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Olive tree (Olea europaea L. subsp. europaea var. europaea) is one of the most important species of the Mediterranean region and one of the most ancient species domesticated. The availability of whole genome assemblies and annotations of olive tree cultivars and oleaster (O. europaea subsp. europaea var. sylvestris) has contributed to a better understanding of genetic and genomic differences between olive tree cultivars. However, compared to other plant species there is still a lack of genomic resources for olive tree popula-tions that span the entire Mediterranean region. In the present study we developed the most complete genomic variation map and the most comprehensive catalog/resource of molecular variation to date for 89 olive tree genotypes originating from the entire Mediterranean basin, revealing the genetic diversity of this commercially significant crop tree and explaining the divergence/similarity among different variants. Addi-tionally, the monumental ancient tree ‘Throuba Naxos’ was studied to characterize the potential origin or routes of olive tree domestication. Several candidate genes known to be associated with key agronomic traits, including olive oil quality and fruit yield, were uncovered by a selective sweep scan to be under selection pressure on all olive tree chromosomes. To further exploit the genomic and phenotypic resources obtained from the current work, genome-wide association analyses were performed for 23 morphological and two agronomic traits. Significant associations were detected for eight traits that provide valuable candidates for fruit tree breeding and for deeper understanding of olive tree biology. info:eu-repo/semantics/publishedVersion

Natural products are not only positioned in the heart of traditional medicine but also in modern medicine as many current drugs are coming from natural sources. Apart from the field of medicine and therapeutics, natural products are broadly used in other industrial fields such as nutrition, skincare products and nanotechnology.The aim of this study was to assess the effects of sweet cherry (Prunus avium L.) fruit extract from the Greek native cultivar 'Vasiliadi', on the human 2D and 3D in vitro models in order to investigate its potential impact on skin. We focused on 2D culture of primary normal human epidermal keratinocytes (NHEK) that were treated with sweet cherry fruit extract. In the first place, we targeted fruit extract potential cytotoxicity by determining ATP intracellular levels. Furthermore, we assessed its potential skin irritability by using 3D skin model. To better understand the bioactivity of sweet cherry fruit. extract, we used qPCR to study the expression of various genes that are implicated in the skin functions. Our experiments showed that sweet cherry fruit extract is non-toxic in 2D keratinocytes culture as well as non-irritant in 3D skin model. Our results revealed that the extract mediated important pathways for the optimum epidermis function such as cell proliferation, immune and inflammatory response.The sweet cherry fruit extracts possesses significant activity in epidermis function without any potential of cytotoxicity or skin irritability, which makes it a rather promising active agent for skincare.

Genome-wide transcriptome analysis provides systems-level insights into plant biology. Due to the limited depth of quantitative proteomics our understanding of gene-protein-complex stoichiometry is largely unknown in plants. Recently, the complexity of the proteome and its cell-/tissue-specific distribution have boosted the research community to the integration of transcriptomics and proteomics landscapes in a proteogenomic approach. Herein, we generated a quantitative proteome and transcriptome abundance atlas of 15 major sweet cherry (Prunus avium L., cv 'Tragana Edessis') tissues represented by 29 247 genes and 7584 proteins. Additionally, 199 984 alternative splicing events, particularly exon skipping and alternative 3' splicing, were identified in 23 383 transcribed regions of the analyzed tissues. Common signatures as well as differences between mRNA and protein quantities, including genes encoding transcription factors and allergens, within and across the different tissues are reported. Using our integrated dataset, we identified key putative regulators of fruit development, notably genes involved in the biosynthesis of anthocyanins and flavonoids. We also provide proteogenomic-based evidence for the involvement of ethylene signaling and pectin degradation in cherry fruit ripening. Moreover, clusters of genes and proteins with similar and different expression and suppression trends across diverse tissues and developmental stages revealed a relatively low RNA abundance-to-protein correlation. The present proteogenomic analysis allows us to identify 17 novel sweet cherry proteins without prior protein-level annotation evidenced in the currently available databases. To facilitate use by the community, we also developed the Sweet Cherry Atlas Database (https://grcherrydb.com/) for viewing and data mining these resources. This work provides new insights into the proteogenomics workflow in plants and a rich knowledge resource for future investigation of gene and protein functions in Prunus species.

