Valorization of Traditional Italian Walnut (Juglans regia L.) Production: Genetic, Nutritional and Sensory Characterization of Locally Grown Varieties in the Trentino Region.

Juglans regia (L.) is cultivated worldwide for its nutrient-rich nuts. In Italy, despite the growing demand, walnut cultivation has gone through a strong decline in recent decades, which led to Italy being among the top five net importing countries. To promote the development of local high-quality Italian walnut production, we devised a multidisciplinary project to highlight the distinctive traits of three varieties grown in the mountainous region Trentino (northeast of Italy): the heirloom 'Bleggiana', a second local accession called local Franquette and the French cultivar 'Lara', recently introduced in the local production to increase yield. The genetic characterization confirmed the uniqueness of 'Bleggiana' and revealed local Franquette as a newly described autochthonous variety, thus named 'Blegette'. The metabolic profiles highlighted a valuable nutritional composition of the local varieties, richer in polyphenols and with a lower ω-6/ω-3 ratio than the commercial 'Lara'. 'Blegette' obtained the highest preference scores from consumers for both the visual aspect and tasting; however, the volatile organic compound profiles did not discriminate among the characterized cultivars. The described local varieties represent an interesting reservoir of walnut genetic diversity and quality properties, which deserve future investigation on agronomically useful traits (e.g., local adaptation and water usage) for a high-quality and sustainable production.

Integration of Infinium and Axiom SNP array data in the outcrossing species Malus × domestica and causes for seemingly incompatible calls.

BACKGROUND: Single nucleotide polymorphism (SNP) array technology has been increasingly used to generate large quantities of SNP data for use in genetic studies. As new arrays are developed to take advantage of new technology and of improved probe design using new genome sequence and panel data, a need to integrate data from different arrays and array platforms has arisen. This study was undertaken in view of our need for an integrated high-quality dataset of Illumina Infinium® 20 K and Affymetrix Axiom® 480 K SNP array data in apple (Malus × domestica). In this study, we qualify and quantify the compatibility of SNP calling, defined as SNP calls that are both accurate and concordant, across both arrays by two approaches. First, the concordance of SNP calls was evaluated using a set of 417 duplicate individuals genotyped on both arrays starting from a set of 10,295 robust SNPs on the Infinium array. Next, the accuracy of the SNP calls was evaluated on additional germplasm (n = 3141) from both arrays using Mendelian inconsistent and consistent errors across thousands of pedigree links. While performing this work, we took the opportunity to evaluate reasons for probe failure and observed discordant SNP calls. RESULTS: Concordance among the duplicate individuals was on average of 97.1% across 10,295 SNPs. Of these SNPs, 35% had discordant call(s) that were further curated, leading to a final set of 8412 (81.7%) SNPs that were deemed compatible. Compatibility was highly influenced by the presence of alternate probe binding locations and secondary polymorphisms. The impact of the latter was highly influenced by their number and proximity to the 3' end of the probe. CONCLUSIONS: The Infinium and Axiom SNP array data were mostly compatible. However, data integration required intense data filtering and curation. This work resulted in a workflow and information that may be of use in other data integration efforts. Such an in-depth analysis of array concordance and accuracy as ours has not been previously described in the literature and will be useful in future work on SNP array data integration and interpretation, and in probe/platform development.

High-quality chromosome-scale assembly of the walnut (Juglans regia L.) reference genome.

BACKGROUND: The release of the first reference genome of walnut (Juglans regia L.) enabled many achievements in the characterization of walnut genetic and functional variation. However, it is highly fragmented, preventing the integration of genetic, transcriptomic, and proteomic information to fully elucidate walnut biological processes. FINDINGS: Here, we report the new chromosome-scale assembly of the walnut reference genome (Chandler v2.0) obtained by combining Oxford Nanopore long-read sequencing with chromosome conformation capture (Hi-C) technology. Relative to the previous reference genome, the new assembly features an 84.4-fold increase in N50 size, with the 16 chromosomal pseudomolecules assembled and representing 95% of its total length. Using full-length transcripts from single-molecule real-time sequencing, we predicted 37,554 gene models, with a mean gene length higher than the previous gene annotations. Most of the new protein-coding genes (90%) present both start and stop codons, which represents a significant improvement compared with Chandler v1.0 (only 48%). We then tested the potential impact of the new chromosome-level genome on different areas of walnut research. By studying the proteome changes occurring during male flower development, we observed that the virtual proteome obtained from Chandler v2.0 presents fewer artifacts than the previous reference genome, enabling the identification of a new potential pollen allergen in walnut. Also, the new chromosome-scale genome facilitates in-depth studies of intraspecies genetic diversity by revealing previously undetected autozygous regions in Chandler, likely resulting from inbreeding, and 195 genomic regions highly differentiated between Western and Eastern walnut cultivars. CONCLUSION: Overall, Chandler v2.0 will serve as a valuable resource to better understand and explore walnut biology.

