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Cocoa black pod caused by Phytophthora is one of the most serious diseases affecting cocoa production. This project set out was to improve our knowledge of the molecular mechanisms involved in Theobroma cacao resistance. We performed a transcriptional screening using macroarrays containing 89 ESTs from subtractive libraries enriched with genes differentially expressed in response to T. cacao/Phytophthora interactions. A gene expression analysis revealed the induction of genes involved in stress signal transduction, general and specific defence genes and genes involved in hormone signalling pathways. RT-qPCR showed that the level expression of two genes in the resistant plant was higher during the first eight hours after pathogen inoculation. Three genes from a family encoding a protease inhibitor displayed different levels of expression depending on resistant and susceptible T. cacao clones. A comparison of promoter sequences revealed a motif present only in genes that were up-expressed in the resistant plant.

Correspondance: pietro.piffanelli@tecnoparco.org; International audience; Introduction. This protocol applies to the characterization of genome structure and evolution of M. acuminata and M. balbisiana genomes; construction of physical maps around markers of interest and QTLs; comparative genomic studies with other monocot species; and study of banana streak virus (BSV) integration patterns in banana nuclear genomes. The principle, key advantages, starting plant material, time required and expected results are presented. Materials and methods. This part describes the required materials, and the protocols for isolation and lysis of banana nuclei; high-molecular-weight (HMW) DNA Hind III digestion and pulsed-field gel electrophoresis (PFGE) analysis; large-scale HMW DNA Hind III digestion and ligation to the pINDIGOBAC–5 vector; and analysis of recombinant BAC clones. It mentions the main problems which can occur. Results. The described protocols will enable the efficient construction of BAC libraries from Musa species. These libraries can be used to construct physical maps and select specific genomic regions to be sequenced.; Introduction. Le protocole s'applique à la caractérisation de la structure du génome et à l'évolution des génomes de M. acuminata et de M. balbisiana ; à la construction de cartes physiques autour des marqueurs d'intérêt et des QTLs ; aux études de génomique comparative avec d'autre espèce de monocotylédones ; à l’étude des modèles d'intégration du virus BSV dans les génomes nucléaires de la banane. Le principe, les principaux avantages, le matériel végétal utilisé, le temps nécessaire et les résultats attendus sont présentés. Matériel et méthodes. Cette partie décrit le matériel de laboratoire nécessaire, et les protocoles permettant l'isolement et la lyse des noyaux de cellules de bananier ; la digestion Hind III de ADN à haut poids moléculaire (HPM) et l’analyse électrophorèse sur gels à champ pulsé ; la digestion Hind III à grande échelle de ADN à HPM et ligation au vecteur pINDIGOBAC-5 ; l’analyse des clones recombinants de BAC. Elle mentionne les principaux problèmes pouvant se poser. Résultats. Les protocoles présentés permettent de construire des bibliothèques de BAC à partir d'espèces du genre Musa. Ces bibliothèques peuvent être utilisées pour construire des cartes physiques et sélectionner des régions du génome à séquencer.

Background The transition from vegetative to floral state in shoot apical meristems (SAM) is a key event in plant development and is of crucial importance for reproductive success. In perennial plants, this event is recurrent during tree life and subject to both within-tree and between-years heterogeneity. In the present study, our goal was to identify candidate processes involved in the repression or induction of flowering in apical buds of adult apple trees. Results Genes differentially expressed (GDE) were examined between trees artificially set in either ‘ON’ or ‘OFF’ situation, and in which floral induction (FI) was shown to be inhibited or induced in most buds, respectively, using qRT-PCR and microarray analysis. From the period of FI through to flower differentiation, GDE belonged to four main biological processes (i) response to stimuli, including response to oxidative stress; (ii) cellular processes, (iii) cell wall biogenesis, and (iv) metabolic processes including carbohydrate biosynthesis and lipid metabolic process. Several key regulator genes, especially TEMPRANILLO (TEM), FLORAL TRANSITION AT MERISTEM (FTM1) and SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) were found differentially expressed. Moreover, homologs of SPL and Leucine-Rich Repeat proteins were present under QTL zones previously detected for biennial bearing. Conclusions This data set suggests that apical buds of ‘ON’ and ‘OFF’ trees were in different physiological states, resulting from different metabolic, hormonal and redox status which are likely to contribute to FI control in adult apple trees. Investigations on carbohydrate and hormonal fluxes from sources to SAM and on cell detoxification process are expected to further contribute to the identification of the underlying physiological mechanisms of FI in adult apple trees. Electronic supplementary material The online version of this article (doi:10.1186/s12870-016-0739-y) contains supplementary material, which is available to authorized users.

