A novel tomato interspecific (Solanum lycopersicum var. cerasiforme and Solanum pimpinellifolium) MAGIC population facilitates trait association and candidate gene discovery in untapped exotic germplasm.

We developed a novel eight-way tomato multiparental advanced generation intercross (MAGIC) population to improve the accessibility of tomato relatives genetic resources to geneticists and breeders. The interspecific tomato MAGIC population (ToMAGIC) was obtained by intercrossing four accessions each of Solanum lycopersicum var. cerasiforme and Solanum pimpinellifolium, which are the weedy relative and the ancestor of cultivated tomato, respectively. The eight exotic ToMAGIC founders were selected based on a representation of the genetic diversity and geographical distribution of the two taxa. The resulting MAGIC population comprises 354 lines, which were genotyped using a new 12k tomato single primer enrichment technology panel and yielded 6488 high-quality single-nucleotide polymorphism (SNPs). The genotyping data revealed a high degree of homozygosity, an absence of genetic structure, and a balanced representation of the founder genomes. To evaluate the potential of the ToMAGIC population, a proof of concept was conducted by phenotyping it for fruit size, plant pigmentation, leaf morphology, and earliness. Genome-wide association studies identified strong associations for the studied traits, pinpointing both previously identified and novel candidate genes near or within the linkage disequilibrium blocks. Domesticated alleles for fruit size were recessive and were found, at low frequencies, in wild/ancestral populations. Our findings demonstrate that the newly developed ToMAGIC population is a valuable resource for genetic research in tomato, offering significant potential for identifying new genes that govern key traits in tomato. ToMAGIC lines displaying a pyramiding of traits of interest could have direct applicability for integration into breeding pipelines providing untapped variation for tomato breeding.

Growth and antioxidant responses to water stress in eggplant MAGIC population parents, F1 hybrids and a subset of recombinant inbred lines.

BACKGROUND: The generation of new eggplant (Solanum melongena L.) cultivars with drought tolerance is a main challenge in the current context of climate change. In this study, the eight parents (seven of S. melongena and one of the wild relative S. incanum L.) of the first eggplant MAGIC (Multiparent Advanced Generation Intercrossing) population, together with four F1 hybrids amongst them, five S5 MAGIC recombinant inbred lines selected for their genetic diversity, and one commercial hybrid were evaluated in young plant stage under water stress conditions (30% field capacity; FC) and control conditions (100% FC). After a 21-day treatment period, growth and biomass traits, photosynthetic pigments, oxidative stress markers, antioxidant compounds, and proline content were evaluated. RESULTS: Significant effects (p < 0.05) were observed for genotype, water treatments and their interaction in most of the traits analyzed. The eight MAGIC population parental genotypes displayed a wide variation in their responses to water stress, with some of them exhibiting enhanced root development and reduced foliar biomass. The commercial hybrid had greater aerial growth compared to root growth. The four F1 hybrids among MAGIC parents differed in their performance, with some having significant positive or negative heterosis in several traits. The subset of five MAGIC lines displayed a wide diversity in their response to water stress. CONCLUSION: The results show that a large diversity for tolerance to drought is available among the eggplant MAGIC materials, which can contribute to developing drought-tolerant eggplant cultivars.

Characterization of Browning, Chlorogenic Acid Content, and Polyphenol Oxidase Activity in Different Varietal Types of Eggplant (Solanum melongena) for Improving Visual and Nutritional Quality.

