Herramientas preliminares para la secuenciación de genomas: Genotecas con vectores de clonación

En este artículo se describen los pasos a seguir haciendo hincapié en los detalles más relevantes y explicando las posibles decisiones que el alumno deberá considerar antes de empezar la realización de las genotecas.Vilanova Navarro, S.; Gadea Vacas, J. (2019). Herramientas preliminares para la secuenciación de genomas: Genotecas con vectores de clonación. http://hdl.handle.net/10251/122006DE

Evaluación de Diferentes Metodologías de Aprendizaje Activo desde el Punto de Vista del Estudiante en la Asignatura Genómica del Grado de Biotecnología

[EN] Biotechnology is a discipline where autonomous learning can be easily implemented. The speed at which strategies and technologies are developed makes mandatory a learning method more focused in “how to learn” than in “what to learn”. The objective of this study is to evaluate the student’s level of satisfaction regarding the learning strategies applied in the subject “Genomics” of the UPV Biotechnology Degree. Among these methodologies, we have used project-based learning, case studies and flip-teaching. Results seem to indicate a high percentaje of satisfaction when these methodologies are applied.[ES] La Biotecnología es una disciplina donde puede implementarse el aprendizaje autónomo. La velocidad a la que las tecnologías y estrategias están cambiando hace necesario un aprendizaje más enfocado a “cómo aprender”, más a “qué aprender”. El objetivo de esta comunicación es evaluar el grado de satisfacción de los estudiantes en referencia a los métodos docentes aplicados en la asignatura Genómica del Grado de Biotecnología de la UPV. Entre estas metodologías, hemos aplicado aprendizaje basado en proyectos, estudio de casos y clase inversa. Los resultados parecen indicar que un alto porcentaje de alumnos está satisfecho con un aprendizaje basado en estas metodologías.Vilanova Navarro, S.; Gadea Vacas, J. (2017). Evaluación de Diferentes Metodologías de Aprendizaje Activo desde el Punto de Vista del Estudiante en la Asignatura Genómica del Grado de Biotecnología. En In-Red 2017. III Congreso Nacional de innovación educativa y de docencia en red. Editorial Universitat Politècnica de València. 372-382. https://doi.org/10.4995/INRED2017.2017.6798OCS37238

Claves y evolución de las tecnologías de Secuenciación Masiva de Segunda Generación

Este artículo docente indica las claves del éxito de las tecnologías de secuenciación masiva de segunda generación y su evolución a lo largo de los últimos años, haciendo referencia a la química de los nucleótidos, la paralelización y las estrategias de secuenciaciónGadea Vacas, J.; Vilanova Navarro, S. (2020). Claves y evolución de las tecnologías de Secuenciación Masiva de Segunda Generación. http://hdl.handle.net/10251/144329DE

Secuenciación Masiva

Este artículo docente explica los pasos principales a realizar en un proceso de secuenciación masiva de segunda generaciónGadea Vacas, J.; Vilanova Navarro, S. (2020). Secuenciación Masiva. http://hdl.handle.net/10251/144323DE

Detection of honey adulteration by conventional and real-time PCR

[EN] This work applies both conventional and real-time PCR DNA amplification techniques for detecting and quantifying rice molasses in honey. Different levels of adulteration were simulated (1, 2, 5, 10, 20, 50%) using commercial rice molasses. Among the different specific genes of rice tested by PCR, the PLD1 primer was the most effective. This allowed the visualization in agarose gel of this type of adulterant up to 5-20%. Moreover, by means of real-time PCR it was possible to distinguish the different levels of rice DNA, and therefore the percentage of adulteration (1-50%). A standard curve built with the DNA serial dilutions of rice genomic DNA concentrations showed that the quantification level was between 2-5%. These results offer compelling evidence that DNA techniques could be useful not only for the detection of adulterations of honey with rice molasses but also for the quantification of levels lower than those of conventional techniques.This study is part of part of the projects funded by the "Agencia Estatal de Investigacion" (AGL2016-77702-R) and by the "Generalitat Valenciana" (AICO/2015/104) of Spain, for which the authors are grateful.Sobrino-Gregorio, L.; Vilanova Navarro, S.; Prohens Tomás, J.; Escriche Roberto, MI. (2019). Detection of honey adulteration by conventional and real-time PCR. Food Control. 95:57-62. https://doi.org/10.1016/j.foodcont.2018.07.037S57629

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)

