Multispectral and thermal infrared data, visual scores for severity of common rust symptoms, and genotypic single nucleotide polymorphism data of three F2-derived biparental doubled-haploid maize populations.

Three F2-derived biparental doubled haploid (DH) maize populations were generated for genetic mapping of resistance to common rust. Each of the three populations has the same susceptible parent, but a different resistance donor parent. Population 1 and 3 consist of 320 lines each, population 2 consists of 260 lines. The DH lines were evaluated for their susceptibility to common rust in two years and with two replications in each year. For phenotyping, a visual score (VS) for susceptibility was assigned. Additionally, unmanned aerial vehicle (UAV) derived multispectral and thermal infrared data was recorded and combined in different vegetation indices ("remote sensing", RS). The DH lines were genotyped with the DarTseq method, to obtain data on single nucleotide polymorphisms (SNPs). After quality control, 9051 markers remained. Missing values were "imputed" by the empirical mean of the marker scores of the respective locus. We used the data for comparison of genome-wide association studies and genomic prediction when based on different phenotyping methods, that is either VS or RS data. The data may be interesting for reuse for instance for benchmarking genomic prediction models, for phytopathological studies addressing common rust, or for specifications of vegetation indices.

Use of remote sensing for linkage mapping and genomic prediction for common rust resistance in maize.

Breeding for disease resistance is a central component of strategies implemented to mitigate biotic stress impacts on crop yield. Conventionally, genotypes of a plant population are evaluated through a labor-intensive process of assigning visual scores (VS) of susceptibility (or resistance) by specifically trained staff, which limits manageable volumes and repeatability of evaluation trials. Remote sensing (RS) tools have the potential to streamline phenotyping processes and to deliver more standardized results at higher through-put. Here, we use a two-year evaluation trial of three newly developed biparental populations of maize doubled haploid lines (DH) to compare the results of genomic analyses of resistance to common rust (CR) when phenotyping is either based on conventional VS or on RS-derived (vegetation) indices. As a general observation, for each population × year combination, the broad sense heritability of VS was greater than or very close to the maximum heritability across all RS indices. Moreover, results of linkage mapping as well as of genomic prediction (GP), suggest that VS data was of a higher quality, indicated by higher -logp values in the linkage studies and higher predictive abilities for genomic prediction. Nevertheless, despite the qualitative differences between the phenotyping methods, each successfully identified the same genomic region on chromosome 10 as being associated with disease resistance. This region is likely related to the known CR resistance locus Rp1. Our results indicate that RS technology can be used to streamline genetic evaluation processes for foliar disease resistance in maize. In particular, RS can potentially reduce costs of phenotypic evaluations and increase trialing capacities.

Genetic variation among elite inbred lines suggests potential to breed for BNI-capacity in maize.

Biological nitrification inhibition (BNI) is a plant function where root systems release antibiotic compounds (BNIs) specifically aimed at suppressing nitrifiers to limit soil-nitrate formation in the root zone. Little is known about BNI-activity in maize (Zea mays L.), the most important food, feed, and energy crop. Two categories of BNIs are released from maize roots; hydrophobic and hydrophilic BNIs, that determine BNI-capacity in root systems. Zeanone is a recently discovered hydrophobic compound with BNI-activity, released from maize roots. The objectives of this study were to understand/quantify the relationship between zeanone activity and hydrophobic BNI-capacity. We assessed genetic variability among 250 CIMMYT maize lines (CMLs) characterized for hydrophobic BNI-capacity and zeanone activity, towards developing genetic markers linked to this trait in maize. CMLs with high BNI-capacity and ability to release zeanone from roots were identified. GWAS was performed using 27,085 SNPs (with unique positions on the B73v.4 reference genome, and false discovery rate = 10), and phenotypic information for BNI-capacity and zeanone production from root systems. Eighteen significant markers were identified; three associated with specific BNI-activity (SBNI), four with BNI-activity per plant (BNIPP), another ten were common between SBNI and BNIPP, and one with zeanone release. Further, 30 annotated genes were associated with the significant SNPs; most of these genes are involved in pathways of "biological process", and one (AMT5) in ammonium regulation in maize roots. Although the inbred lines in this study were not developed for BNI-traits, the identification of markers associated with BNI-capacity suggests the possibility of using these genomic tools in marker-assisted selection to improve hydrophobic BNI-capacity in maize.

Genetic structure analysis and identifying key founder inbred lines in diverse elite sub-tropical maize inbred lines.