Eggplant (Solanum melongena L.) breeding for fruit quality has mostly focused on visual traits and nutritional and bioactive compounds, including chlorogenic acid. However, higher contents of chlorogenic acid may lead to more pronounced fruit flesh browning. We examined a diverse collection of 59 eggplant accessions across five varietal types ('black oval', 'striped', 'anthocyanin-free', 'purple', and 'black elongated') to evaluate the degree of browning, polyphenol oxidase (PPO) activity, and chlorogenic acid (CGA) content. The results reveal moderate correlations among these traits, with no clear differences among the varietal types, suggesting that other factors, including genetic variation, might significantly influence these traits. Notably, 'black oval' accessions demonstrated higher browning and PPO activity, whereas 'striped' accessions showed low variability. The identification of genotypes with lower browning and higher CGA content highlights opportunities for targeted genotype selection to improve eggplant chlorogenic acid content while maintaining low or moderate browning, pointing towards the importance of genetic considerations in breeding strategies to reduce browning and enhance nutritional value.

Nitrogen use efficiency, growth and physiological parameters in different tomato genotypes under high and low N fertilisation conditions.

Identification of novel genotypes with enhanced nitrogen use efficiency (NUE) is a key challenge for a sustainable tomato production. In this respect, the performance of a panel of thirty tomato accessions were evaluated under high (HN; 5 mM N) and low (LN; 0.5 mM N) nitrogen irrigation solutions. For each treatment, when 50% of plants reached the first flower bud stage, plant growth and biomass traits, chlorophyll, flavonol and anthocyanin indexes, nitrogen balance index (NBI), C:N ratio in leaves, stems, and roots, and NUE were evaluated. Significant (p < 0.05) effects were observed for accession, N treatment, and their interaction across all the traits. Under LN, plants showed a delayed development (40 days for HN vs. 65 days for LN) and reduced growth and biomass. On average, LN condition led to 41.8% decrease in nitrogen uptake efficiency (NUpE) but also 189.0% increase in NUtE, resulting in 62.2% overall increase in NUE. A broad range of variation among accessions was observed under both HN and LN conditions. Under LN conditions, chlorophyll index and NBI decreased, while flavonol and anthocyanin indexes increased. Leaf C:N ratio was positively correlated with nitrogen utilisation efficiency (NUtE) in both N treatments. Multi-trait analyses identified top-performing accessions under each condition, allowing to identify one accession among top performers under both conditions. Correlation analysis revealed that high root biomass and leaf C:N ratio are useful markers for selecting high NUE accessions. These findings offer valuable insights for improving tomato NUE under varying nitrogen fertilization conditions and for breeding high-NUE cultivars.

Evaluation of three sets of advanced backcrosses of eggplant with wild relatives from different gene pools under low N fertilization conditions.

The development of new cultivars with improved nitrogen use efficiency (NUE) is key for implementing sustainable agriculture practices. Crop wild relatives (CWRs) provide valuable genetic resources for breeding programs aimed at achieving this goal. In this study, three eggplant (Solanum melongena) accessions together with their advanced backcrosses (ABs; BC3 to BC5 generations) were evaluated for 22 morpho-agronomic, physiological, and NUE traits under low nitrogen (LN) fertilization conditions. The ABs were developed with introgressions from the wild relatives Solanum insanum, Solanum dasyphyllum, and Solanum elaeagnifolium. The AB population comprised a total of 25, 59, and 59 genotypes, respectively, with overall donor wild relative genome coverage percentages of 58.8%, 46.3%, and 99.2%. The three S. melongena recurrent parents were also evaluated under control (normal) N fertilization. Reduction of N fertilization in the parents resulted in decreased chlorophyll content-related traits, aerial biomass, stem diameter, and yield and increased NUE, nitrogen uptake efficiency (NUpE), and nitrogen utilization efficiency (NUtE). However, the decrease in yield was moderate, ranging between 62.6% and 72.6%. A high phenotypic variation was observed within each of the three sets of ABs under LN conditions, with some individuals displaying improved transgressive characteristics over the recurrent parents. Using the single primer enrichment technology 5 k probes platform for high-throughput genotyping, we observed a variable but high degree of recurrent parent genome recovery in the ABs attributable to the lines recombination, allowing the successful identification of 16 quantitative trait loci (QTL). Different allelic effects were observed for the introgressed QTL alleles. Several candidate genes were identified in the QTL regions associated with plant growth, yield, fruit size, and NUE-related parameters. Our results show that eggplant materials with introgressions from CWRs can result in a dramatic impact in eggplant breeding for a more sustainable agriculture.

