Whole-genome re-sequencing of two Italian tomato landraces reveals sequence variations in genes associated with stress tolerance, fruit quality and long shelf-life traits

Tomato is a high value crop and the primary model for fleshy fruit development and ripening. Breeding priorities include increased fruit quality, shelf life and tolerance to stresses. To contribute towards this goal, we re-sequenced the genomes of Corbarino (COR) and Lucariello (LUC) landraces, which both possess the traits of plant adaptation to water deficit, prolonged fruit shelf-life and good fruit quality. Through the newly developed pipeline Reconstructor, we generated the genome sequences of COR and LUC using datasets of 65.8M and 56.4M of 30–150bp paired-end reads, respectively. New contigs including reads that could not be mapped to the tomato reference genome were assembled, and a total of 43, 054 and 44, 579 gene loci were annotated in COR and LUC. Both genomes showed novel regions with similarity to Solanum pimpinellifolium and Solanum pennellii. In addition to small deletions and insertions, 2, 000 and 1, 700 single nucleotide polymorphisms (SNPs) could exert potentially disruptive effects on 1, 371 and 1, 201 genes in COR and LUC, respectively. A detailed survey of the SNPs occurring in fruit quality, shelf life and stress tolerance related-genes identified several candidates of potential relevance. Variations in ethylene response components may concur in determining peculiar phenotypes of COR and LUC

Sequencing and Analysis of Plastid Genome in Mycoheterotrophic Orchid Neottia nidus-avis

Plastids are the semiautonomous organelles that possess their own genome inherited from the cyanobacterial ancestor. The primary function of plastids is photosynthesis so the structure and evolution of plastid genomes are extensively studied in photosynthetic plants. In contrast, little is known about the plastomes of nonphotosynthetic species. In higher plants, plastid genome sequences are available for only three strictly nonphotosynthetic species, the liverwort Aneura mirabilis and two flowering plants, Epifagus virginiana and Rhizanthella gardneri. We report here the complete sequence of a plastid genome of nonphotosynthetic mycoheterotrophic orchid Neottia nidus-avis, determined using 454 pyrosequencing technology. It was found to be reduced in both genome size and gene content; this reduction is however not as drastic as in the other nonphotosynthetic orchid, R. gardneri. Neottia plastome lacks all genes encoding photosynthetic proteins, RNA polymerase subunits but retains most genes of translational apparatus. Those genes that are retained have an increased rate of both synonymous and nonsynonymous substitutions but do not exhibit relaxation of purifying selection either in Neottia or in Rhizanthella

Resequencing

[ES] La revolución que supone la secuenciación de próxima generación está permitiendo la resecuenciación del genoma completo (WGRS) de cientos o incluso miles de ejemplares de cultivos básicos y especies modelo. Con el lanzamiento de su genoma de referencia, progresivamente se están emprendiendo proyectos WGRS también para otras especies de plantas en una amplia variedad de estudios. En berenjena común (Solanum melongena L.), aunque se ha publicado un primer borrador de la secuencia del genoma de referencia, hasta el momento no se han realizado estudios de resecuenciación. En este capítulo presentamos los primeros resultados de la resecuenciación de ocho accesiones, siete de berenjena común y una del pariente silvestre S. incanum L., que corresponden a los progenitores de un cruce multiparental de generación avanzada (MAGIC) población que se encuentra actualmente en desarrollo utilizando la secuencia del genoma de la berenjena recién desarrollada que se presenta en el Cap. 7 de este libro. Se identificaron más de diez millones de polimorfismos entre las accesiones, el 90% de ellos en el S. incanum silvestre relacionado, lo que confirma la erosión genética de la berenjena común cultivada. Entre los progenitores de la población MAGIC, el patrón de distribución de polimorfismos comunes a lo largo de los cromosomas ha revelado posibles huellas de introgresión ancestral de cruces interespecíficos. El conjunto de polimorfismos se ha anotado extensamente y actualmente se está utilizando para análisis adicionales con el fin de genotipar eficientemente la población MAGIC en curso y diseccionar rasgos agronómicos y morfológicos importantes. La información proporcionada en este primer estudio de resecuenciación en berenjena será extremadamente útil para ayudar al fitomejoramiento a desarrollar nuevas variedades mejoradas y resistentes para enfrentar futuras amenazas y desafíos.[EN] The next-generation sequencing revolution is allowing the whole-genome resequencing (WGRS) of hundreds or even thousands of accessions for staple crops and model species. With the release of their reference genome, progressively also other plants, species are undertaking WGRS projects for a broad variety of studies. In common eggplant (Solanum melongena L.), although a first draft of the reference genome sequence has been published, no resequencing studies have been performed so far. In this chapter, we present the first results of the resequencing of eight accessions, seven of common eggplant and one of the wild relative S. incanum L., that correspond to the parents of a multi-parent advanced generation inter-cross (MAGIC) population that is currently under develop- ment using the newly developed eggplant genome sequence presented in Chap. 7 of this book. Over ten million polymorphisms were identified among the accessions, 90% of them in the wild related S. incanum, confirming the genetic erosion of the cultivated common eggplant. Among the MAGIC population parents, the common polymorphism distribu- tion pattern along the chromosomes has revealed possible footprints of ancestral intro- gression from interspecific crosses. The set of polymorphisms has been extensively anno- tated and currently is being used for further analyses in order to efficiently genotype the ongoing MAGIC population and to dissect important agronomic and morphological traits. The information provided in this first resequencing study in eggplant will be extremely helpful to assist plant breeding to develop new improved and resilient varieties to face future threats and challenges.This work has received funding from the European Union’s Horizon 2020 Research and Innovation Programme under grant agreement No 677379 (G2P-SOL project: Linking genetic resources, genomes and phenotypes of Solanaceous crops) and from Spanish Ministerio de Economía, Industria y Competitividad and Fondo Europeo de Desarrollo Regional (grant AGL2015-64755-R from MINECO/FEDER).Prohens Tomás, J.; Vilanova Navarro, S.; Gramazio, P. (2019). Resequencing. En The Eggplant Genome. Springer. 81-89. http://hdl.handle.net/10251/181875S818

