The Chinese pine genome and methylome unveil key features of conifer evolution.

Conifers dominate the world's forest ecosystems and are the most widely planted tree species. Their giant and complex genomes present great challenges for assembling a complete reference genome for evolutionary and genomic studies. We present a 25.4-Gb chromosome-level assembly of Chinese pine (Pinus tabuliformis) and revealed that its genome size is mostly attributable to huge intergenic regions and long introns with high transposable element (TE) content. Large genes with long introns exhibited higher expressions levels. Despite a lack of recent whole-genome duplication, 91.2% of genes were duplicated through dispersed duplication, and expanded gene families are mainly related to stress responses, which may underpin conifers' adaptation, particularly in cold and/or arid conditions. The reproductive regulation network is distinct compared with angiosperms. Slow removal of TEs with high-level methylation may have contributed to genomic expansion. This study provides insights into conifer evolution and resources for advancing research on conifer adaptation and development.

Natural variation in the prolyl 4-hydroxylase gene PtoP4H9 contributes to perennial stem growth in Populus.

Perennial trees must maintain stem growth throughout their entire lifespan to progressively increase in size as they age. The overarching question of the molecular mechanisms that govern stem perennial growth in trees remains largely unanswered. Here we deciphered the genetic architecture that underlies perennial growth trajectories using genome-wide association studies (GWAS) for measures of growth traits across years in a natural population of Populus tomentosa. By analyzing the stem growth trajectory, we identified PtoP4H9, encoding prolyl 4-hydroxylase 9, which is responsible for the natural variation in the growth rate of diameter at breast height (DBH) across years. Quantifying the dynamic genetic contribution of PtoP4H9 loci to stem growth showed that PtoP4H9 played a pivotal role in stem growth regulation. Spatiotemporal expression analysis showed that PtoP4H9 was highly expressed in cambium tissues of poplars of various ages. Overexpression and knockdown of PtoP4H9 revealed that it altered cell expansion to regulate cell wall modification and mechanical characteristics, thereby promoting stem growth in Populus. We showed that natural variation in PtoP4H9 occurred in a BASIC PENTACYSTEINE transcription factor PtoBPC1-binding promoter element controlling PtoP4H9 expression. The geographic distribution of PtoP4H9 allelic variation was consistent with the modes of selection among populations. Altogether, our study provides important genetic insights into dynamic stem growth in Populus, and we confirmed PtoP4H9 as a potential useful marker for breeding or genetic engineering of poplars.

Whole-genome resequencing facilitates the development of a 50K single nucleotide polymorphism genotyping array for Scots pine (Pinus sylvestris L.) and its transferability to other pine species.

Scots pine (Pinus sylvestris L.) is one of the most widespread and economically important conifer species in the world. Applications like genomic selection and association studies, which could help accelerate breeding cycles, are challenging in Scots pine because of its large and repetitive genome. For this reason, genotyping tools for conifer species, and in particular for Scots pine, are commonly based on transcribed regions of the genome. In this article, we present the Axiom Psyl50K array, the first single nucleotide polymorphism (SNP) genotyping array for Scots pine based on whole-genome resequencing, that represents both genic and intergenic regions. This array was designed following a two-step procedure: first, 192 trees were sequenced, and a 430K SNP screening array was constructed. Then, 480 samples, including haploid megagametophytes, full-sib family trios, breeding population, and range-wide individuals from across Eurasia were genotyped with the screening array. The best 50K SNPs were selected based on quality, replicability, distribution across the draft genome assembly, balance between genic and intergenic regions, and genotype-environment and genotype-phenotype associations. Of the final 49 877 probes tiled in the array, 20 372 (40.84%) occur inside gene models, while the rest lie in intergenic regions. We also show that the Psyl50K array can yield enough high-confidence SNPs for genetic studies in pine species from North America and Eurasia. This new genotyping tool will be a valuable resource for high-throughput fundamental and applied research of Scots pine and other pine species.

A transcriptome-based association study of growth, wood quality, and oleoresin traits in a slash pine breeding population.