There is a persistent interest in innovative and multifunctional ingredients in biology research. With regards to this, natural sources have an important role due to their multiple benefits. Thus, this study aims to present the pleiotropic activity of Prunus avium L. extract on human primary fibroblasts for proving its efficacy in dermis-related processes. We focused on the safety and efficacy assessments based on cytotoxicity and gene expression analysis under oxidative stress. Specifically, Prunus avium L. extract was proved non-cytotoxic in human fibroblasts. The gene expression analysis unveiled that this extract has in vitro protective properties on human dermal fibroblasts under oxidative stress related to antioxidant activity, anti-inflammatory response, cell proliferation and cell- aging. Our study demonstrated for the very first time that the Prunus avium L. extract is a multifunctional ingredient as it mediates several human dermis-related in vitro processes highlighting its potential to be used as an active ingredient in skin care products.

Sweet cherry germplasm contains a high variety of phenotypes which are associated with fruit size and shape as well as sugar content, etc. High phenotypic variation can be a result of genetic or epigenetic diversity that may interact through time. Recent studies have provided evidence that besides allelic variation, epiallelic variation can establish new heritable phenotypes. Herein we conducted a genetic and an epigenetic study (using amplified fragment length polymorphism (AFLP) and methylation-sensitive amplified polymorphism (MSAP) markers, respectively), accompanied by phenotypic traits correlation analysis in sweet cherry gene pools. The mean genetic diversity was greater than the epigenetic diversity (hgen = 0.193, hepi = 0.185), while no significant relationship was found between genetic and epigenetic distance according to a Mantel test. Furthermore, according to correlation analyses our results provided evidence that epigenetic diversity in predefined populations of sweet cherry had a stronger impact on phenotypic traits than their rich genetic diversity.

[EN] Background Summer squash (Cucurbita pepo: Cucurbitaceae) are a popular horticultural crop for which there is insufficient genomic and transcriptomic information. Gene expression atlases are crucial for the identification of genes expressed in different tissues at various plant developmental stages. Here, we present the first comprehensive gene expression atlas for a summer squash cultivar, including transcripts obtained from seeds, shoots, leaf stem, young and developed leaves, male and female flowers, fruits of seven developmental stages, as well as primary and lateral roots. Results In total, 27,868 genes and 2352 novel transcripts were annotated from these 16 tissues, with over 18,000 genes common to all tissue groups. Of these, 3812 were identified as housekeeping genes, half of which assigned to known gene ontologies. Flowers, seeds, and young fruits had the largest number of specific genes, whilst intermediate-age fruits the fewest. There also were genes that were differentially expressed in the various tissues, the male flower being the tissue with the most differentially expressed genes in pair-wise comparisons with the remaining tissues, and the leaf stem the least. The largest expression change during fruit development was early on, from female flower to fruit two days after pollination. A weighted correlation network analysis performed on the global gene expression dataset assigned 25,413 genes to 24 coexpression groups, and some of these groups exhibited strong tissue specificity. Conclusions These findings enrich our understanding about the transcriptomic events associated with summer squash development and ripening. This comprehensive gene expression atlas is expected not only to provide a global view of gene expression patterns in all major tissues in C. pepo but to also serve as a valuable resource for functional genomics and gene discovery in Cucurbitaceae., BP was funded by Ministerio de Ciencia, Innovacion y Universidades, cofunded with FEDER funds (Project No. AGL2017-85563-C2-1-R) and by PROMETEO project 2017/078 (to promote excellence groups) by the Conselleria d'Educacio, Investigacio, Cultura vertical bar Esports (Generalitat Valenciana).