Using whole-genome SNP data to reconstruct a large multi-generation pedigree in apple germplasm.

BACKGROUND: Apple (Malus x domestica Borkh.) is one of the most important fruit tree crops of temperate areas, with great economic and cultural value. Apple cultivars can be maintained for centuries in plant collections through grafting, and some are thought to date as far back as Roman times. Molecular markers provide a means to reconstruct pedigrees and thus shed light on the recent history of migration and trade of biological materials. The objective of the present study was to identify relationships within a set of over 1400 mostly old apple cultivars using whole-genome SNP data (~ 253 K SNPs) in order to reconstruct pedigrees. RESULTS: Using simple exclusion tests, based on counting the number of Mendelian errors, more than one thousand parent-offspring relations and 295 complete parent-offspring families were identified. Additionally, a grandparent couple was identified for the missing parental side of 26 parent-offspring pairings. Among the 407 parent-offspring relations without a second identified parent, 327 could be oriented because one of the individuals was an offspring in a complete family or by using historical data on parentage or date of recording. Parents of emblematic cultivars such as 'Ribston Pippin', 'White Transparent' and 'Braeburn' were identified. The overall pedigree combining all the identified relationships encompassed seven generations and revealed a major impact of two Renaissance cultivars of French and English origin, namely 'Reinette Franche' and 'Margil', and one North-Eastern Europe cultivar from the 1700s, 'Alexander'. On the contrary, several older cultivars, from the Middle Ages or the Roman times, had no, or only single, identifiable offspring in the set of studied accessions. Frequent crosses between cultivars originating from different European regions were identified, especially from the nineteenth century onwards. CONCLUSIONS: The availability of over 1400 apple genotypes, previously filtered for genetic uniqueness and providing a broad representation of European germplasm, has been instrumental for the success of this large pedigree reconstruction. It enlightens the history of empirical selection and recent breeding of apple cultivars in Europe and provides insights to speed-up future breeding and selection.

Mapping of fire blight resistance in Malus ×robusta 5 flowers following artificial inoculation.

BACKGROUND: Although the most common path of infection for fire blight, a severe bacterial disease on apple, is via host plant flowers, quantitative trait loci (QTLs) for fire blight resistance to date have exclusively been mapped following shoot inoculation. It is not known whether the same mechanism underlies flower and shoot resistance. RESULTS: We report the detection of a fire blight resistance QTL following independent artificial inoculation of flowers and shoots on two F1 segregating populations derived from crossing resistant Malus ×robusta 5 (Mr5) with susceptible 'Idared' and 'Royal Gala' in experimental orchards in Germany and New Zealand, respectively. QTL mapping of phenotypic datasets from artificial flower inoculation of the 'Idared' × Mr5 population with Erwinia amylovora over several years, and of the 'Royal Gala' × Mr5 population in a single year, revealed a single major QTL controlling floral fire blight resistance on linkage group 3 (LG3) of Mr5. This QTL corresponds to the QTL on LG3 reported previously for the 'Idared' × Mr5 and an 'M9' × Mr5 population following shoot inoculation in the glasshouse. Interval mapping of phenotypic data from shoot inoculations of subsets from both flower resistance populations re-confirmed that the resistance QTL is in the same position on LG3 of Mr5 as that for flower inoculation. These results provide strong evidence that fire blight resistance in Mr5 is controlled by a major QTL on LG3, independently of the mode of infection, rootstock and environment. CONCLUSIONS: This study demonstrates for the first time that resistance to fire blight caused by Erwinia amylovora is independent of the mode of inoculation at least in Malus ×robusta 5.

Pseudo-chromosome-length genome assembly of a double haploid "Bartlett" pear (Pyrus communis L.).

