome (Solanum tuberosum). The two tomato genomes show only 0.6% nucleotide divergence and signs of recent admixture, but show more than 8% divergence from potato, with nine large and several smaller inversions. In contrast to Arabidopsis, but similar to soybean, tomato and potato small RNAs map predominantly to gene-rich chromosomal regions, including gene promoters. The Solanum lineage has experienced two consecutive genome triplications: one that is ancient and shared with rosids, and a more recent one. These triplications set the stage for the neofunctionalization of genes controlling fruit characteristics, such as colour and fleshiness.

Sorghum, an African grass related to sugar cane and maize, is grown for food, feed, fibre and fuel. We present an initial analysis of the ∼730-megabase Sorghum bicolor (L.) Moench genome, placing ∼98% of genes in their chromosomal context using whole-genome shotgun sequence validated by genetic, physical and syntenic information. Genetic recombination is largely confined to about one-third of the sorghum genome with gene order and density similar to those of rice. Retrotransposon accumulation in recombinationally recalcitrant heterochromatin explains the ∼75% larger genome size of sorghum compared with rice. Although gene and repetitive DNA distributions have been preserved since palaeopolyploidization ∼70 million years ago, most duplicated gene sets lost one member before the sorghum–rice divergence. Concerted evolution makes one duplicated chromosomal segment appear to be only a few million years old. About 24% of genes are grass-specific and 7% are sorghum-specific. Recent gene and microRNA duplications may contribute to sorghum’s drought tolerance. The Sorghum bicolor genome sequence is published this week. Sorghum is a cereal grown widely as food, animal feed, fibre and fuel. Tolerant to hot, dry conditions, it is a staple for large populations in the West African Sahel region. Comparisons of the genome with those of maize and rice shed light on the evolution of grasses and of C4 photosynthesis, which is particularly efficient at assimilating carbon at high temperatures. In addition, protein coding genes and miRNAs that could contribute to sorghum's drought tolerance may also be found. Sorghum yield improvement has lagged behind that of other crops and the availability of the genome sequence could provide a vital boost to work on its improvement. Sorghum is an African grass that is grown for food, animal feed and fuel. The current paper presents an initial analysis of the ∼730 megabase genome of Sorghum bicolor. Genome analysis and its comparison with maize and rice shed light on grass genome evolution and also provide insights into the evolution of C4 photosynthesis, as well as protein coding genes and miRNAs that might contribute to sorghum's drought tolerance.

ABSTRACT Despite important strides in marker technologies, the use of marker‐assisted selection has stagnated for the improvement of quantitative traits. Biparental mating designs for the detection of loci affecting these traits (quantitative trait loci [QTL]) impede their application, and the statistical methods used are ill‐suited to the traits' polygenic nature. Genomic selection (GS) has been proposed to address these deficiencies. Genomic selection predicts the breeding values of lines in a population by analyzing their phenotypes and high‐density marker scores. A key to the success of GS is that it incorporates all marker information in the prediction model, thereby avoiding biased marker effect estimates and capturing more of the variation due to small‐effect QTL. In simulations, the correlation between true breeding value and the genomic estimated breeding value has reached levels of 0.85 even for polygenic low heritability traits. This level of accuracy is sufficient to consider selecting for agronomic performance using marker information alone. Such selection would substantially accelerate the breeding cycle, enhancing gains per unit time. It would dramatically change the role of phenotyping, which would then serve to update prediction models and no longer to select lines. While research to date shows the exceptional promise of GS, work remains to be done to validate it empirically and to incorporate it into breeding schemes.

Abstract Background Sequencing technology and assembly algorithms have matured to the point that high-quality de novo assembly is possible for large, repetitive genomes. Current assemblies traverse transposable elements (TEs) and provide an opportunity for comprehensive annotation of TEs. Numerous methods exist for annotation of each class of TEs, but their relative performances have not been systematically compared. Moreover, a comprehensive pipeline is needed to produce a non-redundant library of TEs for species lacking this resource to generate whole-genome TE annotations. Results We benchmark existing programs based on a carefully curated library of rice TEs. We evaluate the performance of methods annotating long terminal repeat (LTR) retrotransposons, terminal inverted repeat (TIR) transposons, short TIR transposons known as miniature inverted transposable elements (MITEs), and Helitrons. Performance metrics include sensitivity, specificity, accuracy, precision, FDR, and F 1 . Using the most robust programs, we create a comprehensive pipeline called Extensive de-novo TE Annotator (EDTA) that produces a filtered non-redundant TE library for annotation of structurally intact and fragmented elements. EDTA also deconvolutes nested TE insertions frequently found in highly repetitive genomic regions. Using other model species with curated TE libraries (maize and Drosophila), EDTA is shown to be robust across both plant and animal species. Conclusions The benchmarking results and pipeline developed here will greatly facilitate TE annotation in eukaryotic genomes. These annotations will promote a much more in-depth understanding of the diversity and evolution of TEs at both intra- and inter-species levels. EDTA is open-source and freely available: https://github.com/oushujun/EDTA .