Slash pine (Pinus elliottii Engelm.) is an important timber and resin species in the United States, China, Brazil and other countries. Understanding the genetic basis of these traits will accelerate its breeding progress. We carried out a genome-wide association study (GWAS), transcriptome-wide association study (TWAS) and weighted gene co-expression network analysis (WGCNA) for growth, wood quality, and oleoresin traits using 240 unrelated individuals from a Chinese slash pine breeding population. We developed high quality 53,229 single nucleotide polymorphisms (SNPs). Our analysis reveals three main results: (1) the Chinese breeding population can be divided into three genetic groups with a mean inbreeding coefficient of 0.137; (2) 32 SNPs significantly were associated with growth and oleoresin traits, accounting for the phenotypic variance ranging from 12.3% to 21.8% and from 10.6% to 16.7%, respectively; and (3) six genes encoding PeTLP, PeAP2/ERF, PePUP9, PeSLP, PeHSP, and PeOCT1 proteins were identified and validated by quantitative real time polymerase chain reaction for their association with growth and oleoresin traits. These results could be useful for tree breeding and functional studies in advanced slash pine breeding program.

Preselection of QTL markers enhances accuracy of genomic selection in Norway spruce.

Genomic prediction (GP) or genomic selection is a method to predict the accumulative effect of all quantitative trait loci (QTLs) in a population by estimating the realized genomic relationships between the individuals and by capturing the linkage disequilibrium between markers and QTLs. Thus, marker preselection is considered a promising method to capture Mendelian segregation effects. Using QTLs detected in a genome-wide association study (GWAS) may improve GP. Here, we performed GWAS and GP in a population with 904 clones from 32 full-sib families using a newly developed 50 k SNP Norway spruce array. Through GWAS we identified 41 SNPs associated with budburst stage (BB) and the largest effect association explained 5.1% of the phenotypic variation (PVE). For the other five traits such as growth and wood quality traits, only 2 - 13 associations were observed and the PVE of the strongest effects ranged from 1.2% to 2.0%. GP using approximately 100 preselected SNPs, based on the smallest p-values from GWAS showed the greatest predictive ability (PA) for the trait BB. For the other traits, a preselection of 2000-4000 SNPs, was found to offer the best model fit according to the Akaike information criterion being minimized. But PA-magnitudes from GP using such selections were still similar to that of GP using all markers. Analyses on both real-life and simulated data also showed that the inclusion of a large QTL SNP in the model as a fixed effect could improve PA and accuracy of GP provided that the PVE of the QTL was ≥ 2.5%.

Genome-wide TCP transcription factors analysis provides insight into their new functions in seasonal and diurnal growth rhythm in Pinus tabuliformis.

BACKGROUND: Pinus tabuliformis adapts to cold climate with dry winter in northern China, serving as important commercial tree species. The TEOSINTE BRANCHED 1, CYCLOIDEA, and PROLIFERATING CELL FACTOR family(TCP)transcription factors were found to play a role in the circadian clock system in Arabidopsis. However, the role of TCP transcription factors in P. tabuliformis remains little understood. RESULTS: In the present study, 43 TCP genes were identified from P. tabuliformis genome database. Based on the phylogeny tree and sequence similarity, the 43 TCP genes were classified into four groups. The motif results showed that different subfamilies indeed contained different motifs. Clade II genes contain motif 1, clade I genes contain motif 1, 8, 10 and clade III and IV contain more motifs, which is consistent with our grouping results. The structural analysis of PtTCP genes showed that most PtTCPs lacked introns. The distribution of clade I and clade II on the chromosome is relatively scattered, while clade III and clade IV is relatively concentrated. Co-expression network indicated that PtTCP2, PtTCP12, PtTCP36, PtTCP37, PtTCP38, PtTCP41 and PtTCP43 were co-expressed with clock genes in annual cycle and their annual cycle expression profiles both showed obvious seasonal oscillations. PtTCP2, PtTCP12, PtTCP37, PtTCP38, PtTCP40, PtTCP41, PtTCP42 and PtTCP43 were co-expressed with clock genes in diurnal cycle. Only the expression of PtTCP42 showed diurnal oscillation. CONCLUSIONS: The TCP gene family, especially clade II, may play an important role in the regulation of the season and circadian rhythm of P. tabuliformis. In addition, the low temperature in winter may affect the diurnal oscillations.

Teasing apart the joint effect of demography and natural selection in the birth of a contact zone.