Insights Into the Adaptation to Greenhouse Cultivation of the Traditional Mediterranean Long Shelf-Life Tomato Carrying the alc Mutation: A Multi-Trait Comparison of Landraces, Selections, and Hybrids in Open Field and Greenhouse

[EN] Long shelf-life tomato (Solanum lycopersicum) landraces, characterized by carrying the alc allele in the NOR. NAC locus, have been traditionally cultivated in the Mediterranean region. These materials are adapted to open field conditions under low input conditions. However, cultivation under greenhouse is expanding fueled by increasing demand of these traditional tomatoes. We hypothesize that the large diversity in the long shelf-life landraces and derived materials can be exploited for adaptation to these new cultivation conditions. We have evaluated 12 varieties (seven landraces, three selections and two hybrids) carrying the alc mutation under open field (OF) and greenhouse (GH) cultivation, and evaluated them for 52 morphological, agronomic, chemical properties, and chemical composition descriptors. All descriptors, except six morphological ones, were variable. The variety effect was the greatest contributor to variation for most morphological traits, as well as for fruit weight, fruit shape, dry matter, and soluble solids content. However, significant environmental and genotype x environment interaction were found for 36 and 42 descriptors, respectively. Fruits from GH plants had lower weight and firmness and were less red than those from OF. On average, in GH yield was 35% lower and daily fruit weight loss in post-harvest 41% higher than in OF. However, fruits from GH had on average higher dry matter and soluble solids contents, antioxidant activity, glucose, fructose, and ascorbic acid concentrations, but lower contents in lycopene and beta-carotene than those from OF. A principal components analysis clearly separated varieties according to the cultivation environment. However, the distribution pattern of varieties within each of the two clusters (GH and OF) was similar, despite the strong G x E interaction for many descriptors. Landraces from the same origin plotted in the same area of each cluster, and selections and hybrids plotted together with the landraces. The results reveal a high impact of the cultivation environment on morphological, agronomic, chemical properties, and chemical composition of Mediterranean long shelf-life traditional tomato varieties. This suggests that breeding programs specifically focused to adaptation to greenhouse conditions should be developed.This work was supported by Associacio de Productors i Comercialitzadors de la Tomata de Penjar d'Alcala de Xivert. Funding was also received from the TRADITOM (Traditional tomato varieties and cultural practices: a case for agricultural diversification with impact on food security and health of European population), G2P-SOL (Linking genetic resources, genomes, and phenotypes of Solanaceous crops) and BRESOV (Breeding for resilient, efficient, and sustainable organic vegetable production) projects. TRADITOM, G2P-SOL, and BRESOV projects have received funding from the European Union's Horizon 2020 research and innovation programme under grant agreements 634561 (TRADITOM), 677379 (G2PSOL), and 774244 (BRESOV). ER is grateful to the Spanish Ministerio de Economia, Industria y Competitividad for a pre-doctoral grant (BES-2016-077482). MP is grateful to Spanish Ministerio de Economia, Industria y Competitividad for a post-doctoral grant within the Juan de la Cierva programme (FCJI-2015-24835) and to Generalitat Valenciana and Fondo Social Europeo for a post-doctoral contract (APOSTD/2018/014).Figás-Moreno, MDR.; Prohens Tomás, J.; Raigón Jiménez, MD.; Pereira-Días, L.; Casanova-Calancha, C.; García-Martínez, MD.; Rosa-Martínez, E.... (2018). Insights Into the Adaptation to Greenhouse Cultivation of the Traditional Mediterranean Long Shelf-Life Tomato Carrying the alc Mutation: A Multi-Trait Comparison of Landraces, Selections, and Hybrids in Open Field and Greenhouse. 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WHOLE-GENOME RE-SEQUENCING OF TWO TOMATO LANDRACES REVEALS SEQUENCE VARIATIONS UNDERPINNING KEY ECONOMICALLY IMPORTANT TRAITS

