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1. A micromethod for high throughput RNA extraction in forest trees

Author

GREGOIRE LE PROVOST, RAÚL HERRERA, JORGE AP PAIVA, PHILIPPE CHAUMEIL, FRANCK SALIN, and CHRISTOPHE PLOMION

Subjects
RNA extraction, micromethod, forest trees, Biology (General), QH301-705.5
Abstract

A large quantity of high quality RNA is often required in the analysis of gene expression. However, RNA extraction from samples taken from woody plants is generally complex, and represents the main limitation to study gene expression, particularly in refractory species like conifers. Standard RNA extraction protocols are available but they are highly time consuming, and not adapted to large scale extraction. Here we present a high-throughput RNA extraction protocol. This protocol was adapted to a micro-scale by modifying the classical cetyltrimethylammonium (CTAB) protocol developed for pine: (i) quantity of material used (100-200 mg of sample), (ii) disruption of samples in microtube using a mechanical tissue disrupter, and (iii) the use of SSTE buffer. One hundred samples of woody plant tissues/organs can be easily treated in two working days. An average of 15 ig of high quality RNA per sample was obtained. The RNA extracted is suitable for applications such as real time reverse transcription polymerase chain reaction, cDNA library construction or synthesis of complex targets for microarray analysis

Published
2007

3. Chloroplast DNA variation in a hyperdiverse tropical tree community

Author

Jean-Marc Frigerio, Rémy J. Petit, Ivan Scotti, Caroline Scotti-Saintagne, Alain Franc, Philippe Chaumeil, Daniel Sabatier, Patrick Léger, Jean-François Molino, Henri Caron, Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), Ecologie des forêts de Guyane (UMR ECOFOG), Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Institut National de la Recherche Agronomique (INRA)-AgroParisTech-Université de Guyane (UG)-Centre National de la Recherche Scientifique (CNRS)-Université des Antilles (UA), Botanique et Modélisation de l'Architecture des Plantes et des Végétations (UMR AMAP), Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Institut National de la Recherche Agronomique (INRA)-Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche pour le Développement (IRD [France-Sud]), from patterns to models in computational biodiversity and biotechnology (PLEIADE), Laboratoire Bordelais de Recherche en Informatique (LaBRI), Université de Bordeaux (UB)-École Nationale Supérieure d'Électronique, Informatique et Radiocommunications de Bordeaux (ENSEIRB)-Centre National de la Recherche Scientifique (CNRS)-Université de Bordeaux (UB)-École Nationale Supérieure d'Électronique, Informatique et Radiocommunications de Bordeaux (ENSEIRB)-Centre National de la Recherche Scientifique (CNRS)-Inria Bordeaux - Sud-Ouest, Institut National de Recherche en Informatique et en Automatique (Inria)-Institut National de Recherche en Informatique et en Automatique (Inria)-Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB)-Institut National de la Recherche Agronomique (INRA), Unité de Recherches Forestières Méditerranéennes (URFM), Institut National de la Recherche Agronomique (INRA), Agence Nationale de la Recherche, Grant/Award Number: ANR-10-LABX-0025, Institut National de la Recherche Agronomique, Grant/Award Number: Bibliotheque du vivant, Speed@ ID and projet innovant EFPA-INRA., ANR-10-LABX-0025,CEBA,CEnter of the study of Biodiversity in Amazonia(2010), Université de Bordeaux (UB)-Institut National de la Recherche Agronomique (INRA), Université de Bordeaux (UB)-Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB)-Institut National de la Recherche Agronomique (INRA)-Inria Bordeaux - Sud-Ouest, Institut National de Recherche en Informatique et en Automatique (Inria)-Institut National de Recherche en Informatique et en Automatique (Inria)-Laboratoire Bordelais de Recherche en Informatique (LaBRI), Centre National de la Recherche Scientifique (CNRS)-École Nationale Supérieure d'Électronique, Informatique et Radiocommunications de Bordeaux (ENSEIRB)-Université Sciences et Technologies - Bordeaux 1-Université Bordeaux Segalen - Bordeaux 2-Centre National de la Recherche Scientifique (CNRS)-École Nationale Supérieure d'Électronique, Informatique et Radiocommunications de Bordeaux (ENSEIRB)-Université Sciences et Technologies - Bordeaux 1-Université Bordeaux Segalen - Bordeaux 2, Ecologie des Forêts Méditerranéennes [Avignon] (URFM 629), ANR-10-LABX-25-01/10-LABX-0025,CEBA,CEnter of the study of Biodiversity in Amazonia(2010), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)-Institut National de la Recherche Agronomique (INRA)-Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Institut de Recherche pour le Développement (IRD [France-Sud]), Université de Bordeaux (UB)-Centre National de la Recherche Scientifique (CNRS)-École Nationale Supérieure d'Électronique, Informatique et Radiocommunications de Bordeaux (ENSEIRB)-Université de Bordeaux (UB)-Centre National de la Recherche Scientifique (CNRS)-École Nationale Supérieure d'Électronique, Informatique et Radiocommunications de Bordeaux (ENSEIRB)-Inria Bordeaux - Sud-Ouest, ANR ‐10 ‐LABX ‐25 ‐01, Université des Antilles (UA)-Université de Guyane (UG)-Centre National de la Recherche Scientifique (CNRS)-AgroParisTech-Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad)-Institut National de la Recherche Agronomique (INRA), Centre National de la Recherche Scientifique (CNRS)-École Nationale Supérieure d'Électronique, Informatique et Radiocommunications de Bordeaux (ENSEIRB)-Université Sciences et Technologies - Bordeaux 1-Université Bordeaux Segalen - Bordeaux 2-Centre National de la Recherche Scientifique (CNRS)-École Nationale Supérieure d'Électronique, Informatique et Radiocommunications de Bordeaux (ENSEIRB)-Université Sciences et Technologies - Bordeaux 1-Université Bordeaux Segalen - Bordeaux 2-Inria Bordeaux - Sud-Ouest, Université de Bordeaux (UB)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université de Bordeaux (UB)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), and Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB)-Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB)

Subjects
0106 biological sciences, Tropical trees, codage, Introgression, [SDV.BID.SPT]Life Sciences [q-bio]/Biodiversity/Systematics, Phylogenetics and taxonomy, 01 natural sciences, DNA barcoding, Genetic diversity, Coalescent theory, chloroplast DNA, hybridization, Original Research, Species diversity, arbre tropical, 0303 health sciences, Vegetal Biology, chloroplaste, Ecology, adn, genetic diversity, [SDV.BV.BOT]Life Sciences [q-bio]/Vegetal Biology/Botanics, French Guiana, Chloroplast DNA, guyane, diversité génétique, introgression, Biology, 010603 evolutionary biology, Intraspecific competition, 03 medical and health sciences, [SDV.EE.ECO]Life Sciences [q-bio]/Ecology, environment/Ecosystems, incomplete lineage sorting, species diversity, tropical trees, Hybridization, Ecology, Evolution, Behavior and Systematics, 030304 developmental biology, Nature and Landscape Conservation, Interspecific competition, 15. Life on land, Incomplete lineage sorting, Evolutionary biology, [SDE.BE]Environmental Sciences/Biodiversity and Ecology, hybridation, Biologie végétale
Abstract

Sequences have been deposited in the R-SYST databank (https://github.com/r-syst/databases/tree/master/r-syst::plants) and NCBI under accession number KX247940–KX249593. The list of studied species, including genus size, seed dispersal syndrome and cpDNA variation, is available at https://github.com/r-syst/databases/tree/master/r-syst::plants. In this list, chloroplast haplotype designation is consistent within each genus.; International audience; We investigate chloroplast DNA variation in a hyperdiverse community of tropical rainforest trees in French Guiana, focusing on patterns of intraspecific and interspecific variation. We test whether a species genetic diversity is higher when it has congeners in the community with which it can exchange genes and if shared haplotypes are more frequent in genetically diverse species, as expected in the presence of introgression. We sampled a total of 1,681 individual trees from 472 species corresponding to 198 genera and sequenced them at a noncoding chloroplast DNA fragment. Polymorphism was more frequent in species that have congeneric species in the study site than in those without congeners (30% vs. 12%). Moreover, more chloroplast haplotypes were shared with congeners in polymorphic species than in monomorphic ones (44% vs. 28%). Despite large heterogeneities caused by genus-specific behaviors in patterns of hybridization, these results suggest that the higher polymorphism in the presence of congeners is caused by local introgression rather than by incomplete lineage sorting. Our findings suggest that introgression has the potential to drive intraspecific genetic diversity in species-rich tropical forests.