Sweet cherries, Prunus avium L. (Rosaceae), are gaining importance due to their perenniallity and nutritional attributes beneficial for human health. Interestingly, sweet cherry cultivars exhibit a wide range of phenotypic diversity in important agronomic traits, such as flowering time and defense reactions against pathogens. In this study, whole-genome resequencing (WGRS) was employed to characterize genetic variation, population structure and allelic variants in a panel of 20 sweet cherry and one wild cherry genotypes, embodying the majority of cultivated Greek germplasm and a representative of a local wild cherry elite phenotype. The 21 genotypes were sequenced in an average depth of coverage of 33.91×. and effective mapping depth, to the genomic reference sequence of ‘Satonishiki’ cultivar, between 22.21× to 36.62×. Discriminant analysis of principal components (DAPC) with SNPs revealed two clusters of genotypes. There was a rapid linkage disequilibrium decay, as the majority of SNP pairs with r2 in near complete disequilibrium (>0.8) were found at physical distances less than 10 kb. Functional analysis of the variants showed that the genomic ratio of non-synonymous/synonymous (dN/dS) changes was 1.78. The higher dN frequency in the Greek cohort of sweet cherry could be the result of artificial selection pressure imposed by breeding, in combination with the vegetative propagation of domesticated cultivars through grafting. The majority of SNPs with high impact (e.g., stop codon gaining, frameshift), were identified in genes involved in flowering time, dormancy and defense reactions against pathogens, providing promising resources for future breeding programs. Our study has established the foundation for further large scale characterization of sweet cherry germplasm, enabling breeders to incorporate diverse germplasm and allelic variants to fine tune flowering and maturity time and disease resistance in sweet cherry cultivars.

Nutrients in the form of fertilizers and/or other additives such as amino acids, dramatically influence plant development and growth, plant nutrient composition and the level of soil pollution. Moreover, the treatment of soil microbiota is emerging as a new strategy in plant breeding to achieve desirable traits. Thus, integrated study of fertilizer application and soil microbiota might lead to a better understanding of soil-plant interactions and inform the design of novel ways to fertilize plants. Herein we report metagenomics data for soil microbiota in lettuce ( Lactuca sativa ) treated with fertilizer, amino acids or their combinations as follows: N-fertilizer + Amino16®, Amino16®, N-fertilizer and no treatment control. Data have been deposited in the NCBI Sequence Read Archive (SRA) (accession number: PRJNA388765).

Molecular taxonomic studies have been performed in the past in order to identify different wheat species and construct a molecular phylogeny. These were based on universal but sufficiently divergent sequences from both the nuclear and chloroplastic genomes of wheat. They included two short plastid sequences from the plastid genes rbcL and matK which have been proposed as the core “barcode” sequences by the “barcoding” guidelines for general plant identification. Historically, in molecular plant taxonomy, plastidic sequences had been favored over nuclear sequences, due to their uniparental inheritance and consequently lower intra-molecular recombination. However recently, the short nuclear sequence from the internal transcribed spacer 2 (ITS2) has been used successfully for the accurate identification of many medicinal and other plant species. Herein, we have used the plastidic matK, rbcL trnL, and the nuclear ITS2 region for the identification of different wheat species of Triticum L. and goatgrass species of Aegilops L. We have successfully discriminated all species that were examined from both genera, thus, validating the ITS2 region as a ‘barcode tool’ for accurate distinction of plants in the genus Triticum L. and Aegilops L. The success rate of PCR amplification and sequencing of the ITS2 region was 100%. We report also that matK, rbcL and trnL regions could not discriminate all species in contrast to the ITS2 region which demonstrated full discriminatory capacity.

The genetic basis of fruit size and shape was investigated for the first time in Cucurbita species and genetic loci associated with fruit morphology have been identified. Although extensive genomic resources are available at present for tomato (Solanum lycopersicum), cucumber (Cucumis sativus), melon (Cucumis melo) and watermelon (Citrullus lanatus), genomic databases for Cucurbita species are limited. Recently, our group reported the generation of pumpkin (Cucurbita pepo) transcriptome databases from two contrasting cultivars with extreme fruit sizes. In the current study we used these databases to perform comparative transcriptome analysis in order to identify genes with potential roles in fruit morphology and fruit size. Differential Gene Expression (DGE) analysis between cv. 'Munchkin' (small-fruit) and cv. 'Big Moose' (large-fruit) revealed a variety of candidate genes associated with fruit morphology with significant differences in gene expression between the two cultivars. In addition, we have set the framework for generating EST-SSR markers, which discriminate different C. pepo cultivars and show transferability to related Cucurbitaceae species. The results of the present study will contribute to both further understanding the molecular mechanisms regulating fruit morphology and furthermore identifying the factors that determine fruit size. Moreover, they may lead to the development of molecular marker tools for selecting genotypes with desired morphological traits.