In the post-genomic era, one of the major challenges is the identification of alleles directly responsible for phenotype variation among different genotypes within the same species. Tomato is a model crop for understanding the development and ripening of climacteric fleshy fruits, and it is also known to be an important source of health-promoting compounds. In addition, cultivated tomato germplasm shows a high phenotypic variation despite its very low genetic diversity. Toward the identification of sequence variations responsible for stress tolerance, high fruit quality and long shelf life, we re-sequenced the genomes of two traditional landraces grown in the Campania region (Southern Italy). Crovarese, belonging to the Corbarino type (COR), and Lucariello (LUC) are typically grown under low water regimes and produce highly appreciated fruits, which can be stored up to 4-8 months. We generated 65.8M and 56.4M of paired-end 30-150 bp reads with an average insert size of 380 bp (± 52bp) and 364 bp (± 49bp) for COR and LUC, respectively. A referenceguided assembly was performed using 'Heinz 1706' as a reference genome. We estimated a mean coverage depth of ~15X for COR and 13X for LUC. Comparing the genomes of COR and LUC with that of 'Heinz 1706' we found a similar distribution of SNPs (68.8% vs. 69.9%, respectively), small deletions (8.9% vs. 8.6%) and small insertions (22.1% vs. 21.3%). Through a de novo assembly of the unmapped reads we identified 29 and 36 new contigs in COR and LUC, respectively. The new contigs could be assigned to the chromosomes thanks to the use of a splitread approach. On average, the contigs inserted in COR were 654bp, whereas those inserted in LUC were 616bp. Using custom RNA-seq data, a total of 43054 and 44576 gene loci were annotated in COR and LUC, corresponding to 62369 and 65094 transcripts, respectively. Among the genes showing a similar structure in COR and LUC compared to 'Heinz 1706', we identified ~2000 and 1700 SNPs causing potentially disruptive effects on the function of 1371 and 1201 genes in COR and LUC, respectively. Interesting GO categories highly represented in genes affected by sequence changes were identified. Major variations were present in stress-responsive genes as well as in fruit quality and development-related genes. From a practical perspective, the identified SNPs and InDels are candidate polymorphisms to track DNA variations associated to key traits of economic interest

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

[EN] 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 19.8x 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.This work has been funded by the European Union's Horizon 2020 Research and Innovation Programme under grant agreement no. 677379 (G2P-SOL project: Linking genetic resources, genomes, and phenotypes of Solanaceous crops), by the Spanish Ministerio de Economia, Industria y Competitividad and Fondo Europeo de Desarrollo Regional/European Regional Development Fund (grant AGL2015-64755-R), and by the Spanish Ministerio de Ciencia, Innovacion y Universidades (MCIU), Agencia Estatal de Investigacion (AEI), and Fondo Europeo de Desarrollo Regional/European Regional Development Fund (grant RTI2018-09592-B-100). PG is grateful to Universitat Politecnica de Valencia and to Japan Society for the Promotion of Science for their respective postdoctoral grants [PAID-10-18 and FY2019 JSPS Postdoctoral Fellowship for Research in Japan (Standard)].Gramazio, P.; Yan, H.; Hasing, T.; Vilanova Navarro, S.; Prohens Tomás, J.; Bombarely, A. (2019). Whole-Genome Resequencing of Seven Eggplant (Solanum melongena) and One Wild Relative (S. incanum) Accessions Provides New Insights and Breeding Tools for Eggplant Enhancement. Frontiers in Plant Science. 10:1-17. https://doi.org/10.3389/fpls.2019.01220S11710Acquadro, A., Barchi, L., Gramazio, P., Portis, E., Vilanova, S., Comino, C., … Lanteri, S. (2017). Coding SNPs analysis highlights genetic relationships and evolution pattern in eggplant complexes. PLOS ONE, 12(7), e0180774. doi:10.1371/journal.pone.0180774Alonso-Blanco, C., Andrade, J., Becker, C., Bemm, F., Bergelson, J., Borgwardt, K. M., … Ding, W. 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Biosynthesis of Salvia specialized metabolites and biotechnological approaches to increase their production

Aromatic Salvia species are particularly valuable for providing several bioactive compounds used as food additives, pigments, cosmetics, perfumes and fine chemicals. Within the Lamiaceae family, the Salvia genus, with more than 900 species, biosynthesizes a plethora of beneficial metabolites including terpenes, steroids and polyphenols. The whole plant can be considered a factory of bioactive compounds, but plant cell and tissue cultures are also an attractive sustainable alternative to cultivation. Salvia cell cultures can readily be initiated from different explants, including leaves, roots, stems, petioles, anthers and seedlings; however high metabolites accumulation in plant tissue and cell culture is a prerequisite for massive production of these bioactive compounds. In this chapter, the occurrence and tissue distribution of specialized metabolites in several Salvia species, especially flavonoids and diterpenoids, will be reviewed along with recent advances in the understanding of biosynthetic pathways as well as regulatory mechanisms leading to their biosynthesis. We will focus on the recent biotechnological approaches aimed at enhancing the final biomass and metabolite accumulation in Salvia cell and tissue cultures. Advances in metabolic engineering strategies will be also summarized, reporting relevant and successful results and potential pitfalls, in order to provide valuable perspectives for design and developing cell and tissue cultures as a reliable and standardized biomass platform for the extraction of Salvia bioactive metabolites