Vast population movements induced by recurrent climatic cycles have shaped the genetic structure of plant species. During glacial periods species were confined to low-latitude refugia from which they recolonized higher latitudes as the climate improved. This multipronged recolonization led to many lineages that later met and formed large contact zones. We utilize genomic data from 5000 Picea abies trees to test for the presence of natural selection during recolonization and establishment of a contact zone in Scandinavia. Scandinavian P. abies is today made up of a southern genetic cluster originating from the Baltics, and a northern one originating from Northern Russia. The contact zone delineating them closely matches the limit between two major climatic regions. We show that natural selection contributed to its establishment and maintenance. First, an isolation-with-migration model with genome-wide linked selection fits the data better than a purely neutral one. Second, many loci show signatures of selection or are associated with environmental variables. These loci, regrouped in clusters on chromosomes, are often related to phenology. Altogether, our results illustrate how climatic cycles, recolonization and selection can establish strong local adaptation along contact zones and affect the genetic architecture of adaptive traits.

MADS-box transcription factors MADS11 and DAL1 interact to mediate the vegetative-to-reproductive transition in pine.

The reproductive transition is an important event that is crucial for plant survival and reproduction. Relative to the thorough understanding of the vegetative phase transition in angiosperms, a little is known about this process in perennial conifers. To gain insight into the molecular basis of the regulatory mechanism in conifers, we used temporal dynamic transcriptome analysis with samples from seven different ages of Pinus tabuliformis to identify a gene module substantially associated with aging. The results first demonstrated that the phase change in P. tabuliformis occurred as an unexpectedly rapid transition rather than a slow, gradual progression. The age-related gene module contains 33 transcription factors and was enriched in genes that belong to the MADS (MCMl, AGAMOUS, DEFICIENS, SRF)-box family, including six SOC1-like genes and DAL1 and DAL10. Expression analysis in P. tabuliformis and a late-cone-setting P. bungeana mutant showed a tight association between PtMADS11 and reproductive competence. We then confirmed that MADS11 and DAL1 coordinate the aging pathway through physical interaction. Overexpression of PtMADS11 and PtDAL1 partially rescued the flowering of 35S::miR156A and spl1,2,3,4,5,6 mutants in Arabidopsis (Arabidopsis thaliana), but only PtMADS11 could rescue the flowering of the ft-10 mutant, suggesting PtMADS11 and PtDAL1 play different roles in flowering regulatory networks in Arabidopsis. The PtMADS11 could not alter the flowering phenotype of soc1-1-2, indicating it may function differently from AtSOC1 in Arabidopsis. In this study, we identified the MADS11 gene in pine as a regulatory mediator of the juvenile-to-adult transition with functions differentiated from the angiosperm SOC1.

Killing two enemies with one stone? Genomics of resistance to two sympatric pathogens in Norway spruce.

Trees must cope with the attack of multiple pathogens, often simultaneously during their long lifespan. Ironically, the genetic and molecular mechanisms controlling this process are poorly understood. The objective of this study was to compare the genetic component of resistance in Norway spruce to Heterobasidion annosum s.s. and its sympatric congener Heterobasidion parviporum. Heterobasidion root- and stem-rot is a major disease of Norway spruce caused by members of the Heterobasidion annosum species complex. Resistance to both pathogens was measured using artificial inoculations in half-sib families of Norway spruce trees originating from central to northern Europe. The genetic component of resistance was analysed using 63,760 genome-wide exome-capture sequenced SNPs and multitrait genome-wide associations. No correlation was found for resistance to the two pathogens; however, associations were found between genomic variants and resistance traits with synergic or antagonist pleiotropic effects to both pathogens. Additionally, a latitudinal cline in resistance in the bark to H. annosum s.s. was found; trees from southern latitudes, with a later bud-set and thicker stem diameter, allowed longer lesions, but this was not the case for H. parviporum. In summary, this study detects genomic variants with pleiotropic effects which explain multiple disease resistance from a genic level and could be useful for selection of resistant trees to both pathogens. Furthermore, it highlights the need for additional research to understand the evolution of resistance traits to multiple pathogens in trees.

The Transcriptional Landscape and Hub Genes Associated with Physiological Responses to Drought Stress in Pinus tabuliformis.