[EN] Cucurbita pepo contains two cultivated subspecies, each of which encompasses four fruit-shape morphotypes (cultivar groups). The Pumpkin, Vegetable Marrow, Cocozelle, and Zucchini Groups are of subsp. pepo and the Acorn, Crookneck, Scallop, and Straightneck Groups are of subsp. ovifera. Recently, a de novo assembly of the C. pepo subsp. pepo Zucchini genome was published, providing insights into its evolution. To expand our knowledge of evolutionary processes within C. pepo and to identify variants associated with particular morphotypes, we performed whole-genome resequencing of seven of these eight C. pepo morphotypes. We report for the first time whole-genome resequencing of the four subsp. pepo (Pumpkin, Vegetable Marrow, Cocozelle, green Zucchini, and yellow Zucchini) morphotypes and three of the subsp. ovifera (Acorn, Crookneck, and Scallop) morphotypes. A high-depth resequencing approach was followed, using the BGISEQ-500 platform that enables the identification of rare variants, with an average of 33.5X. Approximately 94.5% of the clean reads were mapped against the reference Zucchini genome. In total, 3,823,977 high confidence single-nucleotide polymorphisms (SNPs) were identified. Within each accession, SNPs varied from 636,918 in green Zucchini to 2,656,513 in Crookneck, and were distributed homogeneously along the chromosomes. Clear differences between subspecies pepo and ovifera in genetic variation and linkage disequilibrium are highlighted. In fact, comparison between subspecies pepo and ovifera indicated 5710 genes (22.5%) with Fst > 0.80 and 1059 genes (4.1%) with Fst = 1.00 as potential candidate genes that were fixed during the independent evolution and domestication of the two subspecies. Linkage disequilibrium was greater in subsp. ovifera than in subsp. pepo, perhaps reflective of the earlier differentiation of morphotypes within subsp. ovifera. Some morphotype-specific genes have been localized. Our results offer new clues that may provide an improved understanding of the underlying genomic regions involved in the independent evolution and domestication of the two subspecies. Comparisons among SNPs unique to particular subspecies or morphotypes may provide candidate genes responsible for traits of high economic importance., This work has been supported by Hellenic Agricultural Organization (ELGO) Demeter. Furthermore, we thank the Conselleria de Educacio, Investigacio, Cultura i Esport (Generalitat Valenciana) for funding Project Prometeo 2017/078 "Seleccion de Variedades Tradicionales y Desarrollo de Nuevas Variedades de Cucurbitaceas Adaptadas a la Produccion Ecologica". Also, this work was supported by Chiang Mai University.

The phenotypic and genetic analysis of apple (Malus × domestica Borkh) genotypes is essential for breeding species. Information on the morphology and genetic structure of apple offers significant help for germplasm maintenance and selection of suitable material to breed superior cultivars. This study shows the results of an investigation on the morphology and the genetic diversity for 19 apple cultivars, which are preserved in an ex situ collection in Naoussa, Central Macedonia, Greece. Information was recorded over a 5-year period for 47 traits describing plant morphology and phenotype, as well as leaf and fruit quality. Data were analyzed using the principal component analysis (PCA) method. The Euclidean distance metric and the Ward’s agglomeration method were used in an unsupervised hierarchical cluster analysis of all cultivars. The cultivars were grouped into four main clusters, suggesting that the characterized apple collection has a high potential for specific breeding goals. Furthermore, the cultivars were genotyped using seven microsatellite primers. Moderate levels of polymorphism were detected, and 38 distinctive alleles (5.4 alleles per primer pair) were identified. Both multivariate clustering approach (phenotypic data) and the genetic distance clustering approach (genetic data) grouped the apple cultivars according to their type. Hence, these data could be used for protection or patenting processes of existing or new apple cultivars carried out by the EU-Community Plant Variety Office.

