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Made available in DSpace on 2019-10-06T15:42:46Z (GMT). No. of bitstreams: 0 Previous issue date: 2019-04-15 Agropolis Fondation Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Information about population structure and genetic relationships within and among wild and brazilian Coffea arabica L. genotypes is highly relevant to optimize the use of genetic resources for breeding purposes. In this study, we evaluated genetic diversity, clustering analysis based on Jaccard’s coefficient and population structure in 33 genotypes of C. arabica and of three diploid Coffea species (C. canephora, C. eugenioides and C. racemosa) using 30 SSR markers. A total of 206 alleles were identified, with a mean of 6.9 over all loci. The set of SSR markers was able to discriminate all genotypes and revealed that Ethiopian accessions presented higher genetic diversity than commercial varieties. Population structure analysis indicated two genetic groups, one corresponding to Ethiopian accessions and another corresponding predominantly to commercial cultivars. Thirty-four private alleles were detected in the group of accessions collected from West side of Great Rift Valley. We observed a lower average genetic distance of the C. arabica genotypes in relation to C. eugenioides than C. canephora. Interestingly, commercial cultivars were genetically closer to C. eugenioides than C. canephora and C. racemosa. The great allelic richness observed in Ethiopian Arabica coffee, especially in Western group showed that these accessions can be potential source of new alleles to be explored by coffee breeding programs. Instituto Agronômico do Paraná (IAPAR), Rodovia Celso Garcia Cid, 375 Km Centro de Ciências Biológicas Área de Genética e Biologia Molecular Universidade Estadual de Londrina (UEL), CP 10.011 CIRAD UMR AGAP AGAP Univ. Montpellier CIRAD INRA INRIA Montpellier SupAgro Departamento de Agronomia Universidade Estadual de Londrina (UEL), CP 6001 Instituto Agronômico do Paraná (IAPAR) Mestrado em Agricultura Conservacionista, CP 481 Instituto de Biociências de Rio Claro Universidade Estadual Paulista (UNESP) Empresa Brasileira de Pesquisa Agropecuária (EMBRAPA Café) Laboratório de Biotecnologia de Plantas – IAPAR Embrapa Café, Rodovia Celso Garcia Cid, 375 km Instituto de Biociências de Rio Claro Universidade Estadual Paulista (UNESP)

Made available in DSpace on 2018-12-11T17:00:33Z (GMT). No. of bitstreams: 0 Previous issue date: 2016-02-01 Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Studies in diploid parental species of polyploid plants are important to understand their contributions to the formation of plant and species evolution. Coffeaeugenioides is a diploid species that is considered to be an ancestor of allopolyploid Coffeaarabica together with Coffeacanephora. Despite its importance in the evolutionary history of the main economic species of coffee, no study has focused on C. eugenioides molecular genetics. RNA-seq creates the possibility to generate reference transcriptomes and identify coding genes and potential candidates related to important agronomic traits. Therefore, the main objectives were to obtain a global overview of transcriptionally active genes in this species using next-generation sequencing and to analyze specific genes that were highly expressed in leaves and fruits with potential exploratory characteristics for breeding and understanding the evolutionary biology of coffee. A de novo assembly generated 36,935 contigs that were annotated using eight databases. We observed a total of ~5000 differentially expressed genes between leaves and fruits. Several genes exclusively expressed in fruits did not exhibit similarities with sequences in any database. We selected ten differentially expressed unigenes in leaves and fruits to evaluate transcriptional profiles using qPCR. Our study provides the first gene catalog for C. eugenioides and enhances the knowledge concerning the mechanisms involved in the C. arabica homeologous. Furthermore, this work will open new avenues for studies into specific genes and pathways in this species, especially related to fruit, and our data have potential value in assisted breeding applications. Programa de Pós-Graduação em Genética e Biologia Molecular Universidade Estadual de Londrina (UEL) Laboratório de Biotecnologia Vegetal Instituto Agronômico do Paraná (IAPAR) Laboratório de Genômica e Expressão Departamento de Genética Evolução e Bioagentes Instituto de Biologia Universidade Estadual de Campinas (UNICAMP), Campinas Centre de Coopération Internationale en Recherche Agronomique pour le Développement (CIRAD) UMR AGAP, 34398 Embrapa Café Department of Botany Instituto de Biociências de Rio Claro Universidade Estadual Paulista (UNESP) Department of Botany Instituto de Biociências de Rio Claro Universidade Estadual Paulista (UNESP) CNPq: 1002-02 PHEGECO

Made available in DSpace on 2018-11-26T15:46:08Z (GMT). No. of bitstreams: 0 Previous issue date: 2018-01-11 Brazilian Coffee Research Consortium Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) Ciencia sem Fronteiras Brazilian Coffee Research Consortium, INCT Cafe Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Lipids, including the diterpenes cafestol and kahweol, are key compounds that contribute to the quality of coffee beverages. We determined total lipid content and cafestol and kahweol concentrations in green beans and genotyped 107 Coffea arabica accessions, including wild genotypes from the historical FAO collection from Ethiopia. A genome-wide association study was performed to identify genomic regions associated with lipid, cafestol and kahweol contents and cafestol/kahweol ratio. Using the diploid Coffea canephora genome as a reference, we identified 6,696 SNPs. Population structure analyses suggested the presence of two to three groups (K = 2 and K = 3) corresponding to the east and west sides of the Great Rift Valley and an additional group formed by wild accessions collected in western forests. We identified 5 SNPs associated with lipid content, 4 with cafestol, 3 with kahweol and 9 with cafestol/kahweol ratio. Most of these SNPs are located inside or near candidate genes related to metabolic pathways of these chemical compounds in coffee beans. In addition, three trait-associated SNPs showed evidence of directional selection among cultivated and wild coffee accessions. Our results also confirm a great allelic richness in wild accessions from Ethiopia, especially in accessions originating from forests in the west side of the Great Rift Valley. Inst Agron Parana, Lab Biotecnol Vegetal, BR-86047902 Londrina, PR, Brazil CIRAD, UMR AGAP, F-34398 Montpellier, France Empresa Brasileira Pesquisa Agr, BR-0770901 Brasilia, DF, Brazil Univ Estadual Paulista, Inst Biociencias, BR-13506900 Rio Claro, SP, Brazil Univ Montpellier, IRD, CIRAD, IPME, BP 64501, F-34394 Montpellier, France Univ Montpellier, AGAP, CIRAD, INRA,Montpellier SupAgro, Montpellier, France Univ Estadual Paulista, Inst Biociencias, BR-13506900 Rio Claro, SP, Brazil CAPES: 1203-001 CAPES: ANR-10-LABX-0001-01 CAPES: 015/13 Ciencia sem Fronteiras: CAPES PVE 084/13