An improved reference genome for maize, using single-molecule sequencing and high-resolution optical mapping, enables characterization of structural variation and repetitive regions, and identifies lineage expansions of transposable elements that are unique to maize. The maize genome was initially reported in 2009 but with some accuracy limitations. Doreen Ware and colleagues report a new reference genome for maize using single-molecule sequencing and high-resolution optical mapping. The technique shows improvements in the gene space including resolution of gaps and misassemblies and correction of order and orientation of genes. The authors characterize structural variation and repetitive regions, and identify transposable element lineage expansions unique to maize. Complete and accurate reference genomes and annotations provide fundamental tools for characterization of genetic and functional variation1. These resources facilitate the determination of biological processes and support translation of research findings into improved and sustainable agricultural technologies. Many reference genomes for crop plants have been generated over the past decade, but these genomes are often fragmented and missing complex repeat regions2. Here we report the assembly and annotation of a reference genome of maize, a genetic and agricultural model species, using single-molecule real-time sequencing and high-resolution optical mapping. Relative to the previous reference genome3, our assembly features a 52-fold increase in contig length and notable improvements in the assembly of intergenic spaces and centromeres. Characterization of the repetitive portion of the genome revealed more than 130,000 intact transposable elements, allowing us to identify transposable element lineage expansions that are unique to maize. Gene annotations were updated using 111,000 full-length transcripts obtained by single-molecule real-time sequencing4. In addition, comparative optical mapping of two other inbred maize lines revealed a prevalence of deletions in regions of low gene density and maize lineage-specific genes.

We intuitively believe that the dramatic drop in the cost of DNA marker information we have experienced should have immediate benefits in accelerating the delivery of crop varieties with improved yield, quality and biotic and abiotic stress tolerance. But these traits are complex and affected by many genes, each with small effect. Traditional marker-assisted selection has been ineffective for such traits. The introduction of genomic selection (GS), however, has shifted that paradigm. Rather than seeking to identify individual loci significantly associated with a trait, GS uses all marker data as predictors of performance and consequently delivers more accurate predictions. Selection can be based on GS predictions, potentially leading to more rapid and lower cost gains from breeding. The objectives of this article are to review essential aspects of GS and summarize the important take-home messages from recent theoretical, simulation and empirical studies. We then look forward and consider research needs surrounding methodological questions and the implications of GS for long-term selection.

Aluminum (Al) toxicity in acid soils is a significant limitation to crop production worldwide, as approximately 50% of the world's potentially arable soil is acidic. Because acid soils are such an important constraint to agriculture, understanding the mechanisms and genes conferring resistance to Al toxicity has been a focus of intense research interest in the decade since the last article on crop acid soil tolerance was published in this journal. An impressive amount of progress has been made during that time that has greatly increased our understanding of the diversity of Al resistance genes and mechanisms, how resistance gene expression is regulated and triggered by Al and Al-induced signals, and how the proteins encoded by these genes function and are regulated. This review examines the state of our understanding of the physiological, genetic, and molecular bases for crop Al tolerance, looking at the novel Al resistance genes and mechanisms that have been identified over the past ten years. Additionally, it examines how the integration of molecular and genetic analyses of crop Al resistance is starting to be exploited for the improvement of crop plants grown on acid soils via both molecular-assisted breeding and biotechnology approaches.