[EN] 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.This work has been financed by INIA projects RTA2013-00022-C02, RTA2014-00041-C02-02, and RF2010-00025-00-00, FEDER funds. Authors are also grateful to the different Research Institutions, scientists, and breeders, and PDOs and GPIs Regulatory Boards, included on Supplementary Data: Table 1 for providing part of the materials studied here.Pereira-Días, L.; Vilanova Navarro, S.; Fita, A.; Prohens Tomás, J.; Rodríguez Burruezo, A. (2019). 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). Horticulture Research. 6:1-13. https://doi.org/10.1038/s41438-019-0132-8S1136FAO. FAOSTAT Statistics Database. http://www.fao.org/faostat/ (2018). Accessed 20 Aug 2018.Moscone, E. A. et al. 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De novo Transcriptome Assembly and Comprehensive Annotation of Two Tree Tomato Cultivars (Solanum betaceum Cav.) with Different Fruit Color

[EN] The tree tomato (Solanum betaceum Cav.) is an underutilized fruit crop native to the Andean region and phylogenetically related to the tomato and potato. Tree tomato fruits have a high amount of nutrients and bioactive compounds. However, so far there are no studies at the genome or transcriptome level for this species. We performed a de novo assembly and transcriptome annotation for purple-fruited (A21) and an orange-fruited (A23) accessions. A total of 174,252 (A21) and 194,417 (A23) transcripts were assembled with an average length of 851 and 849 bp. A total of 34,636 (A21) and 36,224 (A23) transcripts showed a significant similarity to known proteins. Among the annotated unigenes, 22,096 (A21) and 23,095 (A23) were assigned to the Gene Ontology (GO) term and 14,035 (A21) and 14,540 (A23) were found to have Clusters of Orthologous Group (COG) term classifications. Furthermore, 22,096 (A21) and 23,095 (A23) transcripts were assigned to 155 and 161 (A23) KEGG pathways. The carotenoid biosynthetic process GO terms were significantly enriched in the purple-fruited accession A21. Finally, 68,647 intraspecific single-nucleotide variations (SNVs) and almost 2 million interspecific SNVs were identified. The results of this study provide a wealth of genomic data for the genetic improvement of the tree tomato.Pietro Gramazio is grateful to the Spanish Ministerio de Ciencia e Innovacion for a Juan de laCierva-Formacion post-doctoral grant FJC2019-038921-I funded by MCIN/AEI/10.13039/501100011033)Pacheco, J.; Vilanova Navarro, S.; Grillo-Risco, R.; García-García, F.; Prohens Tomás, J.; Gramazio, P. (2021). De novo Transcriptome Assembly and Comprehensive Annotation of Two Tree Tomato Cultivars (Solanum betaceum Cav.) with Different Fruit Color. Horticulturae. 7(11):1-18. https://doi.org/10.3390/horticulturae7110431S11871

Performance of a Set of Eggplant (Solanum melongena) Lines With Introgressions From Its Wild Relative S. incanum Under Open Field and Screenhouse Conditions and Detection of QTLs

[EN] Introgression lines (ILs) of eggplant (Solanum melongena) represent a resource of high value for breeding and the genetic analysis of important traits. We have conducted a phenotypic evaluation in two environments (open field and screenhouse) of 16 ILs from the first set of eggplant ILs developed so far. Each of the ILs carries a single marker-defined chromosomal segment from the wild eggplant relative S. incanum (accession MM577) in the genetic background of S. melongena (accession AN-S-26). Seventeen agronomic traits were scored to test the performance of ILs compared to the recurrent parent and of identifying QTLs for the investigated traits. Significant morphological differences were found between parents, and the hybrid was heterotic for vigour-related traits. Despite the presence of large introgressed fragments from a wild exotic parent, individual ILs did not display differences with respect to the recipient parent for most traits, although significant genotype x environment interaction (G x E) was detected for most traits. Heritability values for the agronomic traits were generally low to moderate. A total of ten stable QTLs scattered across seven chromosomes was detected. For five QTLs, the S. incanum introgression was associated with higher mean values for plant- and flower-related traits, including vigour prickliness and stigma length. For one flower- and four fruit-related-trait QTLs, including flower peduncle and fruit pedicel lengths and fruit weight, the S. incanum introgression was associated with lower mean values for fruit-related traits. Evidence of synteny to other previously reported in eggplant populations was found for three of the fruit-related QTLs. The other seven stable QTLs are new, demonstrating that eggplant ILs are of great interest for eggplant breeding under different environments.This work was undertaken as part of the initiative "Adapting Agriculture to Climate Change: Collecting, Protecting, and Preparing Crop Wild Relatives", which is supported by the Government of Norway. The project is managed by the Global Crop Diversity Trust with the Millennium Seed Bank of the Royal Botanic Gardens, Kew and implemented in partnership with national and international gene banks and plant breeding institutes around the world. For further information, see the project website: http://www.cwrdiversity.org/.Funding was also received from Spanish Ministerio de Economia, Industria y Competitividad and Fondo Europeo de Desarrollo Regional (grant AGL2015-64755-R from MINECO/FEDER); from Ministerio de Ciencia, Innovacion y Universidades, Agencia Estatal de Investigacion and Fondo Europeo de Desarrollo Regional (grant RTI-2018-094592-B-100 from MCIU/AEI/FEDER, UE); from 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 Vicerrectorado de Investigacion, Innovacion y Transferencia de la Universitat Politecnica de Valencia (Ayuda a Primeros Proyectos de Investigacion; PAID-06-18). Giulio Mangino is grateful to Generalitat Valenciana for a predoctoral grant within the Santiago Grisolia programme (GRISOLIAP/2016/012). Pietro Gramazio is grateful to Japan Society for the Promotion of Science for a postdoctoral grant (P19105, FY2019 JSPS Postdoctoral Fellowship for Research in Japan (Standard)).Mangino, G.; Plazas Ávila, MDLO.; Vilanova Navarro, S.; Prohens Tomás, J.; Gramazio, P. (2020). Performance of a Set of Eggplant (Solanum melongena) Lines With Introgressions From Its Wild Relative S. incanum Under Open Field and Screenhouse Conditions and Detection of QTLs. Agronomy. 10(4):1-15. https://doi.org/10.3390/agronomy10040467S115104FAOSTAThttp://www.fao.org/faostat/Gebhardt, C. (2016). The historical role of species from the Solanaceae plant family in genetic research. Theoretical and Applied Genetics, 129(12), 2281-2294. doi:10.1007/s00122-016-2804-1Hirakawa, H., Shirasawa, K., Miyatake, K., Nunome, T., Negoro, S., Ohyama, A., … Fukuoka, H. (2014). Draft Genome Sequence of Eggplant (Solanum melongena L.): the Representative Solanum Species Indigenous to the Old World. 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Screening of Suitable Plant Regeneration Protocols for Several Capsicum spp. through Direct Organogenesis