The management of diversity for conservation and breeding is of great importance for all plant species and is particularly true in perennial species, such as the coffee Coffea canephora. This species exhibits a large genetic and phenotypic diversity with six different diversity groups. Large field collections are available in the Ivory Coast, Uganda and other Asian, American and African countries but are very expensive and time consuming to establish and maintain in large areas. We propose to improve coffee germplasm management through the construction of genetic core collections derived from a set of 565 accessions that are characterized with 13 microsatellite markers. Core collections of 12, 24 and 48 accessions were defined using two methods aimed to maximize the allelic diversity (Maximization strategy) or genetic distance (Maximum- Length Sub-Tree method). A composite core collection of 77 accessions is proposed for both objectives of an optimal management of diversity and breeding. This core collection presents a gene diversity value of 0.8 and exhibits the totality of the major alleles (i.e., 184) that are present in the initial set. The seven proposed core collections constitute a valuable tool for diversity management and a foundation for breeding programs. The use of these collections for collection management in research centers and breeding perspectives for coffee improvement are discussed.

Base composition is highly variable among and within plant genomes, especially at third codon positions, ranging from GC-poor and homogeneous species to GC-rich and highly heterogeneous ones (particularly Monocots). Consequently, synonymous codon usage is biased in most species, even when base composition is relatively homogeneous. The causes of these variations are still under debate, with three main forces being possibly involved: mutational bias, selection and GC-biased gene conversion (gBGC). So far, both selection and gBGC have been detected in some species but how their relative strength varies among and within species remains unclear. Population genetics approaches allow to jointly estimating the intensity of selection, gBGC and mutational bias. We extended a recently developed method and applied it to a large population genomic dataset based on transcriptome sequencing of 11 angiosperm species spread across the phylogeny. We found that at synonymous positions, base composition is far from mutation-drift equilibrium in most genomes and that gBGC is a widespread and stronger process than selection. gBGC could strongly contribute to base composition variation among plant species, implying that it should be taken into account in plant genome analyses, especially for GC-rich ones., Author summary In protein coding genes, base composition strongly varies within and among plant genomes, especially at positions where changes do not alter the coded protein (synonymous variations). Some species, such as the model plant Arabidopsis thaliana, are relatively GC-poor and homogeneous while others, such as grasses, are highly heterogeneous and GC-rich. The causes of these variations are still debated: are they mainly due to selective or neutral processes? Answering to this question is important to correctly infer whether variations in base composition may have functional roles or not. We extended a population genetics method to jointly estimate the different forces that may affect synonymous variations and applied it to genomic datasets in 11 flowering plant species. We found that GC-biased gene conversion, a neutral process associated with recombination that mimics selection by favouring G and C bases, is a widespread and stronger process than selection and that it could explain the large variation in base composition observed in plant genomes. Our results bear implications for analysing plant genomes and for correctly interpreting what could be functional or not.

Made available in DSpace on 2018-12-11T17:08:57Z (GMT). No. of bitstreams: 0 Previous issue date: 2017-01-01 Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Coffea arabica L. is an important crop in several developing countries. Despite its economic importance, minimal transcriptome data are available for fruit tissues, especially during fruit development where several compounds related to coffee quality are produced. To understand the molecular aspects related to coffee fruit and grain development, we report a large-scale transcriptome analysis of leaf, flower and perisperm fruit tissue development. Illumina sequencing yielded 41,881,572 high-quality filtered reads. De novo assembly generated 65,364 unigenes with an average length of 1,264 bp. A total of 24,548 unigenes were annotated as protein coding genes, including 12,560 full-length sequences. In the annotation process, we identified nine candidate genes related to the biosynthesis of raffinose family oligossacarides (RFOs). These sugars confer osmoprotection and are accumulated during initial fruit development. Four genes from this pathway had their transcriptional pattern validated by quantitative reverse transcription polymerase chain reaction (qRT-PCR). Furthermore, we identified ∼24,000 putative target sites for microRNAs (miRNAs) and 134 putative transcriptionally active transposable elements (TE) sequences in our dataset. This C. arabica transcriptomic atlas provides an important step for identifying candidate genes related to several coffee metabolic pathways, especially those related to fruit chemical composition and therefore beverage quality. Our results are the starting point for enhancing our knowledge about the coffee genes that are transcribed during the flowering and initial fruit development stages. Programa de Pós-GraduacEao em Genética e Biologia Molecular Centro de Ciencias Biológicas Universidade Estadual de Londrina (UEL) Laboratório de Biotecnologia Vegetal Instituto Agronómico do Paraná (IAPAR) Laboratório de Genômica e Expressao Departamento de Genética Evolução e Bioagentes Instituto de Biologia Universidade Estadual de Campinas (UNICAMP) Departamento de Botânica Instituto de Biociências de Rio Claro Universidade Estadual Paulista (UNESP) Centre de Coopération Internationale en Recherche Agronomique Pour le Développement UMR AGAP Programa de Pós Graduação em Agronomia Universidade do Oeste Paulista (UNOESTE) Empresa Brasileira de Pesquisa Agropecuária (Embrapa Café) Departamento de Botânica Instituto de Biociências de Rio Claro Universidade Estadual Paulista (UNESP)

