Merging genotyping-by-sequencing data from two ex situ collections provides insights on the pea evolutionary history

Pea (Pisum sativum L. subsp. sativum) is one of the oldest domesticated species and a widely cultivated legume. In this study, we combined next generation sequencing (NGS) data referring to two genotyping-by-sequencing (GBS) libraries, each one prepared from a different Pisum germplasm collection. The selection of single nucleotide polymorphism (SNP) loci called in both germplasm collections caused some loss of information; however, this did not prevent the obtainment of one of the largest datasets ever used to explore pea biodiversity, consisting of 652 accessions and 22 127 markers. The analysis of population structure reflected genetic variation based on geographic patterns and allowed the definition of a model for the expansion of pea cultivation from the domestication centre to other regions of the world. In genetically distinct populations, the average decay of linkage disequilibrium (LD) ranged from a few bases to hundreds of kilobases, thus indicating different evolutionary histories leading to their diversification. Genome-wide scans resulted in the identification of putative selective sweeps associated with domestication and breeding, including genes known to regulate shoot branching, cotyledon colour and resistance to lodging, and the correct mapping of two Mendelian genes. In addition to providing information of major interest for fundamental and applied research on pea, our work describes the first successful example of integration of different GBS datasets generated from ex situ collections - a process of potential interest for a variety of purposes, including conservation genetics, genome-wide association studies, and breeding

Affordable phenotyping of winter wheat under field and controlled conditions for drought tolerance

Drought stress is one of the key plant stresses reducing grain yield in cereal crops worldwide. Although it is not a breeding target in Northern Europe, the changing climate and the drought of 2018 have increased its significance in the region. A key challenge, therefore, is to identify novel germplasm with higher drought tolerance, a task that will require continuous characterization of a large number of genotypes. The aim of this work was to assess if phenotyping systems with low-cost consumer-grade digital cameras can be used to characterize germplasm for drought tolerance. To achieve this goal, we built a proximal phenotyping cart mounted with digital cameras and evaluated it by characterizing 142 winter wheat genotypes for drought tolerance under field conditions. The same genotypes were additionally characterized for seedling stage traits by imaging under controlled growth conditions. The analysis revealed that under field conditions, plant biomass, relative growth rates, and Normalized Difference Vegetation Index (NDVI) from different growth stages estimated by imaging were significantly correlated to drought tolerance. Under controlled growth conditions, root count at the seedling stage evaluated by imaging was significantly correlated to adult plant drought tolerance observed in the field. Random forest models were trained by integrating measurements from field and controlled conditions and revealed that plant biomass and relative growth rates at key plant growth stages are important predictors of drought tolerance. Thus, based on the results, it can be concluded that the consumer-grade cameras can be key components of affordable automated phenotyping systems to accelerate pre-breeding for drought tolerance

Almond diversity and homozygosity define structure, kinship, inbreeding, and linkage disequilibrium in cultivated germplasm, and reveal genomic associations with nut and seed weight

Almond [Prunus dulcis Miller (D.A. Webb)] is the main tree nut species worldwide. Here, genotyping-by-sequencing (GBS) was applied to 149 almond cultivars from the ex situ collections of the Italian Council for Agricultural Research (CREA) and the Spanish National Research Council (CSIC), leading to the detection of 93,119 single-nucleotide polymorphisms (SNPs). The study of population structure outlined four distinct genetic groups and highlighted diversification between the Mediterranean and Californian gene pools. Data on SNP diversity and runs of homozygosity (ROHs) allowed the definition of kinship, inbreeding, and linkage disequilibrium (LD) decay in almond cultivated germplasm. Four-year phenotypic observations, gathered on 98 cultivars of the CREA collection, were used to perform a genome-wide association study (GWAS) and, for the first time in a crop species, homozygosity mapping (HM), resulting in the identification of genomic associations with nut, shell, and seed weight. Both GWAS and HM suggested that loci controlling nut and seed weight are mostly independent. Overall, this study provides insights on the almond cultivation history and delivers information of major interest for almond genetics and breeding. In a broader perspective, our results encourage the use of ROHs in crop science to estimate inbreeding, choose parental combinations minimizing the risk of inbreeding depression, and identify genomic footprints of selection for specific traits

On the origin of European sheep as revealed by the diversity of the Balkan breeds and by optimizing population-genetic analysis tools

Background: In the Neolithic, domestic sheep migrated into Europe and subsequently spread in westerly and northwesterly directions. Reconstruction of these migrations and subsequent genetic events requires a more detailed characterization of the current phylogeographic differentiation. Results: We collected 50 K single nucleotide polymorphism (SNP) profiles of Balkan sheep that are currently found near the major Neolithic point of entry into Europe, and combined these data with published genotypes from southwest-Asian, Mediterranean, central-European and north-European sheep and from Asian and European mouflons. We detected clines, ancestral components and admixture by using variants of common analysis tools: geography-informative supervised principal component analysis (PCA), breed-specific admixture analysis, across-breed f 4 profiles and phylogenetic analysis of regional pools of breeds. The regional Balkan sheep populations exhibit considerable genetic overlap, but are clearly distinct from the breeds in surrounding regions. The Asian mouflon did not influence the differentiation of the European domestic sheep and is only distantly related to present-day sheep, including those from Iran where the mouflons were sampled. We demonstrate the occurrence, from southeast to northwest Europe, of a continuously increasing ancestral component of up to 20% contributed by the European mouflon, which is assumed to descend from the original Neolithic domesticates. The overall patterns indicate that the Balkan region and Italy served as post-domestication migration hubs, from which wool sheep reached Spain and north Italy with subsequent migrations northwards. The documented dispersal of Tarentine wool sheep during the Roman period may have been part of this process. Our results also reproduce the documented 18th century admixture of Spanish Merino sheep into several central-European breeds. Conclusions: Our results contribute to a better understanding of the events that have created the present diversity pattern, which is relevant for the management of the genetic resources represented by the European sheep population