Conventional and new genetic resources for an eggplant breeding revolution.

Eggplant (Solanum melongena) is a major vegetable crop with great potential for genetic improvement owing to its large and mostly untapped genetic diversity. It is closely related to over 500 species of Solanum subgenus Leptostemonum that belong to its primary, secondary, and tertiary genepools and exhibit a wide range of characteristics useful for eggplant breeding, including traits adaptive to climate change. Germplasm banks worldwide hold more than 19 000 accessions of eggplant and related species, most of which have yet to be evaluated. Nonetheless, eggplant breeding using the cultivated S. melongena genepool has yielded significantly improved varieties. To overcome current breeding challenges and for adaptation to climate change, a qualitative leap forward in eggplant breeding is necessary. The initial findings from introgression breeding in eggplant indicate that unleashing the diversity present in its relatives can greatly contribute to eggplant breeding. The recent creation of new genetic resources such as mutant libraries, core collections, recombinant inbred lines, and sets of introgression lines will be another crucial element and will require the support of new genomics tools and biotechnological developments. The systematic utilization of eggplant genetic resources supported by international initiatives will be critical for a much-needed eggplant breeding revolution to address the challenges posed by climate change.

Comparative Analysis of Phenotypic and Molecular Data on Response to Main Pear Diseases and Pest Attack in a Germplasm Collection.

The pear is an important fruit tree in temperate areas, but due to its sensitivity, fruit yield and quality are often affected by disease and pest attacks. Pear genotypes from a germplasm collection comprising 13 Pyrus species, 17 Romanian varieties, and 50 non-Romanian varieties from a worldwide assortment were investigated in this study. Throughout four years, response to attack of the principal pathogens and pests was investigated phenotypically under natural conditions of infection and infestation. SSR markers were used to analyze the genetic diversity of the genotypes. A standardized method for the evaluation of responses to biotic stressors was proposed, which highlighted significant differences between genotypes. The species and varieties with the lowest degrees of attack (DA%), calculated based on the frequency and intensity of attack, were identified for pear scab (Venturia pyrina), septoria (Septoria pyricola), fire blight (Erwinia amylovora), and psyllids (Psylla sp.). These accessions could provide valuable sources of genes of interest to develop resistant varieties in new pear breeding programs. By combining phenotypic and molecular analyses, significant information was obtained that can be exploited to generate high variability for selection through artificial hybridization by harnessing accessions with complementary molecular fingerprints and high genetic distances.

Mutations in the SmAPRR2 transcription factor suppressing chlorophyll pigmentation in the eggplant fruit peel are key drivers of a diversified colour palette.

Understanding the mechanisms by which chlorophylls are synthesized in the eggplant (Solanum melongena) fruit peel is of great relevance for eggplant breeding. A multi-parent advanced generation inter-cross (MAGIC) population and a germplasm collection have been screened for green pigmentation in the fruit peel and used to identify candidate genes for this trait. A genome-wide association study (GWAS) performed with 420 MAGIC individuals revealed a major association on chromosome 8 close to a gene similar to APRR2. Two variants in SmAPRR2, predicted as having a high impact effect, were associated with the absence of fruit chlorophyll pigmentation in the MAGIC population, and a large deletion of 5.27 kb was found in two reference genomes of accessions without chlorophyll in the fruit peel. The validation of the candidate gene SmAPRR2 was performed by its sequencing in a set of MAGIC individuals and through its de novo assembly in 277 accessions from the G2P-SOL eggplant core collection. Two additional mutations in SmAPRR2 associated with the lack of chlorophyll were identified in the core collection set. The phylogenetic analysis of APRR2 reveals orthology within Solanaceae and suggests that specialization of APRR2-like genes occurred independently in Cucurbitaceae and Solanaceae. A strong geographical differentiation was observed in the frequency of predominant mutations in SmAPRR2, resulting in a lack of fruit chlorophyll pigmentation and suggesting that this phenotype may have arisen and been selected independently several times. This study represents the first identification of a major gene for fruit chlorophyll pigmentation in the eggplant fruit.

