Project description: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. SFP discovery and mapping was achieved using pseudo-testcross screening and selective mapping to simultaneously optimize linkage mapping and microarray costs. SFP genotyping was carried out by hybridizing complementary RNA prepared from 4.5 year-old trees xylem to an SFP array containing 103,000 25-mer oligonucleotide probes representing 20,726 unigenes derived from a modest size expressed sequence tags collection. An SFP-mapping microarray with 43,777 selected candidate SFP probes representing 15,698 genes was subsequently designed and used to genotype SFPs in a larger subset of the segregating population drawn by selective mapping. A total of 1,845 genes were mapped, with 884 of them ordered with high likelihood support on a framework map anchored to 180 microsatellites with average density of 1.2 cM. Using more probes per unigene increased by two-fold the likelihood of detecting segregating SFPs eventually resulting in more genes mapped. In silico validation showed that 87% of the SFPs map to the expected location on the 4.5X draft sequence of the Eucalyptus grandis genome. The Eucalyptus 1,845 gene map is the most highly enriched map for transcriptional information for any forest tree species to date. It represents a major improvement on the number of genes previously positioned on Eucalyptus maps and provides an initial glimpse at the gene space for this global tree genome. A general protocol is proposed to build high-density transcript linkage maps in less characterized plant species by SFP genotyping with a concurrent objective of reducing microarray costs. HIgh-density gene-rich maps represent a powerful resource to assist gene discovery endeavors when used in combination with QTL and association mapping and should be especially valuable to assist the assembly of reference genome sequences soon to come for several plant and animal species.
Project description:Eucalyptus species are widely used in the forestry industry, and a significant increase in the number of sequences available in database repositories has been observed for these species. In proteomics, a protein is identified by correlating the theoretical fragmentation spectrum derived from genomic/transcriptomic data against the experimental fragmentation mass spectrum acquired from large-scale analysis of protein mixtures. Proteogenomics is an alternative approach that can identify novel proteins encoded by regions previously considered as non-coding. This study aimed to confidently identify and confirm the existence of previously unknown protein-coding sequences in the Eucalyptus grandis genome.
Project description: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. SFP discovery and mapping was achieved using pseudo-testcross screening and selective mapping to simultaneously optimize linkage mapping and microarray costs. SFP genotyping was carried out by hybridizing complementary RNA prepared from 4.5 year-old trees xylem to an SFP array containing 103,000 25-mer oligonucleotide probes representing 20,726 unigenes derived from a modest size expressed sequence tags collection. An SFP-mapping microarray with 43,777 selected candidate SFP probes representing 15,698 genes was subsequently designed and used to genotype SFPs in a larger subset of the segregating population drawn by selective mapping. A total of 1,845 genes were mapped, with 884 of them ordered with high likelihood support on a framework map anchored to 180 microsatellites with average density of 1.2 cM. Using more probes per unigene increased by two-fold the likelihood of detecting segregating SFPs eventually resulting in more genes mapped. In silico validation showed that 87% of the SFPs map to the expected location on the 4.5X draft sequence of the Eucalyptus grandis genome. The Eucalyptus 1,845 gene map is the most highly enriched map for transcriptional information for any forest tree species to date. It represents a major improvement on the number of genes previously positioned on Eucalyptus maps and provides an initial glimpse at the gene space for this global tree genome. A general protocol is proposed to build high-density transcript linkage maps in less characterized plant species by SFP genotyping with a concurrent objective of reducing microarray costs. HIgh-density gene-rich maps represent a powerful resource to assist gene discovery endeavors when used in combination with QTL and association mapping and should be especially valuable to assist the assembly of reference genome sequences soon to come for several plant and animal species.
Project description:The expression pattern of transcripts involved in secondary development was determined in four genotypes of E. tereticornis using 8x60K Gene chip. The genome-wide expression data was used to develop a gene co-expression network for xylogenesis. Agilent one-color experiment, Organism: Eucalyptus,Custom Agilent Eucalyptus 8x60k Microarray Gene expression (AMADID: 59849 ) designed by Genotypic Technology Pvt.Ltd.
Project description:Background: Eucalyptus species and interspecific hybrids exhibit valuable growth and wood properties that make them a highly desirable commodity. However, these trees are challenged by a wide array of biotic stresses during their lifetimes. The Eucalyptus grandis reference genome sequence provides a resource to study pest and pathogen defence mechanisms in long-lived woody plants. E. grandis trees are generally susceptible to Chrysoporthe austroafricana, a causal agent of stem cankers on eucalypts. The aim of this study was to characterize the defence response of E. grandis against C. austroafricana. Results: Hormone profiling of susceptible and moderately resistant clonal E. grandis genotypes indicated a reduction in salicylic acid and gibberellic acid levels at 3 days post inoculation. We hypothesized that these signaling pathways may facilitate resistance. To further investigate other defence mechanisms at this time point, transcriptome profiling was performed. This revealed that cell wall modifications and response to oxidative stress form part of the defence responses common to both genotypes, whilst changes in the hormone signaling pathways may contribute to resistance. Additionally the expression of selected candidate defence response genes was induced earlier in moderately resistant trees than in susceptible trees, supporting the hypothesis that a delayed defence response may occur in the susceptible interaction. Conclusion: The ability of a host to fine-tune its defence responses is crucial and the responses identified in this study extends our understanding of plant defence, gained from model systems, to woody perennials. E. grandis trees were stem inoculated with C. austroafricana. RNA was extracted from stem material harvested 3 days post inoculation for transcriptome profiling. Three biological replicates of harvested material was sent for RNA-sequencing
Project description:Background: Eucalyptus species and interspecific hybrids exhibit valuable growth and wood properties that make them a highly desirable commodity. However, these trees are challenged by a wide array of biotic stresses during their lifetimes. The Eucalyptus grandis reference genome sequence provides a resource to study pest and pathogen defence mechanisms in long-lived woody plants. E. grandis trees are generally susceptible to Chrysoporthe austroafricana, a causal agent of stem cankers on eucalypts. The aim of this study was to characterize the defence response of E. grandis against C. austroafricana. Results: Hormone profiling of susceptible and moderately resistant clonal E. grandis genotypes indicated a reduction in salicylic acid and gibberellic acid levels at 3 days post inoculation. We hypothesized that these signaling pathways may facilitate resistance. To further investigate other defence mechanisms at this time point, transcriptome profiling was performed. This revealed that cell wall modifications and response to oxidative stress form part of the defence responses common to both genotypes, whilst changes in the hormone signaling pathways may contribute to resistance. Additionally the expression of selected candidate defence response genes was induced earlier in moderately resistant trees than in susceptible trees, supporting the hypothesis that a delayed defence response may occur in the susceptible interaction. Conclusion: The ability of a host to fine-tune its defence responses is crucial and the responses identified in this study extends our understanding of plant defence, gained from model systems, to woody perennials.