Drought stress has an extensive impact on regulating various physiological, metabolic, and molecular responses. In the present study, the Pinus tabuliformis transcriptome was studied to evaluate the drought-responsive genes using RNA- Sequencing approache. The results depicted that photosynthetic rate and H2O conductance started to decline under drought but recovered 24 h after re-watering; however, the intercellular CO2 concentration (Ci) increased with the onset of drought. We identified 84 drought-responsive transcription factors, 62 protein kinases, 17 transcriptional regulators, and 10 network hub genes. Additionally, we observed the expression patterns of several important gene families, including 2192 genes positively expressed in all 48 samples, and 40 genes were commonly co-expressed in all drought and recovery stages compared with the control samples. The drought-responsive transcriptome was conserved mainly between P. tabuliformis and A. thaliana, as 70% (6163) genes had a homologous in arabidopsis, out of which 52% homologous (3178 genes corresponding to 2086 genes in Arabidopsis) were also drought response genes in arabidopsis. The collaborative network exhibited 10 core hub genes integrating with ABA-dependent and independent pathways closely conserved with the ABA signaling pathway in the transcription factors module. PtNCED3 from the ABA family genes had shown significantly different expression patterns under control, mild, prolonged drought, and recovery stages. We found the expression pattern was considerably increased with the prolonged drought condition. PtNCED3 highly expressed in all drought-tested samples; more interestingly, expression pattern was higher under mild and prolonged drought. PtNCED3 is reported as one of the important regulating enzymes in ABA synthesis. The continuous accumulation of ABA in leaves increased resistance against drought was due to accumulation of PtNCED3 under drought stress in the pine needles.

Evaluation of the efficiency of genomic versus pedigree predictions for growth and wood quality traits in Scots pine.

BACKGROUND: Genomic selection (GS) or genomic prediction is a promising approach for tree breeding to obtain higher genetic gains by shortening time of progeny testing in breeding programs. As proof-of-concept for Scots pine (Pinus sylvestris L.), a genomic prediction study was conducted with 694 individuals representing 183 full-sib families that were genotyped with genotyping-by-sequencing (GBS) and phenotyped for growth and wood quality traits. 8719 SNPs were used to compare different genomic with pedigree prediction models. Additionally, four prediction efficiency methods were used to evaluate the impact of genomic breeding value estimations by assigning diverse ratios of training and validation sets, as well as several subsets of SNP markers. RESULTS: Genomic Best Linear Unbiased Prediction (GBLUP) and Bayesian Ridge Regression (BRR) combined with expectation maximization (EM) imputation algorithm showed slightly higher prediction efficiencies than Pedigree Best Linear Unbiased Prediction (PBLUP) and Bayesian LASSO, with some exceptions. A subset of approximately 6000 SNP markers, was enough to provide similar prediction efficiencies as the full set of 8719 markers. Additionally, prediction efficiencies of genomic models were enough to achieve a higher selection response, that varied between 50-143% higher than the traditional pedigree-based selection. CONCLUSIONS: Although prediction efficiencies were similar for genomic and pedigree models, the relative selection response was doubled for genomic models by assuming that earlier selections can be done at the seedling stage, reducing the progeny testing time, thus shortening the breeding cycle length roughly by 50%.

Effect of number of annual rings and tree ages on genomic predictive ability for solid wood properties of Norway spruce.

BACKGROUND: Genomic selection (GS) or genomic prediction is considered as a promising approach to accelerate tree breeding and increase genetic gain by shortening breeding cycle, but the efforts to develop routines for operational breeding are so far limited. We investigated the predictive ability (PA) of GS based on 484 progeny trees from 62 half-sib families in Norway spruce (Picea abies (L.) Karst.) for wood density, modulus of elasticity (MOE) and microfibril angle (MFA) measured with SilviScan, as well as for measurements on standing trees by Pilodyn and Hitman instruments. RESULTS: GS predictive abilities were comparable with those based on pedigree-based prediction. Marker-based PAs were generally 25-30% higher for traits density, MFA and MOE measured with SilviScan than for their respective standing tree-based method which measured with Pilodyn and Hitman. Prediction accuracy (PC) of the standing tree-based methods were similar or even higher than increment core-based method. 78-95% of the maximal PAs of density, MFA and MOE obtained from coring to the pith at high age were reached by using data possible to obtain by drilling 3-5 rings towards the pith at tree age 10-12. CONCLUSIONS: This study indicates standing tree-based measurements is a cost-effective alternative method for GS. PA of GS methods were comparable with those pedigree-based prediction. The highest PAs were reached with at least 80-90% of the dataset used as training set. Selection for trait density could be conducted at an earlier age than for MFA and MOE. Operational breeding can also be optimized by training the model at an earlier age or using 3 to 5 outermost rings at tree age 10 to 12 years, thereby shortening the cycle and reducing the impact on the tree.

