Estudio genético de fenotipos de interés enológico en levaduras vínicas de Saccharomyces cerevisiae
No hay miniatura disponible
Archivos
Fecha
2013
Autores
Profesor/a Guía
Facultad/escuela
Idioma
es
Título de la revista
ISSN de la revista
Título del volumen
Editor
Universidad Andrés Bello
Nombre de Curso
Licencia CC
Licencia CC
Resumen
Saccharomyces cerevisiae es la principal especie responsable de la
fermentación alcohólica. En esta levadura los fenotipos de interés enológico como el
consumo de nitrógeno son poligénicos y considerados rasgos cuantitativos o rasgos
complejos. Con el objetivo de identificar regiones del genoma (QTLs) y genes
asociados a fenotipos vinculados al consumo de nitrógeno, se realizó un análisis de
ligamiento entre fenotipos y marcadores moleculares, a partir de un cruce entre cepas
filogenéticamente divergentes que comparten una asociación a procesos fermentativos
vínico y sake pero que, sin embargo, presentan diferentes características fenotípicas en
fermentación. 96 segregantes provenientes de este cruce fueron sometidos a
condiciones de fermentación en mosto sintético y, al día 6 de fermentación, fueron
analizadas para el consumo de 20 fuentes de nitrógeno y nitrógeno asimilable por la
levadura (YAN). El análisis de ligamiento entre fenotipo y genotipo permitió mapear
51 QTLs vinculados al consumo de estas fuentes de nitrógeno, determinándose que
una media de 3 QTLs, así como los efectos aditivos y epistáticos entre ellos, explican
las variaciones fenotípicas observadas para estos rasgos. Con el objetivo de validar los
QTLs identificados, se realizó un análisis bioinformático que permitió determinar 40
genes candidatos para explicar las diferencias fenotípicas observadas, de los cuales 9
genes fueron analizados. Estos genes fueron estudiados mediante reciprocidad
hemicigótica, mutando de forma heterocigota cada gen en la cepa híbrida. Este análisis
permitió determinar que las variaciones alélicas en genes vinculados al sistema sensor
de aminoácidos (PTR3), el sistema de fidelización del sistema sensor de aminoácidos (ASI), los reguladores de la represión catabólica del nitrógeno (GATI y GAT2), en
permeasas de amplio espectro (AGPI y AGP3), además de genes vinculados al
metabolismo central del nitrógeno (GLTI) y vinculados a la biosíntesis de metabolitos
derivados de aminoácidos (SPE4), afectan el consumo de fuentes de nitrógeno. El análisis
filogenético de estos genes permitió observar que estos alelos son característicos de cepas
vínicas y de sake. Es más, el análisis de secuencia en ASh y GAT2, dos genes ligados en el
cromosoma XIII que mostraron un alto nivel de pleiotropía antagonista y efectos
epistáticos, permitió determinar una elevada tasa de polimorfismos sinónimos indicativo de
selección balanceadora, sugiriendo que si esta es una característica común en la levadura
los polimorfismos no-neutrales epistáticos pueden ser un contribuyente importante para la
variación genética de rasgos complejos.