Micropropagation of oil palm by somatic embryogenesis produces a proportion of off-type individuals (approximately 5% overall) displaying a homeotically modified flower structure known as mantled. Transformation of the fertile or sterile androecium into carpel-like structures is observed in staminate and pistillate mantled flowers, respectively, resulting in lower oil yields in affected plantations. Given the epigenetic nature of the mantled condition, a gene expression-based approach was used rather than a genetic one to investigate its molecular basis. Suppression subtractive hybridisation (SSH) and macroarray hybridisation were used to compare transcriptome patterns between normal and mantled inflorescences. Two SSH libraries, enriched for complementary deoxyribonucleic acids (cDNAs) of either true-to-type or somaclonal variant material, were generated. Bioinformatic analysis of these two libraries allowed the identification of 1,350 unique sequences and their annotation by a gene ontology-based approach. Macroarray hybridisation was used to compare gene expression between normal and mantled inflorescences, and 32 genes were found to be differentially expressed. The temporal expression patterns of six genes were further investigated in more detail in relation to male and female inflorescence development. Full-length cDNAs were isolated and characterised for two of these genes, EgFB1 and EgRING1, both of which are down-regulated in the mantled inflorescences and both of which encode proteins associated with proteolytic signalling complexes. Our data shed light on gene expression changes associated with the mantled phenotype and have provided novel transcriptome markers which can help to distinguish the abnormal and wild-type inflorescences.

Aeluropus littoralis (Gouan) Parl. is a C4 perennial halophyte monocotyledonous plant belonging to the same family of wheat. Growing as weed in dry salty areas or marshes, it is a salt-secreting, rhizomatous and is used as forage. It is diploid (2n=2X=14) and has a relative small genome of around 342 Mb. A. littoralis is highly salt-tolerant since this plant has the ability to secrete salt. Thus, A. littoralis has the potential to become an important genetic resource for biotechnological strategies to improve salt and drought tolerance in economically important crops such as wheat. We have constructed SSH (Suppression Subtractive Hybridization) cDNA libraries from root (RSD45) and leaf (LSD45) tissues of 45 days old plants grown in the presence of 300 mM NaC1. We have also constructed full-length cDNA library from 15 days old salt stressed (300 mM NaC1) roots (RSTL15). Sequencing revealed 25 and 42 independent transcripts from the RSD45 and LSD45 cDNA libraries respectively, in both cases this was less than 25% of the clones sequenced. In contrast, 425 (60%) of the clones from the RSTL15 library revealed independent transcripts. After comparison with protein databases using BlastX, 335 (68%) ESTs (Expressed Sequence Tag) were classified into putative known functions and unclassified proteins, 59 (12%) have homology only to unidentified homologous sequences. A total of 98 (20%) of the ESTs have no homologies to known sequences in the protein databases which can be considered as novel. © 2007 Elsevier B.V. All rights reserved.