Published
2019
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4. R-Syst::diatom: an open-access and curated barcode database for diatoms and freshwater monitoring

Author

François Keck, Philippe Chaumeil, Alain Franc, Valentin Vasselon, Lenaïg Kermarrec, Frédéric Rimet, Maria Kahlert, Agnès Bouchez, Centre Alpin de Recherche sur les Réseaux Trophiques et Ecosystèmes Limniques (CARRTEL), Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])-Institut National de la Recherche Agronomique (INRA), Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), from patterns to models in computational biodiversity and biotechnology (PLEIADE), Laboratoire Bordelais de Recherche en Informatique (LaBRI), Centre National de la Recherche Scientifique (CNRS)-École Nationale Supérieure d'Électronique, Informatique et Radiocommunications de Bordeaux (ENSEIRB)-Université Sciences et Technologies - Bordeaux 1-Université Bordeaux Segalen - Bordeaux 2-Centre National de la Recherche Scientifique (CNRS)-École Nationale Supérieure d'Électronique, Informatique et Radiocommunications de Bordeaux (ENSEIRB)-Université Sciences et Technologies - Bordeaux 1-Université Bordeaux Segalen - Bordeaux 2-Inria Bordeaux - Sud-Ouest, Institut National de Recherche en Informatique et en Automatique (Inria)-Institut National de Recherche en Informatique et en Automatique (Inria)-Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB)-Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), Asconit Consultants, Swedish University of Agricultural Sciences (SLU), Institut National de la Recherche Agronomique (INRA)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry]), Université de Bordeaux (UB)-École Nationale Supérieure d'Électronique, Informatique et Radiocommunications de Bordeaux (ENSEIRB)-Centre National de la Recherche Scientifique (CNRS)-Université de Bordeaux (UB)-École Nationale Supérieure d'Électronique, Informatique et Radiocommunications de Bordeaux (ENSEIRB)-Centre National de la Recherche Scientifique (CNRS)-Inria Bordeaux - Sud-Ouest, Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB)-Institut National de la Recherche Agronomique (INRA), and Université de Bordeaux (UB)-Centre National de la Recherche Scientifique (CNRS)-École Nationale Supérieure d'Électronique, Informatique et Radiocommunications de Bordeaux (ENSEIRB)-Université de Bordeaux (UB)-Centre National de la Recherche Scientifique (CNRS)-École Nationale Supérieure d'Électronique, Informatique et Radiocommunications de Bordeaux (ENSEIRB)-Inria Bordeaux - Sud-Ouest

Subjects
0106 biological sciences, code barre adn, taxonomie, Sequencing data, Statistics as Topic, pollution de l'eau, Fresh Water, Biology, Barcode, computer.software_genre, 010603 evolutionary biology, 01 natural sciences, DNA barcoding, General Biochemistry, Genetics and Molecular Biology, 18S ribosomal RNA, law.invention, Access to Information, diatomées, law, Databases, Genetic, séquençage, DNA Barcoding, Taxonomic, Data Curation, Phylogeny, 2. Zero hunger, Diatoms, Database, Data curation, Base Sequence, 010604 marine biology & hydrobiology, biology.organism_classification, Freshwater algae, Diatom, Phenotype, Identification (biology), Original Article, [INFO.INFO-BI]Computer Science [cs]/Bioinformatics [q-bio.QM], General Agricultural and Biological Sciences, computer, algorithme, Information Systems, Environmental Monitoring
Abstract

Diatoms are micro-algal indicators of freshwater pollution. Current standardized methodologies are based on microscopic determinations, which is time consuming and prone to identification uncertainties. The use of DNA-barcoding has been proposed as a way to avoid these flaws. Combining barcoding with next-generation sequencing enables collection of a large quantity of barcodes from natural samples. These barcodes are identified as certain diatom taxa by comparing the sequences to a reference barcoding library using algorithms. Proof of concept was recently demonstrated for synthetic and natural communities and underlined the importance of the quality of this reference library. We present an open-access and curated reference barcoding database for diatoms, called R-Syst::diatom, developed in the framework of R-Syst, the network of systematic supported by INRA (French National Institute for Agricultural Research), see http://www.rsyst.inra.fr/en. R-Syst::diatom links DNA-barcodes to their taxonomical identifications, and is dedicated to identify barcodes from natural samples. The data come from two sources, a culture collection of freshwater algae maintained in INRA in which new strains are regularly deposited and barcoded and from the NCBI (National Center for Biotechnology Information) nucleotide database. Two kinds of barcodes were chosen to support the database: 18S (18S ribosomal RNA) and rbcL (Ribulose-1,5-bisphosphate carboxylase/oxygenase), because of their efficiency. Data are curated using innovative (Declic) and classical bioinformatic tools (Blast, classical phylogenies) and up-to-date taxonomy (Catalogues and peer reviewed papers). Every 6 months R-Syst::diatom is updated. The database is available through the R-Syst microalgae website (http://www.rsyst.inra.fr/) and a platform dedicated to next-generation sequencing data analysis, virtual_BiodiversityL@b (https://galaxy-pgtp.pierroton.inra.fr/). We present here the content of the library regarding the number of barcodes and diatom taxa. In addition to these information, morphological features (e.g. biovolumes, chloroplasts…), life-forms (mobility, colony-type) or ecological features (taxa preferenda to pollution) are indicated in R-Syst::diatom.[br/] Database URL: http://www.rsyst.inra.fr/

Published
2016
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5. Proteomic plasticity of two Eucalyptus genotypes under contrasted water regimes in the field

Author

Jean-William Dupuy, Aurélien Barré, Emilie Villar, André Mabialangoma, Anne Marie Lomenech, Jean-Marc Gion, Delphine Vincent, Philippe Chaumeil, Frank Bedon, and Christophe Plomion

Subjects
2. Zero hunger, 0106 biological sciences, Gel electrophoresis, 0303 health sciences, Physiology, Plant Science, 15. Life on land, Biology, 01 natural sciences, Eucalyptus, Phenotype, 03 medical and health sciences, Genotype, Botany, Proteome, Osmoregulation, Adaptation, Cell wall modification, 030304 developmental biology, 010606 plant biology & botany
Abstract

Water deficit affects tree growth and limits wood production. In an attempt to identify the molecular triggers of adaptation mechanisms to water deficit in Eucalyptus, we investigated protein expression patterns of two ecophysiologically contrasted Eucalyptus genotypes. They were grown in the field in either natural conditions or irrigated for 7 weeks during the dry season in the Republic of Congo. At the phenotypic level, genotype (G), treatment (T) and/or G ¥ T interaction effects were observed for aboveand below-ground biomass-related traits. At the molecular level, changes in protein abundance were recorded in leaves (acidic pH 4-7, and basic pH 7-11, proteomes) and stems (acidic proteome) using two-dimensional gel electrophoresis (2-DE). One third of the detected protein spots displayed significantG,T and/orG ¥ T effects, and 158 of them were identified by tandem mass spectrometry (LC-MS/MS) analysis. Thus, several proteins whose molecular plasticity was genetically controlled (i.e. G ¥ T effect) were revealed, highlighting adaptive mechanisms to water deficit specific to each genotype, namely cell wall modification, cell detoxification and osmoregulation. Transcript abundances corresponding to G ¥ T proteins were also investigated by quantitative RT-PCR. These proteins represent relevant targets to improve drought resistance in this ecologically and economically important forest tree genus. (Resume d'auteur)

Published
2011
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6. A micromethod for high throughput RNA extraction in forest trees

Author

Raúl Herrera, Franck Salin, Jorge A. P. Paiva, Christophe Plomion, Philippe Chaumeil, Grégoire Le Provost, Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), and Universidad de Talca

Subjects
0106 biological sciences, arbre forestier, DNA, Complementary, pinus, pin, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Biology, Bioinformatics, 01 natural sciences, General Biochemistry, Genetics and Molecular Biology, Trees, 03 medical and health sciences, chemistry.chemical_compound, Gene Expression Regulation, Plant, Gene expression, microméthode, lcsh:QH301-705.5, ComputingMilieux_MISCELLANEOUS, 030304 developmental biology, Regulation of gene expression, 0303 health sciences, Chromatography, Reverse Transcriptase Polymerase Chain Reaction, Extraction (chemistry), Reproducibility of Results, RNA, CDNA Library Construction, General Medicine, 15. Life on land, Microarray Analysis, RNA extraction, Reverse transcription polymerase chain reaction, micromethod, arn t, Genetic Techniques, chemistry, lcsh:Biology (General), RNA, Plant, Cetrimonium Compounds, extraction, General Agricultural and Biological Sciences, DNA, forest trees, 010606 plant biology & botany
Abstract

A large quantity of high quality RNA is often required in the analysis of gene expression. However, RNA extraction from samples taken from woody plants is generally complex, and represents the main limitation to study gene expression, particularly in refractory species like conifers. Standard RNA extraction protocols are available but they are highly time consuming, and not adapted to large scale extraction. Here we present a high-throughput RNA extraction protocol. This protocol was adapted to a micro-scale by modifying the classical cetyltrimethylammonium (CTAB) protocol developed for pine: (i) quantity of material used (100-200 mg of sample), (ii) disruption of samples in microtube using a mechanical tissue disrupter, and (iii) the use of SSTE buffer. One hundred samples of woody plant tissues/organs can be easily treated in two working days. An average of 15 \ig of high quality RNA per sample was obtained. The RNA extracted is suitable for applications such as real time reverse transcription polymerase chain reaction, cDNA library construction or synthesis of complex targets for microarray analysis.

