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Forest trees cultivars developed by breeders have been increasingly deployed as clones, following long generations of breeding and testing. An established protocol for distinctiveness, uniformity and stability (DUS) testing becomes an essential element for protecting the intellectual property rights associated with these clones. DUS testing with morphological descriptors has, however, shown limitations in categorically distinguishing cultivars, especially with narrow genetic base. DNA fingerprinting based on microsatellite markers has been a powerful tool to discriminate clones. Teak (Tectona grandis) is an economically valuable exotic timber species planted in Brazil. We assessed the individual and combined performance of a selected set of 21 teak microsatellites for identity analysis and parentage testing in a sample of 50 clones planted in Brazil. The clones displayed high genetic diversity attributable to their wide provenance origin. The 21 microsatellites combined provide high power of individual identification with a combined probability of identity of 1.84E-23 for unrelated individuals, and 4.82E-09 for full-sibs, and a power of paternity exclusion higher than 99.99999 % in all testing scenarios. Variable subsets of these 21 markers still provide abundant power of discrimination, although a recommended set should include a minimum of 12 markers with higher information content and reliable genotyping performance. Relatedness and genetic distance analyses revealed unexpected identities or significant recent shared ancestry among otherwise considered unique clones. These results advocate for the importance of including DNA markers, at least as discretionary complementary descriptors to the 22 morphological traits currently adopted for plant variety protection of teak cultivars in Brazil.

The role of natural selection in shaping patterns of diversity is still poorly understood in the Neotropics. We carried out the first genome-wide population genomics study in a Neotropical tree, Handroanthus impetiginosus(Bignoniaceae), sampling 75,838 SNPs by sequence capture in 128 individuals across 13 populations. We found evidences for local adaptation using Bayesian correlations of allele frequency and environmental variables (32 loci in 27 genes) complemented by an analysis of selective sweeps and genetic hitchhiking events using SweepFinder2 (81 loci in 47 genes). Fifteen genes were identified by both approaches. By accounting for population genetic structure, we also found 14 loci with selection signal in a STRUCTURE-defined lineage comprising individuals from five populations, using Outflank. All approaches pinpointed highly diverse and structurally conserved genes affecting plant development and primary metabolic processes. Spatial interpolation forecasted differences in the expected allele frequencies at loci under selection over time, suggesting that H. impetiginosusmay track its habitat during climate changes. However, local adaptation through natural selection may also take place, allowing species persistence due to niche evolution. A high genetic differentiation was seen among the H. impetiginosuspopulations, which, together with the limited power of the experiment, constrains the improved detection of other types of soft selective forces, such as background, balanced, and purifying selection. Small differences in allele frequency distribution among widespread populations and the low number of loci with detectable adaptive sweeps advocate for a polygenic model of adaptation involving a potentially large number of small genome-wide effects.

Terpenes are economically and ecologically important phytochemicals. Their synthesis is controlled by the terpene synthase(TPS) gene family, which is highly diversified throughout the plant kingdom. The plant family Myrtaceae are characterised by especially high terpene concentrations, and considerable variation in terpene profiles. Many Myrtaceae are grown commercially for terpene products including the eucalypts Corymbiaand Eucalyptus. Eucalyptus grandishas the largest TPSgene family of plants currently sequenced, which is largely conserved in the closely related E. globulus. However, the TPSgene family has been well studied only in these two eucalypt species. The recent assembly of two Corymbia citriodorasubsp. variegatagenomes presents an opportunity to examine the conservation of this important gene family across more divergent eucalypt lineages. Manual annotation of the TPSgene family in C. citriodorasubsp. variegatarevealed a similar overall number, and relative subfamily representation, to that previously reported in E. grandisand E. globulus. Many of the TPSgenes were in physical clusters that varied considerably between Eucalyptusand Corymbia, with several instances of translocation, expansion/contraction and loss. Notably, there was greater conservation in the subfamilies involved in primary metabolism than those involved in secondary metabolism, likely reflecting different selective constraints. The variation in cluster size within subfamilies and the broad conservation between the eucalypts in the face of this variation are discussed, highlighting the potential contribution of selection, concerted evolution and stochastic processes. These findings provide the foundation to better understand terpene evolution within the ecologically and economically important Myrtaceae.