South American Leaf Blight (SALB), caused by the ascomycete Pseudocercospora ulei, is responsible for the low productivity of rubber trees in Latin America and is a serious threat to rubber plantations in Asia and Africa, where the rubber trees are derived from highly susceptible clones. Three contrasted genotypes were chosen for their levels of resistance to the pathogen: FX2784 (totally resistant), MDF180 (partially resistant) and PB314 (susceptible). Array analyses were previously performed to identify genes differentially expressed in resistant and susceptible genotypes. Twenty-one genes were selected for further gene expression analysis in non-inoculated and inoculated genotypes from 24 to 216 h post infection (hpi). These genes are involved in ROS production (HbRBOHA, HbRBOHB, HbRBOHC, HbRBOHD), ROS-scavenging systems (cytoplasmic and chloroplastic HbCuZnSOD, HbMnSOD, HbCAT, HbAPX1, HbAPX2, HbMDHAR, HbGCL1, HbGCL2, HbOASTL, HbGPX, HbDHAR), and leaf senescence (HbCASP, HbPCYST, HbWRKY2, HbPLY, HbKAT2). First, a genotype-dependent level of expression was observed. The genes HbRBOHA, HbCuZnSOD cyto, HbCAT, HbGCL and HbWRKY2 were constitutively expressed at lower levels in the MDF180 genotype than in the FX2784 and PB314 genotypes. Conversely, the levels of expression of HbDHAR, HbGPX and HbPCYST were higher in the older, non-inoculated leaves of MDF180. Lower production of ROS and efficient regeneration of reduced ascorbate ensure a balanced redox intracellular state in this genotype. Second, inoculation of the leaves induced few modifications in the expression level of the studied genes. In the MDF180 partially resistant genotype, an increase in the expression level of HbRBOHB, HbRBOHD 48 hpi and a decrease in the expression level of HbDHAR 216 hpi were observed. In the FX2784 totally resistant genotype, an increase in the expression level of HbRBOHD and HbCuZnSOD cyto and a decrease in HbCAT were observed 48 hpi. This transitory variation could be associated with the oxidative burst classically observed in hypersensitive response (HR). The increase in the synthesis of reduced glutathione in this genotype could ensure redox balance and consequently cell homeostasis. In the PB314 susceptible genotype, HbROHC, HbCuZnSOD chloro was up-regulated 216 hpi concomitantly with a decrease in the expression level of HbCAT, consequently causing an accumulation of H2O2 and programmed cell death. The level of expression of a transcription factor, HbWRKY2, was also modulated by the P. ulei infection with early transient up-regulation in the FX2784 totally resistant genotype and permanent up-regulation in the MDF180 partially resistant genotype. These results complement studies on genetic determinism of SALB resistance and a recent publication on Hevea glutathione reductase gene.

Les auteurs ont produit et caractérisé la première banque de BAC (chromosome bactériens artificiels) chez une espèce de bananier (#Musa balbisiana# 'Pisang Klutuk Wulung', d'où le nom de la banque PKW). Un protocole nouveau et amélioré d'isolement des noyaux a été employé pour surmonter les problèmes causés par l'abondance de polyphénols et de polysaccharides dans les tissus foliaires. L'emploi de la cytométrie en flux pour purifier les noyaux a éliminé la contamination aux métabolites secondaires ainsi que l'ADN plastidique. De plus, l'utilité d'un vecteur inductible, pCC1BAC, en vue de l'obtention d'une plus grande quantité d'ADN de BAC a été démontrée. La banque PKW compte l'équivalent de neuf génomes du #M. balbisiana# et la taille moyenne des inserts est de 135 kb. Elle est composée de deux sous-banques; la première (la sous-banque SN avec 24 960 clones) ayant été préparée en suivant un protocole standard amélioré pour l'isolement des noyaux tandis que la seconde (sous-banque FN avec 11 904 clones) a été obtenue à partir des noyaux purifiés par cytométrie en flux. Le criblage de la banque avec 12 sondes RFLP, connues comme étant situées sur huit groupes de liaison chez le #Musa acuminata#, a permis d'identifier une moyenne de 11 clones par sonde. Cette observation permet de valider l'estimé quant à la couverture du génome et indique une forte conservation entre les génomes des deux espèces du genre #Musa#. La localisation sur les chromosomes mitotiques de clones BAC choisis au-moyen d'hybridations FISH a permis de montrer que cette banque de clones BAC constitue une ressource utile pour la cartographie cytogénétique. Dans le cadre d'une démarche de clonage positionne] d'un facteur génétique qui est impliqué dans l'activation de séquence pararétrovirales intégrées appartenant au virus de la striure du bananier genre badnavirus (BSV), le locus BSV exprimé (BEL; "BSV expressed locus") a été délimité physiquement. La banque BAC PKW constitue un outil disponible publiquement et est présentement employé pour révéler les mécanismes d'intégration et d'activation de séquences du BSV et pour étudier la structure et l'évolution du génome du bananier.

