Paul Christou

Catalan Institution for Research and Advanced Studies, Barcino, Catalonia, Spain

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Publications (255)1264.71 Total impact

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    ABSTRACT: Non-alcoholic fatty liver disease (NAFLD) is the most common form of chronic liver disease. Here we show that a mouse model of haploinsufficiency in the lipid and protein phosphatase and tensin homolog protein (PTEN+/−) exhibits hepatomegaly, increased liver lipogenic gene expression (SREBP-1C and PPARγ) and hepatic lesions analogous to human NAFLD. The livers of PTEN+/− mice also contained lower levels of retinoic acid (RA) than normal, similarly to human NAFLD patients. The RA signaling pathway thus offers a novel therapeutic target for the treatment of NAFLD although the impact of nutrition in this context is unclear. We therefore fed PTEN+/− mice for 36 weeks on a diet containing genetically engineered high-carotenoid corn (HCAR) to investigate its potential beneficial effects on the hepatic symptoms of NAFLD. The HCAR diet reduced hepatomegaly and promoted the repartitioning of fatty acids in the liver, away from triacylglycerol storage. At the molecular level, the HCAR diet clearly reduced lipogenic gene expression, boosted catabolism, and increased hepatic RA levels. These results set the stage for human trials to evaluate the use of high-carotenoid foods for the reduction or prevention of steatosis in NAFLD.
    No preview · Article · Jan 2016 · Biochimica et Biophysica Acta (BBA) - Molecular Basis of Disease
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    ABSTRACT: Protein microbicides containing neutralizing antibodies and antiviral lectins may help to reduce the rate of infection with human immunodeficiency virus (HIV) if it is possible to manufacture the components in large quantities at a cost affordable in HIV-endemic regions such as sub-Saharan Africa. We expressed the antiviral lectin griffithsin (GRFT), which shows potent neutralizing activity against HIV, in the endosperm of transgenic rice plants (Oryza sativa), to determine whether rice can be used to produce inexpensive GRFT as a microbicide ingredient. The yield of (OS) GRFT in the best-performing plants was 223 μg/g dry seed weight. We also established a one-step purification protocol, achieving a recovery of 74% and a purity of 80%, which potentially could be developed into a larger-scale process to facilitate inexpensive downstream processing. (OS) GRFT bound to HIV glycans with similar efficiency to GRFT produced in Escherichia coli. Whole-cell assays using purified (OS) GRFT and infectivity assays using crude extracts of transgenic rice endosperm confirmed that both crude and pure (OS) GRFT showed potent activity against HIV and the crude extracts were not toxic towards human cell lines, suggesting they could be administered as a microbicide with only minimal processing. A freedom-to-operate analysis confirmed that GRFT produced in rice is suitable for commercial development, and an economic evaluation suggested that 1.8 kg/ha of pure GRFT could be produced from rice seeds. Our data therefore indicate that rice could be developed as an inexpensive production platform for GRFT as a microbicide component.
    No preview · Article · Jan 2016 · Plant Biotechnology Journal
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    ABSTRACT: In this article, we explore the intellectual property (IP) landscape relevant to the production and commercialization of Carolight(™) , a transgenic multivitamin corn variety created on humanitarian grounds to address micronutrient deficiencies in low-and-middle-income countries. The successful production of this variety requires IP rights risk management because there is a strong protection on inventions and processes via patent portfolios in both developing and industrialized countries. The IP framework is complex, and specialist patent lawyers are usually employed to perform such analysis, but the costs cannot always be met by small, publicly funded projects. We report an alternative strategy, a do-it-yourself patent analysis, to produce a review with limited legal value that can nevertheless lay the foundations for a subsequent more in-depth professional freedom-to-operate opinion.