Published
2007

7. EucaMaps: linking genetic maps and associated QTLs to the Eucalyptus grandis genome

Author

Christophe Plomion, Jean-Marc Gion, Philippe Chaumeil, Biodiversité, Gènes & Communautés (BioGeCo), and Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB)

Subjects
Eucalyptus grandis, Locus des caractères quantitatifs, QTL, Myrtaceae, [SDV]Life Sciences [q-bio], Sélection, Horticulture, Quantitative trait locus, Biology, Logiciel, Genome, F30 - Génétique et amélioration des plantes, Bioinformatique, Genetic linkage, corymbia, Botany, Genetics, Molecular Biology, web-based application, genome, Abiotic component, Whole genome sequencing, Internet, Corymbia, Génome, comparative mapping, Forestry, 15. Life on land, biology.organism_classification, Eucalyptus, Genetic architecture, C30 - Documentation et information, eucalyptus, Evolutionary biology, Gène, Carte génétique, U30 - Méthodes de recherche
Abstract

International audience; EucaMaps is a web-based application that provides genetic linkage maps of Eucalyptus and Corymbia species, their orthologous relationships and the position of 267 published QTLs, hyperlinked to the Eucalyptus grandis genome sequence of BRASUZ1. EucaMaps makes an ideal springboard to relate results on the genetic architecture of several quantitative traits with genomic information using the circular vizualization tool of Circos. Five classes of QTLs (growth, wood quality, tolerance to biotic and abiotic stresses, leaf chemicals, rooting ability) are presented in this web-based application providing a broad overview of major QTL regions in the seven species considered.

Published
2015
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8. Cross-species transferability and mapping of genomic and cDNA SSRs in pines

Author

Christophe Plomion, Virginie Garcia, Philippe Chaumeil, Carmen Collada, A. Guevara, Giovanni G. Vendramin, David Chagné, A. Ramboer, J-M. Frigerio, María-Teresa Cervera, Craig S. Echt, T. E. Richardson, Biodiversité, Gènes et Ecosystèmes (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université Sciences et Technologies - Bordeaux 1, Universidad Politécnica de Madrid (UPM), Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria = National Institute for Agricultural and Food Research and Technology (INIA), Istituto di Genetica Vegetale, Sezione di Firenze, Consiglio Nazionale delle Ricerche [Roma] (CNR), Station de physiologie végétale, Institut National de la Recherche Agronomique (INRA), and New Zealand Forest Research Institute

Subjects
Genetic Markers, 0106 biological sciences, DNA, Complementary, DNA, Plant, PINUS TAEDA, Radiata, UniGene, Genomics, Biology, Polymerase Chain Reaction, PINUS RADIATA, 01 natural sciences, microsatellites, Loss of heterozygosity, 03 medical and health sciences, c-DNA, Trinucleotide Repeats, Gene mapping, Complementary DNA, Genetics, Crosses, Genetic, DNA Primers, Repetitive Sequences, Nucleic Acid, 030304 developmental biology, [SDV.GEN]Life Sciences [q-bio]/Genetics, 0303 health sciences, Base Sequence, PINUS CANARIENSIS, transferability, Chromosome Mapping, PIN MARITIME, food and beverages, General Medicine, SIMPLE SEQUENCE REPEATS, Pinus, biology.organism_classification, genetic variation, Microsatellite, Pinus pinaster, Agronomy and Crop Science, Genome, Plant, Microsatellite Repeats, 010606 plant biology & botany, Biotechnology
Abstract

International audience; Two unigene datasets of Pinus taeda and Pinus pinaster were screened to detect di-, tri- and tetranucleotide repeated motifs using the SSRIT script. A total of 419 simple sequence repeats (SSRs) were identified, from which only 12.8% overlapped between the two sets. The position of the SSRs within their coding sequences were predicted using FrameD. Trinucleotides appeared to be the most abundant repeated motif (63 and 51% in P. taeda and P. pinaster, respectively) and tended to be found within translated regions (76% in both species), whereas dinucleotide repeats were preferentially found within the 5'- and 3'-untranslated regions (75 and 65%, respectively). Fifty-three primer pairs amplifying a single PCR fragment in the source species (mainly P. taeda), were tested for amplification in six other pine species. The amplification rate with other pine species was high and corresponded with the phylogenetic distance between species, varying from 64.6% in P. canariensis to 94.2% in P. radiata. Genomic SSRs were found to be less transferable; 58 of the 107 primer pairs (i.e., 54%) derived from P. radiata amplified a single fragment in P. pinaster. Nine cDNA-SSRs were located to their chromosomes in two P. pinaster linkage maps. The level of polymorphism of these cDNA-SSRs was compared to that of previously and newly developed genomic-SSRs. Overall, genomic SSRs tend to perform better in terms of heterozygosity and number of alleles. This study suggests that useful SSR markers can be developed from pine ESTs

Published
2004
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9. Differential accumulation of soluble proteins in roots of metallicolous and nonmetallicolous populations of Agrostis capillaris L. exposed to Cu

Author

Jean-William Dupuy, Philippe Chaumeil, Michel Mench, Frank Bedon, Elena Hego, Christophe Plomion, Marc Bonneu, Clémence M. Bes, Stéphane Claverol, Aurélien Barré, Patricia M. Palagi, Céline Lalanne, Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), Swiss Institute of Bioinformatics [Lausanne] (SIB), Université de Lausanne (UNIL), Université Sciences et Technologies - Bordeaux 1, and Centre Génomique Fonctionnelle, Plateforme Protéome

Subjects
Proteomics, 0106 biological sciences, Proteome, [SDV]Life Sciences [q-bio], agrostis, Fructose-bisphosphate aldolase, Cu-tolerance, Plant Roots, 01 natural sciences, Biochemistry, Superoxide dismutase, 03 medical and health sciences, chemistry.chemical_compound, Agrostis, Electrophoresis, Gel, Two-Dimensional, Nicotianamine, Trichloroacetic acid, Molecular Biology, Plant Proteins, 030304 developmental biology, Agrostis capillaris, 2. Zero hunger, 0303 health sciences, biology, Superoxide, Methylglyoxal, plant proteomics, biology.organism_classification, Adaptation, Physiological, superoxide dismutase, Oxidative Stress, Solubility, chemistry, biology.protein, Copper, 010606 plant biology & botany
Abstract

International audience; Differential expression of soluble proteins was explored in roots of metallicolous (M) and non-M (NM) plants of Agrostis capillaris L. exposed to increasing Cu to partially identify molecular mechanisms underlying higher Cu tolerance in M plants. Plants were cultivated for 2 months on perlite with a CuSO4 (1–30 μM) spiked-nutrient solution. Soluble proteins extracted by the trichloroacetic acid/acetone procedure were separated with 2DE (linear 4–7 pH gradient). After Coomassie Blue staining and image analysis, 19 proteins differentially expressed were identified using LC-MS/MS and Expressed Sequence Tag (ESTs) databases. At supra-optimal Cu exposure (15–30 μM), glycolysis was likely altered in NM roots with increased production of glycerone-P and methylglyoxal based on overexpression of triosephosphate isomerase and fructose bisphosphate aldolase. Changes in tubulins and higher expressions of 5-methyltetrahydropteroyltriglutamatehomocysteine methyltransferase and S-adenosylmethionine synthase underpinned impacts on the cytoskeleton and stimulation of ethylene metabolism. Increased l-methionine and S-adenosylmethionine amounts may also facilitate production of nicotianamine, which complexes Cu, and of l-cysteine, needed for metallothioneins and GSH. In M roots, the increase of [Cu/Zn] superoxide dismutase suggested a better detoxification of superoxide, when Cu exposure rose. Higher Cu-tolerance of M plants would rather result from simultaneous cooperation of various processes than from a specific mechanism.