Understanding the genetic relationships between the key founder inbred lines and derived inbred lines could provide insight into the breeding history and the structure of genetic diversity of the available elite inbred lines with desirable target traits. The maize improvement program at the International Institute of Tropical Agriculture (IITA) analyzed the pedigree information of 623 sub-tropical maize inbred lines generated at the IITA maize breeding program to identify the key founder inbred lines. We also used 5032 SNP markers to assess the genetic similarities of the founder inbred lines with their progenies subsequently developed for specific target traits. The results of pedigree analysis and SNP markers-based similarity scores identified 20 key founder inbred lines with significant contributions to the development of drought tolerant, early maturing, productive, Striga resistant, provitamin A enriched, and quality protein maize inbred lines. In our breeding program, line TZMi501 belonging to a flint heterotic group (HGA), and TZMi407-S and TZMi214, representing the dent heterotic group (HGB), were identified as the most useful founder inbred lines. The 623 inbred lines were consistently separated into four clusters based on Ward's hierarchical clustering, structure, and principal component analyses, with the 20 founder inbred lines spread into all clusters. The founder inbred lines were more genetically related to the productive inbred lines but showed genetic divergence from the provitamin A enriched inbred lines. These results provide a better understanding of the breeding history of the sub-tropical maize inbred lines to facilitate parental selection aligned to existing heterotic groups for use in breeding programs targeting the improvement of essential traits in maize.

Maize dispersal patterns associated with different types of endosperm and migration of indigenous groups in lowland South America.

BACKGROUND AND AIMS: The lowlands of South America appear to be remarkably important in the evolutionary history of maize, due to new evidence that suggests that maize dispersed from Mexico and arrived in this region in a state of partial domestication. This study aimed to identify dispersal patterns of maize genetic diversity in this part of the continent. METHODS: A total of 170 maize accessions were characterized with 4398 single nucleotide polymorphisms (SNPs) and analysed to determine if maize dispersal was associated with types of endosperm and indigenous language families. KEY RESULTS: Four genetic groups were identified in the discriminant analysis of principal components and five groups in the cluster analysis (neighbour-joining method). The groups were structured according to the predominance of endosperm types (popcorn, floury, flint/semi-flint). Spatial principal component analysis of genetic variation suggests different dispersal patterns for each endosperm type and can be associated with hypotheses of expansions of different indigenous groups. CONCLUSIONS: From a possible origin in Southwestern Amazonia, different maize dispersal routes emerged: (1) towards Northern Amazonia, which continued towards the Caatinga and south-eastern Atlantic Forest (Floury); (2) towards Southern Brazil, passing through the Cerrado and Southern Atlantic Forest reaching the Pampa region (Floury); and (3) along the Atlantic Coast, following Tupi movements originating from two separate expansions: one (Tupinamba) from north to south, and the other (Guarani) in the opposite direction, from south to north (flint, floury and popcorn).

Introgression of Maize Diversity for Drought Tolerance: Subtropical Maize Landraces as Source of New Positive Variants.

Current climate change models predict an increased frequency and intensity of drought for much of the developing world within the next 30 years. These events will negatively affect maize yields, potentially leading to economic and social instability in many smallholder farming communities. Knowledge about the genetic resources available for traits related to drought tolerance has great importance in developing breeding program strategies. The aim of this research was to study a maize landrace introgression panel to identify chromosomal regions associated with a drought tolerance index. For that, we performed Genome-Wide Association Study (GWAS) on 1326 landrace progenies developed by the CIMMYT Genetic Resources Program, originating from 20 landraces populations collected in arid regions. Phenotypic data were obtained from early testcross trials conducted in three sites and two contrasting irrigation environments, full irrigation (well-watered) and reduced irrigation (drought). The populations were genotyped using the DArTSeq® platform, and a final set of 5,695 SNPs markers was used. The genotypic values were estimated using spatial adjustment in a two-stage analysis. First, we performed the individual analysis for each site/irrigation treatment combination. The best linear unbiased estimates (BLUEs) were used to calculate the Harmonic Mean of Relative Performance (HMRP) as a drought tolerance index for each testcross. The second stage was a joint analysis, which was performed using the HMRP to obtain the best linear unbiased predictions (BLUPs) of the index for each genotype. Then, GWAS was performed to determine the marker-index associations and the marker-Grain Yield (GY) associations for the two irrigation treatments. We detected two significant markers associated with the drought-tolerance index, four associated with GY in drought condition, and other four associated with GY in irrigated conditions each. Although each of these markers explained less than 0.1% of the phenotypic variation for the index and GY, we found two genes likely related to the plant response to drought stress. For these markers, alleles from landraces provide a slightly higher yield under drought conditions. Our results indicate that the positive diversity delivered by landraces are still present on the backcrosses and this is a potential breeding strategy for improving maize for drought tolerance and for trait introgression bringing new superior allelic diversity from landraces to breeding populations.