Genotypic variation in Norway spruce correlates to fungal communities in vegetative buds.

The taxonomically diverse phyllosphere fungi inhabit leaves of plants. Thus, apart from the fungi's dispersal capacities and environmental factors, the assembly of the phyllosphere community associated with a given host plant depends on factors encoded by the host's genome. The host genetic factors and their influence on the assembly of phyllosphere communities under natural conditions are poorly understood, especially in trees. Recent work indicates that Norway spruce (Picea abies) vegetative buds harbour active fungal communities, but these are hitherto largely uncharacterized. This study combines internal transcribed spacer sequencing of the fungal communities associated with dormant vegetative buds with a genome-wide association study (GWAS) in 478 unrelated Norway spruce trees. The aim was to detect host loci associated with variation in the fungal communities across the population, and to identify loci correlating with the presence of specific, latent, pathogens. The fungal communities were dominated by known Norway spruce phyllosphere endophytes and pathogens. We identified six quantitative trait loci (QTLs) associated with the relative abundance of the dominating taxa (i.e., top 1% most abundant taxa). Three additional QTLs associated with colonization by the spruce needle cast pathogen Lirula macrospora or the cherry spruce rust (Thekopsora areolata) in asymptomatic tissues were detected. The identification of the nine QTLs shows that the genetic variation in Norway spruce influences the fungal community in dormant buds and that mechanisms underlying the assembly of the communities and the colonization of latent pathogens in trees may be uncovered by combining molecular identification of fungi with GWAS.

Association genetics identifies a specifically regulated Norway spruce laccase gene, PaLAC5, linked to Heterobasidion parviporum resistance.

It is important to improve the understanding of the interactions between the trees and pathogens and integrate this knowledge about disease resistance into tree breeding programs. The conifer Norway spruce (Picea abies) is an important species for the forest industry in Europe. Its major pathogen is Heterobasidion parviporum, causing stem and root rot. In this study, we identified 11 Norway spruce QTLs (Quantitative trait loci) that correlate with variation in resistance to H. parviporum in a population of 466 trees by association genetics. Individual QTLs explained between 2.1 and 5.2% of the phenotypic variance. The expression of candidate genes associated with the QTLs was analysed in silico and in response to H. parviporum hypothesizing that (a) candidate genes linked to control of fungal sapwood growth are more commonly expressed in sapwood, and; (b) candidate genes associated with induced defences are respond to H. parviporum inoculation. The Norway spruce laccase PaLAC5 associated with control of lesion length development is likely to be involved in the induced defences. Expression analyses showed that PaLAC5 responds specifically and strongly in close proximity to the H. parviporum inoculation. Thus, PaLAC5 may be associated with the lignosuberized boundary zone formation in bark adjacent to the inoculation site.

Genetic Improvement of Sawn-Board Stiffness and Strength in Scots Pine (Pinus sylvestris L.).

Given an overall aim of improving Scots pine structural wood quality by selective tree breeding, we investigated the potential of non-destructive acoustic sensing tools to accurately predict wood stiffness (modulus of elasticity, MOE) and strength (modulus of rupture, MOR) of sawn boards. Non-destructive measurements of wood density (DEN), acoustic velocity (VEL) and MOE were carried out at different stages of wood processing chain (standing trees, felled logs and sawn boards), whilst destructively measured stiffness and strength served as benchmark traits. All acoustic based MOE and VEL estimates proved to be good proxies (rA > 0.65) for sawn-board stiffness while MOETREE, VELHIT and resistograph wood density (DENRES) measured on standing trees and MOELOG and VELFAK measured on felled logs well reflected board strength. Individual-tree narrow-sense heritability ( h i 2 ) for VEL, MOE and MOR were weak (0.05-0.26) but were substantially stronger for wood density (0.34-0.40). Moreover, additive genetic coefficients of variation for MOE and MOR were in the range from 5.4% to 9.1%, offering potential targets for exploitation by selective breeding. Consequently, selective breeding based on MOETREE, DENRES or stem straightness (STR) could improve several structural wood traits simultaneously.

Analysis of phenotypic- and Estimated Breeding Values (EBV) to dissect the genetic architecture of complex traits in a Scots pine three-generation pedigree design.