SUMMARYControlled irrigation during the dry period associated with adequate nitrogen (N), phosphorous (P) and potassium (K) fertilization led to the accumulation of biochemical compounds in coffee beans considered as positive precursors of beverage quality. Adult plants of coffee (Coffea arabica ‘Rubi’) were cultivated using different water regimes (WR) and fertilization conditions under the dry climate of the Brazilian Cerrado. Coffee-bean physical characteristics were evaluated as well as biochemical composition by near-infrared analysis. The K treatment mostly affected bean biochemistry, lipid and chlorogenic acid (CGA) contents, which increased with increasing amounts of fertilizer. Caffeine contents increased with higher amounts of N, but no significant effects of P treatment on bean biochemical composition were observed. Sucrose and total lipid contents always appeared higher in beans of non-irrigated plants than those beans from plants grown with continuous irrigation. In contrast, caffeine and CGA contents were higher in beans of irrigated as compared with non-irrigated plants. For the first time, the current results showed that controlled management of irrigation during the dry period associated with reasonable NPK fertilization led to the accumulation of biochemical compounds in coffee beans considered as positive precursors of beverage quality.

Made available in DSpace on 2018-12-11T16:45:05Z (GMT). No. of bitstreams: 0 Previous issue date: 2017-02-01 Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Lipids are among the major chemical compounds present in coffee beans, and they affect the flavor and aroma of the coffee beverage. Coffee oil is rich in kaurene diterpene compounds, mainly cafestol (CAF) and kahweol (KAH), which are related to plant defense mechanisms and to nutraceutical and sensorial beverage characteristics. Despite their importance, the final steps of coffee diterpenes biosynthesis remain unknown. To understand the molecular basis of coffee diterpenes biosynthesis, we report the content dynamics of CAF and KAH in several Coffea arabica tissues and the transcriptional analysis of cytochrome P450 genes (P450). We measured CAF and KAH concentrations in leaves, roots, flower buds, flowers and fruit tissues at seven developmental stages (30–240 days after flowering - DAF) using HPLC. Higher CAF levels were detected in flower buds and flowers when compared to fruits. In contrast, KAH concentration increased along fruit development, peaking at 120 DAF. We did not detect CAF or KAH in leaves, and higher amounts of KAH than CAF were detected in roots. Using P450 candidate genes from a coffee EST database, we performed RT-qPCR transcriptional analysis of leaves, flowers and fruits at three developmental stages (90, 120 and 150 DAF). Three P450 genes (CaCYP76C4, CaCYP82C2 and CaCYP74A1) had transcriptional patterns similar to CAF concentration and two P450 genes (CaCYP71A25 and CaCYP701A3) have transcript accumulation similar to KAH concentration. These data warrant further investigation of these P450s as potential candidate genes involved in the final stages of the CAF and KAH biosynthetic pathways. Instituto Agronômico do Paraná (IAPAR) Laboratório de Biotecnologia Vegetal, CP 481 Universidade Estadual de Londrina (UEL), CP 6001 Instituto Agronômico do Paraná (IAPAR) Laboratório de Ecofisiologia Vegetal, CP 481 Centre de Coopération Internationale en Recherche Agronomique Pour le Développement (CIRAD) UMR AGAP Universidade do Oeste Paulista (UNOESTE), Rodovia Raposo Tavares, Km 572 Universidade Estadual Paulista (UNESP) Instituto de Biociências de Rio Claro, Avenida 24-A, 1515 Empresa Brasileira de Pesquisa Agropecuária (EMBRAPA Café) Universidade Estadual Paulista (UNESP) Instituto de Biociências de Rio Claro, Avenida 24-A, 1515

The aim of this study was to investigate the molecular mechanisms underlying drought acclimation in coffee plants by the identification of candidate genes (CGs) using different approaches. The first approach used the data generated during the Brazilian Coffee expressed sequence tag (EST) project to select 13 CGs by an in silico analysis (electronic northern). The second approach was based on screening macroarrays spotted with plasmid DNA (coffee ESTs) with separate hybridizations using leaf cDNA probes from drought-tolerant and susceptible clones of Coffea canephora var. Conilon, grown under different water regimes. This allowed the isolation of seven additional CGs. The third approach used two-dimensional gel electrophoresis to identify proteins displaying differential accumulation in leaves of drought-tolerant and susceptible clones of C. canephora. Six of them were characterized by MALDI-TOF-MS/MS (matrix-assisted laser desorption-time of flight-tandem mass spectrometry) and the corresponding proteins were identified. Finally, additional CGs were selected from the literature, and quantitative real-time polymerase chain reaction (qPCR) was performed to analyse the expression of all identified CGs. Altogether, >40 genes presenting differential gene expression during drought acclimation were identified, some of them showing different expression profiles between drought-tolerant and susceptible clones. Based on the obtained results, it can be concluded that factors involved a complex network of responses probably involving the abscisic signalling pathway and nitric oxide are major molecular determinants that might explain the better efficiency in controlling stomata closure and transpiration displayed by drought-tolerant clones of C. canephora.