[EN] Peppers (Capsicum spp.) are recalcitrant to in vitro culture regeneration, making the application of in vitro-based breeding strategies difficult. We evaluated the impact of different combinations of auxins, cytokinins and micronutrients on the induction of direct organogenesis in cotyledon and hypocotyl explants of C. annuum, C. baccatum and C. chinense. We found variation in the regeneration response among species and type of explant. In this way, the average numbers of shoots per cotyledon and hypocotyl explant were, respectively, 1.44 and 0.28 for C. annuum, 4.17 and 3.20 for C. baccatum and 0.08 and 0.00 for C. chinense. Out of the six media, the best overall results were obtained with the medium Pep1, which contained 5 mg/L BAP (6-benzylaminopurine), 0.5 mg/L IAA (indole-3-acetic acid) and 0.47 mg/L CuSO4, followed by a subculture in the same medium supplemented with 10 mg/L AgNO3 (medium Pep1.2). The best result for the Pep1 + Pep1.2 medium was obtained for C. baccatum using cotyledon explants, with 8.87 shoots per explant. The explants grown in medium Pep1 + Pep1.2 were the ones with greener tissue, while overall the hypocotyl explants were greener than the cotyledon explants. Our results indicate that there is wide variation among Capsicum species in terms of regeneration. Our results suggest that the synergistic effect of copper and silver resulted in a higher regeneration rate of Capsicum explants. Explants with shoots were transferred to different media for elongation, rooting and acclimatization. Although acclimatized plantlets were obtained for C. baccatum and C. chinense, an improvement in these latter stages would be desirable for a high throughput regeneration pipeline. This work contributes to the improvement of Capsicum regeneration protocols using specific combinations of medium, explant and genotype, reaching the levels of efficiency required for genetic transformation and of gene editing technologies for other crops.This research was funded by Universitat Politecnica de Valencia, research line 20200268.Martínez-López, M.; García-Pérez, A.; Gimeno -Páez, E.; Prohens Tomás, J.; Vilanova Navarro, S.; García-Fortea, E. (2021). Screening of Suitable Plant Regeneration Protocols for Several Capsicum spp. through Direct Organogenesis. Horticulturae. 7(9):1-17. https://doi.org/10.3390/horticulturae7090261S1177

Morphological and molecular diversity in a collection of the Andean tree tomato (Solanum betaceum Cav.)

This contribution has been partially funded by the Instituto Nacional de Investigaciones Agrarias y Agroalimentarias (RF2008-00008-00-00)Acosta-Quezada, P.; Martinez-Laborde, J.; Vilanova Navarro, S.; Prohens Tomás, J. (2011). Morphological and molecular diversity in a collection of the Andean tree tomato (Solanum betaceum Cav.). Bulletin of University of Agricultural Sciences and Veterinary Medicine Cluj-Napoca : Horticulture. 68(1):500-501. http://hdl.handle.net/10251/63083S50050168