Doil Choi and colleagues report the genome sequence of the hot pepper, Capsicum annuum, as well as the resequencing of two cultivated peppers and a wild species, Capsicum chinense. Comparative genomic analysis across Solanaceae provides insights into genome expansion, pungency, ripening and disease resistance in hot peppers. Hot pepper (Capsicum annuum), one of the oldest domesticated crops in the Americas, is the most widely grown spice crop in the world. We report whole-genome sequencing and assembly of the hot pepper (Mexican landrace of Capsicum annuum cv. CM334) at 186.6× coverage. We also report resequencing of two cultivated peppers and de novo sequencing of the wild species Capsicum chinense. The genome size of the hot pepper was approximately fourfold larger than that of its close relative tomato, and the genome showed an accumulation of Gypsy and Caulimoviridae family elements. Integrative genomic and transcriptomic analyses suggested that change in gene expression and neofunctionalization of capsaicin synthase have shaped capsaicinoid biosynthesis. We found differential molecular patterns of ripening regulators and ethylene synthesis in hot pepper and tomato. The reference genome will serve as a platform for improving the nutritional and medicinal values of Capsicum species.

Tomato ripening is a highly coordinated developmental process that coincides with seed maturation. Regulated expression of thousands of genes controls fruit softening as well as accumulation of pigments, sugars, acids, and volatile compounds that increase attraction to animals. A combination of molecular tools and ripening-affected mutants has permitted researchers to establish a framework for the control of ripening. Tomato is a climacteric fruit, with an absolute requirement for the phytohormone ethylene to ripen. This dependence upon ethylene has established tomato fruit ripening as a model system for study of regulation of its synthesis and perception. In addition, several important ripening mutants, including rin, nor, and Cnr, have provided novel insights into the control of ripening processes. Here, we describe how ethylene and the transcription factors associated with the ripening process fit together into a network controlling ripening.

ABSTRACT Minerals and vitamins in food staples eaten widely by the poor may be increased either through conventional plant breeding or through use of transgenic techniques, a process known as biofortification HarvestPlus seeks to develop and distribute cultivars of food staples (rice [ Oryza sativa L.], wheat [ Triticum aestivum L.], maize [ Zea mays L.], cassava [ Manihot esculenta Crantz], pearl millet [ Pennisetum americanum Leeke], beans [ Phaseolus vulgaris L.], sweet potato [ Ipomoea batatas L.]) that are high in Fe, Zn, and provitamin A through an interdisciplinary global alliance of scientific institutions and implementing agencies in developing and developed countries. Biofortified crops offer a rural‐based intervention that, by design, initially reaches these more remote populations, which comprise a majority of the undernourished in many countries, and then penetrates to urban populations as production surpluses are marketed. Thus, biofortification complements fortification and supplementation programs, which work best in centralized urban areas and then reach into rural areas with good infrastructure. Initial investments in agricultural research at a central location can generate high recurrent benefits at low cost as adapted biofortified cultivars become widely available in countries across time at low recurrent costs. Overall, three things must happen for biofortification to be successful. First, the breeding must be successful—high nutrient density must be combined with high yields and high profitability. Second, efficacy must be demonstrated—the micronutrient status of human subjects must be shown to improve when consuming the biofortified cultivars as normally eaten. Third, the biofortified crops must be adopted by farmers and consumed by those suffering from micronutrient malnutrition in significant numbers.

Zhangjun Fei and colleagues report the draft genome of a Chinese elite watermelon inbred line 97103 and resequencing of 20 diverse accessions that represent the three subspecies of Citrullus lunatus. Comparative genome-wide analyses identify the extent of genetic diversity and population structure of watermelon germplasm. Watermelon, Citrullus lanatus, is an important cucurbit crop grown throughout the world. Here we report a high-quality draft genome sequence of the east Asia watermelon cultivar 97103 (2n = 2× = 22) containing 23,440 predicted protein-coding genes. Comparative genomics analysis provided an evolutionary scenario for the origin of the 11 watermelon chromosomes derived from a 7-chromosome paleohexaploid eudicot ancestor. Resequencing of 20 watermelon accessions representing three different C. lanatus subspecies produced numerous haplotypes and identified the extent of genetic diversity and population structure of watermelon germplasm. Genomic regions that were preferentially selected during domestication were identified. Many disease-resistance genes were also found to be lost during domestication. In addition, integrative genomic and transcriptomic analyses yielded important insights into aspects of phloem-based vascular signaling in common between watermelon and cucumber and identified genes crucial to valuable fruit-quality traits, including sugar accumulation and citrulline metabolism.