L’amelioration varietale de Coffea canephora represente un grand defi pour augmenter la productivite de cette espece et assurer un meilleur revenu aux planteurs. Les opportunites sont nombreuses : la diversite genetique de cette espece est bien documentee, les parametres genetiques sont relativement bien etablis pour les principaux caracteres d’interet et les ressources genomiques disponibles s’accumulent. Toutefois, des contraintes sont egalement a considerer : en particulier, l’histoire specifique de la mise en culture et de la selection a conduit a une inegalite des selectionneurs a travers le monde pour l’acces a l’ensemble de la diversite genetique de l’espece. Ainsi, la selection recurrente reciproque (SRR) qui represente une innovation majeure en termes d’utilisation raisonnee des ressources genetiques n’a pu etre menee qu’en Cote d’Ivoire qui seule dispose d’un ensemble complet et reference des ressources genetiques de base de C. canephora. Cette strategie est adaptee a une sortie varietale sous la forme de semences hybrides, plus adaptees a une diffusion de masse que les boutures clonales. En dehors de la Cote d’Ivoire, une selection massale clonale sur des populations en recombinaison plus ou moins bien decrites reste souvent l’axe des programmes de creation varietale. Par ailleurs, le genome de C. canephora est sur le point d’etre sequence et les ressources genomiques sont maintenant suffisantes pour venir en appui des programmes de selection. Les choix strategiques qui permettront d’utiliser au mieux ces ressources au service de l’amelioration genetique doivent etre l’objet d’une reflexion approfondie, tenant compte de l’experience acquise pour d’autres plantes : les cles de ces choix sont discutees.

Coffea canephora Pierre ex Frohener is a perennial plant originated from Africa. Two main groups, Guinean and Congolese, have already been identified within this species. They correspond to main refugia in western and central Africa. In this paper we present the analysis of a region that has not yet been studied, Uganda. Two wild, one feral (once cultivated but abandoned for many years), and two cultivated populations of C. canephora from Uganda were evaluated using 24 microsatellite markers. Basic diversity, dissimilarity and genetic distances between individuals, genetic differentiation between populations, and structure within populations were analysed. Expected heterozygosity was high for wild compartments (0.48 to 0.54) and for cultivated and feral ones (0.57 to 0.59), with the number of private alleles ranging from 12 for cultivated genotypes to 37 for a wild compartment. The Ugandan samples show significant population structuring. We compared the Ugandan populations with a representative sample of known genetic diversity groups within the species using 18 markers. Coffea canephora of Ugandan origin was found to be genetically different from previously identified diversity groups, implying that it forms another diversity group within the species. Given its large distribution and extremely recent domestication, C. canephora can be used to understand the effect of refugia colonization on genetic diversity.

Background Drought is a widespread limiting factor in coffee plants. It affects plant development, fruit production, bean development and consequently beverage quality. Genetic diversity for drought tolerance exists within the coffee genus. However, the molecular mechanisms underlying the adaptation of coffee plants to drought are largely unknown. In this study, we compared the molecular responses to drought in two commercial cultivars (IAPAR59, drought-tolerant and Rubi, drought-susceptible) of Coffea arabica grown in the field under control (irrigation) and drought conditions using the pyrosequencing of RNA extracted from shoot apices and analysing the expression of 38 candidate genes. Results Pyrosequencing from shoot apices generated a total of 34.7 Mbp and 535,544 reads enabling the identification of 43,087 clusters (41,512 contigs and 1,575 singletons). These data included 17,719 clusters (16,238 contigs and 1,575 singletons) exclusively from 454 sequencing reads, along with 25,368 hybrid clusters assembled with 454 sequences. The comparison of DNA libraries identified new candidate genes (n = 20) presenting differential expression between IAPAR59 and Rubi and/or drought conditions. Their expression was monitored in plagiotropic buds, together with those of other (n = 18) candidates genes. Under drought conditions, up-regulated expression was observed in IAPAR59 but not in Rubi for CaSTK1 (protein kinase), CaSAMT1 (SAM-dependent methyltransferase), CaSLP1 (plant development) and CaMAS1 (ABA biosynthesis). Interestingly, the expression of lipid-transfer protein (nsLTP) genes was also highly up-regulated under drought conditions in IAPAR59. This may have been related to the thicker cuticle observed on the abaxial leaf surface in IAPAR59 compared to Rubi. Conclusions The full transcriptome assembly of C. arabica, followed by functional annotation, enabled us to identify differentially expressed genes related to drought conditions. Using these data, candidate genes were selected and their differential expression profiles were confirmed by qPCR experiments in plagiotropic buds of IAPAR59 and Rubi under drought conditions. As regards the genes up-regulated under drought conditions, specifically in the drought-tolerant IAPAR59, several corresponded to orphan genes but also to genes coding proteins involved in signal transduction pathways, as well as ABA and lipid metabolism, for example. The identification of these genes should help advance our understanding of the genetic determinism of drought tolerance in coffee. Electronic supplementary material The online version of this article (doi:10.1186/s12870-016-0777-5) contains supplementary material, which is available to authorized users.

The exploration of accessions originating from Ethiopia is one option available to increase the genetic basis of new coffee cultivars. The aim of this study was to describe the chemical composition variability of Ethiopian coffee accessions maintained in Londrina and to evaluate the year effect on that. We observed no significant interaction between the years of cultivation and the amounts of caffeine, protein, lipids, chlorogenic acids, sucrose, cafestol and kahweol, thereby suggesting that evaluations of one year were reliable to describe this collection. All compounds showed variability, but the highest variability was found in cafestol and kahweol (32.81 and 34.45 %, respectively). Significant correlations among caffeine, chlorogenic acids and protein as well as between sucrose and lipids were observed. A significant negative correlation was found between kahweol and cafestol, and kahweol had positive and negative correlation with caffeine and chlorogenic acids, respectively. Hierarchical clustering analyses identified groups in the population, and the diversity found in one crop year was similar to that for 3 years. Greater diversity of composition was observed among the Western accessions compared to the ones from the East, but no clear separation was found between the accessions of these regions. These accessions are potential sources to increase the genetic diversity to obtain new coffee cultivars.