    Full-text · Article · Oct 2015 · Plant Biotechnology Journal
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    Dataset: Zhu-Korren

    Full-text · Dataset · Sep 2015
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    ABSTRACT: The apocarotenoid crocetin and its glycosylated derivatives, crocins, confer the red colour to saffron. Crocetin biosynthesis in saffron is catalysed by the carotenoid cleavage dioxygenase CCD2 (AIG94929). No homologues have been identified in other plant species due to the very limited presence of crocetin and its derivatives in the plant kingdom.Spring Crocus species with yellow flowers accumulate crocins in the stigma and tepals. Four carotenoid CCDs, namely CaCCD1, CaCCD2 and CaCCD4a/b and CaCCD4c were first cloned and characterized.CaCCD2 was localized in plastids, and a longer CCD2 version, CsCCD2L, was also localized in this compartment. The activity of CaCCD2 was assessed in Escherichia coli and in a stable rice gene function characterization system, demonstrating the production of crocetin in both systems. The expression of all isolated CCDs was evaluated in stigma and tepals at three key developmental stages in relation with apocarotenoid accumulation. CaCCD2 expression parallels crocin accumulation, but C14 apocarotenoids most likely are associated to the CaCCD1 activity in Crocus ancyrensis flowers.The specific CCD2 localization and its membrane interaction will contribute to the development of a better understanding of the mechanism of crocetin biosynthesis and regulation in the chromoplast.
    Full-text · Article · Sep 2015 · New Phytologist
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    ABSTRACT: Although plant biotechnology has been widely investigated for the production of clinical-grade monoclonal antibodies, no antibody products derived from transgenic plants have yet been approved by pharmaceutical regulators for clinical testing. In the Pharma-Planta project, the HIV-neutralizing human monoclonal antibody 2G12 was expressed in transgenic tobacco (Nicotiana tabacum). The scientific, technical and regulatory demands of good manufacturing practice (GMP) were addressed by comprehensive molecular characterization of the transgene locus, confirmation of genetic and phenotypic stability over several generations of transgenic plants, and by establishing standard operating procedures for the creation of a master seed bank, plant cultivation, harvest, initial processing, downstream processing and purification. The project developed specifications for the plant-derived antibody (P2G12) as an active pharmaceutical ingredient (API) based on (i) the guidelines for the manufacture of monoclonal antibodies in cell culture systems; (ii) the draft European Medicines Agency Points to Consider document on quality requirements for APIs produced in transgenic plants; and (iii) de novo guidelines developed with European national regulators. From the resulting process, a GMP manufacturing authorization was issued by the competent authority in Germany for transgenic plant-derived monoclonal antibodies for use in a phase I clinical evaluation. Following preclinical evaluation and ethical approval, a clinical trial application was accepted by the UK national pharmaceutical regulator. A first-in-human, double-blind, placebo-controlled, randomized, dose-escalation phase I safety study of a single vaginal administration of P2G12 was carried out in healthy female subjects. The successful completion of the clinical trial marks a significant milestone in the commercial development of plant-derived pharmaceutical proteins.
    No preview · Article · Jul 2015 · Plant Biotechnology Journal
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    ABSTRACT: Protein microbicides against HIV can help to prevent infection but they are required in large, repetitive doses. This makes current fermenter-based production systems prohibitively expensive. Plants are advantageous as production platforms because they offer a safe, economical and scalable alternative, and cereals such as rice are particularly attractive because they could allow pharmaceutical proteins to be produced economically and on a large scale in developing countries. Pharmaceutical proteins can also be stored as unprocessed seed, circumventing the need for a cold chain. Here, we report the development of transgenic rice plants expressing the HIV-neutralizing antibody 2G12 in the endosperm. Surprisingly for an antibody expressed in plants, the heavy chain was predominantly aglycosylated. Nevertheless, the heavy and light chains assembled into functional antibodies with more potent HIV-neutralizing activity than other plant-derived forms of 2G12 bearing typical high-mannose or plant complex-type glycans. Immunolocalization experiments showed that the assembled antibody accumulated predominantly in protein storage vacuoles but also induced the formation of novel, spherical storage compartments surrounded by ribosomes indicating that they originated from the endoplasmic reticulum. The comparison of wild-type and transgenic plants at the transcriptomic and proteomic levels indicated that endogenous genes related to starch biosynthesis were down-regulated in the endosperm of the transgenic plants, whereas genes encoding prolamin and glutaredoxin-C8 were up-regulated. Our data provide insight into factors that affect the functional efficacy of neutralizing antibodies in plants and the impact of recombinant proteins on endogenous gene expression. © 2015 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.