BACKGROUND: We report an improved assembly and scaffolding of the European pear (Pyrus communis L.) genome (referred to as BartlettDHv2.0), obtained using a combination of Pacific Biosciences RSII long-read sequencing, Bionano optical mapping, chromatin interaction capture (Hi-C), and genetic mapping. The sample selected for sequencing is a double haploid derived from the same "Bartlett" reference pear that was previously sequenced. Sequencing of di-haploid plants makes assembly more tractable in highly heterozygous species such as P. communis. FINDINGS: A total of 496.9 Mb corresponding to 97% of the estimated genome size were assembled into 494 scaffolds. Hi-C data and a high-density genetic map allowed us to anchor and orient 87% of the sequence on the 17 pear chromosomes. Approximately 50% (247 Mb) of the genome consists of repetitive sequences. Gene annotation confirmed the presence of 37,445 protein-coding genes, which is 13% fewer than previously predicted. CONCLUSIONS: We showed that the use of a doubled-haploid plant is an effective solution to the problems presented by high levels of heterozygosity and duplication for the generation of high-quality genome assemblies. We present a high-quality chromosome-scale assembly of the European pear Pyrus communis and demostrate its high degree of synteny with the genomes of Malus x Domestica and Pyrus x bretschneideri.

A Reference Genome Sequence for the European Silver Fir (Abies alba Mill.): A Community-Generated Genomic Resource.

Silver fir (Abies alba Mill.) is a keystone conifer of European montane forest ecosystems that has experienced large fluctuations in population size during during the Quaternary and, more recently, due to land-use change. To forecast the species' future distribution and survival, it is important to investigate the genetic basis of adaptation to environmental change, notably to extreme events. For this purpose, we here provide a first draft genome assembly and annotation of the silver fir genome, established through a community-based initiative. DNA obtained from haploid megagametophyte and diploid needle tissue was used to construct and sequence Illumina paired-end and mate-pair libraries, respectively, to high depth. The assembled A. alba genome sequence accounted for over 37 million scaffolds corresponding to 18.16 Gb, with a scaffold N50 of 14,051 bp. Despite the fragmented nature of the assembly, a total of 50,757 full-length genes were functionally annotated in the nuclear genome. The chloroplast genome was also assembled into a single scaffold (120,908 bp) that shows a high collinearity with both the A. koreana and A. sibirica complete chloroplast genomes. This first genome assembly of silver fir is an important genomic resource that is now publicly available in support of a new generation of research. By genome-enabling this important conifer, this resource will open the gate for new research and more precise genetic monitoring of European silver fir forests.

Development of a highly efficient Axiom™ 70 K SNP array for Pyrus and evaluation for high-density mapping and germplasm characterization.

BACKGROUND: Both a source of diversity and the development of genomic tools, such as reference genomes and molecular markers, are equally important to enable faster progress in plant breeding. Pear (Pyrus spp.) lags far behind other fruit and nut crops in terms of employment of available genetic resources for new cultivar development. To address this gap, we designed a high-density, high-efficiency and robust single nucleotide polymorphism (SNP) array for pear, with the main objectives of conducting genetic diversity and genome-wide association studies. RESULTS: By applying a two-step design process, which consisted of the construction of a first 'draft' array for the screening of a small subset of samples, we were able to identify the most robust and informative SNPs to include in the Applied Biosystems™ Axiom™ Pear 70 K Genotyping Array, currently the densest SNP array for pear. Preliminary evaluation of this 70 K array in 1416 diverse pear accessions from the USDA National Clonal Germplasm Repository (NCGR) in Corvallis, OR identified 66,616 SNPs (93% of all the tiled SNPs) as high quality and polymorphic (PolyHighResolution). We further used the Axiom Pear 70 K Genotyping Array to construct high-density linkage maps in a bi-parental population, and to make a direct comparison with available genotyping-by-sequencing (GBS) data, which suggested that the SNP array is a more robust method of screening for SNPs than restriction enzyme reduced representation sequence-based genotyping. CONCLUSIONS: The Axiom Pear 70 K Genotyping Array, with its high efficiency in a widely diverse panel of Pyrus species and cultivars, represents a valuable resource for a multitude of molecular studies in pear. The characterization of the USDA-NCGR collection with this array will provide important information for pear geneticists and breeders, as well as for the optimization of conservation strategies for Pyrus.