marraccini@cirad.fr; International audience; Sucrose metabolism and the role of sucrose synthase were investigated in the fruit tissues (pericarp, perisperm, and endosperm) of Coffea arabica during development. Acid invertase, sucrose phosphate synthase, and sucrose synthase activities were monitored and compared with the levels of sucrose and reducing sugars. Among these enzymes, sucrose synthase showed the highest activities during the last stage of endosperm and pericarp development and this activity paralleled closely the accumulation of sucrose in these tissues at this stage. Carbon partitioning in fruits was studied by pulse-chase experiments with C-14-sugars and revealed high rates of sucrose turnover in perisperm and endosperm tissues. Additional feeding experiments with (CO2)-C-14 showed that leaf photosynthesis contributed more to seed development than the pericarp in terms of photosynthate supply to the endosperm. Sugar analysis, feeding experiments, and histological studies indicated that the perisperm plays an important role in this downloading process. It was observed that the perisperm presents a transient accumulation of starch which is degraded as the seed develops. Two full-length cDNAs (CaSUS1 and CaSUS2) and the complete gene sequence of the latter were also isolated. They encode sucrose synthase isoforms that are phylogenetically distinct, indicating their involvement in different physiological functions during cherry development. Contrasting expression patterns were observed for CaSUS1 and CaSUS2 in perisperm, endosperm, and pericarp tissues: CaSUS1 mRNAs accumulated mainly during the early development of perisperm and endosperm, as well as during pericarp growing phases, whereas those of CaSUS2 paralleled sucrose synthase activity in the last weeks of pericarp and endosperm development. Taken together, these results indicate that sucrose synthase plays an important role in sugar metabolism during sucrose accumulation in the coffee fruit

Eleven mating designs, 6 top-crosses and 5 North Carolina II, were used to estimate narrow sense, broad sense and within-cross heritability of Coffea canephora yield in Cote d'Ivoire. The objective was to predict achievable genetic gains according to different breeding strategies. Narrow sense, broad sense, family and within-cross heritabilities were 0.32, 0.43, 0.83 and 0.22, on average respectively. The percentage of additivity was high (81% on average). A degree of variation was nevertheless observed for estimates over trials. This was found to be mainly due to parent sampling in each trial. By theoretical computation, the minimum number of parents required to obtain optimum precision was found to be around 30. Predicted genetic gains for yield through clonal selection at a selection level of 1% ranged from 7 to 28% over the best hybrid in each trial, suggesting thus clones still interesting for breeders. The consequences for coffee breeding are discussed.

Genetically-modified coffee clones (GMCs) were presented in a previous work. They were created from a single commercial clone of Coffea canephora Pierre (clone 126). Therefore they all have the same genotype, except for the localization of the transgenic insertions. They synthesize the Bacillus thuringiensis endotoxin Cry1Ac against Leucoptera coffeella Gue´- rin-Me´neville, a secondary pest of C. canephora and one of the main pest of C. arabica in South America. The synthetic Cry1Ac gene is regulated by the EF1a- A1 promoter of Arabidopsis thaliana (L.) Heynh. They were tested in an experimental field for their resistance against the pest insect in a previous work. In the present work, levels of Cry1Ac were measured in the leaves. The insecticidal protein was evenly distributed in all the leaves. Cry1Ac levels were measured once in the coffee plants of a sensitive GMC and of 14 resistant ones grown in the experimental field and in plants grown in a greenhouse. Some resistant GMCs contained higher levels but it is not possible to confirm that it would be enough for a sustainable resistance to the pest. Growth speed was variable in the plot. The correlation with plant height and other indicators of plant growth was examined. Cry1Ac levels in the GMCs were negatively correlated with growth speed. The latter was not statistically influenced neither by Cry1Ac synthesis nor by the genetic modification in itself as seen by comparing the GMCs and the unmodified control clone 126. Hence the conclusion is that the growth conferred to field-grown plants by environmental factors and especially the soil was probably the underlying cause of the negative correlation. Other field experiments would be necessary in order to confirm this result. It would be important that the genetic construct inserted in these GMCs and mainly the EF1a-A1 promoter of the Cry1Ac gene be reconsidered since Cry1Ac levels might be too low to provide efficient and sustainable protection against L. coffeella in a highly favourable environment for coffee plants. Alternatives are discussed.

Coffee (Coffea sp.) is a perennial plant widely cultivated in many tropical countries. It is a cash crop for millions of small farmers in these areas. As for other tree species, coffee has long breeding cycles, which makes conventional breeding programs time-consuming. For that matter, genetic transformation can be an effective way to introduce a desired trait in elite varieties or for functional genomics. In this chapter, we describe two highly efficient and reliable Agrobacterium-mediated transformation techniques developed for the C. arabica cultivated species: (1) A. tumefaciens to study and introduce genes conferring resistance/tolerance to biotic (coffee leaf rust, insects) and abiotic stress (drought, heat, seed desiccation) in fully transformed plants and (2) A. rhizogenes to study candidate gene expression for nematode resistance in transformed roots.