Genetic diversity and population structure of early and extra-early maturing maize germplasm adapted to sub-Saharan Africa.

BACKGROUND: Assessment and effective utilization of genetic diversity in breeding programs is crucial for sustainable genetic improvement and rapid adaptation to changing breeding objectives. During the past two decades, the commercialization of the early and extra-early maturing cultivars has contributed to rapid expansion of maize into different agro-ecologies of sub-Saharan Africa (SSA) where maize has become an important component of the agricultural economy and played a vital role in food and nutritional security. The present study aimed at understanding the population structure and genetic variability among 439 early and extra-early maize inbred lines developed from three narrow-based and twenty-seven broad-based populations by the International Iinstitute of Tropical Agriculture Maize Improvement Program (IITA-MIP). These inbreds were genotyped using 9642 DArTseq-based single nucleotide polymorphism (SNP) markers distributed uniformly throughout the maize genome. RESULTS: About 40.8% SNP markers were found highly informative and exhibited polymorphic information content (PIC) greater than 0.25. The minor allele frequency and PIC ranged from 0.015 to 0.500 and 0.029 to 0.375, respectively. The STRUCTURE, neighbour-joining phylogenetic tree and principal coordinate analysis (PCoA) grouped the inbred lines into four major classes generally consistent with the selection history, ancestry and kernel colour of the inbreds but indicated a complex pattern of the genetic structure. The pattern of grouping of the lines based on the STRUCTURE analysis was in concordance with the results of the PCoA and suggested greater number of sub-populations (K = 10). Generally, the classification of the inbred lines into heterotic groups based on SNP markers was reasonably reliable and in agreement with defined heterotic groups of previously identified testers based on combining ability studies. CONCLUSIONS: Complete understanding of potential heterotic groups would be difficult to portray by depending solely on molecular markers. Therefore, planned crosses involving representative testers from opposing heterotic groups would be required to refine the existing heterotic groups. It is anticipated that the present set of inbreds could contribute new beneficial alleles for population improvement, development of hybrids and lines with potential to strengthen future breeding programs. Results of this study would help breeders in formulating breeding strategies for genetic enhancement and sustainable maize production in SSA.

Diversity analysis of 80,000 wheat accessions reveals consequences and opportunities of selection footprints.

Undomesticated wild species, crop wild relatives, and landraces represent sources of variation for wheat improvement to address challenges from climate change and the growing human population. Here, we study 56,342 domesticated hexaploid, 18,946 domesticated tetraploid and 3,903 crop wild relatives in a massive-scale genotyping and diversity analysis. Using DArTseqTM technology, we identify more than 300,000 high-quality SNPs and SilicoDArT markers and align them to three reference maps: the IWGSC RefSeq v1.0 genome assembly, the durum wheat genome assembly (cv. Svevo), and the DArT genetic map. On average, 72% of the markers are uniquely placed on these maps and 50% are linked to genes. The analysis reveals landraces with unexplored diversity and genetic footprints defined by regions under selection. This provides fertile ground to develop wheat varieties of the future by exploring specific gene or chromosome regions and identifying germplasm conserving allelic diversity missing in current breeding programs.

The Development of Quality Control Genotyping Approaches: A Case Study Using Elite Maize Lines.