Saccharomyces cerevisiae is the main species responsible for alcoholic fermentation. In this yeast phenotypes of oenological interest such as nitrogen uptake are polygenic and considered quantitative or complex traits. With the aim of identifying genome regions (QTLs) and genes associated to phenotypes linked to nitrogen consumption, a linkage analysis was carried out between phenotypes and molecular markers from a cross between phylogenetically divergent strains. Although these strains share an association to wine and sake fermentative processes, they have different phenotypic characteristics during the fermentation. From this crossbreed, 96 segregants were fermented in synthetic wine must and analyzed for the uptake of 20 nitrogen sources and yeast asimilable nitrogen (YAN) at day 6 of the fermentation. Linkage analysis between phenotype and genotype allowed us to map 51 QTLs linked to the consumption of these nitrogen sources determining that an average of 3 QTLs, as well as the additive and epistatic effects among them explain the observed phenotypic variation for these traits. In order to validate the QTLs identified, a bioinformatic analysis carried out identified 40 candidate genes to explain the observed phenotypic differences, from which 9 were analyzed. These genes were studied by reciprocal hemizygotic analysis where each gene was mutated in a heterozygous manner in the hybrid strain. This analysis revealed that the allelic variations in genes associated to the amino acid sensor system (PTR3), the system ensuring the fidelity in the amino acid sensor system (Ash), regulators of nitrogen catabolite repression (GATT and GAT2), broad spectrum permeases (AGPI and AGP3), genes associated to the central nitrogen metabolism (GLTI) and the biosynthesis of amino acid derived metabolites (SPE4), affect the uptake of nitrogen sources. Phylogenetic analysis of these genes showed that these alleles are typical from wine and sake strains. Moreover, sequence analysis of ASh and GAT2, two genes linked on chromosome XIII displaying a high level of antagonistic pleiotropy and epistatic effects, revealed a high rate of synonym polymorphisms indicative of balancing selection. Therefore, if this feature is common in yeast, non-neutral epistatic polymorphisms may be an outstanding contributor to the genetic variation of complex traits.
Saccharomyces cerevisiae is the main species responsible for alcoholic fermentation. In this yeast phenotypes of oenological interest such as nitrogen uptake are polygenic and considered quantitative or complex traits. With the aim of identifying genome regions (QTLs) and genes associated to phenotypes linked to nitrogen consumption, a linkage analysis was carried out between phenotypes and molecular markers from a cross between phylogenetically divergent strains. Although these strains share an association to wine and sake fermentative processes, they have different phenotypic characteristics during the fermentation. From this crossbreed, 96 segregants were fermented in synthetic wine must and analyzed for the uptake of 20 nitrogen sources and yeast asimilable nitrogen (YAN) at day 6 of the fermentation. Linkage analysis between phenotype and genotype allowed us to map 51 QTLs linked to the consumption of these nitrogen sources determining that an average of 3 QTLs, as well as the additive and epistatic effects among them explain the observed phenotypic variation for these traits. In order to validate the QTLs identified, a bioinformatic analysis carried out identified 40 candidate genes to explain the observed phenotypic differences, from which 9 were analyzed. These genes were studied by reciprocal hemizygotic analysis where each gene was mutated in a heterozygous manner in the hybrid strain. This analysis revealed that the allelic variations in genes associated to the amino acid sensor system (PTR3), the system ensuring the fidelity in the amino acid sensor system (Ash), regulators of nitrogen catabolite repression (GATT and GAT2), broad spectrum permeases (AGPI and AGP3), genes associated to the central nitrogen metabolism (GLTI) and the biosynthesis of amino acid derived metabolites (SPE4), affect the uptake of nitrogen sources. Phylogenetic analysis of these genes showed that these alleles are typical from wine and sake strains. Moreover, sequence analysis of ASh and GAT2, two genes linked on chromosome XIII displaying a high level of antagonistic pleiotropy and epistatic effects, revealed a high rate of synonym polymorphisms indicative of balancing selection. Therefore, if this feature is common in yeast, non-neutral epistatic polymorphisms may be an outstanding contributor to the genetic variation of complex traits.
Notas
Tesis (Bioquímico, Magíster en Bioquímica)
Esta tesis se realizó en el Laboratorio de Microbiología Aplicada y Biotecnología de la Facultad Tecnológica de la Universidad de Santiago de Chile y fue financiada por el proyecto FONDECYT W 110059: "Genetic and Molecular Improvement of Yeast for Wine Making".
Esta tesis se realizó en el Laboratorio de Microbiología Aplicada y Biotecnología de la Facultad Tecnológica de la Universidad de Santiago de Chile y fue financiada por el proyecto FONDECYT W 110059: "Genetic and Molecular Improvement of Yeast for Wine Making".
Palabras clave
Genética, Fenotipo, Levaduras, Composición, Saccharomyces Cerevisiae