A bacterial artificial chromosome (BAC) library for banana was constructed from leaves of the wild diploid 'Calcutta 4' clone (Musa acuminata subsp. Burmannicoides 2n = 2 x = 22). 'Calcutta 4' is widely used in breeding programs for its resistance to the current major disease of banana and is being used to build a genetic reference map of banana. As banana leaves are particularly rich in polyphenols and polysaccharides a protocol was adapted to isolate intact nuclei and high-molecular-weight (HMW) DNA. A total of 55,152 clones with an average insert size of 100 kb were picked. The frequency of BAC clones carrying inserts derived from chloroplast and mitochondrial DNA was estimated to be 1.5%. The coverage of the library is equivalent to 9.0-times the haploid genome. The BAC library was screened with 13 RFLP probes belonging to the 8 linkage groups of the consensus molecular map of banana. A total of 135 clones were identified giving an average of 10.38 clones for each locus. This BAC library will be a valuable starting tool for many of the goals of the recently emerged International Musa Genomic Consortium. One of our initial objectives will be to develop a banana physical map by BAC-FISH (fluorescent in situ hybridization) viewing the characterization of translocation break points.

Ostreococcus tauri is a marine photosynthetic picoeukaryote presenting a minimal cellular organization with one nucleus, one chloroplast, and one mitochondrion. It has the smallest genome described among free-living eukaryotic cells, and we showed by pulsed-field gel electrophoresis (PFGE) that it is divided between 15 bands ranging from 1.2 to 0.15 Mb, giving a total size of 9.7 Mb. A Bacterial Artificial Chromosome (BAC) library was prepared from genomic DNA extracted from a culture of O. tauri. A total of 2457 clones was obtained with an average insert size of around 70 kb, representing an 18-fold coverage of the genome. The library was spotted on high density filters, and several probes of coding sequences were hybridized to both the high density BAC library filters and directly to the dried PFGE gels of the O. tauri genomic DNA. These hybridizations allowed a preliminary organization of the library and the localization of several markers on the chromosomes. Randomly selected fragments were also sequenced, representing 12% of the O. tauri genome. Many sequences showed significant similarities in data banks, mainly with plant and algae sequences. About 1000 coding sequences could be identified. These data confirmed the position of O. tauri in the green lineage and the hypothesis of a very compact organization of its genome.

International audience; Theobroma cacao L. is a major cash crop for tropical countries, providing incomes for 14 million small farmers. Establishing sustainable disease resistance and maintaining cocoa qualities are among the major objectives of breeding programs. To enrich the high-density genetic map, useful for all cocoa genetic studies, with gene-based markers, a recently produced large EST resource was mined to develop expressed sequence tag-based simple sequence repeat markers (EST-SSRs) defined in genes with a putative known function. A set of 174 polymorphic EST-SSRs was identified from a selection of 314 non-redundant EST-SSRs with a putative known function. Of them, 115 loci were mapped on the cocoa reference map. This new map contains 582 codominant markers arranged in ten linkage groups corresponding to the haploid number of chromosomes. An average interval between markers of 1.3 cM was found, with approximately one SSR every 2 cM. This new set of EST-SSRs includes 14 candidate genes for plant resistance or cocoa qualities. The percentage of polymorphic SSRs varied depending on the different gene regions from which they originated, with respectively 54%, 69%, and 82% of polymorphic EST-SSRs originating from coding sequences, and from the non-coding untranslated 5'UTR and 3'UTR regions. This new map contains a set of 384 SSR markers that are easily transferable across different mapping populations and useful for all genetic analyses in T. cacao. The new set of EST-SSRs will be a useful tool for studying the functional diversity of populations and for carrying out association mapping studies.

