A high quality draft consensus sequence of the genome of a heterozygous grapevine variety

Background. Worldwide, grapes and their derived products have a large market. The cultivated grape species Vitis vinifera has potential to become a model for fruit trees genetics. Like many plant species, it is highly heterozygous, which is an additional challenge to modern whole genome shotgun sequencing. In this paper a high quality draft genome sequence of a cultivated clone of V. vinifera Pinot Noir is presented. Principal Findings. We estimate the genome size of V. vinifera to be 504.6 Mb. Genomic sequences corresponding to 477.1 Mb were assembled in 2,093 metacontigs and 435.1 Mb were anchored to the 19 linkage groups (LGs). The number of predicted genes is 29,585, of which 96.1% were assigned to LGs. This assembly of the grape genome provides candidate genes implicated in traits relevant to grapevine cultivation, such as those influencing wine quality, via secondary metabolites, and those connected with the extreme susceptibility of grape to pathogens. Single nucleotide polymorphism (SNP) distribution was consistent with a diffuse haplotype structure across the genome. Of around 2,000,000 SNPs, 1,751,176 were mapped to chromosomes and one or more of them were identified in 86.7% of anchored genes. The relative age of grape duplicated genes was estimated and this made possible to reveal a relatively recent Vitisspecific large scale duplication event concerning at least 10 chromosomes (duplication not reported before). Conclusions. Sanger shotgun sequencing and highly efficient sequencing by synthesis (SBS), together with dedicated assembly programs, resolved a complex heterozygous genome. A consensus sequence of the genome and a set of mapped marker loci were generated. Homologous chromosomes of Pinot Noir differ by 11.2% of their DNA (hemizygous DNA plus chromosomal gaps). SNP markers are offered as a tool with the potential of introducing a new era in the molecular breeding of grape

Comparison of different methods for SNP detection in grapevine

Single nucleotide polymorphisms (SNPs) are the most abundant of all markers, both in animal and plant genomes. In crops and tree species considerable investment has been recently made on this genomic technology. While large-scale characterisation of SNPs by high-throughput techniques is possible, such highthroughput platforms are not available to all plant breeding laboratories. This report compares alternative multi-purpose and affordable methods for SNP assay in grapevine (Vitis spp.). In particular, the efficiency, sensitivity and reliability of single-strand conformation polymorphism (SSCP) on both non-denaturant gels and fluorescence-based capillary electrophoresis are compared with minisequencing (single nucleotide extension reaction). The results indicate that when multiplexing in combination with minisequencing is a mid-throughput, reliable and flexible technique for the detection of SNPs and can therefore be used effectively to improve marker assisted breeding in grapevine.

A RAPD, AFLP and SSR linkage map, and QTL analysis in European beech (Fagus sylvatica L.)

The genetic linkage map of European beech ( Fagus sylvatica L.) that we report here is the first to our knowledge. Based on a total of 312 markers (28 RAPDs, 274 AFLPs, 10 SSRs) scored in 143 individuals from a F(1) full-sib family. Two maps (one for each parent) were constructed according to a "two-way pseudo-testcross" mapping strategy. In the male map 119 markers could be clustered in 11 major groups (971 cM), while in the female map 132 markers were distributed in 12 major linkage groups (844 cM). In addition, four and one minor linkage groups (doublets and triplets) were obtained for the male and female map respectively. The two maps cover about 82% and 78% of the genome. Based on the position of 15 AFLP and 2 SSR loci segregating in both parents, seven homologous linkage groups could be identified. In the same pedigree we investigated the association with genetic markers of several quantitative traits: leaf area, leaf number and shape in 2 different years, specific leaf area, leaf carbon-isotope discrimination and tree height. A composite interval-mapping approach was used to estimate the number of QTLs, the amount of variation explained by each of them, and their position on the genetic linkage maps. Eight QTLs associated with leaf traits were found that explained between 15% and 35% of the trait variation, five on the female map and three on the male map

