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Tesis Doctoral UNIVERSIDAD DE CÓRDOBA FACULTAD DE CIENCIAS DEPARTAMENTO DE BIOQUÍMICA Y BIOLOGÍA MOLECULAR TESIS DOCTORAL "FUNCTIONAL CHARACTERIZATION OF STRAWBERRY ( FRAGARIA x ANANASSA ) FRUIT- SPECIFIC AND RIPENING-RELATED GENES INVOLVED IN AROMA AND ANTHOCHYANINS BIOSYNTHESIS" Memoria de Tesis Doctoral presentada por Guadalupe Cumplido Laso, Licenciada en Biología, para optar al grado de Doctor por la Universidad de Córdoba con la mención de Doctorado Internacional . Córdoba, Diciembre de 2012 TITULO: FUNCTIONAL CHARACTERIZATION OF STRAWBERRY (FRAGARIA x ANANASSA) FRUIT-SPECIFIC AND RIPENING-RELATED GENES INVOLVED IN AROMA AND ANTHOCHYANINS BIOSYNTHESIS AUTOR: GUADALUPE CUMPLIDO LASO © Edita: Servicio de Publicaciones de la Universidad de Córdoba. Campus de Rabanales Ctra. Nacional IV, Km. 396 A 14071 Córdoba www.uco.es/publicaciones [email protected] TÍTULO DE LA TESIS: “ Functional characterization of strawberry ( Fragaria x ananassa ) fruit-specific and ripening-related genes involved in aroma and anthocyanins biosynthesis ” DOCTORANDO/A: Guadalupe Cumplido Laso INFORME RAZONADO DEL/DE LOS DIRECTOR/ES DE LA TESIS (se hará mención a la evolución y desarrollo de la tesis, así como a trabajos y publicaciones derivados de la misma). La Lda. Guadalupe Cumplido Laso ha desarrollado en el seno del grupo BIO-278 liderado por el Dr. Juan Muñoz Blanco el trabajo de investigación llamado “ Functional characterization of strawberry ( Fragaria x ananassa ) fruit-specific and ripening- related genes involved in aroma and anthocyanins biosynthesis ” que constituye el tema de su tesis doctoral. Este trabajo de investigación ha sido dirigido y supervisado por el Dr. Juan Muñoz Blanco y la Dra. Rosario Blanco Portales, ambos miembros del Departamento de Bioquímica y Biología Molecular de la Universidad de Córdoba. Durante el periodo de investigación en el que se ha desarrollado esta tesis, la Lda. Cumplido Laso, además de haber desarrollado un correcto aprendizaje en el laboratorio que la ha permitido adquirir una amplia experiencia en diferentes técnicas de Biología Molecular (Microarrays, extracción de RNA, amplificación de RNA, generación de construcciones mediante tecnología Gateway, obtención e inducción de proteínas recombinantes, transformación de plantas de fresa de forma estable y transitoria, cultivo “in vitro” de plantas de fresa y las demás mostradas en el apartado de materiales y métodos de la tesis presentada) ha realizado tres estancias cortas (13 meses en total) en el grupo de investigación dirigido por el Dr. Willfried Schwab en el Departamento “Biomolecular food technology” de la Universidad Técnica de Munich (TUM), donde se realizaron parte de los análisis presentados en la tesis. Así, durante el desarrollo de esta tesis doctoral se han aislado varios genes, FaAAT2 y FaMyb10 , directamente relacionados con la producción del aroma y la síntesis de antocianinas respectivamente, en el fruto de fresa. El hecho de que ambos genes estén involucrados en el desarrollo de dos características tan determinantes de las propiedades organolépticas finales del fruto de fresa hace este trabajo de investigación especialmente interesante desde el punto de vista biotecnológico y abre las puertas a la obtención en el futuro de frutos de fresa mejorados. Por otra parte, los resultados mostrados en la tesis y las colaboraciones realizadas por la Lda. Cumplido-Laso dentro del grupo BIO-278 y con otros grupos de investigación de la Universidad de Málaga y de Munich han dado lugar a 11 comunicaciones a congresos, nacionales e internacionales, además de a tres publicaciones (dos de ellas bajo revisión) en revistas de alto índice de impacto en el campo de la investigación en plantas superiores que avalan su interés para la comunidad científica. Así, las comunicaciones presentadas a congresos han sido: “Searching for the Starwberry genes by functional genomic as valuable biotechnological tools”. Muñoz-Blanco, José Luis Caballero, Rosario Blanco-Portales, Mari Luz Bellido Cabello de Alba, Guadalupe Cumplido Laso , Bridget Moreno Suárez, Carmen García Limones, Francisco Amil-Ruíz, Sonia Encinas Villarejo, Aureliano Bombarelly, José Sánchez Sevilla and Victoriano Valpuesta. VI Internacional Strawberry Symposium ISHS (2008) (Huelva). Comunicación oral. “Caracterización molecular de genes relacionados en procesos de resistencia a patógenos y en la maduración del fruto de fresa”. Bridget Moreno-Suárez, Guadalupe Cumplido-Laso , Laura Medina Puche, M Luz Bellido Cabello de Alba, José Luis Caballero, Juan Muñoz-Blanco y Rosario Blanco Portales. IX Reunión de Biología Molecular de Plantas (2008) (Santiago de Compostela). “Estudios de expresión y función fisiológica de una alcohol acil transferasa (FaAAT2) del fruto de fresa ( Fragaria x ananassa )”. Guadalupe Cumplido-Laso , Laura Medina Puche, Bridget Moreno-Suárez, MLuz Bellido Cabello de Alba, Carmen García-Limones, Nicolás García Caparrós, Jose Luis Caballero, Juan Muñoz-Blanco y Rosario Blanco Portales. IX Reunión de Biología Molecular de Plantas (2008) (Santiago de Compostela). “Estudios funcionales de genes potencialmente reguladores del proceso de maduración de la fresa (Fragaria x ananassa)”. Rosario Blanco Portales, Guadalupe Cumplido-Laso , Bridget Moreno-Suárez, Laura Medina Puche, Carmen García-Limones, M Luz Bellido Cabello de Alba, Enriqueta Moyano Cañete, José Luis Caballero, Juan Muñoz-Blanco. IX Reunión de Biología Molecular de Plantas (2008) (Santiago de Compostela). “MADS-BOX genes envolved in strawberry fruit ripening: identification of putative target genes regulated by FAMADS1 using microarrays” . M Luz Bellido Cabello de Alba, Rosario Blanco Portales, Enriqueta Moyano Cañete, Carmen García-Limones, Guadalupe Cumplido-Laso , Bridget Moreno-Suárez, Laura Medina Puche, José Luis Caballero y Juan Muñoz-Blanco. IX Reunión de Biología Molecular de Plantas (2008) (Santiago de Compostela). “Determinación del papel que desempeña el ácido abscísico en la maduarción de la fresa mediante estudios funcionales de los genes NCEDs”. Laura Medina Puche, Rosario Blanco Portales, Guadalupe Cumplido-Laso , Bridget Moreno-Suárez, M Luz Bellido Cabello de Alba, Carmen García-Limones, Nicolás García Caparrós, José Luis Caballero y Juan Muñoz-Blanco. IX Reunión de Biología Molecular de Plantas (2008) (Santiago de Compostela). “Using of an fruit derived ESTs microarray platform to discover new candidate genes with potential biotechnological roles in strawberry fruit development, ripening and senescence”. Blanco-Portales R, Bellido-Cabello de Alba M Luz, Cumplido-Laso G , Medina-Puche, L, González A, Trelles O, Schwab W, Caballero JL, Muñoz-Blanco J. Plant GEMs (2009) (Lisboa) “Caracterización funcional de un gen que codifica a una alcohol acil trasferasa (Faaat2) de fruto de fresa (Fragaria x ananassa). Guadalupe Cumplido-Laso , Laura Medina-Puche, Bridget Moreno-Suárez, Enriqueta Moyano-Cañete, Wilfried Schwab, Juan Muñoz-Blanco, Rosario Blanco-Portales. XXXIII Congreso de la Sociedad Española de Bioquímica y Biología Molecular (SEBBM) (2010) (Córdoba). Comunicación oral. “Estudios funcionales de una aldo-ceto reductasa (FaAKR3) en el proceso de maduración del fruto de fresa (Fragaria x ananassa)”. José Antonio Mérida, Henrik Keränen, Laura Medina, Guadalupe Cumplido-Laso , Rosario Blanco-Portales, Juan Muñoz, Enriqueta Moyano. XXXIII Congreso de la Sociedad Española de Bioquímica y Biología Molecular (SEBBM) (2010) (Córdoba). “Using of an strawberry oligo microarray platform to discover new candidate genes with potential biotechnological roles in strawberry fruit development, ripening and senescence”. Blanco-Portales, Rosario; Moyano, Enriqueta; Cumplido-Laso, Guadalupe ; Medina- Puche, Laura; Trelles, Oswaldo; Caballero, José Luis; Muñoz-Blanco, Juan. 28th International Horticultural Congress. IHC (2010) (Lisboa). “Using of an derived ESTs microarray platform to discover new candidate genes with potential biotechnological roles in strawberry fruit development, ripening and senescence ”. Blanco-Portales, Rosario; Moyano, Enriqueta; Cumplido-Laso, Guadalupe ; Medina- Puche, Laura; Muñoz-Mérida, Antonio; Trelles, Oswaldo; Schwab, Wilfried; Caballero, José Luis; Muñoz-Blanco, Juan. 28th International Horticultural Congress. IHC (2010) (Lisboa). Charla invitada. Los artículos de investigación publicados o bajo revisión han sido: “The fruit ripening-related gene FaAAT2 encodes an acyl transferase involved in strawberry aroma biogénesis”. Cumplido-Laso G , Medina-Puche L, Moyano E, Hoffmann T, Sinz Q, Ring L, Studart-Wittkowski C, Caballero JL, Schwab W, Muñoz- Blanco J, Blanco-Portales R. (2012). Journal of Experimental Botany, 63(11): 4275-90. Índice de impacto: 5,3 “FaMYB10 plays a major role in the regulation of the flavonoid/phenylpropanoid metabolism during the ripening of Fragaria x ananassa fruits” . Guadalupe Cumplido Laso ; Laura Medina Puche; Francisco Amil Ruiz; Bridget Moreno Suarez; Ludwig Ring; Antonio Rodriguez Franco; José Luis Caballero; Wilfried Schwab; Juan Muñoz Blanco; Rosario Blanco Portales. Plant Physiology (bajo revisión). Índice de impacto: 7,05. A mi familia AGRADECIMIENTOS Durante este período de tiempo he tenido la suerte de conocer a muchas personas que han contribuido a sacar este trabajo adelante, por lo que quiero mostrar a través de algunas palabras todo mi agradecimiento. En primer lugar quiero agradecer a mis directores de Tesis, Juan Muñoz Blanco y Rosario Blanco Portales, por haber apostado por mí, darme la oportunidad de trabajar con ellos y adentrarme en el mundo de la ciencia.
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