Biological Traits and Genetic Relationships Amongst Cultivars of Three Species of Tagetes (Asteraceae).

Marigolds (Tagetes spp.) are multifunctional flowering plants belonging to the Asteraceae family, well-known and widespread for their ornamental value and many other uses. In this study, morphological differences and genetic relationships among 21 cultivars of three species of marigold (Tagetes patula, T. erecta and T. tenuifolia) were analysed. Results have revealed obvious differences among genotypes, starting from the morphological characteristics of the seeds and their capacity to germinate to adult plant morphological characteristics, both between cultivars and species. The genotypic differences were manifested in considerable variation in the development of phenological stages and the main morphological traits of plants and flowers. PCA and hierarchical clustering analyses of morphological traits revealed a homogeneous grouping of cultivars within each species, except for Orion, belonging to T. patula, which was closer to T. erecta cultivars. A subset of 13 cultivars from the three species was subjected to SSR analysis, revealing considerable genetic diversity and good separation between T. patula on the one side and T. erecta and T. tenuifolia on the other. The observed heterozygosity was much lower than the expected heterozygosity, revealing a high degree of fixation. The results reveal that the three species evaluated have considerable morphological and genetic diversity, which has important implications for assessing genetic diversity, conserving germplasm and selecting parents for new breeding works in marigolds.

Newly Developed MAGIC Population Allows Identification of Strong Associations and Candidate Genes for Anthocyanin Pigmentation in Eggplant.

Multi-parent advanced generation inter-cross (MAGIC) populations facilitate the genetic dissection of complex quantitative traits in plants and are valuable breeding materials. We report the development of the first eggplant MAGIC population (S3 Magic EGGplant InCanum, S3MEGGIC; 8-way), constituted by the 420 S3 individuals developed from the intercrossing of seven cultivated eggplant (Solanum melongena) and one wild relative (S. incanum) parents. The S3MEGGIC recombinant population was genotyped with the eggplant 5k probes SPET platform and phenotyped for anthocyanin presence in vegetative plant tissues (PA) and fruit epidermis (FA), and for the light-insensitive anthocyanic pigmentation under the calyx (PUC). The 7,724 filtered high-confidence single-nucleotide polymorphisms (SNPs) confirmed a low residual heterozygosity (6.87%), a lack of genetic structure in the S3MEGGIC population, and no differentiation among subpopulations carrying a cultivated or wild cytoplasm. Inference of haplotype blocks of the nuclear genome revealed an unbalanced representation of the founder genomes, suggesting a cryptic selection in favour or against specific parental genomes. Genome-wide association study (GWAS) analysis for PA, FA, and PUC detected strong associations with two myeloblastosis (MYB) genes similar to MYB113 involved in the anthocyanin biosynthesis pathway, and with a COP1 gene which encodes for a photo-regulatory protein and may be responsible for the PUC trait. Evidence was found of a duplication of an ancestral MYB113 gene with a translocation from chromosome 10 to chromosome 1 compared with the tomato genome. Parental genotypes for the three genes were in agreement with the identification of the candidate genes performed in the S3MEGGIC population. Our new eggplant MAGIC population is the largest recombinant population in eggplant and is a powerful tool for eggplant genetics and breeding studies.

Ploidy Modification for Plant Breeding Using In Vitro Organogenesis: A Case in Eggplant.

