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Functional analysis of vitamin E biosynthesis in tomato fruits by a gfp‐based VIGS system‐based VIGS system Leandro Quadrana1, Ramon Asis3, Mariana Lopez1, Juliana Almeida2, Alisdair R. Fernie4, Magdalena Rossi2, Fernando Carrari1. Rossi2, Fernando Carrari1. 1, Ramon Asis3, Mariana Lopez1, Juliana Almeida2, Alisdair R. Fernie4, Magdalena Rossi2, Fernando Carrari1.2, Fernando Carrari1. 1Instituto de Biotecnologia, Instituto Nacional de Tecnologia Agropecuaria (IB‐INTA), and Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), PO Box 25, B1712WAA Castelar, Argentina (partner group of the Max Planck Institute for Molecular Plant Physiology, Potsdam‐Golm, Germany). Investigaciones Científicas y Técnicas (CONICET), PO Box 25, B1712WAA Castelar, Argentina (partner group of the Max Planck Institute for Molecular Plant Physiology, Potsdam‐Golm, Germany). Instituto de Biotecnologia, Instituto Nacional de Tecnologia Agropecuaria (IB‐INTA), and Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), PO Box 25, B1712WAA Castelar, Argentina (partner group of the Max Planck Institute for Molecular Plant Physiology, Potsdam‐Golm, Germany).‐Golm, Germany). 2Departamento de Botanica‐IB‐USP, 277, 05508‐900, São Paulo, SP, Brazil.Departamento de Botanica‐IB‐USP, 277, 05508‐900, São Paulo, SP, Brazil. 3CIBICI, Facultad de Ciencias Quimicas Universidad Nacional de Cordoba, CC 5000, Cordoba, Argentina.CIBICI, Facultad de Ciencias Quimicas Universidad Nacional de Cordoba, CC 5000, Cordoba, Argentina. 4Max Planck Institute for Molecular Plant Physiology, Golm, GermanyMax Planck Institute for Molecular Plant Physiology, Golm, Germany lquadrana@cnia.inta.gov.ar Vitamin E comprises a family of amphiphilic antioxidants incluiding tocopherols and tocotrienols, which are exclusively synthesized by photosynthetic organisms. Four isomers: alfa, beta, gamma and delta tocopherol are produced in tomato. Alfa‐ tocopherol is the most abundant in fruits and has the mayor vitamin E related activity. Even when most of the enzymes of Vitamin E biosynthesis were characterised in Arabidopis, the understanding of this biosynthetic pathway in tomato is scarce. By screening the Solanum pennellii introgression line (IL) population we found that ripe fruits from IL 8‐2, 8‐2‐1, 9‐2 and 9‐2‐6 present significant alterations in their tocopherol profiles. Moreover, genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding 8‐2, 8‐2‐1, 9‐2 and 9‐2‐6 present significant alterations in their tocopherol profiles. Moreover, genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding scarce. By screening the Solanum pennellii introgression line (IL) population we found that ripe fruits from IL 8‐2, 8‐2‐1, 9‐2 and 9‐2‐6 present significant alterations in their tocopherol profiles. Moreover, genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding 8‐2, 8‐2‐1, 9‐2 and 9‐2‐6 present significant alterations in their tocopherol profiles. Moreover, genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding has the mayor vitamin E related activity. Even when most of the enzymes of Vitamin E biosynthesis were characterised in Arabidopis, the understanding of this biosynthetic pathway in tomato is scarce. By screening the Solanum pennellii introgression line (IL) population we found that ripe fruits from IL 8‐2, 8‐2‐1, 9‐2 and 9‐2‐6 present significant alterations in their tocopherol profiles. Moreover, genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding 8‐2, 8‐2‐1, 9‐2 and 9‐2‐6 present significant alterations in their tocopherol profiles. Moreover, genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding scarce. By screening the Solanum pennellii introgression line (IL) population we found that ripe fruits from IL 8‐2, 8‐2‐1, 9‐2 and 9‐2‐6 present significant alterations in their tocopherol profiles. Moreover, genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding 8‐2, 8‐2‐1, 9‐2 and 9‐2‐6 present significant alterations in their tocopherol profiles. Moreover, genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding ‐ tocopherol is the most abundant in fruits and has the mayor vitamin E related activity. Even when most of the enzymes of Vitamin E biosynthesis were characterised in Arabidopis, the understanding of this biosynthetic pathway in tomato is scarce. By screening the Solanum pennellii introgression line (IL) population we found that ripe fruits from IL 8‐2, 8‐2‐1, 9‐2 and 9‐2‐6 present significant alterations in their tocopherol profiles. Moreover, genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding 8‐2, 8‐2‐1, 9‐2 and 9‐2‐6 present significant alterations in their tocopherol profiles. Moreover, genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding scarce. By screening the Solanum pennellii introgression line (IL) population we found that ripe fruits from IL 8‐2, 8‐2‐1, 9‐2 and 9‐2‐6 present significant alterations in their tocopherol profiles. Moreover, genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding 8‐2, 8‐2‐1, 9‐2 and 9‐2‐6 present significant alterations in their tocopherol profiles. Moreover, genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding Arabidopis, the understanding of this biosynthetic pathway