Published
2014
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10. A next-generation sequencing approach to river biomonitoring using benthic diatoms

Author

Jean-Marc Frigerio, Philippe Chaumeil, Frédéric Rimet, Lenaïg Kermarrec, Alain Franc, Jean-François Humbert, Agnès Bouchez, Asconit Consultants, Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), Centre Alpin de Recherche sur les Réseaux Trophiques et Ecosystèmes Limniques (CARRTEL), Institut National de la Recherche Agronomique (INRA)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry]), Institut d'Ecologie et des Sciences de l'Environnement de Paris, Institut National de la Recherche Agronomique (INRA)-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut de Recherche pour le Développement (IRD [France-Ouest])-Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12)-Centre National de la Recherche Scientifique (CNRS)-Ecole Normale Supérieure Paris-Saclay (ENS Paris Saclay), Ecology of Forests, Grassland and Freshwaters Division of Institut national de la recherche agronomique (INRA), Plant Health and Environment Division of INRA, French National Agency for Water and Aquatic Environments (ONEMA), and French National Agency for Research and Technology (ANRT)

Subjects
0106 biological sciences, 0303 health sciences, Ecology, [SDV]Life Sciences [q-bio], fungi, 15. Life on land, Aquatic Science, Biology, Ribosomal RNA, biology.organism_classification, 010603 evolutionary biology, 01 natural sciences, DNA sequencing, Molecular ecology, 03 medical and health sciences, Taxon, Diatom, Evolutionary biology, Pyrosequencing, Identification (biology), Bioindicator, Ecology, Evolution, Behavior and Systematics, 030304 developmental biology
Abstract

International audience; Diatoms are main bioindicators used to assess the ecological quality of rivers, but their identification is difficult and time-consuming. Next Generation Sequencing (NGS) can be used to study communities of microorganisms, so we carried out a test of the reliability of 454 pyrosequencing for estimating diatom inventories in environmental samples. We used small subunit ribosomal deoxyribonucleic acid (SSU rDNA), ribulose-1, 5-bisphosphate carboxylase (rbcL), and cytochrome oxidase I (COI) markers and examined reference libraries to define thresholds between the intra- and interspecific and intra- and intergeneric genetic distances. Based on tests of 1 mock community, we used a threshold of 99% identity for SSU rDNA and rbcL sequences to study freshwater diatoms at the species level. We applied 454 pyrosequencing to 4 contrasting environmental samples (with one in duplicate), assigned taxon names to environmental sequences, and compared the qualitative and quantitative molecular inventories to those obtained by microscopy. Species richness detected by microscopy was always higher than that detected by pyrosequencing. Some morphologically detected taxa may have been persistent frustules from dead cells. Some taxa detected by molecular analysis were not detected by morphology and vice versa. The main source of divergence appears to be inadequate taxonomic coverage in DNA reference libraries. Only a small percentage of species (but almost all genera) in morphological inventories were included in DNA reference libraries. DNA reference libraries contained a smaller percentage of species from tropical (27.1–38.1%) than from temperate samples (53.7–77.8%). Agreement between morphological and molecular inventories was better for species with relative abundance >1% than for rare species. The rbcL marker appeared to provide more reproducible results (94.9% species similarity between the 2 duplicates) and was very useful for molecular identification, but procedural standardization is needed. The water-quality ranking assigned to a site via the Pollution Sensitivity diatom index was the same whether calculated with molecular or morphological data. Pyrosequencing is a promising approach for detecting all species, even rare ones, once reference libraries have been developed.

Published
2014
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11. R-SYST network : a French consortium for barcoding and species identification

Author

Agnès Bouchez, Marc Buee, Philippe Chaumeil, Valerie Laval, Marion Le Saux, Marie-Anne Auger-Rozenberg, Alain Franc, Centre Alpin de Recherche sur les Réseaux Trophiques et Ecosystèmes Limniques (CARRTEL), Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])-Institut National de la Recherche Agronomique (INRA), Interactions Arbres-Microorganismes (IAM), Université de Lorraine (UL)-Institut National de la Recherche Agronomique (INRA), Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), BIOlogie et GEstion des Risques en agriculture (BIOGER), Institut National de la Recherche Agronomique (INRA)-AgroParisTech, Institut de Recherche en Horticulture et Semences (IRHS), AGROCAMPUS OUEST-Institut National de la Recherche Agronomique (INRA)-Université d'Angers (UA), Unité de recherche Zoologie Forestière (UZF), Institut National de la Recherche Agronomique (INRA), AgroParisTech-Institut National de la Recherche Agronomique (INRA), Institut National de la Recherche Agronomique (INRA)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry]), Institut National de la Recherche Agronomique (INRA)-Université de Lorraine (UL), Université d'Angers (UA)-Institut National de la Recherche Agronomique (INRA)-AGROCAMPUS OUEST, Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), and Unité de recherche Zoologie Forestière (URZF)

Subjects
[SDE.MCG]Environmental Sciences/Global Changes, graphe clustering, phylogeography, plant associated bacteria, barcoding, diatom, tree, network, taxonomic classification, sequence database, insect, fungi, [SDE.BE]Environmental Sciences/Biodiversity and Ecology, systematics
Abstract

absent

Published
2013

12. Proteomic characterization of phenotypic plasticity between metallicolous and non-metallicolous populations of Agrostis capillaris L. exposed to Cu

Author

Elena Hego, Clémence Bes, Franck Bedon, Michel Mench, Palagi, P. M., Philippe Chaumeil, Barre, A., Claverol, S., W Dupuy, J., Bonneu, M., Céline Lalanne, christophe plomion, Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), Swiss Institute of Bioinformatics [Lausanne] (SIB), Université de Lausanne (UNIL), Université de Bordeaux Ségalen [Bordeaux 2], International Conference on the Biogeochemistry of Trace Elements (ICOBTE). INT., ProdInra, Migration, and Hasselt University (UHasselt). Hasselt, BEL.

Subjects
Agrostis capillaris, [SDV] Life Sciences [q-bio], tolerance, [SDV]Life Sciences [q-bio], phytoremediation, root, superoxide dismutase, ComputingMilieux_MISCELLANEOUS
Abstract

International audience

Published
2013

13. Next-generation sequencing to inventory taxonomic diversity in eukaryotic communities: a test for freshwater diatoms

Author

Lenaïg Kermarrec, Agnès Bouchez, Jean-François Humbert, Frédéric Rimet, Philippe Chaumeil, Alain Franc, Centre Alpin de Recherche sur les Réseaux Trophiques et Ecosystèmes Limniques (CARRTEL), Institut National de la Recherche Agronomique (INRA)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry]), Asconit Consultants, Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), Biogéochimie et écologie des milieux continentaux (Bioemco), École normale supérieure - Paris (ENS-PSL), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut de Recherche pour le Développement (IRD)-Institut National de la Recherche Agronomique (INRA)-Université Pierre et Marie Curie - Paris 6 (UPMC)-AgroParisTech-Centre National de la Recherche Scientifique (CNRS), Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])-Institut National de la Recherche Agronomique (INRA), Centre National de la Recherche Scientifique (CNRS)-AgroParisTech-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Recherche Agronomique (INRA)-École normale supérieure - Paris (ENS Paris), and Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)

Subjects
0106 biological sciences, benthic diatom diversity, Ribulose-Bisphosphate Carboxylase, species inventories, [SDV]Life Sciences [q-bio], Molecular Sequence Data, Fresh Water, Biology, 01 natural sciences, Genome, DNA, Mitochondrial, DNA, Ribosomal, DNA sequencing, law.invention, Molecular ecology, Electron Transport Complex IV, 03 medical and health sciences, law, environmental communities, Genetics, RNA, Ribosomal, 18S, molecular ecology, Ecology, Evolution, Behavior and Systematics, Polymerase chain reaction, 030304 developmental biology, Diatoms, 0303 health sciences, 010604 marine biology & hydrobiology, Genetic Variation, High-Throughput Nucleotide Sequencing, high-throughput sequencing, biology.organism_classification, DNA extraction, Diatom, Evolutionary biology, Metagenomics, [SDE]Environmental Sciences, Pyrosequencing, Metagenome, [SDE.BE]Environmental Sciences/Biodiversity and Ecology, Biotechnology
Abstract

International audience; The recent emergence of barcoding approaches coupled to those of next-generation sequencing (NGS) has raised new perspectives for studying environmental communities. In this framework, we tested the possibility to derive accurate inventories of diatom communities from pyrosequencing outputs with an available DNA reference library. We used three molecular markers targeting the nuclear, chloroplast and mitochondrial genomes (SSU rDNA, rbcL and cox1) and three samples of a mock community composed of 30 known diatom strains belonging to 21 species. In the goal to detect methodological biases, one sample was constituted directly from pooled cultures, whereas the others consisted of pooled PCR products. The NGS reads obtained by pyrosequencing (Roche 454) were compared first to a DNA reference library including the sequences of all the species used to constitute the mock community, and second to a complete DNA reference library with a larger taxonomic coverage. A stringent taxonomic assignation gave inventories that were compared to the real one. We detected biases due to DNA extraction and PCR amplification that resulted in false-negative detection. Conversely, pyrosequencing errors appeared to generate false positives, especially in case of closely allied species. The taxonomic coverage of DNA reference libraries appears to be the most crucial factor, together with marker polymorphism which is essential to identify taxa at the species level. RbcL offers a high resolving power together with a large DNA reference library. Although needing further optimization, pyrosequencing is suitable for identifying diatom assemblages and may find applications in the field of freshwater biomonitoring.