Quality control (QC) of germplasm identity and purity is a critical component of breeding and conservation activities. SNP genotyping technologies and increased availability of markers provide the opportunity to employ genotyping as a low-cost and robust component of this QC. In the public sector available low-cost SNP QC genotyping methods have been developed from a very limited panel of markers of 1,000 to 1,500 markers without broad selection of the most informative SNPs. Selection of optimal SNPs and definition of appropriate germplasm sampling in addition to platform section impact on logistical and resource-use considerations for breeding and conservation applications when mainstreaming QC. In order to address these issues, we evaluated the selection and use of SNPs for QC applications from large DArTSeq data sets generated from CIMMYT maize inbred lines (CMLs). Two QC genotyping strategies were developed, the first is a "rapid QC", employing a small number of SNPs to identify potential mislabeling of seed packages or plots, the second is a "broad QC", employing a larger number of SNP, used to identify each germplasm entry and to measure heterogeneity. The optimal marker selection strategies combined the selection of markers with high minor allele frequency, sampling of clustered SNP in proportion to marker cluster distance and selecting markers that maintain a uniform genomic distribution. The rapid and broad QC SNP panels selected using this approach were further validated using blind test assessments of related re-generation samples. The influence of sampling within each line was evaluated. Sampling 192 individuals would result in close to 100% possibility of detecting a 5% contamination in the entry, and approximately a 98% probability to detect a 2% contamination of the line. These results provide a framework for the establishment of QC genotyping. A comparison of financial and time costs for use of these approaches across different platforms is discussed providing a framework for institutions involved in maize conservation and breeding to assess the resource use effectiveness of QC genotyping. Application of these research findings, in combination with existing QC approaches, will ensure the regeneration, distribution and use in breeding of true to type inbred germplasm. These findings also provide an effective approach to optimize SNP selection for QC genotyping in other species.

Genomic Prediction of Gene Bank Wheat Landraces.

This study examines genomic prediction within 8416 Mexican landrace accessions and 2403 Iranian landrace accessions stored in gene banks. The Mexican and Iranian collections were evaluated in separate field trials, including an optimum environment for several traits, and in two separate environments (drought, D and heat, H) for the highly heritable traits, days to heading (DTH), and days to maturity (DTM). Analyses accounting and not accounting for population structure were performed. Genomic prediction models include genotype × environment interaction (G × E). Two alternative prediction strategies were studied: (1) random cross-validation of the data in 20% training (TRN) and 80% testing (TST) (TRN20-TST80) sets, and (2) two types of core sets, "diversity" and "prediction", including 10% and 20%, respectively, of the total collections. Accounting for population structure decreased prediction accuracy by 15-20% as compared to prediction accuracy obtained when not accounting for population structure. Accounting for population structure gave prediction accuracies for traits evaluated in one environment for TRN20-TST80 that ranged from 0.407 to 0.677 for Mexican landraces, and from 0.166 to 0.662 for Iranian landraces. Prediction accuracy of the 20% diversity core set was similar to accuracies obtained for TRN20-TST80, ranging from 0.412 to 0.654 for Mexican landraces, and from 0.182 to 0.647 for Iranian landraces. The predictive core set gave similar prediction accuracy as the diversity core set for Mexican collections, but slightly lower for Iranian collections. Prediction accuracy when incorporating G × E for DTH and DTM for Mexican landraces for TRN20-TST80 was around 0.60, which is greater than without the G × E term. For Iranian landraces, accuracies were 0.55 for the G × E model with TRN20-TST80. Results show promising prediction accuracies for potential use in germplasm enhancement and rapid introgression of exotic germplasm into elite materials.

Exploring and Mobilizing the Gene Bank Biodiversity for Wheat Improvement.

Identifying and mobilizing useful genetic variation from germplasm banks to breeding programs is an important strategy for sustaining crop genetic improvement. The molecular diversity of 1,423 spring bread wheat accessions representing major global production environments was investigated using high quality genotyping-by-sequencing (GBS) loci, and gene-based markers for various adaptive and quality traits. Mean diversity index (DI) estimates revealed synthetic hexaploids to be genetically more diverse (DI= 0.284) than elites (DI = 0.267) and landraces (DI = 0.245). GBS markers discovered thousands of new SNP variations in the landraces which were well known to be adapted to drought (1273 novel GBS SNPs) and heat (4473 novel GBS SNPs) stress environments. This may open new avenues for pre-breeding by enriching the elite germplasm with novel alleles for drought and heat tolerance. Furthermore, new allelic variation for vernalization and glutenin genes was also identified from 47 landraces originating from Iraq, Iran, India, Afghanistan, Pakistan, Uzbekistan and Turkmenistan. The information generated in the study has been utilized to select 200 diverse gene bank accessions to harness their potential in pre-breeding and for allele mining of candidate genes for drought and heat stress tolerance, thus channeling novel variation into breeding pipelines. This research is part of CIMMYT's ongoing 'Seeds of Discovery' project visioning towards the development of high yielding wheat varieties that address future challenges from climate change.

The genome of Eucalyptus grandis.