    No preview · Article · Apr 2015 · Plant Biotechnology Journal
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    ABSTRACT: The aim of this study was to assess whether endosperm-specific carotenoid biosynthesis influenced core metabolic processes in maize embryo and endosperm and how global seed metabolism adapted to this expanded biosynthetic capacity. Although enhancement of carotenoid biosynthesis was targeted to the endosperm of maize kernels, a concurrent up-regulation of sterol and fatty acid biosynthesis in the embryo was measured. Targeted terpenoid analysis, and non-targeted metabolomic, proteomic, and transcriptomic profiling revealed changes especially in carbohydrate metabolism in the transgenic line. In-depth analysis of the data, including changes of metabolite pools and increased enzyme and transcript concentrations, gave a first insight into the metabolic variation precipitated by the higher up-stream metabolite demand by the extended biosynthesis capacities for terpenoids and fatty acids. An integrative model is put forward to explain the metabolic regulation for the increased provision of terpenoid and fatty acid precursors, particularly glyceraldehyde 3-phosphate and pyruvate or acetyl-CoA from imported fructose and glucose. The model was supported by higher activities of fructokinase, glucose 6-phosphate isomerase, and fructose 1,6-bisphosphate aldolase indicating a higher flux through the glycolytic pathway. Although pyruvate and acetyl-CoA utilization was higher in the engineered line, pyruvate kinase activity was lower. A sufficient provision of both metabolites may be supported by a by-pass in a reaction sequence involving phosphoenolpyruvate carboxylase, malate dehydrogenase, and malic enzyme. © The Author 2015. Published by Oxford University Press on behalf of the Society for Experimental Biology.
    Full-text · Article · Mar 2015 · Journal of Experimental Botany
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    ABSTRACT: Carotenoids are health-promoting organic molecules that act as antioxidants and essential nutrients. We show that chickens raised on a diet enriched with an engineered corn variety containing very high levels of four key carotenoids (β-carotene, lycopene, zeaxanthin and lutein) are healthy and accumulate more bioavailable carotenoids in peripheral tissues, muscle, skin and fat, and more retinol in the liver, than birds fed on standard corn diets (including commercial corn supplemented with colour additives). Birds were challenged with the protozoan parasite Eimeria tenella and those on the high-carotenoid diet grew normally, suffered only mild disease symptoms (diarrhoea, footpad dermatitis and digital ulcers) and had lower faecal oocyst counts than birds on the control diet. Our results demonstrate that carotenoid-rich corn maintains poultry health and increases the nutritional value of poultry products without the use of feed additives. © 2015 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.
    Full-text · Article · Mar 2015 · Plant Biotechnology Journal
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    ABSTRACT: Plant metabolic pathways are complex and often feature multiple levels of regulation. Until recently, metabolic engineering in plants relied on the laborious testing of ad hoc modifications to achieve desirable changes in the metabolic profile. However, technological advances in data mining, modeling, multigene engineering and genome editing are now taking away much of the guesswork by allowing the impact of modifications to be predicted more accurately. In this review we discuss recent developments in knowledge-based metabolic engineering strategies, that is the gathering and mining of genomic, transcriptomic, proteomic and metabolomic data to generate models of metabolic pathways that help to define and refine optimal intervention strategies. Copyright © 2014 Elsevier Ltd. All rights reserved.
    Full-text · Article · Nov 2014 · Current Opinion in Biotechnology
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    Sol M Rivera · Paul Christou · Ramon Canela-Garayoa
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    ABSTRACT: The present review compiles positive MS fragmentation data of selected carotenoids obtained using various ionization techniques and matrices. In addition, new experimental data from the analysis of carotenoids in transgenic maize and rice callus are provided. Several carotenes and oxygen-functionalized carotenoids containing epoxy, hydroxyl, and ketone groups were ionized by atmospheric pressure chemical ionization (APCI)-tandem mass spectrometry (MS/MS) in positive ion mode. Thus, on the basis of the information obtained from the literature and our own experiments, we identified characteristic carotenoid ions that can be associated to functional groups in the structures of these compounds. In addition, pigments with a very similar structure were differentiated through comparison of the intensities of their fragments. The data provide a basis for the structural elucidation of carotenoids by mass spectrometry (MS). © 2013 Wiley Periodicals, Inc. Mass Spec Rev. 9999: 1-20, 2013.