A SNP transferability survey within the genus Vitis

<p>Abstract</p> <p>Background</p> <p>Efforts to sequence the genomes of different organisms continue to increase. The DNA sequence is usually decoded for one individual and its application is for the whole species. The recent sequencing of the highly heterozygous <it>Vitis vinifera </it>L. cultivar Pinot Noir (clone ENTAV 115) genome gave rise to several thousand polymorphisms and offers a good model to study the transferability of its degree of polymorphism to other individuals of the same species and within the genus.</p> <p>Results</p> <p>This study was performed by genotyping 137 SNPs through the SNPlex™ Genotyping System (Applied Biosystems Inc.) and by comparing the SNPlex sequencing results across 35 (of the 137) regions from 69 grape accessions. A heterozygous state transferability of 31.5% across the unrelated cultivars of <it>V. vinifera</it>, of 18.8% across the wild forms of <it>V. vinifera</it>, of 2.3% among non-<it>vinifera Vitis </it>species, and of 0% with <it>Muscadinia rotundifolia </it>was found. In addition, mean allele frequencies were used to evaluate SNP informativeness and develop useful subsets of markers.</p> <p>Conclusion</p> <p>Using SNPlex application and corroboration from the sequencing analysis, the informativeness of SNP markers from the heterozygous grape cultivar Pinot Noir was validated in <it>V. vinifera </it>(including cultivars and wild forms), but had a limited application for non-<it>vinifera Vitis </it>species where a resequencing strategy may be preferred, knowing that homology at priming sites is sufficient. This work will allow future applications such as mapping and diversity studies, accession identification and genomic-research assisted breeding within <it>V. vinifera</it>.</p

Development and Evaluation of a 9K SNP Addition to the Peach Ipsc 9K SNP Array v1

The IPSC 9K peach SNP array released by the international community has been a valuable tool in research and application. Even though majority of SNPs (84%) were polymorphic in the evaluation panels there were many genomic regions with low coverage, including those important for breeding. The existing peach array has been updated with 9K additional SNPs covering previously identified gaps and including recently identified SNPs important for breeding. SNPs (1,808,996) identified by sequencing 49 genomes of additional peach accessions were used as the main source of additional SNPs. Focal point strategy was used to select 8,971 SNPs within 40kb window from the 2,821 focal points distributed across the genome. Additional 129 SNPs were chosen to saturate either regions important for breeding or close the gaps larger than 100kb. The array was validated with 1,770 peach and 26 Prunus accessions (almond, plum, apricot, wild relatives). The add-on contained 7,862 SNPs evenly spread across 8 peach pseudo-molecules with only one SNP positioned on scaffold 13 covering 224.99Mbp of peach genome. The 9K add-on improved the 9K peach array by increasing the total number of usable SNPs by 7,206. The number of SNPs per chromosome increased on average by 50% with only on average 0.18% increase in total physical coverage. Number of gaps larger than 0.3 Mbp was reduced to 2 one on each chromosome 3 and 8. Overall genotyping efficiency in all material was &gt;90% except in almond, 82%. Number of informative markers, assessed by ASSIsT software, were highest in peach 64% and lowest in almond 10%, with 61% of markers being informative in wild Prunus (12) and 35% in apricot (4) and 2 - 33% in Japanese and European plum, respectively. Among 36.2% discarded markers 33% were monomorphic and 30% shifted homozygous in material used. Those markers could be informative in different background raising total number of informative markers. Ann addition of new SNPs to array improved the density and usefulness of the array in Prunus species. The practical applications of new 16K Illumina SNP peach array will be discussed. Specified Source(s) of Funding: USDA-NIFA-SCRI-Ros- BREED (2014-51181-22378

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 (~253K 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 19th 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