Abstract: Genomic selection (GS) involves selection decisions based on genomic breeding values estimated as the sum of the effects of genome-wide markers capturing most quantitative trait loci (QTL) for the target trait(s). GS is revolutionizing breeding practice in domestic animals. The same approach and concepts can be readily applied to forest tree breeding where long generation times and late expressing complex traits are also a challenge. GS in forest trees would have additional advantages: large training populations can be easily assembled and accurately phenotyped for several traits, and the extent of linkage disequilibrium (LD) can be high in elite populations with small effective population size (N e) frequently used in advanced forest tree breeding programs. Deterministic equations were used to assess the impact of LD (modeled by N e and intermarker distance), the size of the training set, trait heritability, and the number of QTL on the predicted accuracy of GS. Results indicate that GS has the potential to radically improve the efficiency of tree breeding. The benchmark accuracy of conventional BLUP selection is reached by GS even at a marker density ~2 markers/cM when N e ≤ 30, while up to 20 markers/cM are necessary for larger N e. Shortening the breeding cycle by 50% with GS provides an increase ≥100% in selection efficiency. With the rapid technological advances and declining costs of genotyping, our cautiously optimistic outlook is that GS has great potential to accelerate tree breeding. However, further simulation studies and proof-of-concept experiments of GS are needed before recommending it for operational implementation.

Abstract: Eucalypts are keystone species in their natural ranges and are extensively planted worldwide for high-quality woody biomass. A novel set of 21 polymorphic and interspecifically transferable microsatellite markers based on tetra-, penta- and hexanucleotide repeats were developed and tested for high-precision genotyping of species of Eucalyptus. These microsatellites were characterized in population samples of four species, Eucalyptus grandis, Eucalyptus globulus, Eucalyptus urophylla, and Eucalyptus camaldulensis, representing three phylogenetic sections of subgenus Symphyomyrtus. These markers provide a clear advantage for accurate allele calling due to their larger allele size difference. Two multiplexed microsatellite combinations, a 14-locus/four-dye and an 18-locus/five-dye set, analyzable in single lanes were designed, providing resolution and throughput analogous to those routinely used in human DNA profiling. This set of microsatellites was shown to have high resolution for clone fingerprinting, inter-individual genetic distance estimation, species distinction, and assignment of hybrid individuals to their most likely ancestral species. These systems will be particularly useful for comparative population genetics and molecular breeding applications that require consistent allele calling across different points in time or laboratories.

Abstract: In this report the major locus for Puccinia psidii rust resistance, Ppr1, was positioned on the reference genetic map for Eucalyptus. Additionally, its position was validated by association genetics in a related and two unrelated pedigrees involving different Eucalyptus grandis resistant trees crossed to individuals of two other species, Eucalyptus tereticornis and Eucalyptus camaldulensis. These results are consistent with the hypothesis that Ppr1 controls a large proportion of the variation for rust resistance, strengthening its role as a major locus in Eucalyptus and providing its unequivocal genomic position on linkage group 3. A localized map with 19 microsatellite loci was built around Ppr1. Multiallelic profiles were observed at several mapped microsatellites suggesting recent tandem duplications in the genomic landscape surrounding Ppr1. Markers EMBRA125 and EMBRA1071 flank Ppr1 at 9.5% and 7% recombination, respectively, and were found to be in linkage equilibrium in a E. grandis breeding population, consistent with the expectations in outcrossed Eucalyptus. Their potential use for MAS will specifically be directed to identifying resistant offspring of P. psidii resistant parent trees that are heterozygous at Ppr1. In these circumstances, a significant amount of LD is expected to occur between specific alleles at flanking microsatellites and the resistance allele at Ppr1. Moreover, the positional information of Ppr1 paves the way for prospective undertakings in this genomic region with the upcoming availability of a draft genome for E. grandis.

The genetic diversity of a comprehensive germplasm collection involving 192 Jatropha curcasL. accessions collected throughout Brazil, spanning a wide latitudinal range from the states of Maranhão (1°49′ S, 44°52′ W) to Rio Grande do Sul (29°33′ S, 55°07′ W), was studied with 96 random amplified polymorphic DNA (RAPD) primers and six selected microsatellite markers. Only 23 of the 381 replicated RAPD markers and one microsatellite were polymorphic. Surprisingly, all accessions were homozygous at all but one microsatellite, in contrast with the outcrossing mating system reported for the species, suggesting that J. curcasnot only supports selfing but possibly breeds by geitonogamy. Similarity based clustering revealed only 43 unique multilocus profiles in the 192 accessions. The probabilities of accessions with indistinguishable multilocus profiles being true duplicates varied between 83 and 99%. No relationship between clustering of accessions and geographic origin was observed, suggesting that J. curcashas experienced a widespread dispersion across regions by seeds and possibly vegetative propagules. The narrow genetic base and extent of potentially duplicated accessions likely reflects a recent common ancestry, drift, and intensive selection of the currently cultivated materials since the time of introduction. This result highlights an urgent need for the introduction of new and diverse accessions to this germplasm collection if Brazil is to drive and sustain successful breeding programs.