    Full-text · Article · Sep 2014 · Mass Spectrometry Reviews
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    Maite Sabalza · Paul Christou · Teresa Capell
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    ABSTRACT: Molecular pharming is a cost-effective platform for the production of recombinant proteins in plants. Although the biopharmaceutical industry still relies on a small number of standardized fermentation-based technologies for the production of recombinant proteins there is now a greater awareness of the advantages of molecular pharming particularly in niche markets. Here we discuss some of the technical, economic and regulatory barriers that constrain the clinical development and commercialization of plant-derived pharmaceutical proteins. We also discuss strategies to increase productivity and product quality/homogeneity. The advantages of whole plants should be welcomed by the industry because this will help to reduce the cost of goods and therefore expand the biopharmaceutical market into untapped sectors.
    Full-text · Article · Jul 2014 · Biotechnology Letters
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    ABSTRACT: Carotenoids are nutritionally-beneficial organic tetraterpenoid pigments synthesized mainly by plants, bacteria and fungi. Although research has focused on the production of carotenoids in staple crops to improve nutritional welfare in developing countries, there is also an enormous market for carotenoids in the industrialized world, where they are produced both as commodities and luxury goods targeted at the pharmaceutical, nutraceutical, food/feed additive, cosmetics and fine chemicals sectors. Carotenoids are economically valuable because they have diverse bioactive and chemical properties. Some are essential nutrients (e.g. β-carotene), others are antioxidants with specific roles (e.g. lutein and zeaxanthin) or general health-promoting roles that reduce the risk or progression of diseases associated with oxidative stress (e.g. lycopene), and still others are natural pigments (e.g. astaxanthin, which is added to fish feed to impart a desirable pink flesh color). Even carotenoid degradation products, such as damascones and damascenones, are used as fragrances in the perfumes industry. Here we discuss the importance of carotenoids in different market sectors, review current methods for commercial production and its regulation, summarize the most relevant patents and consider evidence supporting the health claims made by different industry sectors, focusing on case studies representing the most commercially valuable carotenoids on the market: β-carotene, lycopene, lutein, zeaxanthin and astaxanthin.
    Full-text · Article · Jul 2014 · Phytochemistry Reviews
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    E Vamvaka · R M Twyman · P Christou · T Capell
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    ABSTRACT: The population of sub-Saharan Africa is at risk from multiple, poverty-related endemic diseases. HIV and malaria are the most prevalent, but they disproportionately affect different groups of people, i.e. HIV predominantly affects sexually-active adults whereas malaria has a greater impact on children and pregnant women. Nevertheless, there is a significant geographical and epidemiological overlap which results in bidirectional and synergistic interactions with important consequences for public health. The immunosuppressive effects of HIV increase the risk of infection when individuals are exposed to malaria parasites and also the severity of malaria symptoms. Similarly, acute malaria can induce a temporary increase in the HIV viral load. HIV is associated with a wide range of opportunistic infections that can be misdiagnosed as malaria, resulting in the wasteful misuse of antimalarial drugs and a failure to address the genuine cause of the disease. There is also a cumulative risk of toxicity when antiretroviral and antimalarial drugs are given to the same patients. Synergistic approaches involving the control of malaria as a strategy to fight HIV/AIDS and vice versa are therefore needed in co-endemic areas. Plant biotechnology has emerged as a promising approach to tackle poverty-related diseases because plant-derived drugs and vaccines can be produced inexpensively in developing countries and may be distributed using agricultural infrastructure without the need for a cold chain. Here we explore some of the potential contributions of plant biotechnology and its integration into broader multidisciplinary public health programs to combat the two diseases in developing countries.