Coffee, tea, and chocolate converge Caffeine has evolved multiple times among plant species, but no one knows whether these events involved similar genes. Denoeud et al. sequenced the Coffea canephora (coffee) genome and identified a conserved gene order (see the Perspective by Zamir). Although this species underwent fewer genome duplications than related species, the relevant caffeine genes experienced tandem duplications that expanded their numbers within this species. Scientists have seen similar but independent expansions in distantly related species of tea and cacao, suggesting that caffeine might have played an adaptive role in coffee evolution. Science , this issue p. 1181 ; see also p. 1124

We searched for quantitative trait loci (QTL) associated with the palm oil fatty acid composition of mature fruits of the oil palm E. guineensis Jacq. in comparison with its wild relative E. oleifera (H.B.K) Corte´ s. The oil palm cross LM2T x DA10D between two heterozygous parents was considered in our experiment as an intraspecific representative of E. guineensis. Its QTLs were compared to QTLs published for the same traits in an interspecific Elaeis pseudo-backcross used as an indirect representative of E. oleifera. Few correlations were found in E. guineensis between pulp fatty acid proportions and yield traits, allowing for the rather independent selection of both types of traits. Sixteen QTLs affecting palm oil fatty acid proportions and iodine value were identified in oil palm. The phenotypic variation explained by the detected QTLs was low to medium in E. guineensis, ranging between 10% and 36%. The explained cumulative variation was 29% for palmitic acid C16:0 (one QTL), 68% for stearic acid C18:0 (two QTLs), 50% for oleic acid C18:1 (three QTLs), 25% for linoleic acid C18:2 (one QTL), and 40% (two QTLs) for the iodine value. Good marker co-linearity was observed between the intraspecific and interspecific Simple Sequence Repeat (SSR) linkage maps. Specific QTL regions for several traits were found in each mapping population. Our comparative QTL results in both E. guineensis and interspecific materials strongly suggest that, apart from two common QTL zones, there are two specific QTL regions with major effects, which might be one in E. guineensis, the other in E. oleifera, which are independent of each other and harbor QTLs for several traits, indicating either pleiotropic effects or linkage. Using QTL maps connected by highly transferable SSR markers, our study established a good basis to decipher in the future such hypothesis at the Elaeis genus level.

Near-infrared spectroscopy (NIRS) is among the many tools available to study the biochemical diversity of coffee species. This technique is inexpensive, fast and accurate, and it requires only small amounts of samples. The aim of this study was to use NIRS to estimate the amount of diterpenes (cafestol and kahweol) in green coffee. To construct the prediction model, 126 Ethiopian accessions coffee collection and 44 modern cultivars were analyzed. The total sample set was split into two groups as follows: a group of 130 samples for calibration and a group of 40 samples for the validation step. Reference values of cafestol and kahweol were determined by high performance liquid chromatography (HPLC). Cafestol values ranged from 182.62 g to 1308.62 mg 100 g − 1 , and kahweol values ranged from 182.69 to 1265.41 mg 100 g − 1 . To improve the quality of the calibration step, a pretreatment with the second derivative was applied to smooth the raw spectra. The prediction models of cafestol and kahweol were developed using the modified partial least squares regression (mPLS). The performance of these models was evaluated by the ratio of performance deviation (RPD) and R 2 parameters, obtained by the ratio of the NIR prediction data and the corresponding reference data. The prediction models of cafestol (RPD = 2.74; R 2 = 0.89) and kahweol (RPD = 2.2; R 2 = 0.88) confirm the validity of NIRS analysis to determine diterpenes contents in green coffee.

The recurrent selection schemes recommended by CIRAD for 6 tropical tree crops are presented and compared. Breeding programmes are carried out under cooperation between CIRAD and its partners in regions where the crops are grown. The crops are cacao, coffee, rubber tree, oil palm, coconut and eucalyptus. After a short look at the background, the reasons behind the options chosen are given and the main characteristics of the schemes are described and illustrated. The state of progress and some major results are discussed. Over and above the differences linked to plant biology, the genetic diversity available and the type of varietal output, common characteristics are emphasized. The time taken for each generation means simplifying the intercrossing phases between successive breeding cycles, and thus, the parents tested are heterozygous: whenever possible, a clonal varietal output is an additional source of progress. The use of biotechnologies to study diversity is a valuable asset, and their application for early selection (QTL) is also promising. The preponderance of combining ability coexists with marked heterosis, which poses the problem of the latter's origin. As a result, all of the schemes described, but one, are reciprocal recurrent selection schemes.

Breeding of the allogamous Coffea canephora species in Cote d'Ivoire is based, from 1984 onward, on a reciprocal recurrent selection scheme exploiting the good performance of crosses between two genetically distinct populations (‘Guinean’ and ‘Congolese’). Results are presented here on intergroup cross progenies evaluated on individual plant basis over a six year period in three fully randomized trials planted in 1985 and 1986. Yields of the bi-parental crosses varied from 16 to 1407% of the mean of the two commercial clonal cultivars and the most productive crosses were also the most vigourous. Selection of individual trees and cross progenies, based on a combined individual-family index, was carried out for high yield, good vigour of young plants and moderate canopy diameter of adult trees. Expected genetic gains, as estimated in relation to the mean of all crosses, for selection of 5% of the best trees, were high for yield (over 60%), moderate for young plant vigour (14 to 18%), and, as intended, low for canopy diameter. In addition, selection by independent culling for leaf rust resistance and bean weight was carried out. As a result, 6 promising crosses with yield similar or higher than those of the control cultivars, were selected as candidates for commercial use, as well as 65 individual trees, within 19 different crosses. Expected genetic gain for yield for these selected trees is about 60% compared to the average yield of the most productive control clone. Continuation of the programme is discussed, with specific attention to further progress expected for quality traits.

Background A reciprocal recurrent selection program has been under way for the Coffea canephora coffee tree for approximately thirty years in the Ivory Coast. Association genetics would help to speed up this program by more rapidly selecting zones of interest in the genome. However, prior to any such studies, the linkage disequilibrium (LD) needs to be assessed between the markers on the genome. These data are essential for guiding association studies. Results This article describes the first results of an LD assessment in a coffee tree species. Guinean and Congolese breeding populations of C. canephora have been used for this work, with the goal of identifying ways of using these populations in association genetics. We identified changes in the LD along the genome within the different C. canephora diversity groups. In the different diversity groups studied, the LD was variable. Some diversity groups displayed disequilibria over long distances (up to 25 cM), whereas others had disequilibria not exceeding 1 cM. We also discovered a fine structure within the Guinean group. Conclusions Given these results, association studies can be used within the species C. canephora. The coffee recurrent selection scheme being implemented in the Ivory Coast can thus be optimized. Lastly, our results could be used to improve C. arabica because one of its parents is closely related to C. canephora.

