The application of quantitative genetics in plant and animal breeding has largely focused on additive models, which may also capture dominance and epistatic effects. Partitioning genetic variance into its additive and non-additive components using pedigree-based models (P-BLUP) is difficult with most commonly available family structures. However, the availability of dense panels of molecular markers makes possible the use of additive and dominance realized genomic relationships for the estimation of variance components and the prediction of genetic values (G-BLUP). We evaluated height data from a multi-family population of the tree species Pinus taeda with a systematic series of models accounting for additive, dominance and first order epistatic interactions (additive-by-additive, dominance-by-dominance, and additive-by-dominance), using either pedigree- or marker-based information. We show that, compared with the pedigree, use of realized genomic relationships in marker-base models yields a substantially more precise separation of additive and non-additive components of genetic variance. We conclude that the marker-based relationship matrices in a model including additive and non-additive effects performed better, improving breeding value prediction. Moreover, our results suggest that, for tree height in this population, the additive and non-additive components of genetic variance are similar in magnitude. This novel result improves our current understanding of the genetic control and architecture of a quantitative trait and should be considered when developing breeding strategies.
Genetically improving constitutive resin canal development in Pinus stems may enhance the capacity to synthesize terpenes for bark beetle resistance, chemical feedstocks, and biofuels. To discover genes that potentially regulate axial resin canal number (RCN), single nucleotide polymorphisms (SNPs) in 4027 genes were tested for association with RCN in two growth rings and three environments in a complex pedigree of 520 Pinus taeda individuals (CCLONES). The map locations of associated genes were compared with RCN quantitative trait loci (QTLs) in a (P. taeda × Pinus elliottii) × P. elliottii pseudo-backcross of 345 full-sibs (BC1). Resin canal number was heritable (h(2) ˜ 0.12-0.21) and positively genetically correlated with xylem growth (rg ˜ 0.32-0.72) and oleoresin flow (rg ˜ 0.15-0.51). Sixteen well-supported candidate regulators of RCN were discovered in CCLONES, including genes associated across sites and ages, unidirectionally associated with oleoresin flow and xylem growth, and mapped to RCN QTLs in BC1. Breeding is predicted to increase RCN 11% in one generation and could be accelerated with genomic selection at accuracies of 0.45-0.52 across environments. There is significant genetic variation for RCN in loblolly pine, which can be exploited in breeding for elevated terpene content.
The large genome size of many species hinders the development and application of genomic tools to study them. For instance, loblolly pine (Pinus taeda L.), an ecologically and economically important conifer, has a large and yet uncharacterized genome of 21.7 Gbp. To characterize the pine genome, we performed exome capture and sequencing of 14 729 genes derived from an assembly of expressed sequence tags. Efficiency of sequence capture was evaluated and shown to be similar across samples with increasing levels of complexity, including haploid cDNA, haploid genomic DNA and diploid genomic DNA. However, this efficiency was severely reduced for probes that overlapped multiple exons, presumably because intron sequences hindered probe:exon hybridizations. Such regions could not be entirely avoided during probe design, because of the lack of a reference sequence. To improve the throughput and reduce the cost of sequence capture, a method to multiplex the analysis of up to eight samples was developed. Sequence data showed that multiplexed capture was reproducible among 24 haploid samples, and can be applied for high-throughput analysis of targeted genes in large populations. Captured sequences were de novo assembled, resulting in 11 396 expanded and annotated gene models, significantly improving the knowledge about the pine gene space. Interspecific capture was also evaluated with over 98% of all probes designed from P. taeda that were efficient in sequence capture, were also suitable for analysis of the related species Pinus elliottii Engelm.
Genome structure variation, including copy number variation and presence/absence variation, comprises a large extent of maize genetic diversity; however, its effect on phenotypes remains largely unexplored. Here, we describe how copy number variation underlies a rare allele that contributes to maize aluminum (Al) tolerance. Al toxicity is the primary limitation for crop production on acid soils, which make up 50% of the worlds potentially arable lands. In a recombinant inbred line mapping population, copy number variation of the Al tolerance gene multidrug and toxic compound extrusion 1 (MATE1) is the basis for the quantitative trait locus of largest effect on phenotypic variation. This expansion in MATE1 copy number is associated with higher MATE1 expression, which in turn results in superior Al tolerance. The three MATE1 copies are identical and are part of a tandem triplication. Only three maize inbred lines carrying the three-copy allele were identified from maize and teosinte diversity panels, indicating that copy number variation for MATE1 is a rare, and quite likely recent, event. These maize lines with higher MATE1 copy number are also Al-tolerant, have high MATE1 expression, and originate from regions of highly acidic soils. Our findings show a role for copy number variation in the adaptation of maize to acidic soils in the tropics and suggest that genome structural changes may be a rapid evolutionary response to new environments.