Apple whole genome sequences: recent advances and new prospects.

In 2010, a major scientific milestone was achieved for tree fruit crops: publication of the first draft whole genome sequence (WGS) for apple (Malus domestica). This WGS, v1.0, was valuable as the initial reference for sequence information, fine mapping, gene discovery, variant discovery, and tool development. A new, high quality apple WGS, GDDH13 v1.1, was released in 2017 and now serves as the reference genome for apple. Over the past decade, these apple WGSs have had an enormous impact on our understanding of apple biological functioning, trait physiology and inheritance, leading to practical applications for improving this highly valued crop. Causal gene identities for phenotypes of fundamental and practical interest can today be discovered much more rapidly. Genome-wide polymorphisms at high genetic resolution are screened efficiently over hundreds to thousands of individuals with new insights into genetic relationships and pedigrees. High-density genetic maps are constructed efficiently and quantitative trait loci for valuable traits are readily associated with positional candidate genes and/or converted into diagnostic tests for breeders. We understand the species, geographical, and genomic origins of domesticated apple more precisely, as well as its relationship to wild relatives. The WGS has turbo-charged application of these classical research steps to crop improvement and drives innovative methods to achieve more durable, environmentally sound, productive, and consumer-desirable apple production. This review includes examples of basic and practical breakthroughs and challenges in using the apple WGSs. Recommendations for "what's next" focus on necessary upgrades to the genome sequence data pool, as well as for use of the data, to reach new frontiers in genomics-based scientific understanding of apple.

A new genomic tool for walnut (Juglans regia L.): development and validation of the high-density Axiom™ J. regia 700K SNP genotyping array.

Over the last 20 years, global production of Persian walnut (Juglans regia L.) has grown enormously, likely reflecting increased consumption due to its numerous benefits to human health. However, advances in genome-wide association (GWA) studies and genomic selection (GS) for agronomically important traits in walnut remain limited due to the lack of powerful genomic tools. Here, we present the development and validation of a high-density 700K single nucleotide polymorphism (SNP) array in Persian walnut. Over 609K high-quality SNPs have been thoroughly selected from a set of 9.6 m genome-wide variants, previously identified from the high-depth re-sequencing of 27 founders of the Walnut Improvement Program (WIP) of University of California, Davis. To validate the effectiveness of the array, we genotyped a collection of 1284 walnut trees, including 1167 progeny of 48 WIP families and 26 walnut cultivars. More than half of the SNPs (55.7%) fell in the highest quality class of 'Poly High Resolution' (PHR) polymorphisms, which were used to assess the WIP pedigree integrity. We identified 151 new parent-offspring relationships, all confirmed with the Mendelian inheritance test. In addition, we explored the genetic variability among cultivars of different origin, revealing how the varieties from Europe and California were differentiated from Asian accessions. Both the reconstruction of the WIP pedigree and population structure analysis confirmed the effectiveness of the Applied Biosystems™ Axiom™ J. regia 700K SNP array, which initiates a novel genomic and advanced phase in walnut genetics and breeding.

An integrated approach for increasing breeding efficiency in apple and peach in Europe.

Despite the availability of whole genome sequences of apple and peach, there has been a considerable gap between genomics and breeding. To bridge the gap, the European Union funded the FruitBreedomics project (March 2011 to August 2015) involving 28 research institutes and private companies. Three complementary approaches were pursued: (i) tool and software development, (ii) deciphering genetic control of main horticultural traits taking into account allelic diversity and (iii) developing plant materials, tools and methodologies for breeders. Decisive breakthroughs were made including the making available of ready-to-go DNA diagnostic tests for Marker Assisted Breeding, development of new, dense SNP arrays in apple and peach, new phenotypic methods for some complex traits, software for gene/QTL discovery on breeding germplasm via Pedigree Based Analysis (PBA). This resulted in the discovery of highly predictive molecular markers for traits of horticultural interest via PBA and via Genome Wide Association Studies (GWAS) on several European genebank collections. FruitBreedomics also developed pre-breeding plant materials in which multiple sources of resistance were pyramided and software that can support breeders in their selection activities. Through FruitBreedomics, significant progresses were made in the field of apple and peach breeding, genetics, genomics and bioinformatics of which advantage will be made by breeders, germplasm curators and scientists. A major part of the data collected during the project has been stored in the FruitBreedomics database and has been made available to the public. This review covers the scientific discoveries made in this major endeavour, and perspective in the apple and peach breeding and genomics in Europe and beyond.