The grape is one of the earliest domesticated fruit crops and, since antiquity, it has been widely cultivated and prized for its fruit and wine. Here, we characterize genome-wide patterns of genetic variation in over 1,000 samples of the domesticated grape, Vitis vinifera subsp. vinifera, and its wild relative, V. vinifera subsp. sylvestris from the US Department of Agriculture grape germplasm collection. We find support for a Near East origin of vinifera and present evidence of introgression from local sylvestris as the grape moved into Europe. High levels of genetic diversity and rapid linkage disequilibrium (LD) decay have been maintained in vinifera, which is consistent with a weak domestication bottleneck followed by thousands of years of widespread vegetative propagation. The considerable genetic diversity within vinifera, however, is contained within a complex network of close pedigree relationships that has been generated by crosses among elite cultivars. We show that first-degree relationships are rare between wine and table grapes and among grapes from geographically distant regions. Our results suggest that although substantial genetic diversity has been maintained in the grape subsequent to domestication, there has been a limited exploration of this diversity. We propose that the adoption of vegetative propagation was a double-edged sword: Although it provided a benefit by ensuring true breeding cultivars, it also discouraged the generation of unique cultivars through crosses. The grape currently faces severe pathogen pressures, and the long-term sustainability of the grape and wine industries will rely on the exploitation of the grape's tremendous natural genetic diversity.

We report de novo genome assemblies, transcriptomes, annotations, and methylomes for the 26 inbreds that serve as the founders for the maize nested association mapping population. The number of pan-genes in these diverse genomes exceeds 103,000, with approximately a third found across all genotypes. The results demonstrate that the ancient tetraploid character of maize continues to degrade by fractionation to the present day. Excellent contiguity over repeat arrays and complete annotation of centromeres revealed additional variation in major cytological landmarks. We show that combining structural variation with single-nucleotide polymorphisms can improve the power of quantitative mapping studies. We also document variation at the level of DNA methylation and demonstrate that unmethylated regions are enriched for cis-regulatory elements that contribute to phenotypic variation.

Ensembl (https://www.ensembl.org) is a freely available genomic resource that has produced high-quality annotations, tools, and services for vertebrates and model organisms for more than two decades. In recent years, there has been a dramatic shift in the genomic landscape, with a large increase in the number and phylogenetic breadth of high-quality reference genomes, alongside major advances in the pan-genome representations of higher species. In order to support these efforts and accelerate downstream research, Ensembl continues to focus on scaling for the rapid annotation of new genome assemblies, developing new methods for comparative analysis, and expanding the depth and quality of our genome annotations. This year we have continued our expansion to support global biodiversity research, doubling the number of annotated genomes we support on our Rapid Release site to over 1700, driven by our close collaboration with biodiversity projects such as Darwin Tree of Life. We have also strengthened support for key agricultural species, including the first regulatory builds for farmed animals, and have updated key tools and resources that support the global scientific community, notably the Ensembl Variant Effect Predictor. Ensembl data, software, and tools are freely available.

ABSTRACT Simulation and empirical studies of genomic selection (GS) show accuracies sufficient to generate rapid genetic gains. However, with the increased popularity of GS approaches, numerous models have been proposed and no comparative analysis is available to identify the most promising ones. Using eight wheat ( Triticum aestivum L.), barley ( Hordeum vulgare L.), Arabidopsis thaliana (L.) Heynh., and maize ( Zea mays L.) datasets, the predictive ability of currently available GS models along with several machine learning methods was evaluated by comparing accuracies, the genomic estimated breeding values (GEBVs), and the marker effects for each model. While a similar level of accuracy was observed for many models, the level of overfitting varied widely as did the computation time and the distribution of marker effect estimates. Our comparisons suggested that GS in plant breeding programs could be based on a reduced set of models such as the Bayesian Lasso, weighted Bayesian shrinkage regression (wBSR, a fast version of BayesB), and random forest (RF) (a machine learning method that could capture nonadditive effects). Linear combinations of different models were tested as well as bagging and boosting methods, but they did not improve accuracy. This study also showed large differences in accuracy between subpopulations within a dataset that could not always be explained by differences in phenotypic variance and size. The broad diversity of empirical datasets tested here adds evidence that GS could increase genetic gain per unit of time and cost.