The use of antimitotic agents such as colchicine has been common to obtain polyploid organisms. However, this approach entails certain problems, from its toxicity to the operators for being carcinogenic compounds to the instability of the individuals obtained, and the consequent reversion to its original ploidy because the individuals obtained in most cases are chimeric. In vitro culture allows taking advantage of the full potential offered by the cellular totipotence of plant organisms. Based on this, we present a new in vitro culture protocol to obtain polyploid organisms using zeatin riboside (ZR) and eggplant as a model organism. Flow cytometry is used to identify tetraploid regenerants. The regeneration of whole plants from the appropriate tissues using ZR allowed developing polyploid individuals in eggplant, a crop that tends to be recalcitrant to in vitro organogenesis. Thanks to the use of the polysomatic pattern of the explants, we have been able to develop a methodology that allows to obtain stable non-chimeric polyploid individuals from organogenic processes.

Morphoagronomic characterization and whole-genome resequencing of eight highly diverse wild and weedy S. pimpinellifolium and S. lycopersicum var. cerasiforme accessions used for the first interspecific tomato MAGIC population.

The wild Solanum pimpinellifolium (SP) and the weedy S. lycopersicum var. cerasiforme (SLC) are largely unexploited genetic reservoirs easily accessible to breeders, as they are fully cross-compatible with cultivated tomato (S. lycopersicum var. lycopersicum). We performed a comprehensive morphological and genomic characterization of four wild SP and four weedy SLC accessions, selected to maximize the range of variation of both taxa. These eight accessions are the founders of the first tomato interspecific multi-parent advanced generation inter-cross (MAGIC) population. The morphoagronomic characterization was carried out with 39 descriptors to assess plant, inflorescence, fruit and agronomic traits, revealing the broad range of diversity captured. Part of the morphological variation observed in SP was likely associated to the adaptation of the accessions to different environments, while in the case of SLC to both human activity and adaptation to the environment. Whole-genome resequencing of the eight accessions revealed over 12 million variants, ranging from 1.2 to 1.9 million variants in SLC and from 3.1 to 4.8 million in SP, being 46.3% of them (4,897,803) private variants. The genetic principal component analysis also confirmed the high diversity of SP and the complex evolutionary history of SLC. This was also reflected in the analysis of the potential footprint of common ancestors or old introgressions identified within and between the two taxa. The functional characterization of the variants revealed a significative enrichment of GO terms related to changes in cell walls that would have been negatively selected during domestication and breeding. The comprehensive morphoagronomic and genetic characterization of these accessions will be of great relevance for the genetic analysis of the first interspecific MAGIC population of tomato and provides valuable knowledge and tools to the tomato community for genetic and genomic studies and for breeding purposes.

A Deep Learning-Based System (Microscan) for the Identification of Pollen Development Stages and Its Application to Obtaining Doubled Haploid Lines in Eggplant.

The development of double haploids (DHs) is a straightforward path for obtaining pure lines but has multiple bottlenecks. Among them is the determination of the optimal stage of pollen induction for androgenesis. In this work, we developed Microscan, a deep learning-based system for the detection and recognition of the stages of pollen development. In a first experiment, the algorithm was developed adapting the RetinaNet predictive model using microspores of different eggplant accessions as samples. A mean average precision of 86.30% was obtained. In a second experiment, the anther range to be cultivated in vitro was determined in three eggplant genotypes by applying the Microscan system. Subsequently, they were cultivated following two different androgenesis protocols (Cb and E6). The response was only observed in the anther size range predicted by Microscan, obtaining the best results with the E6 protocol. The plants obtained were characterized by flow cytometry and with the Single Primer Enrichment Technology high-throughput genotyping platform, obtaining a high rate of confirmed haploid and double haploid plants. Microscan has been revealed as a tool for the high-throughput efficient analysis of microspore samples, as it has been exemplified in eggplant by providing an increase in the yield of DHs production.

The Dawn of the Age of Multi-Parent MAGIC Populations in Plant Breeding: Novel Powerful Next-Generation Resources for Genetic Analysis and Selection of Recombinant Elite Material.