Eucalypts are the world's most widely planted hardwood trees. Their outstanding diversity, adaptability and growth have made them a global renewable resource of fibre and energy. We sequenced and assembled >94% of the 640-megabase genome of Eucalyptus grandis. Of 36,376 predicted protein-coding genes, 34% occur in tandem duplications, the largest proportion thus far in plant genomes. Eucalyptus also shows the highest diversity of genes for specialized metabolites such as terpenes that act as chemical defence and provide unique pharmaceutical oils. Genome sequencing of the E. grandis sister species E. globulus and a set of inbred E. grandis tree genomes reveals dynamic genome evolution and hotspots of inbreeding depression. The E. grandis genome is the first reference for the eudicot order Myrtales and is placed here sister to the eurosids. This resource expands our understanding of the unique biology of large woody perennials and provides a powerful tool to accelerate comparative biology, breeding and biotechnology.

Genomic characterization of DArT markers based on high-density linkage analysis and physical mapping to the Eucalyptus genome.

Diversity Arrays Technology (DArT) provides a robust, high throughput, cost-effective method to query thousands of sequence polymorphisms in a single assay. Despite the extensive use of this genotyping platform for numerous plant species, little is known regarding the sequence attributes and genome-wide distribution of DArT markers. We investigated the genomic properties of the 7,680 DArT marker probes of a Eucalyptus array, by sequencing them, constructing a high density linkage map and carrying out detailed physical mapping analyses to the Eucalyptus grandis reference genome. A consensus linkage map with 2,274 DArT markers anchored to 210 microsatellites and a framework map, with improved support for ordering, displayed extensive collinearity with the genome sequence. Only 1.4 Mbp of the 75 Mbp of still unplaced scaffold sequence was captured by 45 linkage mapped but physically unaligned markers to the 11 main Eucalyptus pseudochromosomes, providing compelling evidence for the quality and completeness of the current Eucalyptus genome assembly. A highly significant correspondence was found between the locations of DArT markers and predicted gene models, while most of the 89 DArT probes unaligned to the genome correspond to sequences likely absent in E. grandis, consistent with the pan-genomic feature of this multi-Eucalyptus species DArT array. These comprehensive linkage-to-physical mapping analyses provide novel data regarding the genomic attributes of DArT markers in plant genomes in general and for Eucalyptus in particular. DArT markers preferentially target the gene space and display a largely homogeneous distribution across the genome, thereby providing superb coverage for mapping and genome-wide applications in breeding and diversity studies. Data reported on these ubiquitous properties of DArT markers will be particularly valuable to researchers working on less-studied crop species who already count on DArT genotyping arrays but for which no reference genome is yet available to allow such detailed characterization.

A reference linkage map for Eucalyptus.

BACKGROUND: Genetic linkage maps are invaluable resources in plant research. They provide a key tool for many genetic applications including: mapping quantitative trait loci (QTL); comparative mapping; identifying unlinked (i.e. independent) DNA markers for fingerprinting, population genetics and phylogenetics; assisting genome sequence assembly; relating physical and recombination distances along the genome and map-based cloning of genes. Eucalypts are the dominant tree species in most Australian ecosystems and of economic importance globally as plantation trees. The genome sequence of E. grandis has recently been released providing unprecedented opportunities for genetic and genomic research in the genus. A robust reference linkage map containing sequence-based molecular markers is needed to capitalise on this resource. Several high density linkage maps have recently been constructed for the main commercial forestry species in the genus (E. grandis, E. urophylla and E. globulus) using sequenced Diversity Arrays Technology (DArT) and microsatellite markers. To provide a single reference linkage map for eucalypts a composite map was produced through the integration of data from seven independent mapping experiments (1950 individuals) using a marker-merging method. RESULTS: The composite map totalled 1107 cM and contained 4101 markers; comprising 3880 DArT, 213 microsatellite and eight candidate genes. Eighty-one DArT markers were mapped to two or more linkage groups, resulting in the 4101 markers being mapped to 4191 map positions. Approximately 13% of DArT markers mapped to identical map positions, thus the composite map contained 3634 unique loci at an average interval of 0.31 cM. CONCLUSION: The composite map represents the most saturated linkage map yet produced in Eucalyptus. As the majority of DArT markers contained on the map have been sequenced, the map provides a direct link to the E. grandis genome sequence and will serve as an important reference for progressing eucalypt research.

Genomic selection for growth and wood quality in Eucalyptus: capturing the missing heritability and accelerating breeding for complex traits in forest trees.