More accurate and precise phenotyping strategies are necessary to empower high-resolution linkage mapping and genome-wide association studies and for training genomic selection models in plant improvement. Within this framework, the objective of modern phenotyping is to increase the accuracy, precision and throughput of phenotypic estimation at all levels of biological organization while reducing costs and minimizing labor through automation, remote sensing, improved data integration and experimental design. Much like the efforts to optimize genotyping during the 1980s and 1990s, designing effective phenotyping initiatives today requires multi-faceted collaborations between biologists, computer scientists, statisticians and engineers. Robust phenotyping systems are needed to characterize the full suite of genetic factors that contribute to quantitative phenotypic variation across cells, organs and tissues, developmental stages, years, environments, species and research programs. Next-generation phenotyping generates significantly more data than previously and requires novel data management, access and storage systems, increased use of ontologies to facilitate data integration, and new statistical tools for enhancing experimental design and extracting biologically meaningful signal from environmental and experimental noise. To ensure relevance, the implementation of efficient and informative phenotyping experiments also requires familiarity with diverse germplasm resources, population structures, and target populations of environments. Today, phenotyping is quickly emerging as the major operational bottleneck limiting the power of genetic analysis and genomic prediction. The challenge for the next generation of quantitative geneticists and plant breeders is not only to understand the genetic basis of complex trait variation, but also to use that knowledge to efficiently synthesize twenty-first century crop varieties.

ABSTRACT Advancements in genotyping are rapidly decreasing marker costs and increasing genome coverage. This is facilitating the use of marker‐assisted selection (MAS) in plant breeding. Commonly employed MAS strategies, however, are not well suited for agronomically important complex traits, requiring extra time for field‐based phenotyping to identify agronomically superior lines. Genomic selection (GS) is an emerging alternative to MAS that uses all marker information to calculate genomic estimated breeding values (GEBVs) for complex traits. Selections are made directly on GEBV without further phenotyping. We developed an analytical framework to (i) compare gains from MAS and GS for complex traits and (ii) provide a plant breeding context for interpreting results from studies on GEBV accuracy. We designed MAS and GS breeding strategies with equal budgets for a high‐investment maize ( Zea mays L.) program and a low‐investment winter wheat ( Triticum aestivum L.) program. Results indicate that GS can outperform MAS on a per‐year basis even at low GEBV accuracies. Using a previously reported GEBV accuracy of 0.53 for net merit in dairy cattle, expected annual gain from GS exceeded that of MAS by about threefold for maize and twofold for winter wheat. We conclude that if moderate selection accuracies can be achieved, GS could dramatically accelerate genetic gain through its shorter breeding cycle.

Zea mays is an important genetic model for elucidating transcriptional networks. Uncertainties about the complete structure of mRNA transcripts limit the progress of research in this system. Here, using single-molecule sequencing technology, we produce 111,151 transcripts from 6 tissues capturing ∼70% of the genes annotated in maize RefGen_v3 genome. A large proportion of transcripts (57%) represent novel, sometimes tissue-specific, isoforms of known genes and 3% correspond to novel gene loci. In other cases, the identified transcripts have improved existing gene models. Averaging across all six tissues, 90% of the splice junctions are supported by short reads from matched tissues. In addition, we identified a large number of novel long non-coding RNAs and fusion transcripts and found that DNA methylation plays an important role in generating various isoforms. Our results show that characterization of the maize B73 transcriptome is far from complete, and that maize gene expression is more complex than previously thought.

Metarhizium anisopliae, the type species of the anamorph entomopathogenic genus Metarhizium, is currently composed of four varieties, including the type variety, and had been demonstrated to be closely related to M. taii, M. pingshaense and M. guizhouense. In this study we evaluate phylogenetic relationships within the M. anisopliae complex, identify monophyletic lineages and clarify the species taxonomy. To this end we have employed a multigene phylogenetic approach using near-complete sequences from nuclear encoded EF-1alpha, RPB1, RPB2 and beta-tubulin gene regions and evaluated the morphology of these taxa, including ex-type isolates whenever possible. The phylogenetic and in some cases morphological evidence supports the monophyly of nine terminal taxa in the M. anisopliae complex that we recognize as species. We propose to recognize at species rank M. anisopliae, M. guizhouense, M. pingshaense, M. acridum stat. nov., M. lepidiotae stat. nov. and M. majus stat. nov. In addition we describe the new species M. globosum and M. robertsii, resurrect the name M. brunneum and show that M. taii is a later synonym of M. guizhouense.