Rapidly enhancing oleoresin production in conifer stems through genomic selection and genetic engineering may increase resistance to bark beetles and terpenoid yield for liquid biofuels. We integrated association genetic and genomic prediction analyses of oleoresin flow (g 24 h(-1)) using 4854 single nucleotide polymorphisms (SNPs) in expressed genes within a pedigreed population of loblolly pine (Pinus taeda) that was clonally replicated at three sites in the southeastern United States. Additive genetic variation in oleoresin flow (h(2) ? 0.12-0.30) was strongly correlated between years in which precipitation varied (r(a) ? 0.95), while the genetic correlation between sites declined from 0.8 to 0.37 with increasing differences in soil and climate among sites. A total of 231 SNPs were significantly associated with oleoresin flow, of which 81% were specific to individual sites. SNPs in sequences similar to ethylene signaling proteins, ABC transporters, and diterpenoid hydroxylases were associated with oleoresin flow across sites. Despite this complex genetic architecture, we developed a genomic prediction model to accelerate breeding for enhanced oleoresin flow that is robust to environmental variation. Results imply that breeding could increase oleoresin flow 1.5- to 2.4-fold in one generation.
In a context of climate change, phenotypic plasticity provides long-lived species, such as trees, with the means to adapt to environmental variations occurring within a single generation. In eucalyptus plantations, water availability is a key factor limiting productivity. However, the molecular mechanisms underlying the adaptation of eucalyptus to water shortage remain unclear. In this study, we compared the molecular responses of two commercial eucalyptus hybrids during the dry season. Both hybrids differ in productivity when grown under water deficit.
In light of impending water and arable land shortages, population growth and climate change, it is more important than ever to examine how forest tree domestication can be accelerated to sustainably meet future demands for wood, biomass, paper, fuel and biomaterials. Because of long breeding cycles, tree domestication cannot be rapidly achieved through traditional genetic improvement methods alone. Integrating modern genetic and genomic techniques with conventional breeding will expedite tree domestication. Breeders will only embrace these technologies if they are cost-effective and readily accessible, and forest landowners will only adopt end-products that meet with regulatory approval and public acceptance. All parties involved must work together to achieve these objectives for the benefit of society.
Technological advances are progressively increasing the application of genomics to a wider array of economically and ecologically important species. High-density maps enriched for transcribed genes facilitate the discovery of connections between genes and phenotypes. We report the construction of a high-density linkage map of expressed genes for the heterozygous genome of Eucalyptus using Single Feature Polymorphism (SFP) markers.
High-throughput SNP genotyping has become an essential requirement for molecular breeding and population genomics studies in plant species. Large scale SNP developments have been reported for several mainstream crops. A growing interest now exists to expand the speed and resolution of genetic analysis to outbred species with highly heterozygous genomes. When nucleotide diversity is high, a refined diagnosis of the target SNP sequence context is needed to convert queried SNPs into high-quality genotypes using the Golden Gate Genotyping Technology (GGGT). This issue becomes exacerbated when attempting to transfer SNPs across species, a scarcely explored topic in plants, and likely to become significant for population genomics and inter specific breeding applications in less domesticated and less funded plant genera.
Characterized by distinctive evolutionary adaptations, spiders provide a comprehensive system for evolutionary and developmental studies of anatomical organs, including silk and venom production. Here we performed cDNA sequencing using massively parallel sequencers (454 GS-FLX Titanium) to generate ?80,000 reads from the spinning gland of Actinopus spp. (infraorder: Mygalomorphae) and Gasteracantha cancriformis (infraorder: Araneomorphae, Orbiculariae clade). Actinopus spp. retains primitive characteristics on web usage and presents a single undifferentiated spinning gland while the orbiculariae spiders have seven differentiated spinning glands and complex patterns of web usage. MIRA, Celera Assembler and CAP3 software were used to cluster NGS reads for each spider. CAP3 unigenes passed through a pipeline for automatic annotation, classification by biological function, and comparative transcriptomics. Genes related to spider silks were manually curated and analyzed. Although a single spidroin gene family was found in Actinopus spp., a vast repertoire of specialized spider silk proteins was encountered in orbiculariae. Astacin-like metalloproteases (meprin subfamily) were shown to be some of the most sampled unigenes and duplicated gene families in G. cancriformis since its evolutionary split from mygalomorphs. Our results confirm that the evolution of the molecular repertoire of silk proteins was accompanied by the (i) anatomical differentiation of spinning glands and (ii) behavioral complexification in the web usage. Finally, a phylogenetic tree was constructed to cluster most of the known spidroins in gene clades. This is the first large-scale, multi-organism transcriptome for spider spinning glands and a first step into a broad understanding of spider web systems biology and evolution.