RAPD markers were used to investigate the distribution of genetic variability in a germplasm collection of the dioecious tropical tree Aroeira (Myracrodruon urundeuva) composed of open pollinated maternal half-sib families collected from trees at least 300 meters apart and grouped in nine collection areas throughout its geographical range in Brazil. Open pollinated families from nearby trees from two natural populations were also included in the study. A Principal Coordinate Analysis (PCO) based on genetic similarities estimated with 83 RAPD markers showed no defined clustering among individuals from the same collection areas. However, a matrix correlation test showed low, but significant correlation between genetic and geographic distances (r = 0.17; p < 0.02). An AMOVA showed that 92% of the genetic variation is found within collection areas. Significant variation was found both between regions and between areas within regions although only 3.54 and 4.44% of the variation was found at these levels respectively. An AMOVA on the individuals from the two natural populations resulted in a pattern of distribution congruent to the one found for the collection areas, with 97% of variability within populations. An analysis of 10 open-pollinated half-sib families revealed that between 75% and 89% of the variability is contained within families, suggesting that seeds from one tree represent a significant proportion of the variability found in the population. These results indicate that, either for the establishment of in situgenetic reserves or for ex situconservation, efforts should be directed to the conservation of several individual trees or collection of open pollinated seed families in a few distant sites. In addition, if resources are available to enlarge the genetic base of the existing collection, expeditions should prioritize collection sites located in areas at significant distances to those already sampled.

A total of 1,037 individuals living in five different sociogeographic regions of Brazil were studied in relation to 12 short tandem repeat polymorphisms. The objective was to assess the degree of European, African, and Amerindian contributions to their gene pools. Although most of the genetic variability was found within regions, significant differences were also observed between regions. The estimated relative proportions of the above-indicated continental contributions showed intermediate values between those obtained with uniparental (mtDNA, Y-chromosome) data, and a higher percentage of European heritage as compared to previous autosome results. A north–south trend of increasing European contribution was also found, in agreement with the history of the Brazilian population. Am. J. Hum. Biol. 15:824–834, 2003. © 2003 Wiley-Liss, Inc.

A total of 1,037 individuals living in five different sociogeographic regions of Brazil were studied in relation to 12 short tandem repeat polymorphisms. The objective was to assess the degree of European, African, and Amerindian contributions to their gene pools. Although most of the genetic variability was found within regions, significant differences were also observed between regions. The estimated relative proportions of the above‐indicated continental contributions showed intermediate values between those obtained with uniparental (mtDNA, Y‐chromosome) data, and a higher percentage of European heritage as compared to previous autosome results. A north–south trend of increasing European contribution was also found, in agreement with the history of the Brazilian population. Am. J. Hum. Biol. 15:824–834, 2003. © 2003 Wiley‐Liss, Inc.

We describe a simple and cost-effective method for the synthesis of an internal fluorescently labeled DNA standard for fragment sizing using an automated DNA sequencer. A set of primer pairs labeled with ROX was developed to amplify 12 DNA fragments, 58–417 bp, derived from a conserved region of plant chloroplast DNA. These amplified fragments were mixed together, constituting a fluorescent internal DNA size marker. The precision of the size standard was evaluated by estimating the size of 20 alleles that were amplified at four dinucleotide microsatellite loci with the synthesized size standard and the commercial internal sizing standard, GeneScan®Rox-500. A number of intra-gel and inter-gel comparisons were run, and an analysis of variance was carried out. No significant difference was observed between the size estimates obtained with the synthesized DNA standard and the commercial standard. This facile and general PCR-based method for the synthesis of internal standards allows for significant savings in the implementation of large genotyping experiments using microsatellite or AFLP markers.

Abstract: In cassava, apomixis fixes heterosis and avoids transmission of systemic pathogens which complicate vegetative propagation of the crop. A combination of evidence from maternal inheritance of RAPD markers and the structure of the embryonic sac in large progeny sets of two distinct genotypes have further confirmed the occurrence of apomixis in cassava. We could advance further on earlier reports of the detection of apomixis in four ways: (1) we could arrive at an estimate of the rate of facultative apomixis in the range of 2%; (2) we detected the occurrence of apomixis in a second genotype, derived from a different interspecific cross; (3) apomictic behavior was demonstrated in an F1 individual and (4) parallel embryonic evidence was generated that corroborate the potential occurrence of apomixis by apospory. The fact that apomixis was detected in an F1 interspecific hybrid hints to the possibility of directly transferring genes for apomixis from a wild relative to cultivated cassava.