The compelling need to increase global agricultural production requires new breeding approaches that facilitate exploiting the diversity available in the plant genetic resources. Multi-parent advanced generation inter-cross (MAGIC) populations are large sets of recombinant inbred lines (RILs) that are a genetic mosaic of multiple founder parents. MAGIC populations display emerging features over experimental bi-parental and germplasm populations in combining significant levels of genetic recombination, a lack of genetic structure, and high genetic and phenotypic diversity. The development of MAGIC populations can be performed using "funnel" or "diallel" cross-designs, which are of great relevance choosing appropriate parents and defining optimal population sizes. Significant advances in specific software development are facilitating the genetic analysis of the complex genetic constitutions of MAGIC populations. Despite the complexity and the resources required in their development, due to their potential and interest for breeding, the number of MAGIC populations available and under development is continuously growing, with 45 MAGIC populations in different crops being reported here. Though cereals are by far the crop group where more MAGIC populations have been developed, MAGIC populations have also started to become available in other crop groups. The results obtained so far demonstrate that MAGIC populations are a very powerful tool for the dissection of complex traits, as well as a resource for the selection of recombinant elite breeding material and cultivars. In addition, some new MAGIC approaches that can make significant contributions to breeding, such as the development of inter-specific MAGIC populations, the development of MAGIC-like populations in crops where pure lines are not available, and the establishment of strategies for the straightforward incorporation of MAGIC materials in breeding pipelines, have barely been explored. The evidence that is already available indicates that MAGIC populations will play a major role in the coming years in allowing for impressive gains in plant breeding for developing new generations of dramatically improved cultivars.

SILEX: a fast and inexpensive high-quality DNA extraction method suitable for multiple sequencing platforms and recalcitrant plant species.

BACKGROUND: The use of sequencing and genotyping platforms has undergone dramatic improvements, enabling the generation of a wealth of genomic information. Despite this progress, the availability of high-quality genomic DNA (gDNA) in sufficient concentrations is often a main limitation, especially for third-generation sequencing platforms. A variety of DNA extraction methods and commercial kits are available. However, many of these are costly and frequently give either low yield or low-quality DNA, inappropriate for next generation sequencing (NGS) platforms. Here, we describe a fast and inexpensive DNA extraction method (SILEX) applicable to a wide range of plant species and tissues. RESULTS: SILEX is a high-throughput DNA extraction protocol, based on the standard CTAB method with a DNA silica matrix recovery, which allows obtaining NGS-quality high molecular weight genomic plant DNA free of inhibitory compounds. SILEX was compared with a standard CTAB extraction protocol and a common commercial extraction kit in a variety of species, including recalcitrant ones, from different families. In comparison with the other methods, SILEX yielded DNA in higher concentrations and of higher quality. Manual extraction of 48 samples can be done in 96 min by one person at a cost of 0.12 €/sample of reagents and consumables. Hundreds of tomato gDNA samples obtained with either SILEX or the commercial kit were successfully genotyped with Single Primer Enrichment Technology (SPET) with the Illumina HiSeq 2500 platform. Furthermore, DNA extracted from Solanum elaeagnifolium using this protocol was assessed by Pulsed-field gel electrophoresis (PFGE), obtaining a suitable size ranges for most sequencing platforms that required high-molecular-weight DNA such as Nanopore or PacBio. CONCLUSIONS: A high-throughput, fast and inexpensive DNA extraction protocol was developed and validated for a wide variety of plants and tissues. SILEX offers an easy, scalable, efficient and inexpensive way to extract DNA for various next-generation sequencing applications including SPET and Nanopore among others.

Fostering Conservation via an Integrated Use of Conventional Approaches and High-Throughput SPET Genotyping: A Case Study Using the Endangered Canarian Endemics Solanum lidii and S. vespertilio (Solanaceae).