We sequenced and assembled the genome of Theobroma cacao, an economically important tropical fruit tree crop that is the source of chocolate. The assembly corresponds to 76% of the estimated genome size and contains almost all previously described genes, with 82% of them anchored on the 10 T. cacao chromosomes. Analysis of this sequence information highlighted specific expansion of some gene families during evolution, for example flavonoid-related genes. It also provides a major source of candidate genes for T. cacao disease resistance and quality improvement. Based on the inferred paleohistory of the T. cacao genome, we propose an evolutionary scenario whereby the ten T. cacao chromosomes were shaped from an ancestor through eleven chromosome fusions. The T. cacao genome can be considered as a simple living relic of higher plant evolution.

International audience; We sequenced and assembled the draft genome of Theobroma cacao, an economically important tropical-fruit tree crop that is the source of chocolate. This assembly corresponds to 76% of the estimated genome size and contains almost all previously described genes, with 82% of these genes anchored on the 10 T. cacao chromosomes. Analysis of this sequence information highlighted specific expansion of some gene families during evolution, for example, flavonoid-related genes. It also provides a major source of candidate genes for T. cacao improvement. Based on the inferred paleohistory of the T. cacao genome, we propose an evolutionary scenario whereby the ten T. cacao chromosomes were shaped from an ancestor through eleven chromosome fusions.

Little is known about the early green developmental stage of banana fruit before the commercial harvesting stage. In this study, we demonstrate that banana fruit (cv Cavendish) grown under our pedoclimatical conditions undergoes changes in ethylene responsiveness between 40 (immature fruit unable to respond to ethylene), 60 and 90 DAF (days after flowering; early and late mature fruit able to respond to ethylene, respectively). Further, we have combined subtractive suppression hybridization (SSH) and macro-array hybridization to construct four different SSH libraries comprising a total of 3072 clones and identify 876 clones that are differentially expressed during fruit ripening. Some of these positive clones were partially sequenced to generate ESTs. Sequence analysis revealed that 163 clones on 177 (92%) presented a high similarity with different genes in the database and were related to various plant mechanisms. Northern blot analysis and real-time quantitative PCR conducted on 24 selected clones showed that the subtraction worked properly and led to get more insights into the early green developmental stages of banana. These genes will contribute to increase pools of public EST collections of banana for identification of candidate genes and the providing of molecular markers usable to improve banana fruit quality throughout conventional breeding or biotechnology approaches. © 2007 Elsevier Ireland Ltd. All rights reserved.