As the second species used for commercial coffee, evaluation of Coffea canephora Pierre ex Froehner population genetics is a challenging task for coffee breeding. This study examined the pattern of genetic variability and genetic relationships of cultivated and wild populations of C. canephora sampled across the Guineo-Congolese region of Africa and some improved populations maintained in field genebanks. A total of 293 individuals, sampled from 17 populations, were genotyped with 39 nuclear microsatellite markers. Genetic diversity and structure were investigated with both a model-based and a graphical approach; isolation by distance was also tested. Relationships between the diversity clusters are discussed with regard to differentiation due to several glacial refuges during the Last Glacial Maximum (LGM). High genetic diversity within C. canephora is confirmed with a mean number of alleles of 11.85 per marker, a mean gene diversity of 0.72 and a mean observed heterozygosity of 0.36. An overall structure of two main groups (Guinean and Congolese) subdivided in six subgroups (2 for the Guinean and 4 for the Congolese) was found, including one described for the first time in the Guinean group. A fine structure within the Guinean group was also newly detected. Genetic structure of C. canephora appears to be consistent with its geographic repartition at the continent scale. Structure of diversity was found in accordance with localizations of refuge zones during LGM and migration from this period. Results from this genetic structure study raise our capabilities to better manage and use the collections of genetics resources for breeding purposes. Those results will be used in future association studies to optimize the number of genotypes to be phenotyped.

Coffee is one of the world’s most important agricultural commodities. Coffee belongs to the Rubiaceae family in the euasterid I clade of dicotyledonous plants, to which the Solanaceae family also belongs. Two bacterial artificial chromosome (BAC) libraries of a homozygous doubled haploid plant of Coffea canephora were constructed using two enzymes, HindIII and BstYI. A total of 134,827 high quality BAC-end sequences (BESs) were generated from the 73,728 clones of the two libraries, and 131,412 BESs were conserved for further analysis after elimination of chloroplast and mitochondrial sequences. This corresponded to almost 13 % of the estimated size of the C. canephora genome. 6.7 % of BESs contained simple sequence repeats, the most abundant (47.8 %) being mononucleotide motifs. These sequences allow the development of numerous useful marker sites. Potential transposable elements (TEs) represented 11.9 % of the full length BESs. A difference was observed between the BstYI and HindIII libraries (14.9 vs. 8.8 %). Analysis of BESs against known coding sequences of TEs indicated that 11.9 % of the genome corresponded to known repeat sequences, like for other flowering plants. The number of genes in the coffee genome was estimated at 41,973 which is probably overestimated. Comparative genome mapping revealed that microsynteny was higher between coffee and grapevine than between coffee and tomato or Arabidopsis. BESs constitute valuable resources for the first genome wide survey of coffee and provide new insights into the composition and evolution of the coffee genome.

IATE Axe 4 : Biotechnologie microbienne et enzymatique des lipides et des agropolymères; We chose an Elaeis interspecific pseudo-backcross of first generation (E. oleifera x E. guineensis) x E. guineensis to identify quantitative trait loci (QTLs) for fatty acid composition of palm oil. A dense microsatellite linkage map of 362 loci spanned 1.485 cM, representing the 16 pairs of homologous chromosomes in the Elaeis genus from which we traced segregating alleles from both E. oleifera and E. guineensis grandparents. The relative linear orders of mapped loci suggested the probable absence of chromosome rearrangements between the E. oleifera and E. guineensis genomes. A total of 19 QTL associated to the palm oil fatty acid composition were evidenced. The QTL positions and the species origin as well as the estimated effects of the QTL marker alleles were in coherence with the knowledge of the oil biosynthesis pathway in plants and with the individual phenotypic correlations between the traits. The mapping of chosen Elaeis key genes related to oleic acid C18:1, using intra-gene SNPs, supported several QTLs underlying notably FATA and SAD enzymes. The high number of hyper-variable SSR loci of known relative linear orders and the QTL information make these resources valuable for such mapping study in other Elaeis breeding materials.

Knowledge on heritability is essential for selecting varieties resistant against coffee wilt disease caused by Fusarium xylarioides, which is currently devastating coffee in East and Central Africa. Variability of the resistance against coffee wilt disease in Coffea canephora and its inheritance were investigated in three experiments corresponding to a clonal trial involving 20 clones, a 10- parent partial diallel progeny, and a half-sib progeny test. There were significant quantitative genetic variations among the clones and progenies, suggesting polygenic control of the resistance. Around 50-65 % tree mortality was the optimal disease level for calculating heritability and genetic gains. General and specific combining abilities calculated within the optimal disease range for partial diallel analysis were significant. Broad-sense heritability for the same analysis and same disease range was moderate (0.329), and corresponding narrow-sense heritability was low (0.112). Broad-sense heritability from clones in the field at the same disease range was also moderate (0.333). Narrow-sense heritability from regression of half-sib progeny means onto parent means in the field and screen house was 0.183 and 0.369, respectively. Selecting tolerant clones for improvement against the disease is possible, and genetic gains are possible by using tolerant parents in breeding programs.