Islands provide unique opportunities to integrated research approaches to study evolution and conservation because boundaries are circumscribed, geological ages are often precise, and many taxa are greatly imperiled. We combined morphological and hybridization studies with high-throughput genotyping platforms to streamline relationships in the endangered monophyletic and highly diverse lineage of Solanum in the Canarian archipelago, where three endemic taxa are currently recognized. Inter-taxa hybridizations were performed, and morphological expression was assessed with a common-garden approach. Using the eggplant Single Primer Enrichment Technology (SPET) platform with 5,093 probes, 74 individuals of three endemic taxa (Solanum lidii, S. vespertilio subsp. vespertilio, and S. vespertilio subsp. doramae) were sampled for SNPs. While morphological and breeding studies showed clear distinctions and some continuous variation, inter-taxon hybrids were fertile and heterotic for vigor traits. SPET genotyping revealed 1,421 high-quality SNPs and supported four, not three, distinct taxonomic entities associated with post-emergence geological, ecological and geographic factors of the islands. Given the lack of barriers to hybridization among all the taxa and their molecular differences, great care must be taken in population management. Conservation strategies must take account of the sexual and breeding systems and genotypic distribution among populations to successfully conserve and restore threatened/endangered island taxa, as exemplified by Solanum on the Canary Islands.

A highly efficient organogenesis protocol based on zeatin riboside for in vitro regeneration of eggplant.

BACKGROUND: Efficient organogenesis induction in eggplant (Solanum melongena L.) is required for multiple in vitro culture applications. In this work, we aimed at developing a universal protocol for efficient in vitro regeneration of eggplant mainly based on the use of zeatin riboside (ZR). We evaluated the effect of seven combinations of ZR with indoleacetic acid (IAA) for organogenic regeneration in five genetically diverse S. melongena and one S. insanum L. accessions using two photoperiod conditions. In addition, the effect of six different concentrations of indolebutyric acid (IBA) in order to promote rooting was assessed to facilitate subsequent acclimatization of plants. The ploidy level of regenerated plants was studied. RESULTS: In a first experiment with accessions MEL1 and MEL3, significant (p < 0.05) differences were observed for the four factors evaluated for organogenesis from cotyledon, hypocotyl and leaf explants, with the best results obtained (9 and 11 shoots for MEL1 and MEL3, respectively) using cotyledon tissue, 16 h light / 8 h dark photoperiod conditions, and medium E6 (2 mg/L of ZR and 0 mg/L of IAA). The best combination of conditions was tested in the other four accessions and confirmed its high regeneration efficiency per explant when using both cotyledon and hypocotyl tissues. The best rooting media was R2 (1 mg/L IBA). The analysis of ploidy level revealed that between 25 and 50% of the regenerated plantlets were tetraploid. CONCLUSIONS: An efficient protocol for organogenesis of both cultivated and wild accessions of eggplant, based on the use of ZR, is proposed. The universal protocol developed may be useful for fostering in vitro culture applications in eggplant requiring regeneration of plants and, in addition, allows developing tetraploid plants without the need of antimitotic chemicals.

Whole-Genome Resequencing of Seven Eggplant (Solanum melongena) and One Wild Relative (S. incanum) Accessions Provides New Insights and Breeding Tools for Eggplant Enhancement.

Whole-genome resequencing provides information of great relevance for crop genetics, evolution, and breeding. Here, we present the first whole-genome resequencing study using seven eggplant (Solanum melongena) and one wild relative (Solanum incanum) accessions. These eight accessions were selected for displaying a high phenotypic and genetic diversity and for being the founder parents of an eggplant multiparent advanced generation intercrosses population. By resequencing at an average depth of 19.8× and comparing to the high-quality reference genome "67/3" over 10 million high-reliable polymorphisms were discovered, of which over 9 million (84.5%) were single nucleotide polymorphisms and more than 700,000 (6.5%) InDels. However, while for the S. melongena accessions, the variants identified ranged from 0.8 to 1.3 million, over 9 million were detected for the wild S. incanum. This confirms the narrow genetic diversity of the domesticated eggplant and suggests that introgression breeding using wild relatives can efficiently contribute to broadening the genetic basis of this crop. Differences were observed among accessions for the enrichment in genes regulating important biological processes. By analyzing the distribution of the variants, we identified the potential footprints of old introgressions from wild relatives that can help to unravel the unclear domestication and breeding history. The comprehensive annotation of these eight genomes and the information provided in this study represents a landmark in eggplant genomics and allows the development of tools for eggplant genetics and breeding.