Background The Lepidoptera Spodoptera frugiperda is a pest which causes widespread economic damage on a variety of crop plants. It is also well known through its famous Sf9 cell line which is used for numerous heterologous protein productions. Species of the Spodoptera genus are used as model for pesticide resistance and to study virus host interactions. A genomic approach is now a critical step for further new developments in biology and pathology of these insects, and the results of ESTs sequencing efforts need to be structured into databases providing an integrated set of tools and informations. Description The ESTs from five independent cDNA libraries, prepared from three different S. frugiperda tissues (hemocytes, midgut and fat body) and from the Sf9 cell line, are deposited in the database. These tissues were chosen because of their importance in biological processes such as immune response, development and plant/insect interaction. So far, the SPODOBASE contains 29,325 ESTs, which are cleaned and clustered into non-redundant sets (2294 clusters and 6103 singletons). The SPODOBASE is constructed in such a way that other ESTs from S. frugiperda or other species may be added. User can retrieve information using text searches, pre-formatted queries, query assistant or blast searches. Annotation is provided against NCBI, UNIPROT or Bombyx mori ESTs databases, and with GO-Slim vocabulary. Conclusion The SPODOBASE database provides integrated access to expressed sequence tags (EST) from the lepidopteran insect Spodoptera frugiperda. It is a publicly available structured database with insect pest sequences which will allow identification of a number of genes and comprehensive cloning of gene families of interest for scientific community. SPODOBASE is available from URL: http://bioweb.ensam.inra.fr/spodobase

Corresponding author: E-mail: thierry.leroy@cirad.fr; International audience; The first bacterial artificial chromosome (BAC) library of Robusta coffee (Coffea canephora) was constructed, with the aim of developing molecular resources to study the genome structure and evolution of this perennial crop. Clone 126, which is highly productiveand confers good technological and organoleptic qualities of beverage, was chosen for development of this library. The BAC library contains 55,296 clones, with an average insert size of 135 Kb per plasmid, therefore representing theoretically nine haploid genome equivalents of C. canephora. Its validation was achieved with a set of 13 genetically anchored single-copy and 4 duplicated RFLP probes and yielded on average 9 BAC clones per probe. Screening of this BAC library was also carried out with partial cDNA probes coding for enzymes of sugar metabolism like invertases and sucrose synthase, with the aim of characterizing the organization and promoter structure of this important class of genes. It was shown that genes for both cell wall and vacuolar forms of invertases were probably unique in the Robusta genome whereas sucrose synthase was encoded by at least two genes. One of them (CcSUS1) was cloned and sequenced, showing that our BAC library is a valuable tool to rapidly identify genes of agronomic interest or linked to cup quality in C. canephora.

A library of 29 482 T-DNA enhancer trap lines has been generated in rice cv. Nipponbare. The regions flanking the T-DNA left border from the first 12 707 primary transformants were systematically isolated by adapter anchor PCR and sequenced. A survey of the 7480 genomic sequences larger than 30 bp (average length 250 bp), representing 56.4% of the total readable sequences and matching the rice bacterial artificial chromosome/ phage artificial chromosome (BAC/PAC) sequences assembled in pseudomolecules allowed the assigning of 6645 (88.8%) T-DNA insertion sites to at least one position in the rice genome of cv. Nipponbare. T-DNA insertions appear to be rather randomly distributed over the 12 rice chromosomes, with a slightly higher insertion frequency in chromosomes 1, 2, 3 and 6. The distribution of 723 independent T-DNA insertions along the chromosome 1 pseudomolecule did not differ significantly from that of the predicted coding sequences in exhibiting a lower insertion density around the centromere region and a higher density in the subtelomeric regions where the gene density is higher. Further establishment of density graphs of T-DNA inserts along the recently released 12 rice pseudomolecules confirmed this non-uniform chromosome distribution. T-DNA appeared less prone to hot spots and cold spots of integration when compared with those revealed by a concurrent assignment of the Tos17 retrotransposon flanking sequences deposited in the National Center for Biotechnology Information (NCBI). T-DNA inserts rarely integrated into repetitive sequences. Based on the predicted gene annotation of chromosome 1, preferential insertion within the first 250 bp from the putative ATG start codon has been observed. Using 4 kb of sequences surrounding the insertion points, 62% of the sequences showed significant similarity to gene encoding known proteins (E-value

