Linkage disequilibrium (LD) mapping using natural populations results in higher resolution of markertrait associations compared to family-based quantitative trait locus (QTL) studies. Depending on the extent of LD, it is possible to identify alleles within candidate genes associated with a trait. Analysis of a natural mutant in Arabidopsis has shown that mutations in cinnamoyl CoA reductase (CCR), a key lignin gene, affect physical properties of the secondary cell wall such as stiffness and strength. Using this gene, we tested whether LD mapping could identify alleles associated with microfibril angle (MFA), a wood quality trait affecting stiffness and strength of wood. We identified 25 common single-nucleotide polymorphism (SNP) markers in the CCR gene in Eucalyptus nitens. Using single-marker and haplotype analyses in 290 trees from a E. nitens natural population, two haplotypes significantly associated with MFA were found. These results were confirmed in two full-sib families of E. nitens and Eucalyptus globulus. In an effort to understand the functional significance of the SNP markers, we sequenced the cDNA clones and identified an alternatively spliced variant from the significant haplotype region. This study demonstrates that LD mapping can be used to identify alleles associated with wood quality traits in natural populations of trees.

Forest trees are ideally suited to association mapping due to their high levels of diversity and low genomic linkage disequilibrium. Using an association mapping approach, single-nucleotide polymorphism (SNP) markers influencing quantitative variation in wood quality were identified in a natural population of Pinus radiata. Of 149 sites examined, 10 demonstrated significant associations (P , 0.05, q , 0.1) with one or more traits after accounting for population structure and experimentwise error. Without accounting for marker interactions, phenotypic variation attributed to individual SNPs ranged from 2 to 6.5%. Undesirable negative correlations between wood quality and growth were not observed, indicating potential to break negative correlations by selecting for individual SNPs in breeding programs. Markers that yielded significant associations were reexamined in an Australian land race. SNPs from three genes (PAL1, PCBER, and SUSY) yielded significant associations. Importantly, associations with two of these genes validated associations with density previously observed in the discovery population. In both cases, decreased wood density was associated with the minor allele, suggesting that these SNPs may be under weak negative purifying selection for density in the natural populations. These results demonstrate the utility of LD mapping to detect associations, even when the power to detect SNPs with small effect is anticipated to be low. N UMEROUS traits of agronomic importance are demonstrated to be under genetic control (Keurentjes et al. 2008), and there is considerable interest in characterizing the causative polymorphisms underlying their quantitative variation. In forest trees, quantitative variation in traits such as mechanical and pulping properties of wood, growth, cold hardiness, and drought acclimation are likely to result from allelic variation within multiple genes (Neale and Savolainen 2004;Oraguzie and Wilcox 2007). Because of the commercial importance of radiata pine (Pinus radiata D. Don), we are exploring the molecular basis of variation in its wood properties using an association genetics approach.Radiata pine wood properties are variable in domesticated populations and exhibit a quantitative mode of inheritance with high heritability, indicating a strong underlying genetic component. High heritabilities have been observed for solid wood traits including density, cellulose microfibril angle (MFA), and modulus of elasticy (MOE) (Baltunis et al. 2007) and for carbohydrate composition, pulp yield, fiber length, and perimeter (Evans et al. 1997;Kibblewhite 1999); and genetic control has been established for several important production traits (Kumar 2004;Dungey et al. 2006;Gapare et al. 2006;Wu et al. 2007).It is anticipated that variation in wood quality will be dependent on variation in numerous genes involved in xylogenesis. Studies of gene expression in developing xylem have revealed genes involved in spatially and temporally regulated processes such as cambial division, cell different...

A large full-sib family of radiata pine ( Pinus radiata Donn. ex D. Don) was used for quantitative trait locus (QTL) detection and independent verification. QTL detection experiments were carried out for juvenile wood density (JWD) and stem diameter at breast height (DBH) using selective genotyping. Evenly spaced RFLP and microsatellite markers were selected from an existing linkage map. QTLs were verified in an independent set of progeny from the same family. Based on map location, at least eight QTL positions for JWD and two for DBH were detected and verified. The percent variance accounted for by the markers ranged from 0.78% to 3.58%, suggesting a genomic architecture of many genes with small effect. Two unrelated "bridging" families were chosen as candidates for marker-aided selection (MAS), and six microsatellite markers showing an association with JWD or DBH were tested in these families. Of these, four markers showed a consistent association with JWD in one or both of the bridging families. Results from this study provide a basis for MAS in P. radiata.

Six related radiata pine ( Pinus radiata) full-sib families were used to detect and independently verify quantitative trait loci (QTLs) for resistance to Dothistroma needle blight, caused by Dothistroma septospora. The detection families had from 26 to 30 individuals each, and had either a common maternal (31053) or paternal (31032) parent; one family (cross 4) consisted of progeny from both parents, 31053 x 31032. Approximately 200 additional progeny from cross 4 were clonally replicated and planted at two sites, with at least five to seven ramets of each individual per site. Marker segregation data were collected from a total of 250 RFLP and microsatellite markers, and single factor ANOVAs were conducted separately for each family and marker. A number of putative associations were observed, some across more than one family. Permutation tests were used to confirm expected probabilities of multiple associations based on chance alone. Seven markers representing at least four QTLs for resistance to Dothistroma were identified as being significant in more than one family; one of these was significant at P<0.05 in three families and highly significant at P<0.01 in a fourth. Further confirmation was obtained by testing those markers that were significant in more than one of the detection families (or highly significant in cross 4) in the clonally replicated progeny from cross 4. Four QTL positions were verified in the clonal populations, with a total percent variation accounted for of 12.5.

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