Beans are one of the most important staple crops in the world. Runner bean (Phaseolus coccineus L.) is a small-scale agriculture crop compared to common bean (Phaseolusvulgaris). Beans have been introduced to Europe from the Central America to Europe and since then they have been scattered to different geographical regions. This has resulted in the generation of numerous local cultivars and landraces with distinguished characters and adaptive potential. To identify and characterize the underlying genomic variation of two very closely related runner bean cultivars, we performed RNA-Seq with de novo transcriptome assembly in two landraces of P. coccineus, ‘Gigantes’ and ‘Elephantes’ phenotypically distinct, differing in seed size and shape. The cleaned reads generated 37,379 and 37,774 transcripts for ‘Gigantes’ and ‘Elephantes,’ respectively. A total of 1896 DEGs were identified between the two cultivars, 1248 upregulated in ‘Elephantes’ and 648 upregulated in ‘Gigantes.’ A significant upregulation of defense-related genes was observed in ‘Elephantes,’ among those, numerous members of the AP2-EREBP, WRKY, NAC, and bHLH transcription factor families. In total, 3956 and 4322 SSRs were identified in ‘Gigantes’ and ‘Elephantes,’ respectively. Trinucleotide repeats were the most dominant repeat motif, accounting for 41.9% in ‘Gigantes’ and 40.1% in ‘Elephantes’ of the SSRs identified, followed by dinucleotide repeats (29.1% in both cultivars). Additionally, 19,281 putative SNPs were identified, among those 3161 were non-synonymous, thus having potential functional implications. High-confidence non-synonymous SNPs were successfully validated with an HRM assay, which can be directly adopted for P. coccineus molecular breeding. These results significantly expand the number of polymorphic markers within P. coccineus genus, enabling the robust identification of runner bean cultivars, the construction of high-resolution genetic maps, potentiating genome-wide association studies. They finally contribute to the genetic reservoir for the improvement of the closely related and intercrossable Phaseolus vulgaris.

An increasing demand for cherry production (Prunus avium L.) in Greece led to the development of new high quality sweet cherry cultivars. Self-incompatibility in cherry is one of the most challenging issues for the species’ cultivation and top breeding priority. Τhe present study focuses on the development of new hybrids with improved traits such as productivity, fruit size, organoleptic characteristics and self-compatibility. For this purpose, thirty different cultivars were crossed and produced hybrids that were evaluated according to 34 morpho-physiological characteristics. The results were analyzed using the XLSTAT (version 2014.1) software and a dendrogram was constructed using the agglomerative hierarchical clustering method. Optimal hybrid clustering was achieved when characteristics of great economic importance such as fruit shape and size, growth habit and days to blooming were included in the analysis. Based on the results, new sweet cherry hybrids with the special character of self-compatibility were developed. Our findings provide crucial new information for sweet cherry future breeding programs and cultivation.

Barbary fig ( Opuntia ficus indica ) could be an economically important species as it could be an alternative crop extremely tolerant to dry condition and water deficiency. Moreover, it could be used in alternative sustainable cultivation systems and landscape conservation. In this work, we report the analysis of the genetic diversity of Greek Barbary fig genotypes using ISSR molecular markers. Six primers were screened to assess their ability to detect polymorphisms within twenty-two Barbary fig accessions and generated 57 markers (bands), with an average of 9.5 markers per primer. The percentage of polymorphic bands (50.21%) and the resolving power (R P ) (28.85) showed the efficiency of the used primers. Mean values for GD (gene diversity) and I (Shannon index) were found as 0.215 and 0.355, respectively. The revealed ISSR markers allow distinguishing all accessions analyzed except for one case. UPGMA dendrogram and PCoA (Principal Coordinate Analysis) were performed to access patterns of diversity among genotypes. The high genetic diversity existing in the Greek germplasm suggests that it would be beneficial to utilize this pool in Barbary fig breeding programs and germplasm management activities.

Olea europaea L. has been cultivated in the Mediterranean region for thousands of years and is of major economic importance. The origin of olive cultivars remains as complex to trace as their identification. Thus, their molecular characterization and discrimination will enable olive germplasm management. In addition, it would be a useful tool for authentication of olive products. High-resolution melting (HRM) analysis, coupled with five microsatellite markers, was integrated to facilitate molecular identification and characterization of main O. europaea cultivars collected from the National Olive Tree Germplasm Collection established in Chania, Greece. The five microsatellite loci used were highly informative and generated a unique melting curve profile for each of the 47 cultivars and for each microsatellite tested. In particular, three microsatellite markers (DCA03, DCA09 and DCA17), which generated 29 HRM profiles, were sufficient to genotype all the olive cultivars studied, highlighting their potential use for cultivar identification. Furthermore, this assay provided a flexible, cost-effective and closed-tube microsatellite genotyping method well suited for molecular characterization of olive cultivars.