Single Primer Enrichment Technology (SPET) for High-Throughput Genotyping in Tomato and Eggplant Germplasm.

Single primer enrichment technology (SPET) is a new, robust, and customizable solution for targeted genotyping. Unlike genotyping by sequencing (GBS), and like DNA chips, SPET is a targeted genotyping technology, relying on the sequencing of a region flanking a primer. Its reliance on single primers, rather than on primer pairs, greatly simplifies panel design, and allows higher levels of multiplexing than PCR-based genotyping. Thanks to the sequencing of the regions surrounding the target SNP, SPET allows the discovery of thousands of closely linked, novel SNPs. In order to assess the potential of SPET for high-throughput genotyping in plants, a panel comprising 5k target SNPs, designed both on coding regions and introns/UTRs, was developed for tomato and eggplant. Genotyping of two panels composed of 400 tomato and 422 eggplant accessions, comprising both domesticated material and wild relatives, generated a total of 12,002 and 30,731 high confidence SNPs, respectively, which comprised both target and novel SNPs in an approximate ratio of 1:1.6, and 1:5.5 in tomato and eggplant, respectively. The vast majority of the markers was transferrable to related species that diverged up to 3.4 million years ago (Solanum pennellii for tomato and S. macrocarpon for eggplant). Maximum Likelihood phylogenetic trees and PCA outputs obtained from the whole dataset highlighted genetic relationships among accessions and species which were congruent with what was previously reported in literature. Better discrimination among domesticated accessions was achieved by using the target SNPs, while better discrimination among wild species was achieved using the whole SNP dataset. Our results reveal that SPET genotyping is a robust, high-throughput technology for genetic fingerprinting, with a high degree of cross-transferability between crops and their cultivated and wild relatives, and allows identification of duplicates and mislabeled accessions in genebanks.

Genetic diversity, population structure, and relationships in a collection of pepper (Capsicum spp.) landraces from the Spanish centre of diversity revealed by genotyping-by-sequencing (GBS).

Pepper (Capsicum spp.) is one of the most important vegetable crops; however, pepper genomic studies lag behind those of other important Solanaceae. Here we present the results of a high-throughput genotyping-by-sequencing (GBS) study of a collection of 190 Capsicum spp. accessions, including 183 of five cultivated species (C. annuum, C. chinense, C. frutescens, C. baccatum, and C. pubescens) and seven of the wild form C. annuum var. glabriusculum. Sequencing generated 6,766,231 high-quality read tags, of which 40.7% were successfully aligned to the reference genome. SNP calling yielded 4083 highly informative segregating SNPs. Genetic diversity and relationships of a subset of 148 accessions, of which a complete passport information was available, was studied using principal components analysis (PCA), discriminant analysis of principal components (DAPC), and phylogeny approaches. C. annuum, C. baccatum, and C. chinense were successfully separated by all methods. Our population was divided into seven clusters by DAPC, where C. frutescens accessions were clustered together with C. chinense. C. annuum var. glabriusculum accessions were spread into two distinct genetic pools, while European accessions were admixed and closely related. Separation of accessions was mainly associated to differences in fruit characteristics and origin. Phylogeny studies showed a close relation between Spanish and Mexican accessions, supporting the hypothesis that the first arose from a main genetic flow from the latter. Tajima's D statistic values were consistent with positive selection in the C. annuum clusters, possibly related to domestication or selection towards traits of interest. This work provides comprehensive and relevant information on the origin and relationships of Spanish landraces and for future association mapping studies in pepper.