Aluminum (Al) toxicity is one of the most important yield-limiting factors of many crops worldwide. The primary symptom of Al toxicity syndrome is the inhibition of root growth leading to poor water and nutrient absorption. Al tolerance has been extensively studied using hydroponic experiments. However, unlike soil conditions, this method does not address all of the components that are necessary for proper root growth and development. In the present study, we grew two maize genotypes with contrasting tolerance to Al in soil containing toxic levels of Al and then compared their transcriptomic responses.
A fundamental goal of systems biology is to identify genetic elements that contribute to complex phenotypes and to understand how they interact in networks predictive of system response to genetic variation. Few studies in plants have developed such networks, and none have examined their conservation among functionally specialized organs. Here we used genetical genomics in an interspecific hybrid population of the model hardwood plant Populus to uncover transcriptional networks in xylem, leaves, and roots. Pleiotropic eQTL hotspots were detected and used to construct coexpression networks a posteriori, for which regulators were predicted based on cis-acting expression regulation. Networks were shown to be enriched for groups of genes that function in biologically coherent processes and for cis-acting promoter motifs with known roles in regulating common groups of genes. When contrasted among xylem, leaves, and roots, transcriptional networks were frequently conserved in composition, but almost invariably regulated by different loci. Similarly, the genetic architecture of gene expression regulation is highly diversified among plant organs, with less than one-third of genes with eQTL detected in two organs being regulated by the same locus. However, colocalization in eQTL position increases to 50% when they are detected in all three organs, suggesting conservation in the genetic regulation is a function of ubiquitous expression. Genes conserved in their genetic regulation among all organs are primarily cis regulated (approximately 92%), whereas genes with eQTL in only one organ are largely trans regulated. Trans-acting regulation may therefore be the primary driver of differentiation in function between plant organs.
Aluminum (Al) toxicity is a major worldwide constraint to crop productivity on acidic soils. Al becomes soluble at low pH, inhibiting root growth and severely reducing yields. Maize is an important staple food and commodity crop in acidic soil regions, especially in South America and Africa where these soils are very common. Al exclusion and intracellular tolerance have been suggested as two important mechanisms for Al tolerance in maize, but little is known about the underlying genetics.
Martian regolith (unconsolidated surface material) is a potential medium for plant growth in bioregenerative life support systems during manned missions on Mars. However, hydrated magnesium sulfate mineral levels in the regolith of Mars can reach as high as 10 wt%, and would be expected to be highly inhibitory to plant growth.
We describe a high-throughput method for estimating cell-wall chemistry traits using analytical pyrolysis. The instrument used to perform the high-throughput cell-wall chemistry analysis consists of a commercially available pyrolysis unit and autosampler coupled to a custom-built molecular beam mass spectrometer. The system is capable of analyzing approximately 42 biomass samples per hour. Lignin content and syringyl to guaiacol (S/G) ratios can be estimated directly from the spectra and differences in cell wall chemistry in large groups of samples can easily be identified using multivariate statistical data analysis methods. The utility of the system is demonstrated on a set of 800 greenhouse-grown poplar trees grown under two contrasting nitrogen treatments. High-throughput analytical pyrolysis was able to determine that the lignin content varied between 13 and 28% and the S/G ratio ranged from 0.5 to 1.5. There was more cell-wall chemistry variation in the plants grown under high nitrogen conditions than trees grown under nitrogen-deficiency conditions. Analytical pyrolysis allows the user to rapidly screen large numbers of samples at low cost, using very little sample material while producing reliable and reproducible results.