Published
2013
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14. Mérignac (33), Occupation des berges de la Devèze de l'époque gauloise à l'époque moderne : Rapport de fouille

Author

Béague, Nadine, Jean-Philippe, Chaumeil, Dietsch-Sellami, Marie-France, Xavier, Boës, Institut national de recherches archéologiques préventives (Inrap), Identités, Territoires, Expressions, Mobilités (ITEM), Université de Pau et des Pays de l'Adour (UPPA), and Inrap GSO

Subjects
bois, ossements animaux, [SHS.ARCHEO]Humanities and Social Sciences/Archaeology and Prehistory, chaussée, fouille, aménagement de berge, pieu, hydraulique, xylologie, objet métallique, carpologie, maison, géologie, palynologie, céramique moderne, cours d'eau, rivière
Abstract

La fouille archéologique de l’Îlot 3 de Mérignac a été motivée par la découverte lors du diagnostic de plusieurs types d’aménagements de berges de la Devèze, rivière actuellement canalisée à environ 50 m au sud du site. Le milieu humide de la fouille a permis la conservation de bois échoués en bordure de rivière, d’un système de maintien de berge en pieux d’aulne, et d’une surélévation ou bordure de chaussée en pieux de pin. Plusieurs états du cours d’eau ont été perçus, notamment la berge naturelle du cours de la Devèze avant les modifications d’époque moderne et quelques éléments d’occupation d’époque romaine ont pu être reconnue. La surprise a été, à l’échelle de la fouille d’une superficie de 268 m² de pouvoir mettre en évidence un paléochenal d’une orientation complètement différente de celle du cours actuel de la Devèze. Pour le moment et en raison justement de la petite taille de la fenêtre ouverte sur ce secteur, il est impossible de répondre aux nombreuses questions que soulève la présence de ce cours d’eau inédit. Grâce à la découverte d’éléments de charpente en chêne échoués au fond, il est acquis que ce chenal se comble dans la deuxième moitié du Ier siècle avant J.C. Ce comblement est suivi d’une tentative d’assainissement du secteur sous la forme de fossés d’écoulement, s’accompagnant d’un alignement de pieux en aulne qui permettait le maintien de la berge ou le franchissement des fossés. Ces aménagements devaient sans doute se rattacher à un habitat ou un lieu de culte antique situé à l’emplacement de l’actuelle église Saint-Vincent. Ils sont abandonnés au bout de peu de temps et le lieu est de nouveau investi par l’eau. La période médiévale n’est absolument pas représentée sur la fouille, alors que l’église Saint-Vincent est un lieu très fréquenté dès le Haut-Moyen Age, mais il est vrai que l’édifice religieux est édifié hors de portée des crues de la rivière. L’analyse des multiples prélèvements de pollens réalisés dans les sédiments de bords de berge et dans le colmatage du paléochenal, ainsi que l’étude des graines et des bois permettent de reconstituer le paysage pour les époques où le site est fréquenté et pour lesquelles il y a des indices chronologiques.

Published
2013

15. E-Biothon : Une plate-forme pour accélérer les recherches en biologie, santé et environnement

Author

Nicolas Bard, Sylvie Boin, François Bothorel, Philippe Chaumeil, Philippe Collinet, Michel Daydé, Benjamin Depardon, Frédéric Desprez, Marie Flé, Alain Franc, Jean-Marc Frigerio, Olivier Gascuel, Stéphane Guindon, Jean-François Gibrat, Denis Girou, Pierre-François Lavallée, Vincent LEFORT, Gilles Lesage, Marc Rugeri, Evelyne Ruinet, Christophe Seguin, Sylvie Thérond, Laboratoire de l'Informatique du Parallélisme (LIP), École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de Recherche en Informatique et en Automatique (Inria)-Centre National de la Recherche Scientifique (CNRS), IBM France, IBM France (IBM), IBM-IBM, Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), Institut du développement et des ressources en informatique scientifique (IDRIS), Centre National de la Recherche Scientifique (CNRS), Algorithmes Parallèles et Optimisation (IRIT-APO), Institut de recherche en informatique de Toulouse (IRIT), Université Toulouse Capitole (UT Capitole), Université de Toulouse (UT)-Université de Toulouse (UT)-Université Toulouse - Jean Jaurès (UT2J), Université de Toulouse (UT)-Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS)-Institut National Polytechnique (Toulouse) (Toulouse INP), Université de Toulouse (UT)-Toulouse Mind & Brain Institut (TMBI), Université Toulouse - Jean Jaurès (UT2J), Université de Toulouse (UT)-Université de Toulouse (UT)-Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT)-Université Toulouse Capitole (UT Capitole), Université de Toulouse (UT), Institut National Polytechnique (Toulouse) (Toulouse INP), SysFera [Villeurbanne], Algorithms and Software Architectures for Distributed and HPC Platforms (AVALON), Inria Grenoble - Rhône-Alpes, Institut National de Recherche en Informatique et en Automatique (Inria)-Institut National de Recherche en Informatique et en Automatique (Inria)-Laboratoire de l'Informatique du Parallélisme (LIP), Université de Lyon-Université de Lyon-Institut National de Recherche en Informatique et en Automatique (Inria)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS), Méthodes et Algorithmes pour la Bioinformatique (MAB), Laboratoire d'Informatique de Robotique et de Microélectronique de Montpellier (LIRMM), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Unité Mathématique Informatique et Génome (MIG), Institut National de la Recherche Agronomique (INRA), Groupe d'Interet Scientifique (GIS). FRA., École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), UMR BioGeCo, INRA & Université Bordeaux, Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB)-Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), IDRIS, Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS), Université Toulouse 1 Capitole (UT1)-Université Toulouse - Jean Jaurès (UT2J)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Institut National Polytechnique (Toulouse) (Toulouse INP), Université Fédérale Toulouse Midi-Pyrénées, Université de Lyon-Université de Lyon-Institut National de Recherche en Informatique et en Automatique (Inria)-Centre National de la Recherche Scientifique (CNRS)-École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), Mathématique Informatique et Génome, Institut National de la Recherche Agronomique (INRA)-Institut National de la Recherche Agronomique (INRA), Université Toulouse Le Mirail (Toulouse 2) (UTM), Institut National de Recherche en Informatique et en Automatique (Inria), Université Montpellier 2 - Sciences et Techniques (UM2), Université Toulouse 1 Capitole (UT1), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Université Toulouse - Jean Jaurès (UT2J)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Institut National Polytechnique (Toulouse) (Toulouse INP), Université Fédérale Toulouse Midi-Pyrénées-Université Toulouse 1 Capitole (UT1), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Centre National de la Recherche Scientifique (CNRS)-Université de Lyon-Institut National de Recherche en Informatique et en Automatique (Inria)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-École normale supérieure - Lyon (ENS Lyon), and Université de Lyon-École normale supérieure - Lyon (ENS Lyon)-Centre National de la Recherche Scientifique (CNRS)-Université de Lyon-Université Claude Bernard Lyon 1 (UCBL)

Subjects
taxonomie, [SDV]Life Sciences [q-bio], plateforme informatique, épidémiologie, [INFO]Computer Science [cs], [MATH]Mathematics [math], [INFO.INFO-DC]Computer Science [cs]/Distributed, Parallel, and Cluster Computing [cs.DC], recherche, bioinformatique, biodiversité
Abstract

National audience; E-Biothon : Une plate-forme pour accélérer les recherches en biologie, santé et environnement

Published
2013

16. New Biomonitoring Approaches based on Next Generation Sequencing : a Test for Freshwater Diatom Communities

Author

Lenaïg Kermarrec, Frédéric Rimet, Alain Franc, Philippe Chaumeil, Jean Francois Humbert, Agnès Bouchez, Centre Alpin de Recherche sur les Réseaux Trophiques et Ecosystèmes Limniques (CARRTEL), Institut National de la Recherche Agronomique (INRA)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry]), Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), Biogéochimie et écologie des milieux continentaux (Bioemco), Centre National de la Recherche Scientifique (CNRS)-AgroParisTech-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Recherche Agronomique (INRA)-École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL), Ghent University [Belgium] (UGENT). National Botanic Garden of Belgium, Ghent, BEL., École normale supérieure - Paris (ENS-PSL), and Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut de Recherche pour le Développement (IRD)-Institut National de la Recherche Agronomique (INRA)-Université Pierre et Marie Curie - Paris 6 (UPMC)-AgroParisTech-Centre National de la Recherche Scientifique (CNRS)