As a result of this work, we were able to characterize seven indigenous to Greece Salvia officinalis populations using genetic and metabolomic tools. These tools can be used to select the most promising genotypes, capable to design future breeding programs for high valuable varieties. An initial investigation was carried out to compare the genetic and metabolic diversity in S. officinalis grown in Greece and to discern the relationship between the two sets of data. Analysis of inter-simple sequence repeats (ISSR) revealed significant genetic differences among seven sage populations, which were grouped into three main clusters according to an UPGMA ISSR data-based dendrogram and Principle Coordinate Analysis. 80 loci were scored of which up to 90% were polymorphic at species level. According to the composition of their essential oil, the populations were classified into two chemotypes: 1.8 cineole/α-thujone and α-thujone/1.8 cineole. Additionally, a targeted ultra performance liquid chromatography (UPLC–MS/MS) method was used to qualify and quantify phenolic compounds in methanolic extracts of the seven sage genotypes according to which they were districted in six clusters among the sage populations. The main compounds characterizing the seven genotypes were rosmarinic acid and carnosol, followed by apigenin-7-O-glucoside (Ap7glc), and luteolin-7-O-glucoside (Lu7glc). The correlation between matrices obtained from ISSR data and metabolic profiles was non-significant. However, based on the differences in metabolic fingerprint, we aimed to define populations using as main selection criteria the high polyphenol content and desired essential oil composition, using state to the art analytical tools for the identification of parent lines for breeding programs.

Single nucleotide polymorphisms (SNPs) provide an important tool for cultivar identification in studies of genetic diversity, but until now, the time-consuming and costly nature of DNA sequencing has limited the identification of new markers. Herein, we describe the application of high-resolution melting (HRM), a recent enhancement to traditional DNA melting analysis, for the characterization of polymerase chain reaction products and the identification of nine gene-based SNPs for distinguishing the main Greek sweet cherry cultivars. The expected heterozygosity value of nine SNPs averaged at 0.518. The combined power of discrimination for the SNP markers was 0.999969. The ability of HRM to accurately discern nucleotide changes in a DNA sequence makes it a cost- and time-effective alternative to traditional sequencing for the detection of gene-based SNPs.

Identification of genotypes in Sideritis is complicated owing to the morphological similarity and common occurrence of natural hybridisation within Sideritis species. Species- and genotype-specific DNA markers are very useful for plant identification, breeding and preservation programs. Herein, a real-time polymerase chain reaction (PCR) of ITS2 barcode region coupled with high resolution melting-curve (HRM) analysis was evaluated for an accurate, rapid and sensitive tool for species identification focusing on seven Sideritis species growing in Greece. The HRM assay developed in this study is a rapid and straightforward method for the identification and discrimination of the investigated Sideritis species. This assay is simple compared to other genotyping methods as it does not require DNA sequencing or post-PCR processing. Therefore, this method offers a new alternative for rapid detection of Sideritis species.

Cucurbita pepo (squash, pumpkin, gourd), a worldwide-cultivated vegetable of American origin, is extremely variable in fruit characteristics. However, the information associated with genes and genetic markers for pumpkin is very limited. In order to identify new genes and to develop genetic markers, we performed a transcriptome analysis (RNA-Seq) of two contrasting pumpkin cultivars. Leaves and female flowers of cultivars, ‘Big Moose’ with large round fruits and ‘Munchkin’ with small round fruits, were harvested for total RNA extraction. We obtained a total of 6 GB (Big Moose; http://www.ncbi.nlm.nih.gov/Traces/sra/?run=SRR3056882) and 5 GB (Munchkin; http://www.ncbi.nlm.nih.gov/Traces/sra/?run=SRR3056883) sequence data (NCBI SRA database SRX1502732 and SRX1502735, respectively), which correspond to 18,055,786 and 14,824,292 150-base reads. After quality assessment, the clean sequences where 17,995,932 and 14,774,486 respectively. The numbers of total transcripts for ‘Big Moose’ and ‘Munchkin’ were 84,727 and 68,051, respectively. TransDecoder identified possible coding regions in assembled transcripts. This study provides transcriptome data for two contrasting pumpkin cultivars, which might be useful for genetic marker development and comparative transcriptome analyses. Keywords: RNA-Seq, Pumpkin, Contrasting cultivars, Cucurbita pepo