Genome-Wide Association Mapping of Flowering and Ripening Periods in Apple.

Deciphering the genetic control of flowering and ripening periods in apple is essential for breeding cultivars adapted to their growing environments. We implemented a large Genome-Wide Association Study (GWAS) at the European level using an association panel of 1,168 different apple genotypes distributed over six locations and phenotyped for these phenological traits. The panel was genotyped at a high-density of SNPs using the Axiom®Apple 480 K SNP array. We ran GWAS with a multi-locus mixed model (MLMM), which handles the putatively confounding effect of significant SNPs elsewhere on the genome. Genomic regions were further investigated to reveal candidate genes responsible for the phenotypic variation. At the whole population level, GWAS retained two SNPs as cofactors on chromosome 9 for flowering period, and six for ripening period (four on chromosome 3, one on chromosome 10 and one on chromosome 16) which, together accounted for 8.9 and 17.2% of the phenotypic variance, respectively. For both traits, SNPs in weak linkage disequilibrium were detected nearby, thus suggesting the existence of allelic heterogeneity. The geographic origins and relationships of apple cultivars accounted for large parts of the phenotypic variation. Variation in genotypic frequency of the SNPs associated with the two traits was connected to the geographic origin of the genotypes (grouped as North+East, West and South Europe), and indicated differential selection in different growing environments. Genes encoding transcription factors containing either NAC or MADS domains were identified as major candidates within the small confidence intervals computed for the associated genomic regions. A strong microsynteny between apple and peach was revealed in all the four confidence interval regions. This study shows how association genetics can unravel the genetic control of important horticultural traits in apple, as well as reduce the confidence intervals of the associated regions identified by linkage mapping approaches. Our findings can be used for the improvement of apple through marker-assisted breeding strategies that take advantage of the accumulating additive effects of the identified SNPs.

High-quality de novo assembly of the apple genome and methylome dynamics of early fruit development.

Using the latest sequencing and optical mapping technologies, we have produced a high-quality de novo assembly of the apple (Malus domestica Borkh.) genome. Repeat sequences, which represented over half of the assembly, provided an unprecedented opportunity to investigate the uncharacterized regions of a tree genome; we identified a new hyper-repetitive retrotransposon sequence that was over-represented in heterochromatic regions and estimated that a major burst of different transposable elements (TEs) occurred 21 million years ago. Notably, the timing of this TE burst coincided with the uplift of the Tian Shan mountains, which is thought to be the center of the location where the apple originated, suggesting that TEs and associated processes may have contributed to the diversification of the apple ancestor and possibly to its divergence from pear. Finally, genome-wide DNA methylation data suggest that epigenetic marks may contribute to agronomically relevant aspects, such as apple fruit development.

Integrated QTL detection for key breeding traits in multiple peach progenies.

BACKGROUND: Peach (Prunus persica (L.) Batsch) is a major temperate fruit crop with an intense breeding activity. Breeding is facilitated by knowledge of the inheritance of the key traits that are often of a quantitative nature. QTLs have traditionally been studied using the phenotype of a single progeny (usually a full-sib progeny) and the correlation with a set of markers covering its genome. This approach has allowed the identification of various genes and QTLs but is limited by the small numbers of individuals used and by the narrow transect of the variability analyzed. In this article we propose the use of a multi-progeny mapping strategy that used pedigree information and Bayesian approaches that supports a more precise and complete survey of the available genetic variability. RESULTS: Seven key agronomic characters (data from 1 to 3 years) were analyzed in 18 progenies from crosses between occidental commercial genotypes and various exotic lines including accessions of other Prunus species. A total of 1467 plants from these progenies were genotyped with a 9 k SNP array. Forty-seven QTLs were identified, 22 coinciding with major genes and QTLs that have been consistently found in the same populations when studied individually and 25 were new. A substantial part of the QTLs observed (47%) would not have been detected in crosses between only commercial materials, showing the high value of exotic lines as a source of novel alleles for the commercial gene pool. Our strategy also provided estimations on the narrow sense heritability of each character, and the estimation of the QTL genotypes of each parent for the different QTLs and their breeding value. CONCLUSIONS: The integrated strategy used provides a broader and more accurate picture of the variability available for peach breeding with the identification of many new QTLs, information on the sources of the alleles of interest and the breeding values of the potential donors of such valuable alleles. These results are first-hand information for breeders and a step forward towards the implementation of DNA-informed strategies to facilitate selection of new cultivars with improved productivity and quality.