    Full-text · Article · Mar 2014 · Biotechnology advances
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    ABSTRACT: Metabolic engineering can be used to modulate endogenous metabolic pathways in plants or introduce new metabolic capabilities in order to increase the production of a desirable compound or reduce the accumulation of an undesirable one. In practice, there are several major challenges that need to be overcome, such as gaining enough knowledge about the endogenous pathways to understand the best intervention points, identifying and sourcing the most suitable metabolic genes, expressing those genes in such a way as to produce a functional enzyme in a heterologous background, and, finally, achieving the accumulation of target compounds without harming the host plant. This article discusses the strategies that have been developed to engineer complex metabolic pathways in plants, focusing on recent technological developments that allow the most significant bottlenecks to be overcome. Expected final online publication date for the Annual Review of Plant Biology Volume 65 is April 29, 2014. Please see for revised estimates.
    Full-text · Article · Feb 2014 · Annual Review of Plant Biology
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    ABSTRACT: The do-it-yourself patent search is a useful alternative to professional patent analysis particularly in the context of publicly funded projects where funds for IP activities may be limited. As a case study, we analysed patents related to the engineering of terpenoid indole alkaloid (TIA) metabolism in plants. We developed a focused search strategy to remove redundancy and reduce the workload without missing important and relevant patents. This resulted in the identification of approximately 50 key patents associated with TIA metabolic engineering in plants, which could form the basis of a more detailed freedom-to-operate analysis. The structural elements of this search strategy could easily be transferred to other contexts, making it a useful generic model for publicly funded research projects.
    Full-text · Article · Feb 2014 · Plant Biotechnology Journal
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    ABSTRACT: Rice endosperm is devoid of carotenoids because the initial biosynthetic steps are absent. The early carotenogenesis reactions were constituted through co-transformation of endosperm-derived rice callus with phytoene synthase and phytoene desaturase transgenes. Subsequent steps in the pathway such as cyclization and hydroxylation reactions were catalyzed by endogenous rice enzymes in the endosperm. The carotenoid pathway was extended further by including a bacterial ketolase gene able to form astaxanthin, a high value carotenoid which is not a typical plant carotenoid. In addition to astaxanthin and precursors, a carotenoid accumulated in the transgenic callus which did not fit into the pathway to astaxanthin. This was subsequently identified as 4-keto-α-carotene by HPLC co-chromatography, chemical modification, mass spectrometry and the reconstruction of its biosynthesis pathway in Escherichia coli. We postulate that this keto carotenoid is formed from α-carotene which accumulates by combined reactions of the heterologous gene products and endogenous rice endosperm cyclization reactions.
    Full-text · Article · Jan 2014 · Phytochemistry
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    ABSTRACT: The food production and processing value chain is under pressure from all sides—increasing demand driven by a growing and more affluent population; dwindling resources caused by urbanization, land erosion, pollution and competing agriculture such as biofuels; and increasing constraints on production methods driven by consumers and regulators demanding higher quality, reduced chemical use, and most of all environmentally beneficial practices ‘from farm to fork’. This pressure can only be addressed by developing efficient and sustainable agricultural practices that are harmonized throughout the value chain, so that renewable resources can be exploited without damaging the environment. Bridges must, therefore, be built between the diverse areas within the food production and processing value chain, including bridges between different stages of production, between currently unlinked agronomic practices, and between the different levels and areas of research to achieve joined-up thinking within the industry, so that the wider impact of different technologies, practices and materials on productivity and sustainability is understood at the local, regional, national and global scales. In this article, we consider the challenges at different stages and levels of the value chain and how new technologies and strategies could be used to build bridges and achieve more sustainable food/feed production in the future.