We have constructed and validated the first cocoa ( Theobroma cacao L.) BAC library, with the aim of developing molecular resources to study the structure and evolution of the genome of this perennial crop. This library contains 36,864 clones with an average insert size of 120 kb, representing approximately ten haploid genome equivalents. It was constructed from the genotype Scavina-6 (Sca-6), a Forastero clone highly resistant to cocoa pathogens and a parent of existing mapping populations. Validation of the BAC library was carried out with a set of 13 genetically-anchored single copy and one duplicated markers. An average of nine BAC clones per probe was identified, giving an initial experimental estimation of the genome coverage represented in the library. Screening of the library with a set of resistance gene analogues (RGAs), previously mapped in cocoa and co-localizing with QTL for resistance to Phytophthora traits, confirmed at the physical level the tight clustering of RGAs in the cocoa genome and provided the first insights into the relationships between genetic and physical distances in the cocoa genome. This library represents an available BAC resource for structural genomic studies or map-based cloning of genes corresponding to important QTLs for agronomic traits such as resistance genes to major cocoa pathogens like Phytophthora spp ( palmivora and megakarya), Crinipellis perniciosa and Moniliophthora roreri.

This study reports on expression analysis associated with molecular systems biology of cacao- Moniliophthora perniciosa interaction. Gene expression data were obtained for two cacao genotypes (TSH1188, resistant; Catongo, susceptible) challenged or not with the fungus M. perniciosa and collected at three time points through disease. Using expression analysis, we identified 154 and 227 genes that are differentially expressed in TSH1188 and Catongo, respectively. The expression of some of these genes was confirmed by RT-qPCR. Physical protein-protein interaction (PPPI) networks of Arabidopsis thaliana orthologous proteins corresponding to resistant and susceptible interactions were obtained followed by cluster and gene ontology analyses. The integrated analysis of gene expression and systems biology allowed designing a general scheme of major mechanisms associated with witches' broom disease resistance/susceptibility. In this sense, the TSH1188 cultivar shows strong production of ROS and elicitors at the beginning of the interaction with M. perniciosa followed by resistance signal propagation and ROS detoxification. On the other hand, the Catongo genotype displays defense mechanisms that include the synthesis of some defense molecules but without success in regards to elimination of the fungus. This phase is followed by the activation of protein metabolism which is achieved with the production of proteasome associated with autophagy as a precursor mechanism of PCD. This work also identifies candidate genes for further functional studies and for genetic mapping and marker assisted selection.

Background Theobroma cacao L., is a tree originated from the tropical rainforest of South America. It is one of the major cash crops for many tropical countries. T. cacao is mainly produced on smallholdings, providing resources for 14 million farmers. Disease resistance and T. cacao quality improvement are two important challenges for all actors of cocoa and chocolate production. T. cacao is seriously affected by pests and fungal diseases, responsible for more than 40% yield losses and quality improvement, nutritional and organoleptic, is also important for consumers. An international collaboration was formed to develop an EST genomic resource database for cacao. Results Fifty-six cDNA libraries were constructed from different organs, different genotypes and different environmental conditions. A total of 149,650 valid EST sequences were generated corresponding to 48,594 unigenes, 12,692 contigs and 35,902 singletons. A total of 29,849 unigenes shared significant homology with public sequences from other species. Gene Ontology (GO) annotation was applied to distribute the ESTs among the main GO categories. A specific information system (ESTtik) was constructed to process, store and manage this EST collection allowing the user to query a database. To check the representativeness of our EST collection, we looked for the genes known to be involved in two different metabolic pathways extensively studied in other plant species and important for T. cacao qualities: the flavonoid and the terpene pathways. Most of the enzymes described in other crops for these two metabolic pathways were found in our EST collection. A large collection of new genetic markers was provided by this ESTs collection. Conclusion This EST collection displays a good representation of the T. cacao transcriptome, suitable for analysis of biochemical pathways based on oligonucleotide microarrays derived from these ESTs. It will provide numerous genetic markers that will allow the construction of a high density gene map of T. cacao. This EST collection represents a unique and important molecular resource for T. cacao study and improvement, facilitating the discovery of candidate genes for important T. cacao trait variation.