QTL and candidate gene mapping for polyphenolic composition in apple fruit

<p>Abstract</p> <p>Background</p> <p>The polyphenolic products of the phenylpropanoid pathway, including proanthocyanidins, anthocyanins and flavonols, possess antioxidant properties that may provide health benefits. To investigate the genetic architecture of control of their biosynthesis in apple fruit, various polyphenolic compounds were quantified in progeny from a 'Royal Gala' × 'Braeburn' apple population segregating for antioxidant content, using ultra high performance liquid chromatography of extracts derived from fruit cortex and skin.</p> <p>Results</p> <p>Construction of genetic maps for 'Royal Gala' and 'Braeburn' enabled detection of 79 quantitative trait loci (QTL) for content of 17 fruit polyphenolic compounds. Seven QTL clusters were stable across two years of harvest and included QTLs for content of flavanols, flavonols, anthocyanins and hydroxycinnamic acids. Alignment of the parental genetic maps with the apple whole genome sequence <it>in silico </it>enabled screening for co-segregation with the QTLs of a range of candidate genes coding for enzymes in the polyphenolic biosynthetic pathway. This co-location was confirmed by genetic mapping of markers derived from the gene sequences. <it>Leucoanthocyanidin reductase </it>(<it>LAR1</it>) co-located with a QTL cluster for the fruit flavanols catechin, epicatechin, procyanidin dimer and five unknown procyanidin oligomers identified near the top of linkage group (LG) 16, while <it>hydroxy cinnamate/quinate transferase </it>(<it>HCT</it>/<it>HQT</it>) co-located with a QTL for chlorogenic acid concentration mapping near the bottom of LG 17.</p> <p>Conclusion</p> <p>We conclude that <it>LAR1 </it>and <it>HCT</it>/<it>HQT </it>are likely to influence the concentration of these compounds in apple fruit and provide useful allele-specific markers for marker assisted selection of trees bearing fruit with healthy attributes.</p

Valorization of traditional Italian walnut (Juglans regia L.) production: genetic, nutritional and sensory characterization of locally grown varieties in the Trentino region

15openYesJuglans 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.Di Pierro, Erica A.; Franceschi, Pietro; Endrizzi, Isabella; Farneti, Brian; Poles, Lara; Masuero, Domenico; Khomenko, Iuliia; Trenti, Francesco; Marrano, Annarita; Vrhovsek, Urska; Gasperi, Flavia; Biasioli, Franco; Guella, Graziano; Bianco, Luca; Troggio, MichelaDi Pierro, E.A.; Franceschi, P.; Endrizzi, I.; Farneti, B.; Poles, L.; Masuero, D.; Khomenko, I.; Trenti, F.; Marrano, A.; Vrhovsek, U.; Gasperi, F.; Biasioli, F.; Guella, G.; Bianco, L.; Troggio, M

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

Genetic Variability Study in a Wide Germplasm of Domesticated Peach Through High Throughput

Peach (Prunus persica (L.) Batsch) is one of the most economically important fruit crops in temperate areas. Classical fruit tree breeding is generally slow and inefficient. Molecular markers could improve its efficiency but, although nowadays many Mendelian traits are mapped in peach and SSR markers have been found to be linked to some of the key major genes, its use in breeding programs is still limited. Main reasons for that are insufficient linkage between the markers and the genes and the lack of markers suitable for medium-high degree of multiplexing. To address this limitation, about 1,300 peach cultivars were genotyped with the 9K peach SNP chip (Verde et al. 2012) in the frame of FruitBreedomics project. This germplasm was chosen to be representative of the genetic diversity present in five germplasm collection in Europe and in China. Out of the 8144SNPs present in the chip, about 4300 were positively genotyped and used for the further analysis. The average number of heterozygous loci in the genotyped accessions was 1186 (spanning from 13 to 2775). The preliminary results of the population structure reveal three main subpopulations and the presence of high number of admixed individuals. LD seems to decay at distance longer than ca. 1 Mb. These results will be instrumental for implementing LD-based mapping of QTLs and genes in peach