The Peach v2.0 release: high-resolution linkage mapping and deep resequencing improve chromosome-scale assembly and contiguity.

BACKGROUND: The availability of the peach genome sequence has fostered relevant research in peach and related Prunus species enabling the identification of genes underlying important horticultural traits as well as the development of advanced tools for genetic and genomic analyses. The first release of the peach genome (Peach v1.0) represented a high-quality WGS (Whole Genome Shotgun) chromosome-scale assembly with high contiguity (contig L50 214.2 kb), large portions of mapped sequences (96%) and high base accuracy (99.96%). The aim of this work was to improve the quality of the first assembly by increasing the portion of mapped and oriented sequences, correcting misassemblies and improving the contiguity and base accuracy using high-throughput linkage mapping and deep resequencing approaches. RESULTS: Four linkage maps with 3,576 molecular markers were used to improve the portion of mapped and oriented sequences (from 96.0% and 85.6% of Peach v1.0 to 99.2% and 98.2% of v2.0, respectively) and enabled a more detailed identification of discernible misassemblies (10.4 Mb in total). The deep resequencing approach fixed 859 homozygous SNPs (Single Nucleotide Polymorphisms) and 1347 homozygous indels. Moreover, the assembled NGS contigs enabled the closing of 212 gaps with an improvement in the contig L50 of 19.2%. CONCLUSIONS: The improved high quality peach genome assembly (Peach v2.0) represents a valuable tool for the analysis of the genetic diversity, domestication, and as a vehicle for genetic improvement of peach and related Prunus species. Moreover, the important phylogenetic position of peach and the absence of recent whole genome duplication (WGD) events make peach a pivotal species for comparative genomics studies aiming at elucidating plant speciation and diversification processes.

A high-density, multi-parental SNP genetic map on apple validates a new mapping approach for outcrossing species.

Quantitative trait loci (QTL) mapping approaches rely on the correct ordering of molecular markers along the chromosomes, which can be obtained from genetic linkage maps or a reference genome sequence. For apple (Malus domestica Borkh), the genome sequence v1 and v2 could not meet this need; therefore, a novel approach was devised to develop a dense genetic linkage map, providing the most reliable marker-loci order for the highest possible number of markers. The approach was based on four strategies: (i) the use of multiple full-sib families, (ii) the reduction of missing information through the use of HaploBlocks and alternative calling procedures for single-nucleotide polymorphism (SNP) markers, (iii) the construction of a single backcross-type data set including all families, and (iv) a two-step map generation procedure based on the sequential inclusion of markers. The map comprises 15 417 SNP markers, clustered in 3 K HaploBlock markers spanning 1 267 cM, with an average distance between adjacent markers of 0.37 cM and a maximum distance of 3.29 cM. Moreover, chromosome 5 was oriented according to its homoeologous chromosome 10. This map was useful to improve the apple genome sequence, design the Axiom Apple 480 K SNP array and perform multifamily-based QTL studies. Its collinearity with the genome sequences v1 and v3 are reported. To our knowledge, this is the shortest published SNP map in apple, while including the largest number of markers, families and individuals. This result validates our methodology, proving its value for the construction of integrated linkage maps for any outbreeding species.

Development and validation of the Axiom(®) Apple480K SNP genotyping array.