Subjects
comparaison de séquences, diatomées, eau douce, évaluation de la qualité, qualité environnementale, [SDV]Life Sciences [q-bio], outil moléculaire, [SDE]Environmental Sciences, marquage génétique, identification moléculaire, ComputingMilieux_MISCELLANEOUS, rivière
Abstract

International audience

Published
2012

17. A pipeline for automatic taxonomic community inventories on data from NGS : an example with freshwater diatoms

Author

Lenaïg Kermarrec, Philippe Chaumeil, Frédéric Rimet, Jean-Marc Frigerio, Valerie Laval, Marc-Henri Lebrun, Agnès Bouchez, Alain Franc, Centre Alpin de Recherche sur les Réseaux Trophiques et Ecosystèmes Limniques (CARRTEL), Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])-Institut National de la Recherche Agronomique (INRA), Biodiversité, Gènes et Communautés, Institut National de la Recherche Agronomique (INRA), BIOlogie et GEstion des Risques en agriculture (BIOGER), AgroParisTech-Institut National de la Recherche Agronomique (INRA), Institut National de la Recherche Agronomique (INRA)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry]), Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), Institut National de la Recherche Agronomique (INRA)-AgroParisTech, and University of Adelaide. Adélaide, AUS.

Subjects
comparaison de séquences, [SDV.SA]Life Sciences [q-bio]/Agricultural sciences, diatomées, eau douce, évaluation de la qualité, qualité environnementale, marquage génétique
Abstract

absent

Published
2011

18. Proteomic plasticity of two Eucalyptus genotypes under contrasted water regimes in the field

Author

Frank, Bedon, Emilie, Villar, Delphine, Vincent, Jean-William, Dupuy, Anne-Marie, Lomenech, André, Mabialangoma, Philippe, Chaumeil, Aurélien, Barré, Christophe, Plomion, and Jean-Marc, Gion

Subjects
Proteomics, Eucalyptus, Genotype, Plant Stems, Proteome, Water, Hydrogen-Ion Concentration, Adaptation, Physiological, Droughts, Plant Leaves, Phenotype, Congo, Gene Expression Regulation, Plant, Stress, Physiological, Electrophoresis, Gel, Two-Dimensional, Biomass, Seasons, Plant Proteins
Abstract

Water deficit affects tree growth and limits wood production. In an attempt to identify the molecular triggers of adaptation mechanisms to water deficit in Eucalyptus, we investigated protein expression patterns of two ecophysiologically contrasted Eucalyptus genotypes. They were grown in the field in either natural conditions or irrigated for 7 weeks during the dry season in the Republic of Congo. At the phenotypic level, genotype (G), treatment (T) and/or G × T interaction effects were observed for above- and below-ground biomass-related traits. At the molecular level, changes in protein abundance were recorded in leaves (acidic pH 4-7, and basic pH 7-11, proteomes) and stems (acidic proteome) using two-dimensional gel electrophoresis (2-DE). One third of the detected protein spots displayed significant G, T and/or G × T effects, and 158 of them were identified by tandem mass spectrometry (LC-MS/MS) analysis. Thus, several proteins whose molecular plasticity was genetically controlled (i.e. G × T effect) were revealed, highlighting adaptive mechanisms to water deficit specific to each genotype, namely cell wall modification, cell detoxification and osmoregulation. Transcript abundances corresponding to G × T proteins were also investigated by quantitative RT-PCR. These proteins represent relevant targets to improve drought resistance in this ecologically and economically important forest tree genus.

Published
2011

19. Development and implementation of a highly-multiplexed SNP array for genetic mapping in maritime pine and comparative mapping with loblolly pine

Author

Juan P. Jaramillo-Correa, Yves Van de Peer, María Teresa Cervera, Jill L. Wegrzyn, Philippe Chaumeil, Christophe Plomion, Diana Zelenika, Jean-Marc Frigerio, David B. Neale, Anne Boland, Pauline Garnier-Géré, Antoine Kremer, Santiago C. González-Martínez, Emilie Chancerel, Andrew J. Eckert, Yao-Cheng Lin, Grégoire Le Provost, Pierre Rouzé, Camille Lepoittevin, Delphine Grivet, Christophe Boury, Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), Center for Plant Systems Biology (PSB Center), Vlaams Instituut voor Biotechnologie [Ghent, Belgique] (VIB), Instituto de Ecologia, Universidad Nacional Autónoma de México (UNAM), University of California [Davis] (UC Davis), University of California, Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria = National Institute for Agricultural and Food Research and Technology (INIA), Department of Forest Ecology and GeneticsForest Research Centre, Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria, Universidad Nacional Autónoma de México = National Autonomous University of Mexico (UNAM), and University of California (UC)

Subjects
0106 biological sciences, PIN A L'ENCENS, PINUS TAEDA, PIN MARITIME, CARTOGRAPHIE GENETIQUE, POLYMORPHISME NUCLÉOTIDIQUE SIMPLE, EXTRACTION D'ADN, 01 natural sciences, TAEDA L, PROTEIN MARKERS, Oligonucleotide Array Sequence Analysis, Expressed Sequence Tags, Genetics, 0303 health sciences, Expressed sequence tag, pinus pinaster, CONSTRUCTION, qtl, Chromosome Mapping, VITIS-VINIFERA L, Tag SNP, Pedigree, SNP genotyping, diversité génétique, SINGLE NUCLEOTIDE POLYMORPHISMS, génotype, Research Article, sélection assistée par marqueurs, Biotechnology, SNP array, Genotype, lcsh:QH426-470, lcsh:Biotechnology, Biology, Quantitative trait locus, Polymorphism, Single Nucleotide, LINKAGE MAP, 03 medical and health sciences, Gene mapping, RAPD, lcsh:TP248.13-248.65, ALGORITHM, Genotyping, 030304 developmental biology, Comparative genomics, [SDV.GEN]Life Sciences [q-bio]/Genetics, PINASTER AIT, Biology and Life Sciences, 15. Life on land, Pinus, lcsh:Genetics, GENOME LENGTH, 010606 plant biology & botany
Abstract

Background Single nucleotide polymorphisms (SNPs) are the most abundant source of genetic variation among individuals of a species. New genotyping technologies allow examining hundreds to thousands of SNPs in a single reaction for a wide range of applications such as genetic diversity analysis, linkage mapping, fine QTL mapping, association studies, marker-assisted or genome-wide selection. In this paper, we evaluated the potential of highly-multiplexed SNP genotyping for genetic mapping in maritime pine (Pinus pinaster Ait.), the main conifer used for commercial plantation in southwestern Europe. Results We designed a custom GoldenGate assay for 1,536 SNPs detected through the resequencing of gene fragments (707 in vitro SNPs/Indels) and from Sanger-derived Expressed Sequenced Tags assembled into a unigene set (829 in silico SNPs/Indels). Offspring from three-generation outbred (G2) and inbred (F2) pedigrees were genotyped. The success rate of the assay was 63.6% and 74.8% for in silico and in vitro SNPs, respectively. A genotyping error rate of 0.4% was further estimated from segregating data of SNPs belonging to the same gene. Overall, 394 SNPs were available for mapping. A total of 287 SNPs were integrated with previously mapped markers in the G2 parental maps, while 179 SNPs were localized on the map generated from the analysis of the F2 progeny. Based on 98 markers segregating in both pedigrees, we were able to generate a consensus map comprising 357 SNPs from 292 different loci. Finally, the analysis of sequence homology between mapped markers and their orthologs in a Pinus taeda linkage map, made it possible to align the 12 linkage groups of both species. Conclusions Our results show that the GoldenGate assay can be used successfully for high-throughput SNP genotyping in maritime pine, a conifer species that has a genome seven times the size of the human genome. This SNP-array will be extended thanks to recent sequencing effort using new generation sequencing technologies and will include SNPs from comparative orthologous sequences that were identified in the present study, providing a wider collection of anchor points for comparative genomics among the conifers.