The genetic control of carbon allocation and partitioning in woody perennial plants is poorly understood despite its importance for carbon sequestration, biofuels and other wood-based industries. It is also unclear how environmental cues, such as nitrogen availability, impact the genes that regulate growth, biomass allocation and wood composition in trees. We phenotyped 396 clonally replicated genotypes of an interspecific pseudo-backcross pedigree of Populus for wood composition and biomass traits in above- and below-ground organs. The loci that regulate growth, carbon allocation and partitioning under two nitrogen conditions were identified, defining the contribution of environmental cues to their genetic control. Sixty-three quantitative trait loci were identified for the 20 traits analyzed. The majority of quantitative trait loci are specific to one of the two nitrogen treatments, demonstrating significant nitrogen-dependent genetic control. A highly significant genetic correlation was observed between plant growth and lignin/cellulose composition, and quantitative trait loci co-localization identified the genomic position of potential pleiotropic regulators. Pleiotropic loci linking higher growth rates to wood with less lignin are excellent targets to engineer tree germplasm improved for pulp, paper and cellulosic ethanol production. The causative genes are being identified with a genetical genomics approach.
Microarrays have demonstrated significant power for genome-wide analyses of gene expression, and recently have also revolutionized the genetic analysis of segregating populations by genotyping thousands of loci in a single assay. Although microarray-based genotyping approaches have been successfully applied in yeast and several inbred plant species, their power has not been proven in an outcrossing species with extensive genetic diversity. Here we have developed methods for high-throughput microarray-based genotyping in such species using a pseudo-backcross progeny of 154 individuals of Populus trichocarpa and P. deltoides analyzed with long-oligonucleotide in situ-synthesized microarray probes. Our analysis resulted in high-confidence genotypes for 719 single-feature polymorphism (SFP) and 1014 gene expression marker (GEM) candidates. Using these genotypes and an established microsatellite (SSR) framework map, we produced a high-density genetic map comprising over 600 SFPs, GEMs and SSRs. The abundance of gene-based markers allowed us to localize over 35 million base pairs of previously unplaced whole-genome shotgun (WGS) scaffold sequence to putative locations in the genome of P. trichocarpa. A high proportion of sampled scaffolds could be verified for their placement with independently mapped SSRs, demonstrating the previously un-utilized power that high-density genotyping can provide in the context of map-based WGS sequence reassembly. Our results provide a substantial contribution to the continued improvement of the Populus genome assembly, while demonstrating the feasibility of microarray-based genotyping in a highly heterozygous population. The strategies presented are applicable to genetic mapping efforts in all plant species with similarly high levels of genetic diversity.
Growth traits in trees are fundamental components of adaptation in a forest ecosystem and of productivity in planted forests. A number of processes determine tree growth, which are controlled by genetic and epigenetic factors that respond dynamically to environmental signals throughout centuries. Advances in genomics have allowed an increased comprehension of the complex mechanisms of tree growth and adaptation. Yet, the application of genomics to improving forest productivity and sustainability still entails capturing a large proportion of the total genetic variation controlling the component traits. Nonetheless, genetics and genomics are unifying disciplines that will serve well to dissect the variables and mechanisms of tree growth and development.
Loblolly pine (Pinus taeda L.) is an important conifer for which a suite of genomic resources is being generated for comparative studies. Despite recent attempts to sequence the large genome of conifers, their assembly and the positioning of genes remains largely incomplete. The inter-specific synteny in pines suggests that a gene-based map would be useful to support genome assemblies and analysis of conifers. To establish a reference gene-based genetic map we performed exome sequencing of 14,729 genes on a mapping population of 72 haploid samples generating a resource of 7,434 sequence variants segregating for 3,787 genes. Most markers are single nucleotide polymorphisms, although short insertions/deletions and multiple nucleotide polymorphisms were also utilized. Marker segregation in the population was used to generate a high-density, gene-based genetic map. 2,841 genes were mapped to pines 12 linkage groups with an average of one marker every 0.58 cM. Capture data was utilized to detect gene presence/absence variations and position 65 genes on the map. We compared the marker order of genes previously mapped in loblolly pine and found high agreement. We estimated that 4,123 genes had enough sequencing depth for reliable detection of markers, suggesting a high marker conversation rate of 92% (3,787 / 4,123). This is possible because a significant portion of the gene is captured and sequenced, increasing the chances of identifying a polymorphic site for characterization and mapping. This sub-centiMorgan genetic map provides a valuable resource for gene positioning on chromosomes and guide for the assembly of a reference pine genome.
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