    No preview · Article · Dec 2013 · Molecular Breeding
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    ABSTRACT: We have developed an assay based on rice embryogenic callus for the rapid functional characterization of metabolic genes. We validated the assay using a selection of well-characterized genes with known functions in the carotenoid biosynthesis pathway, allowing the rapid visual screening of callus phenotypes based on tissue color. We were then able to use the system to identify the functions of two uncharacterized genes: a chemically-synthesized β-carotene ketolase gene optimized for maize codon usage; and a wild-type Arabidopsis thaliana ortholog of the cauliflower Orange gene. In contrast to previous reports, we found that the wild-type Orange allele was sufficient to induce chromoplast differentiation. We also found that chromoplast differentiation could be induced by increasing the availability of precursors and thus driving flux through the pathway, even in the absence of Orange. Remarkably, we found that diverse endosperm-specific promoters were highly active in rice callus despite their restricted activity in mature plants. Our callus system provides a unique opportunity to predict the impact of metabolic engineering in complex pathways and provides a starting point for quantitative modeling and the rational design of engineering strategies using synthetic biology. We discuss the impact of our data on the analysis and engineering of the carotenoid biosynthesis pathway. This article is protected by copyright. All rights reserved.
    Full-text · Article · Nov 2013 · The Plant Journal
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    ABSTRACT: Over the last two decades, many carotenogenic genes have been cloned and used to generate metabolically-engineered plants producing higher levels of carotenoids. However, comparatively little is known about the regulation of endogenous carotenogenic genes in higher plants, and this restricts our ability to predict how engineered plants will perform in terms of carotenoid content and composition. During petal development in the Great Yellow Gentian (Gentiana lutea), carotenoid accumulation, the formation of chromoplasts and the upregulation of several carotenogenic genes are temporally coordinated. We investigated the regulatory mechanisms responsible for this coordinated expression by isolating five G. lutea carotenogenic gene (GlPDS, GlZDS, GlLYCB, GlBCH and GlLYCE) promoters by inverse PCR. Each promoter was sufficient for developmentally-regulated expression of the gusA reporter gene following transient expression in tomato (Solanum lycopersicum cv. Micro-Tom). Interestingly, the GlLYCB and GlBCH promoters drove high levels of gusA expression in chromoplast-containing mature green fruits, but low levels in chloroplast-containing immature green fruits, indicating a strict correlation between promoter activity, tomato fruit development and chromoplast differentiation. As well as core promoter elements such as TATA and CAAT boxes, all five promoters together with previously characterized GlZEP promoter contained three common cis-regulatory motifs involved in the response to methyl jasmonate (CGTCA) and ethylene (ATCTA), and required for endosperm expression (Skn-1_motif, GTCAT). These shared common cis-acting elements may represent binding sites for transcription factors responsible for co-regulation. Our data provide insight into the regulatory basis of the coordinated upregulation of carotenogenic gene expression during flower development in G. lutea.
    Full-text · Article · Nov 2013 · Physiologia Plantarum

Publication Stats

14k Citations
1,264.71 Total Impact Points


  • 2008-2015
    • Catalan Institution for Research and Advanced Studies
      Barcino, Catalonia, Spain
  • 2005-2015
    • Universitat de Lleida
      • Department of Vegetal Production and Forestry Science
      Lérida, Catalonia, Spain
  • 1994-2013
    • John Innes Centre
      • Department of Cell and Developmental Biology
      Norwich, England, United Kingdom
  • 2003-2006
    • Fraunhofer Institute for Molecular Biology and Applied Ecology IME
      • Department of Plant Biotechnology
      Aachen, North Rhine-Westphalia, Germany
  • 2004
    • RWTH Aachen University
      • Institute of Biology VII (Molecular Biotechnology)
      Aachen, North Rhine-Westphalia, Germany
    • Institute of Molecular Biology
      Mayence, Rheinland-Pfalz, Germany
    • University of Minnesota Duluth
      Duluth, Minnesota, United States
  • 2002
    • University of California, Davis
      • Department of Plant Biology
      Davis, California, United States
  • 2001
    • Université de Technologie de Compiègne
      Compiègne, Picardie, France
  • 2000
    • Durham University
      • School of Biological and Biomedical Sciences
      Durham, England, United Kingdom
  • 1986-1990
    • University of Wisconsin - Green Bay
      Green Bay, Wisconsin, United States