In forest tree breeding, family-based Quantitative Trait Loci (QTL) studies are valuable as methods to dissect the complexity of a trait and as a source of candidate genes. In the field of conifer research, our study contributes to the evaluation of phenotypic and predicted breeding values for the identification of QTL linked to complex traits in a three-generation pedigree population in Scots pine (Pinus sylvestris L.). A total of 11 470 open pollinated F2-progeny trees established at three different locations, were measured for growth and adaptive traits. Breeding values were predicted for their 360 mothers, originating from a single cross of two grand-parents. A multilevel LASSO association analysis was conducted to detect QTL using genotypes of the mothers with the corresponding phenotypes and Estimated Breeding Values (EBV). Different levels of genotype-by-environment (G × E) effects among sites at different years, were detected for survival and height. Moderate-to-low narrow sense heritabilities and EBV accuracies were found for all traits and all sites. We identified 18 AFLPs and 12 SNPs to be associated with QTL for one or more traits. 62 QTL were significant with percentages of variance explained ranging from 1.7 to 18.9%. In those cases where the same marker was associated to a phenotypic or an ebvQTL, the ebvQTL always explained higher proportion of the variance, maybe due to the more accurate nature of Estimated Breeding Values (EBV). Two SNP-QTL showed pleiotropic effects for traits related with hardiness, seed, cone and flower production. Furthermore, we detected several QTL with significant effects across multiple ages, which could be considered as strong candidate loci for early selection. The lack of reproducibility of some QTL detected across sites may be due to environmental heterogeneity reflected by the genotype- and QTL-by-environment effects.

Increased Prediction Ability in Norway Spruce Trials Using a Marker X Environment Interaction and Non-Additive Genomic Selection Model.

A genomic selection study of growth and wood quality traits is reported based on control-pollinated Norway spruce families established in 2 Northern Swedish trials at 2 locations using exome capture as a genotyping platform. Nonadditive effects including dominance and first-order epistatic interactions (including additive-by-additive, dominance-by-dominance, and additive-by-dominance) and marker-by-environment interaction (M×E) effects were dissected in genomic and phenotypic selection models. Genomic selection models partitioned additive and nonadditive genetic variances more precisely than pedigree-based models. In addition, predictive ability in GS was substantially increased by including dominance and slightly increased by including M×E effects when these effects are significant. For velocity, response to genomic selection per year increased up to 78.9/80.8%, 86.9/82.9%, and 91.3/88.2% compared with response to phenotypic selection per year when genomic selection was based on 1) main marker effects (M), 2) M + M×E effects (A), and 3) A + dominance effects (AD) for sites 1 and 2, respectively. This indicates that including M×E and dominance effects not only improves genetic parameter estimates but also when they are significant may improve the genetic gain. For tree height, Pilodyn, and modulus of elasticity (MOE), response to genomic selection per year improved up to 68.9%, 91.3%, and 92.6% compared with response to phenotypic selection per year, respectively.Subject Area: Quantitative genetics and Mendelian inheritance.

Accuracy of genomic selection for growth and wood quality traits in two control-pollinated progeny trials using exome capture as the genotyping platform in Norway spruce.

BACKGROUND: Genomic selection (GS) can increase genetic gain by reducing the length of breeding cycle in forest trees. Here we genotyped 1370 control-pollinated progeny trees from 128 full-sib families in Norway spruce (Picea abies (L.) Karst.), using exome capture as genotyping platform. We used 116,765 high-quality SNPs to develop genomic prediction models for tree height and wood quality traits. We assessed the impact of different genomic prediction methods, genotype-by-environment interaction (G × E), genetic composition, size of the training and validation set, relatedness, and number of SNPs on accuracy and predictive ability (PA) of GS. RESULTS: Using G matrix slightly altered heritability estimates relative to pedigree-based method. GS accuracies were about 11-14% lower than those based on pedigree-based selection. The efficiency of GS per year varied from 1.71 to 1.78, compared to that of the pedigree-based model if breeding cycle length was halved using GS. Height GS accuracy decreased to more than 30% while using one site as training for GS prediction and using this model to predict the second site, indicating that G × E for tree height should be accommodated in model fitting. Using a half-sib family structure instead of full-sib structure led to a significant reduction in GS accuracy and PA. The full-sib family structure needed only 750 markers to reach similar accuracy and PA, as compared to 100,000 markers required for the half-sib family, indicating that maintaining the high relatedness in the model improves accuracy and PA. Using 4000-8000 markers in full-sib family structure was sufficient to obtain GS model accuracy and PA for tree height and wood quality traits, almost equivalent to that obtained with all markers. CONCLUSIONS: The study indicates that GS would be efficient in reducing generation time of breeding cycle in conifer tree breeding program that requires long-term progeny testing. The sufficient number of trees within-family (16 for growth and 12 for wood quality traits) and number of SNPs (8000) are required for GS with full-sib family relationship. GS methods had little impact on GS efficiency for growth and wood quality traits. GS model should incorporate G × E effect when a strong G × E is detected.