in tomato is scarce. By screening the Solanum pennellii introgression line (IL) population we found that ripe fruits from IL 8‐2, 8‐2‐1, 9‐2 and 9‐2‐6 present significant alterations in their tocopherol profiles. Moreover, genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding 8‐2, 8‐2‐1, 9‐2 and 9‐2‐6 present significant alterations in their tocopherol profiles. Moreover, genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding Solanum pennellii introgression line (IL) population we found that ripe fruits from IL 8‐2, 8‐2‐1, 9‐2 and 9‐2‐6 present significant alterations in their tocopherol profiles. Moreover, genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding ‐2, 8‐2‐1, 9‐2 and 9‐2‐6 present significant alterations in their tocopherol profiles. Moreover, genomic analyses allowed identification of slvte1, slvte4 and slvte3, orthologs to the corresponding Arabidopsis genes onto the regions spanning the mentioned introgressions. In order to study the function of these genes in tomato, we designed a suitable platform for functional studies based on viral induced gene silencing (VIGS). To overcome the well known limitations imposed by this system (ie. patchy phenotypes, pleiotropic effects of the tracking marker), we first analysed the impact on fruit metabolism of two reporter genes, the phytoene desaturase (pds) and gfp in WT and transgenic GFP plants respectively. GC‐MS metabolite profiles demonstrated that pds silencing produces dramatic alterations on pigments contents (carotenoids and tocopherols) in both mature green and ripe tomato fruits. By contrast, gfp silencing produces alterations in neither primary metabolism nor pigments accumulation, making it a suitable system for tocopherol metabolism studies. Results obtained by silencing tocopherol genes in fruits will be discussed. demonstrated that pds silencing produces dramatic alterations on pigments contents (carotenoids and tocopherols) in both mature green and ripe tomato fruits. By contrast, gfp silencing produces alterations in neither primary metabolism nor pigments accumulation, making it a suitable system for tocopherol metabolism studies. Results obtained by silencing tocopherol genes in fruits will be discussed. In order to study the function of these genes in tomato, we designed a suitable platform for functional studies based on viral induced gene silencing (VIGS). To overcome the well known limitations imposed by this system (ie. patchy phenotypes, pleiotropic effects of the tracking marker), we first analysed the impact on fruit metabolism of two reporter genes, the phytoene desaturase (pds) and gfp in WT and transgenic GFP plants respectively. GC‐MS metabolite profiles demonstrated that pds silencing produces dramatic alterations on pigments contents (carotenoids and tocopherols) in both mature green and ripe tomato fruits. By contrast, gfp silencing produces alterations in neither primary metabolism nor pigments accumulation, making it a suitable system for tocopherol metabolism studies. Results obtained by silencing tocopherol genes in fruits will be discussed. demonstrated that pds silencing produces dramatic alterations on pigments contents (carotenoids and tocopherols) in both mature green and ripe tomato fruits. By contrast, gfp silencing produces alterations in neither primary metabolism nor pigments accumulation, making it a suitable system for tocopherol metabolism studies. Results obtained by silencing tocopherol genes in fruits will be discussed. genes onto the regions spanning the mentioned introgressions. In order to study the function of these genes in tomato, we designed a suitable platform for functional studies based on viral induced gene silencing (VIGS). To overcome the well known limitations imposed by this system (ie. patchy phenotypes, pleiotropic effects of the tracking marker), we first analysed the impact on fruit metabolism of two reporter genes, the phytoene desaturase (pds) and gfp in WT and transgenic GFP plants respectively. GC‐MS metabolite profiles demonstrated that pds silencing produces dramatic alterations on pigments contents (carotenoids and tocopherols) in both mature green and ripe tomato fruits. By contrast, gfp silencing produces alterations in neither primary metabolism nor pigments accumulation, making it a suitable system for tocopherol metabolism studies. Results obtained by silencing tocopherol genes in fruits will be discussed. demonstrated that pds silencing produces dramatic alterations on pigments contents (carotenoids and tocopherols) in both mature green and ripe tomato fruits. By contrast, gfp silencing produces alterations in neither primary metabolism nor pigments accumulation, making it a suitable system for tocopherol metabolism studies. Results obtained by silencing tocopherol genes in fruits will be discussed. ‐MS metabolite profiles demonstrated that pds silencing produces dramatic alterations on pigments contents (carotenoids and tocopherols) in both mature green and ripe tomato fruits. By contrast, gfp silencing produces alterations in neither primary metabolism nor pigments accumulation, making it a suitable system for tocopherol metabolism studies. Results obtained by silencing tocopherol genes in fruits will be discussed.