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Casuarina is a widely cultivated plantation tree species in coastal India, primarily due to its fast growth, high productivity and suitable for pulp and paper production. However, genetic studies of Casuarina have been hindered by lack of genomic resources and genetic markers. Knowledge of the genetic diversity and population structure of Casuarina germplasms will provide the basis for utilizing and improving resource in the breeding program. Keeping this in view, in the present study, we have identified a total of 11,503 simple sequence repeat (SSR) makers from 86,415 expressed sequence tags (ESTs) of Casuarina equisetifolia and C. junghuhniana after redundancy elimination. Dinucleotide repeats were the most abundant accounting for 72.5 % of all microsatellites, followed by trimer (23.4 %), hexamer (1.7 %), tetramer (1.5 %), and very few pentamer (0.6 %) repeats. Of these, 50 markers were used to estimate genetic diversity and population structure among 96 accessions of C. cunninghamiana and C. junghuhniana. EST-SSR markers revealed high level of polymorphism, detecting a total of 829 alleles with an average of 17 alleles per locus. Polymorphic information content (PIC) values ranged from 0.32 to 0.93, with an average of 0.78 per locus. The average observed (H o ) and expected heterozygosity (H e ) obtained was high and fairly similar in C. cunninghamiana and C. junghuhniana, thereby suggesting highly heterogeneous nature of Casuarina. Population structure using a Bayesian model-based clustering approach identified clear delineation between C. cunninghamiana and C junghuhniana. Further, these markers were also evaluated in four species of Casuarina confirming high rate of cross-species transferability. The results of this study can provide valuable insights for genetic and genomic research in Casuarina.

Traits that differentiate cross-fertile plant species can be dissected by genetic linkage analysis in interspecific hybrids. Such studies have been greatly facilitated in Eucalyptus tree species by the recent development of Diversity Arrays Technology (DArT) markers. DArT is an affordable, high-throughput marker technology for the construction of high-density genetic linkage maps. Eucalyptus grandis and Eucalyptus urophylla are commonly used to produce fast-growing, disease tolerant hybrids for clonal eucalypt plantations in tropical and subtropical regions. We analysed 7,680 DArT markers in an F2 pseudo-backcross mapping pedigree based on an F1 hybrid clone of E. grandis and E. urophylla. A total of 2,440 markers (31.7%) were polymorphic and could be placed in linkage maps of the F1 hybrid and two pure-species backcross parents. An integrated genetic linkage map was constructed for the pedigree resulting in 11 linkage groups (n = 11) with 2,290 high-confidence (LOD ≥ 3.0) markers and a total map length of 1,107.6 cM. DNA sequence analysis of the mapped DArT marker fragments revealed that 43% were located in protein coding regions and 90% could be placed in the recently completed draft genome assembly of E. grandis. Together with the anchored genomic sequence information, this linkage map will allow detailed genetic dissection of quantitative traits and hybrid fitness characters segregating in the F2 progeny and will facilitate the development of markers for molecular breeding in Eucalyptus.

International audience; 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.

Background F1 hybrid clones of Eucalyptus grandis and E. urophylla are widely grown for pulp and paper production in tropical and subtropical regions. Volume growth and wood quality are priority objectives in Eucalyptus tree improvement. The molecular basis of quantitative variation and trait expression in eucalypt hybrids, however, remains largely unknown. The recent availability of a draft genome sequence (http://www.phytozome.net) and genome-wide genotyping platforms, combined with high levels of genetic variation and high linkage disequilibrium in hybrid crosses, greatly facilitate the detection of quantitative trait loci (QTLs) as well as underlying candidate genes for growth and wood property traits. In this study, we used Diversity Arrays Technology markers to assess the genetic architecture of volume growth (diameter at breast height, DBH) and wood basic density in four-year-old progeny of an interspecific backcross pedigree of E. grandis and E. urophylla. In addition, we used Illumina RNA-Seq expression profiling in the E. urophylla backcross family to identify cis- and trans-acting polymorphisms (eQTLs) affecting transcript abundance of genes underlying QTLs for wood basic density. Results A total of five QTLs for DBH and 12 for wood basic density were identified in the two backcross families. Individual QTLs for DBH and wood basic density explained 3.1 to 12.2% of phenotypic variation. Candidate genes underlying QTLs for wood basic density on linkage groups 8 and 9 were found to share trans-acting eQTLs located on linkage groups 4 and 10, which in turn coincided with QTLs for wood basic density suggesting that these QTLs represent segregating components of an underlying transcriptional network. Conclusion This is the first demonstration of the use of next-generation expression profiling to quantify transcript abundance in a segregating tree population and identify candidate genes potentially affecting wood property variation. The QTLs identified in this study provide a resource for identifying candidate genes and developing molecular markers for marker-assisted breeding of volume growth and wood basic density. Our results suggest that integrated analysis of transcript and trait variation in eucalypt hybrids can be used to dissect the molecular basis of quantitative variation in wood property traits.

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.

Background E. grandis is used extensively for the production of pulp and paper due to its rapid growth, good form and ease of vegetative propagation. E. urophylla exhibits tolerance to fungal diseases that limit the growth of E. grandis in tropical and subtropical regions. Interspecific hybrids of these two species are, therefore, commonly used to produce fast-growing, disease tolerant hybrids for clonal eucalypt plantations in tropical and subtropical regions (e.g., South Africa, Congo and Brazil). These hybrids often exhibit superior growth and wood quality compared to the pure species, but the underlying genetic basis of the observed hybrid superiority remains unclear. Phenotypic variation observed in interspecific mapping populations has been used to identify QTLs in several genetic linkage studies in Eucalyptus. However, QTL intervals have generally been wide (20 to 30 cM) and may include several hundred genes. To bridge the gap between fine mapping and QTL validation studies, the expression levels of genes and metabolites in individuals from the segregating population can be treated as quantitative traits and used for eQTL and mQTL mapping, respectively. Co-localization of wood property, expression and metabolite QTLs will facilitate the identification of positional candidate genes and other components of regulatory networks underlying phenotypic variation. Methods To identify genetic factors controlling growth and wood property traits in Eucalyptus, an F2 pseudo-backcross mapping family derived from a cross between an F1 hybrid (GUSAP1, E. grandisxE. urophylla, Sappi Forest Research) and an E. urophylla parent (USAP1), consisting of 555 individuals, was used for genetic linkage map construction using microsatellite and DArT markers. Phenotypic trait assessment included physical measurements of tree diameter and wood density performed on 319 three-year-old individuals. Klason (acid-soluble & -insoluble) lignin and cell wall sugar content were determined for a selection of 100 backcross progeny and used for near-infrared analysis (NIRA) calibration. NIRA predictions for glucose, xylose, arabinose, cellulose and total lignin content, as well as pulp yield were made for all 315 individuals. Total lignin and S:G ratios were also separately measured for the 315 individuals. Immature xylem tissues, collected from 192 backcross progeny, were used for metabolite profiling (ORNL, Oak Ridge, TN) and Illumina mRNA-Seq (PE50, BGI Americas) quantification of transcript levels.