Cultivated apple (Malus × domestica Borkh.) is one of the most important fruit crops in temperate regions, and has great economic and cultural value. The apple genome is highly heterozygous and has undergone a recent duplication which, combined with a rapid linkage disequilibrium decay, makes it difficult to perform genome-wide association (GWA) studies. Single nucleotide polymorphism arrays offer highly multiplexed assays at a relatively low cost per data point and can be a valid tool for the identification of the markers associated with traits of interest. Here, we describe the development and validation of a 487K SNP Affymetrix Axiom(®) genotyping array for apple and discuss its potential applications. The array has been built from the high-depth resequencing of 63 different cultivars covering most of the genetic diversity in cultivated apple. The SNPs were chosen by applying a focal points approach to enrich genic regions, but also to reach a uniform coverage of non-genic regions. A total of 1324 apple accessions, including the 92 progenies of two mapping populations, have been genotyped with the Axiom(®) Apple480K to assess the effectiveness of the array. A large majority of SNPs (359 994 or 74%) fell in the stringent class of poly high resolution polymorphisms. We also devised a filtering procedure to identify a subset of 275K very robust markers that can be safely used for germplasm surveys in apple. The Axiom(®) Apple480K has now been commercially released both for public and proprietary use and will likely be a reference tool for GWA studies in apple.

Whole-Genome Analysis of Diversity and SNP-Major Gene Association in Peach Germplasm.

Peach was domesticated in China more than four millennia ago and from there it spread world-wide. Since the middle of the last century, peach breeding programs have been very dynamic generating hundreds of new commercial varieties, however, in most cases such varieties derive from a limited collection of parental lines (founders). This is one reason for the observed low levels of variability of the commercial gene pool, implying that knowledge of the extent and distribution of genetic variability in peach is critical to allow the choice of adequate parents to confer enhanced productivity, adaptation and quality to improved varieties. With this aim we genotyped 1,580 peach accessions (including a few closely related Prunus species) maintained and phenotyped in five germplasm collections (four European and one Chinese) with the International Peach SNP Consortium 9K SNP peach array. The study of population structure revealed the subdivision of the panel in three main populations, one mainly made up of Occidental varieties from breeding programs (POP1OCB), one of Occidental landraces (POP2OCT) and the third of Oriental accessions (POP3OR). Analysis of linkage disequilibrium (LD) identified differential patterns of genome-wide LD blocks in each of the populations. Phenotypic data for seven monogenic traits were integrated in a genome-wide association study (GWAS). The significantly associated SNPs were always in the regions predicted by linkage analysis, forming haplotypes of markers. These diagnostic haplotypes could be used for marker-assisted selection (MAS) in modern breeding programs.

ASSIsT: an automatic SNP scoring tool for in- and outbreeding species.

UNLABELLED: ASSIsT (Automatic SNP ScorIng Tool) is a user-friendly customized pipeline for efficient calling and filtering of SNPs from Illumina Infinium arrays, specifically devised for custom genotyping arrays. Illumina has developed an integrated software for SNP data visualization and inspection called GenomeStudio (GS). ASSIsT builds on GS-derived data and identifies those markers that follow a bi-allelic genetic model and show reliable genotype calls. Moreover, ASSIsT re-edits SNP calls with null alleles or additional SNPs in the probe annealing site. ASSIsT can be employed in the analysis of different population types such as full-sib families and mating schemes used in the plant kingdom (backcross, F1, F2), and unrelated individuals. The final result can be directly exported in the format required by the most common software for genetic mapping and marker-trait association analysis. ASSIsT is developed in Python and runs in Windows and Linux. AVAILABILITY AND IMPLEMENTATION: The software, example data sets and tutorials are freely available at http://compbiotoolbox.fmach.it/assist/. CONTACT: eric.vandeweg@wur.nl.

New candidate genes for the fine regulation of the colour of grapes.

In the last decade, great progress has been made in clarifying the main determinants of anthocyanin accumulation in grape berry skin. However, the molecular details of the fine variation among cultivars, which ultimately contributes to wine typicity, are still not completely understood. To shed light on this issue, the grapes of 170 F1 progeny from the cross 'Syrah'×'Pinot Noir' were characterized at the mature stage for the content of 15 anthocyanins during four growing seasons. This huge data set was used in combination with a dense genetic map to detect genomic regions controlling the anthocyanin pathway both at key enzymatic points and at particular branches. Genes putatively involved in fine tuning the global regulation of anthocyanin biosynthesis were identified by exploring the gene predictions in the QTL (quantitative trait locus) confidence intervals and their expression profile during berry development in offspring with contrasting anthocyanin accumulation. New information on some aspects which had scarcely been investigated so far, such as anthocyanin transport into the vacuole, or completely neglected, such as acylation, is provided. These genes represent a valuable resource in grapevine molecular-based breeding programmes to improve both fruit and wine quality and to tailor wine sensory properties according to consumer demand.