Genotyping-by-Sequencing in Vigna unguiculata Landraces and Its Utility for Assessing Taxonomic Relationships

Genotyping by sequencing (GBS) was used to analyze relationships among cowpea and asparagus bean landraces from southern Italy and to assess the utility of this technology to study taxonomy in a wider panel, including V. unguiculata cultigroups, subspecies, and other Vigna species. The analysis of SNPs derived from GBS highlighted that, among the cowpea landraces, the African samples were separated from the other material, while, for the Italian landraces, a certain clustering depending on seed color/pattern was observed in the dendrogram. When examining the V. unguiculata species complex, a clear separation between the two groups of wild subspecies, i.e., the allogamous wild perennials and the perennial out/inbreds, could be observed, the former representing the more ancestral wild progenitors of V. unguiculata. The species V. vexillata appeared more closely related to V. unguiculata than to the other Vigna species analyzed

Genotyping-by-Sequencing in Vigna unguiculata Landraces and Its Utility for Assessing Taxonomic Relationships

Genotyping by sequencing (GBS) was used to analyze relationships among cowpea and asparagus bean landraces from southern Italy and to assess the utility of this technology to study taxonomy in a wider panel, including V. unguiculata cultigroups, subspecies, and other Vigna species. The analysis of SNPs derived from GBS highlighted that, among the cowpea landraces, the African samples were separated from the other material, while, for the Italian landraces, a certain clustering depending on seed color/pattern was observed in the dendrogram. When examining the V. unguiculata species complex, a clear separation between the two groups of wild subspecies, i.e., the allogamous wild perennials and the perennial out/inbreds, could be observed, the former representing the more ancestral wild progenitors of V. unguiculata. The species V. vexillata appeared more closely related to V. unguiculata than to the other Vigna species analyzed

Recommendations for choosing the genotyping method and best practices for quality control in crop genome-wide association studies

High-throughput genotyping is boosting genome-wide association studies (GWAS) in crop species, ultimately leading to the identification of single nucleotide polymorphisms (SNPs) and genes associated with economically important traits. Choosing a cost-effective genotyping method for crop GWAS requires careful examination of several aspects, namely the purpose and the scale of the study, crop-specific genomic features, and technical and economic matters associated with each genotyping option. Once genotypic data have been obtained, quality control (QC) procedures must be applied to avoid bias and false signals in genotype-phenotype association tests. QC for human GWAS has been extensively reviewed, however QC for crop GWAS may require different actions, depending on the mating system and breeding activities. Here we review most popular genotyping methods based on next generation sequencing (NGS) and array hybridization, and provide observations that should guide the investigator in the choice of the genotyping method for crop GWAS. We define best practises to perform QC, including actions that are specific for crop species. Finally, we provide an overview of bioinformatics tools that can be used to accomplish all needed tasks. Overall, this work aims to provide guidelines to harmonize those procedures leading to SNP datasets ready for crop GWAS

Genotyping-by-Sequencing Defines Genetic Structure within the “Acquaviva” Red Onion Landrace

Genetic structure and distinctive features of landraces, such as adaptability to local agro-ecosystems and specific qualitative profiles, can be substantially altered by the massive introduction of allochthonous germplasm. The landrace known as “Cipolla rossa di Acquaviva” (Acquaviva red onion, further referred to as ARO) is traditionally cultivated and propagated in a small area of the Apulia region (southern Italy). However, the recent rise of its market value and cultivation area is possibly causing genetic contamination with foreign propagating material. In this work, genotyping-by-sequencing (GBS) was used to characterize genetic variation of seven onion populations commercialized as ARO, as well as one population of the landrace “Montoro” (M), which is phenotypically similar, but originates from another cultivation area and displays different qualitative features. A panel of 5011 SNP markers was used to perform parametric and non-parametric genetic structure analyses, which supported the hypothesis of genetic contamination of germplasm commercialized as ARO with a gene pool including the M landrace. Four ARO populations formed a core genetic group, homogeneous and clearly distinct from the other ARO and M populations. Conversely, the remaining three ARO populations did not display significant differences with the M population. A set of private alleles for the ARO core genetic group was identified, indicating the possibility to trace the ARO landrace by means of a SNP-based molecular barcode. Overall, the results of this study provide a framework for further breeding activities and the traceability of the ARO landrace

Supplementary Material. Detection and distribution of two alleles of the Sk gene, controlling the kernel taste in almond [Prunus dulcis Miller (D.A. Webb)]

Supplementary Figures and Tables. Supplementary Figure 1. ADMIXTURE cross-validation (CV) error estimates for a number of ancestral populations (K) ranging from 1 to 10. The red triangle indicates the lowest CV error detected for K=4 Supplementary Table 1. Results from genotyping 134 almond cultivars with the marker assays developed in this study. T1036 and G989 are indicative of the Sk-1 and Sk-2 alleles, respectively. Genotypic calls from the KASP and dual label assays were fully consistent, thus they are reported as a single column for each nucleotide position. Genotyping of cultivars indicated in bold was confirmed by Sanger sequencingPeer reviewe