Implications of accounting for marker-based population structure in the quantitative genetic evaluation of genetic parameters related to growth and wood properties in Norway spruce.

BACKGROUND: Forest geneticists typically use provenances to account for population differences in their improvement schemes; however, the historical records of the imported materials might not be very precise or well-aligned with the genetic clusters derived from advanced molecular techniques. The main objective of this study was to assess the impact of marker-based population structure on genetic parameter estimates related to growth and wood properties and their trade-offs in Norway spruce, by either incorporating it as a fixed effect (model-A) or excluding it entirely from the analysis (model-B). RESULTS: Our results indicate that models incorporating population structure significantly reduce estimates of additive genetic variance, resulting in substantial reduction of narrow-sense heritability. However, these models considerably improve prediction accuracies. This was particularly significant for growth and solid-wood properties, which showed to have the highest population genetic differentiation (QST) among the studied traits. Additionally, although the pattern of correlations remained similar across the models, their magnitude was slightly lower for models that included population structure as a fixed effect. This suggests that selection, consistently performed within populations, might be less affected by unfavourable genetic correlations compared to mass selection conducted without pedigree restrictions. CONCLUSION: We conclude that the results of models properly accounting for population structure are more accurate and less biased compared to those neglecting this effect. This might have practical implications for breeders and forest managers where, decisions based on imprecise selections can pose a high risk to economic efficiency.

Transcriptome profiling of radiata pine branches reveals new insights into reaction wood formation with implications in plant gravitropism.

BACKGROUND: Formation of compression (CW) and opposite wood (OW) in branches and bent trunks is an adaptive feature of conifer trees in response to various displacement forces, such as gravity, wind, snow and artificial bending. Several previous studies have characterized tracheids, wood and gene transcription in artificially or naturally bent conifer trunks. These studies have provided molecular basis of reaction wood formation in response to bending forces and gravity stimulus. However, little is known about reaction wood formation and gene transcription in conifer branches under gravity stress. In this study SilviScan® technology was used to characterize tracheid and wood traits in radiate pine (Pinus radiata D. Don) branches and genes differentially transcribed in CW and OW were investigated using cDNA microarrays. RESULTS: CW drastically differed from OW in tracheids and wood traits with increased growth, thicker tracheid walls, larger microfibril angle (MFA), higher density and lower stiffness. However, CW and OW tracheids had similar diameters in either radial or tangential direction. Thus, gravity stress largely influenced wood growth, secondary wall deposition, cellulose microfibril orientation and wood properties, but had little impact on primary wall expansion. Microarray gene transcription revealed about 29% of the xylem transcriptomes were significantly altered in CW and OW sampled in both spring and autumn, providing molecular evidence for the drastic variation in tracheid and wood traits. Genes involved in cell division, cellulose biosynthesis, lignin deposition, and microtubules were mostly up-regulated in CW, conferring its greater growth, thicker tracheid walls, higher density, larger MFA and lower stiffness. However, genes with roles in cell expansion and primary wall formation were differentially transcribed in CW and OW, respectively, implicating their similar diameters of tracheid walls and different tracheid lengths. Interestingly, many genes related to hormone and calcium signalling as well as various environmental stresses were exclusively up-regulated in CW, providing important clues for earlier molecular signatures of reaction wood formation under gravity stimulus. CONCLUSIONS: The first comprehensive investigation of tracheid characteristics, wood properties and gene transcription in branches of a conifer species revealed more accurate and new insights into reaction wood formation in response to gravity stress. The identified differentially transcribed genes with diverse functions conferred or implicated drastic CW and OW variation observed in radiata pine branches. These genes are excellent candidates for further researches on the molecular mechanisms of reaction wood formation with a view to plant gravitropism.