Published
2011
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20. Efficiency of 3 genetic markers to determine the composition of a diatom assemblage using next generation sequencing

Author

Lenaïg Kermarrec, Frédéric Rimet, Alain Franc, Philippe Chaumeil, Jean Francois Humbert, Agnès Bouchez, Centre Alpin de Recherche sur les Réseaux Trophiques et Ecosystèmes Limniques (CARRTEL), Institut National de la Recherche Agronomique (INRA)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry]), Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), Biogéochimie et écologie des milieux continentaux (Bioemco), Centre National de la Recherche Scientifique (CNRS)-AgroParisTech-Université Pierre et Marie Curie - Paris 6 (UPMC)-Institut National de la Recherche Agronomique (INRA)-École normale supérieure - Paris (ENS Paris), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL), University of Adelaide. Adélaide, AUS., École normale supérieure - Paris (ENS-PSL), and Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Institut de Recherche pour le Développement (IRD)-Institut National de la Recherche Agronomique (INRA)-Université Pierre et Marie Curie - Paris 6 (UPMC)-AgroParisTech-Centre National de la Recherche Scientifique (CNRS)

Subjects
comparaison de séquences, protiste, diatomées, eau douce, évaluation de la qualité, qualité environnementale, [SDV]Life Sciences [q-bio], [SDE]Environmental Sciences, marquage génétique, ComputingMilieux_MISCELLANEOUS
Abstract

International audience

Published
2011

21. Current trends in microsatellite genotyping

Author

Franck Salin, Rémy J. Petit, Thibaut Malausa, Olivier Lepais, Patrick Léger, Stefanie Wagner, Erwan Guichoux, Philippe Chaumeil, Lélia Lagache, Camille Lepoittevin, E. Revardel, Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), Centre de recherche Pernod Ricard, Steinmann Institut, Rheinische Friedrich-Wilhelms-Universität Bonn, School of Biological and Environmental Sciences, University of Stirling, Interactions Biotiques et Santé Végétale, and Institut National de la Recherche Agronomique (INRA)

Subjects
0106 biological sciences, Genetic Markers, Genotype, MULTIPLEXING, Population, Computational biology, Biology, CARTOGRAPHIE GENETIQUE, 010603 evolutionary biology, 01 natural sciences, Polymerase Chain Reaction, 03 medical and health sciences, NEXTGEN SEQUENCING, Multiplex polymerase chain reaction, QUALITY CONTROL, Genetics, medicine, Multiplex, Genetic Testing, education, BINNING, HIGH THROUGHPUT, Genotyping, Ecology, Evolution, Behavior and Systematics, Selection (genetic algorithm), 030304 developmental biology, Genetic testing, [SDV.EE]Life Sciences [q-bio]/Ecology, environment, 0303 health sciences, education.field_of_study, medicine.diagnostic_test, Computational Biology, High-Throughput Nucleotide Sequencing, SSR, Microsatellite, GENETIQUE DES POPULATIONS, CONTRÔLE DE QUALITÉ, Biotechnology, Microsatellite Repeats
Abstract

International audience; Microsatellites have been popular molecular markers ever since their advent in the late eighties. Despite growing competition from new genotyping and sequencing techniques, the use of these versatile and cost-effective markers continues to increase, boosted by successive technical advances. First, methods for multiplexing PCR have considerably improved over the last years, thereby decreasing genotyping costs and increasing throughput. Second, next-generation sequencing technologies allow the identification of large numbers of microsatellite loci at reduced cost in non-model species. As a consequence, more stringent selection of loci is possible, thereby further enhancing multiplex quality and efficiency. However, current practices are lagging behind. By surveying recently published population genetic studies relying on simple sequence repeats, we show that more than half of the studies lack appropriate quality controls and do not make use of multiplex PCR. To make the most of the latest technical developments, we outline the need for a well-established strategy including standardized high-throughput bench protocols and specific bioinformatic tools, from primer design to allele calling.

Published
2011
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22. Interaction between environmental factors affects the accumulation of root proteins in hydroponically grown Eucalyptus globulus (Labill.)

Author

Jean-William Dupuy, Christophe Plomion, Aurélien Barré, Isabel Feito, Jean-Marc Gion, Frank Bedon, Philippe Chaumeil, Anne Marie Lomenech, Juan Majada, Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), SERIDA, Forestry Section, Partenaires INRAE, Université de Bordeaux Ségalen [Bordeaux 2], Centre de Coopération Internationale en Recherche Agronomique pour le Développement (Cirad), INRA, and ERA-NET

Subjects
0106 biological sciences, Proteomics, Identification, Proteome, Physiology, [SDV]Life Sciences [q-bio], F62 - Physiologie végétale - Croissance et développement, Plant Science, Endoplasmic Reticulum, 01 natural sciences, Plant Roots, Polyethylene Glycols, WATER POTENTIAL, Cutting, Hydroponics, Tandem Mass Spectrometry, Rosaniline Dyes, Electrophoresis, Gel, Two-Dimensional, Plant Proteins, Gel electrophoresis, 0303 health sciences, Eucalyptus, Pulp (paper), Agriculture, Adaptation, Physiological, Droughts, Physiologie végétale, Eucalyptus globulus, Shoot, Osmotic shock, F60 - Physiologie et biochimie végétale, Réticulum endoplasmique, Biology, engineering.material, Environment, OSMOTIC STRESS, 03 medical and health sciences, ROOT, Osmotic Pressure, Stress, Physiological, Botany, Genetics, 030304 developmental biology, Endoplasmic reticulum, Water, 15. Life on land, biology.organism_classification, 2-DE, K10 - Production forestière, Stress osmotique, Spain, Électrophorèse, engineering, Protéine végétale, 010606 plant biology & botany, Abscisic Acid, Racine
Abstract

International audience; Eucalyptus globulus (Labill.) is used for pulp and paper production worldwide. In this report we studied changes in protein expression in one osmotically stressed elite clone widely used in industrial plantations in Spain. High molecular weight polyethylene glycol (PEG) was used as an osmoticum in the growing medium. Roots of rooted cuttings were sampled after 3 and 36 h of treatment. Water potential and abscissic acid content were measured in shoot and root apices to characterize the physiological states of the plants. Total soluble proteins from roots were extracted and separated using two-dimensional gel electrophoresis (2-DE). Gels were stained with Coomassie brillant blue for quantitative analysis of protein accumulation. From a total of 406 reproducible spots, 34 were found to be differentially expressed depending on treatment (osmotic versus control condition) and/or stress duration (3 h versus 36 h), and were further characterized by tandem mass spectrometry. Several proteins were reliably identified including adenosine kinase, actin, stress-related proteins as well as proteins associated to cellular processes, among which some residents of the endoplasmic reticulum. This study constitutes the first investigation of the root proteome in this important forest tree genus.

Published
2011
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23. Plasticity of maritime pine (Pinus pinaster Ait.) wood forming tissues during a growing season

Author

G. Le Provost, Jorge A. P. Paiva, Pedro Fevereiro, D. Da Silva-Perez, Philippe Chaumeil, Alexandre Bosc, Pauline Garnier-Géré, Christophe Plomion, José Graça, Sara Santos, José Carlos Rodrigues, Ana Alves, Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), Universidade Nova de Lisboa = NOVA University Lisbon (NOVA), Instituto de Investigação Científica Tropical (IICT), Departamento de Engenharia Florestal, Instituto Superior de Agronomia, Institut Technologique Forêt Cellulose Bois-construction Ameublement (FCBA), Écologie fonctionnelle et physique de l'environnement (EPHYSE), and Institut National de la Recherche Agronomique (INRA)

Subjects
0106 biological sciences, Candidate gene, Physiology, Climate, Rain, Growing season, Plant Science, Biology, 01 natural sciences, Polymerase Chain Reaction, MOLECULAR PLASTICITY, METABOLOME, Transcriptome, 03 medical and health sciences, Cell Wall, Xylem, WOOD FORMATION, Botany, LATE-WOOD, Cluster Analysis, RNA, Messenger, TRANSCRIPTOME, Gene, 030304 developmental biology, Transpiration, 0303 health sciences, Principal Component Analysis, EARLY-WOOD, Gene Expression Profiling, Temperature, PIN MARITIME, Plant Transpiration, 15. Life on land, biology.organism_classification, Pinus, Phenotype, Wood, [SDV.BV.PEP]Life Sciences [q-bio]/Vegetal Biology/Phytopathology and phytopharmacy, PINUS PINASTER, Pinus pinaster, Seasons, 010606 plant biology & botany
Abstract

International audience; The seasonal effect is the most significant external source of variation affecting vascular cambial activity and the development of newly divided cells, and hence wood properties. Here, the effect of edapho-climatic conditions on the phenotypic and molecular plasticity of differentiating secondary xylem during a growing season was investigated. Wood-forming tissues of maritime pine (Pinus pinaster) were collected from the beginning to the end of the growing season in 2003. Data from examination of fibre morphology, Fourier-transform infrared spectroscopy (FTIR), analytical pyrolysis, and gas chromatography/mass spectrometry (GC/MS) were combined to characterize the samples. Strong variation was observed in response to changes in edapho-climatic conditions. A genomic approach was used to identify genes differentially expressed during this growing season. Out of 3512 studied genes, 19% showed a significant seasonal effect. These genes were clustered into five distinct groups, the largest two representing genes over-expressed in the early- or late-wood-forming tissues, respectively. The other three clusters were characterized by responses to specific edapho-climatic conditions. This work provides new insights into the plasticity of the molecular machinery involved in wood formation, and reveals candidate genes potentially responsible for the phenotypic differences found between early- and late-wood.