• Genomic selection (GS) is expected to cause a paradigm shift in tree breeding by improving its speed and efficiency. By fitting all the genome-wide markers concurrently, GS can capture most of the 'missing heritability' of complex traits that quantitative trait locus (QTL) and association mapping classically fail to explain. Experimental support of GS is now required. • The effectiveness of GS was assessed in two unrelated Eucalyptus breeding populations with contrasting effective population sizes (N(e) = 11 and 51) genotyped with > 3000 DArT markers. Prediction models were developed for tree circumference and height growth, wood specific gravity and pulp yield using random regression best linear unbiased predictor (BLUP). • Accuracies of GS varied between 0.55 and 0.88, matching the accuracies achieved by conventional phenotypic selection. Substantial proportions (74-97%) of trait heritability were captured by fitting all genome-wide markers simultaneously. Genomic regions explaining trait variation largely coincided between populations, although GS models predicted poorly across populations, likely as a result of variable patterns of linkage disequilibrium, inconsistent allelic effects and genotype × environment interaction. • GS brings a new perspective to the understanding of quantitative trait variation in forest trees and provides a revolutionary tool for applied tree improvement. Nevertheless population-specific predictive models will likely drive the initial applications of GS in forest tree breeding.

Population genetic analysis and phylogeny reconstruction in Eucalyptus (Myrtaceae) using high-throughput, genome-wide genotyping.

A set of over 8000 Diversity Arrays Technology (DArT) markers was tested for its utility in high-resolution population and phylogenetic studies across a range of Eucalyptus taxa. Small-scale population studies of Eucalyptus camaldulensis, Eucalyptus cladocalyx, Eucalyptus globulus, Eucalyptus grandis, Eucalyptus nitens, Eucalyptus pilularis and Eucalyptus urophylla demonstrated the potential of genome-wide genotyping with DArT markers to differentiate species, to identify interspecific hybrids and to resolve biogeographic disjunctions within species. The population genetic studies resolved geographically partitioned clusters in E. camaldulensis, E. cladocalyx, E. globulus and E. urophylla that were congruent with previous molecular studies. A phylogenetic study of 94 eucalypt species provided results that were largely congruent with traditional taxonomy and ITS-based phylogenies, but provided more resolution within major clades than had been obtained previously. Ascertainment bias (the bias introduced in a phylogeny from using markers developed in a small sample of the taxa that are being studied) was not detected. DArT offers an unprecedented level of resolution for population genetic, phylogenetic and evolutionary studies across the full range of Eucalyptus species.

A high-density Diversity Arrays Technology (DArT) microarray for genome-wide genotyping in Eucalyptus.

BACKGROUND: A number of molecular marker technologies have allowed important advances in the understanding of the genetics and evolution of Eucalyptus, a genus that includes over 700 species, some of which are used worldwide in plantation forestry. Nevertheless, the average marker density achieved with current technologies remains at the level of a few hundred markers per population. Furthermore, the transferability of markers produced with most existing technology across species and pedigrees is usually very limited. High throughput, combined with wide genome coverage and high transferability are necessary to increase the resolution, speed and utility of molecular marker technology in eucalypts. We report the development of a high-density DArT genome profiling resource and demonstrate its potential for genome-wide diversity analysis and linkage mapping in several species of Eucalyptus. FINDINGS: After testing several genome complexity reduction methods we identified the PstI/TaqI method as the most effective for Eucalyptus and developed 18 genomic libraries from PstI/TaqI representations of 64 different Eucalyptus species. A total of 23,808 cloned DNA fragments were screened and 13,300 (56%) were found to be polymorphic among 284 individuals. After a redundancy analysis, 6,528 markers were selected for the operational array and these were supplemented with 1,152 additional clones taken from a library made from the E. grandis tree whose genome has been sequenced. Performance validation for diversity studies revealed 4,752 polymorphic markers among 174 individuals. Additionally, 5,013 markers showed segregation when screened using six inter-specific mapping pedigrees, with an average of 2,211 polymorphic markers per pedigree and a minimum of 859 polymorphic markers that were shared between any two pedigrees. CONCLUSIONS: This operational DArT array will deliver 1,000-2,000 polymorphic markers for linkage mapping in most eucalypt pedigrees and thus provide high genome coverage. This array will also provide a high-throughput platform for population genetics and phylogenetics in Eucalyptus. The transferability of DArT across species and pedigrees is particularly valuable for a large genus such as Eucalyptus and will facilitate the transfer of information between different studies. Furthermore, the DArT marker array will provide a high-resolution link between phenotypes in populations and the Eucalyptus reference genome, which will soon be completed.