Published
2008
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24. Using transcriptome analysis to identify osmotic stress candidate genes in maritime pine (Pinus pinaster Ait.)

Author

Jean-Marc Frigerio, Christian Dubos, Philippe Chaumeil, Franck Salin, Garcia, V., Barre, A., Daruvar, A., christophe plomion, Biodiversité, Gènes et Ecosystèmes (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université Sciences et Technologies - Bordeaux 1, Centre de Bioinformatique de Bordeaux (CBIB), CGFB, and ProdInra, Migration

Subjects
[SDV] Life Sciences [q-bio], [SDV]Life Sciences [q-bio], PIN MARITIME, ComputingMilieux_MISCELLANEOUS, OSMOTIC STRESS, CANDIDATE GENES
Abstract

International audience

Published
2002

25. Approches moléculaires de la qualité du bois chez le pin maritime

Author

christophe plomion, Le Provost, G., Jorge Almiro Paiva, Gion, J. M., Céline Lalanne, Pionneau, C., Delphine Madur, Pot, D., Chagné, D., Frigerio, J. M., Garcia, V., Hélène Dumazet, Philippe Chaumeil, Brach, J., Rodrigues, J. C., ProdInra, Migration, Unité de recherches forestières (BORDX PIERR UR ), and Institut National de la Recherche Agronomique (INRA)

Subjects
[SDV] Life Sciences [q-bio], [SDV]Life Sciences [q-bio], PIN MARITIME, BIOLOGIE MOLECULAIRE, ComputingMilieux_MISCELLANEOUS, GENOMIQUE
Abstract

National audience

Published
2002

26. Analysis of expressed sequence tags in maritime pine (Pinus pinaster Aiton)

Author

Grégoire Le Provost, Jean-Marc Frigerio, Philippe Chaumeil, Saneyoshi Ueno, Camille Lepoittevin, Jorge Almiro Paiva, Patrick Léger, Franck Salin, Maite Cervera, Nacer Mohellibi, Francisco Cantón, Christophe Klopp, christophe plomion, Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), Instituto de Biologia Experimental e Tecnológica (IBET), Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria = National Institute for Agricultural and Food Research and Technology (INIA), Université de Bordeaux Ségalen [Bordeaux 2], Universidad de Málaga [Málaga] = University of Málaga [Málaga], Unité de Biométrie et Intelligence Artificielle (UBIA), and Institut National de la Recherche Agronomique (INRA)

Subjects
GENOMIC RESOURCE, PINUS PINASTER, [SDV]Life Sciences [q-bio], PIN MARITIME, EST, [INFO]Computer Science [cs], MARITIME PINE, [MATH]Mathematics [math], RESSOURCE GENOMIQUE, ComputingMilieux_MISCELLANEOUS
Abstract

International audience

27. metaMatch: un algorithme pour l'assignation taxonomique en métagénomique

Author

Jean-Marc Frigerio, Philippe Chaumeil, Pierre Gay, Lenaïg Kermarrec, Frédéric Rimet, Agnès Bouchez, Alain Franc, Biodiversité, Gènes & Communautés (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université de Bordeaux (UB), Mésocentre de Calcul Intensif Aquitain (MCIA), Université Sciences et Technologies - Bordeaux 1-Université de Bordeaux (UB)-Université de Pau et des Pays de l'Adour (UPPA)-Université Victor Segalen - Bordeaux 2, Centre Alpin de Recherche sur les Réseaux Trophiques et Ecosystèmes Limniques (CARRTEL), Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])-Institut National de la Recherche Agronomique (INRA), and Institut National de la Recherche Agronomique (INRA)-Université Savoie Mont Blanc (USMB [Université de Savoie] [Université de Chambéry])

Subjects
[INFO.INFO-DC]Computer Science [cs]/Distributed, Parallel, and Cluster Computing [cs.DC]
Abstract

Community ecology faces a new challenge as the next-generation sequencing approaches can yield data from hundreds of microbial community samples. This way, combined with accurate and reliable taxonomic assessment, yields hundreds of new data that will contribute to a better understanding of community assemblies formed under various environmental and historical conditions. Algorithms classifying sequences by comparison to a reference library are the most widely used tools for assessing community composition of environmental samples. However, as they are computationally intensive, almost all these algorithms (most standard being BLAST and similar offsprings) use heuristics designed to speed up the database exploration phase, at the cost of being less strict with the quality of the match between a query and a reference. This problem is naturally distributable, as all comparisons (query, reference) are independent. Here, we present a tool enabling comparisons between queries ( say, one million reads) and reference sequences (say, several thousands), and its implementation on two infrastructures: a cluster in MCIA (Mésocentre de Calcul Intensif en Aquitaine) and a production grid EGI. We show how tracking the large number of jobs generated was nearly impossible with gLite, and how this problem could be solved using Dirac. We compare time and quality between a run on Avakas and on the grid EGI. As a perspective, we will develop a user friendly interface enabling this tool to be used routinely on the grid as a diagnostic for a user not acquainted with computing subtleties of the grid.

28. Automated SNP detection in expressed sequence tags: statistical considerations and application to maritime pine sequences

Author

Olivier Cantin, Pauline Garnier-Géré, Patrick Léger, Philippe Chaumeil, Jean-Marc Frigerio, Frédéric Laigret, David Pot, Frank Bedon, Christophe Plomion, Virginie Garcia, Loïck Le Dantec, Antoine de Daruvar, David Chagné, Unité de recherches Espèces Fruitières et Vigne (UREFV), Institut National de la Recherche Agronomique (INRA), Centre de Bioinformatique de Bordeaux (CBIB), CGFB, Biodiversité, Gènes et Ecosystèmes (BioGeCo), Institut National de la Recherche Agronomique (INRA)-Université Sciences et Technologies - Bordeaux 1, Station de physiologie végétale, Elion Kivolola, Léa Sylvanie, Laboratoire Bordelais de Recherche en Informatique (LaBRI), and Université de Bordeaux (UB)-Centre National de la Recherche Scientifique (CNRS)-École Nationale Supérieure d'Électronique, Informatique et Radiocommunications de Bordeaux (ENSEIRB)

Subjects
0106 biological sciences, IN SILICO DETECTION, Sequence analysis, [INFO.INFO-OH]Computer Science [cs]/Other [cs.OH], SNP, Single-nucleotide polymorphism, Plant Science, Biology, Polymorphism, Single Nucleotide, 01 natural sciences, 03 medical and health sciences, Gene Frequency, Genetics, Point Mutation, EST, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Phrap, Allele frequency, Alleles, 030304 developmental biology, Expressed Sequence Tags, 0303 health sciences, Expressed sequence tag, Reproducibility of Results, PIN MARITIME, Sequence Analysis, DNA, General Medicine, GENETIQUE, 15. Life on land, Tag SNP, Pinus, SNP genotyping, [INFO.INFO-OH] Computer Science [cs]/Other [cs.OH], ANALYSE, Agronomy and Crop Science, Algorithms, 010606 plant biology & botany
Abstract

International audience; We developed an automated pipeline for the detection of single nucleotide polymorphisms (SNPs) in expressed sequence tag (EST) data sets, by combining three DNA sequence analysis programs:Phred, Phrap and PolyBayes. This application requires access to the individual electrophoregram traces. First, a reference set of 65 SNPs was obtained from the sequencing of 30 gametes in 13 maritime pine (Pinus pinaster Ait.) gene fragments (6671 bp), resulting in a frequency of 1 SNP every 102.6 bp. Second, parameters of the three programs were optimized in order to retrieve as many true SNPs, while keeping the rate of false positive as low as possible. Overall, the efficiency of detection of true SNPs was 83.1%. However, this rate varied largely as a function of the rare SNP allele frequency: down to 41% for rare SNP alleles (frequency ` 10%), up to 98% for allele frequencies above 10%. Third, the detection method was applied to the 18498 assembled maritime pine (Pinus pinaster Ait.) ESTs, allowing to identify a total of 1400 candidate SNPs, in contigs containing between 4 and 20 sequence reads. These genetic resources, described for the first time in a forest tree species, were made available at http://www.pierroton.inra/genetics/Pinesnps. We also derived an analytical expression for the SNP detection probability as a function of the SNP allele frequency, the number of haploid genomes used to generate the EST sequence database, and the sample size of the contigs considered for SNP detection. The frequency of the SNP allele was shown to be the main factor influencing the probability of SNP detection

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