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R. Leroy, Thierry Seguin, Marc Roeckel-Drevet, Patricia Pujade-Renaud, Valerie IFC (Institut Francais du Caoutchouc, Paris, France); Michelin (Clermont-Ferrand, France); SIPH ("Societe Internationale de Plantations d'Hevas", Courbevoie, France); SOCFIN ("Societe Financiere des Caoutchoucs", Bruxelles, Belgium) This work was supported in part by a grant from the IFC (Institut Francais du Caoutchouc, Paris, France) and the companies Michelin (Clermont-Ferrand, France), SIPH ("Societe Internationale de Plantations d'Hevas", Courbevoie, France) and SOCFIN ("Societe Financiere des Caoutchoucs", Bruxelles, Belgium). We thank Boris Fumanal and Jean-Stephane Venisse for their valuable comments. Springer New york Si; Corynespora cassiicola is the causal agent of Corynespora Leaf Fall (CLF) disease. CLF is one of the most important fungal diseases of rubber trees in Asia and Africa but disease outbreaks have not been reported in South America. Cassiicolin, a small cysteine-rich glycoprotein secreted by the pathogenic isolate CCP, was previously identified as a potential disease effector in rubber tree. Recently, the cassiicolin-encoding gene () was characterized and shown to be expressed in the early phase of infection. In this study, we investigated whether previously undetected strains of are present in South American rubber plantations by examining the fungal endophyte population found in asymptomatic rubber tree leaves. Four isolates were identified as . Genes encoding new forms of the cassiicolin precursor protein (Cas3 and Cas4) were identified from these isolates. Three of four isolates were able to induce symptoms on the cultivar they were isolated from in a detached leaf assay, with different kinetics and intensities. One isolate had the same pathogenicity profile as the pathogenic isolate CCP; the other two isolates developed symptoms late during the course of infection, suggesting saprotrophic capabilities. However, no or transcripts could be detected upon inoculation with the endophytic isolates, whereas the reference gene was expressed upon inoculation with the CCP isolate. This work demonstrated that is present in South America in an endophytic form and that it may evolve from an endophytic to a saprophytic or even potentially pathogenic life style.

UMR 1334 AGAP : Equipe AFEF 'Architecture et Fonctionnement des Espèces fruitières' ; Team AFFS 'Architecture and Functioning of Fruit Species' Contact: valerie.pujade-renaud@cirad.fr; Corynespora Leaf Fall (CLF) is a major disease of rubber tree (Hevea brasiliensis) caused by the Ascomycota Corynespora cassiicola. Here we describe the cloning and characterization of a gene encoding cassiicolin (Cas), a glycosylated cystein-rich small secreted protein (SSP) identified as a potential CLF disease effector in rubber tree. Three isolates with contrasted levels of aggressiveness were analyzed comparatively. The cassiicolin gene was detected - and the toxin successfully purified - from the isolates with high and medium aggressiveness (CCP and CCAM3 respectively) but not from the isolate with the lowest aggressiveness (CCAM1), suggesting the existence of a different disease effector in the later. CCP and CCAM3 carried strictly identical cassiicolin genes and produced toxins of identical mass, as evidence by mass spectrometry analysis, thus suggesting conserved post-translational modifications in addition to sequence identity. The differences in aggressiveness between CCP and CCAM3 may be attributed to differences in cassiicolin transcript levels rather than qualitative variations in cassiicolin structure. Cassiicolin may play an important role in the early phase of infection since a peak of cassiicolin transcripts occurred in 1 or 2 days after inoculation (before the occurrence of the first symptoms), in both the tolerant and the susceptible cultivars.

Background Understanding the genetic elements that contribute to key aspects of coffee biology will have an impact on future agronomical improvements for this economically important tree. During the past years, EST collections were generated in Coffee, opening the possibility to create new tools for functional genomics. Results The "PUCE CAFE" Project, organized by the scientific consortium NESTLE/IRD/CIRAD, has developed an oligo-based microarray using 15,721 unigenes derived from published coffee EST sequences mostly obtained from different stages of fruit development and leaves in Coffea Canephora (Robusta). Hybridizations for two independent experiments served to compare global gene expression profiles in three types of tissue matter (mature beans, leaves and flowers) in C. canephora as well as in the leaves of three different coffee species (C. canephora, C. eugenoides and C. arabica). Microarray construction, statistical analyses and validation by Q-PCR analysis are presented in this study. Conclusion We have generated the first 15 K coffee array during this PUCE CAFE project, granted by Génoplante (the French consortium for plant genomics). This new tool will help study functional genomics in a wide range of experiments on various plant tissues, such as analyzing bean maturation or resistance to pathogens or drought. Furthermore, the use of this array has proven to be valid in different coffee species (diploid or tetraploid), drastically enlarging its impact for high-throughput gene expression in the community of coffee research.

Coffea canephora breeding requires combining sustainable productivity with improved technological and cup quality characteristics. Beverage quality is a complex and subjective trait, and breeding for this trait is time consuming and depends on knowledge of the genetics of its components. A highly variable C. canephora progeny resulting from an intraspecific cross was assessed for 63 traits over 5 years. To identify quantitative trait loci (QTLs) controlling agronomic, technological, and quality-related traits, a genetic map comprising 236 molecular markers was constructed, and composite interval mapping was performed. Beverage quality was evaluated in relation to biochemical and cup tasting traits. QTLs were identified for almost half of the traits evaluated, with effects ranging from 6% to 80% of phenotypic variation. Most of them present a consistent detection over years. The strongest QTLs explained a high percentage of the variation for yield in 2006 (34% to 57%), bean size (25% to 35%), content of chlorogenic acids (22% to 35%), sucrose and trigonelline content (29% to 81%), and acidity and bitterness of coffee beverages (30% to 55%). Regions of the C. canephora genome influencing beverage quality were identified. Five QTL zones were co-localized with candidate genes related to the biosynthesis of the analyzed traits: two genes coding for caffeine biosynthesis, one gene implicated in the biosynthesis of chlorogenic acids, and two genes implicated in sugar metabolism. This is one of the first studies on the identification of QTLs combining agronomic and quality traits in coffee. The high variability of quality traits within C. canephora and the presence of consistent QTLs offer breeders a promising tool to improve coffee cup quality.

Genetic parameters were estimated in trials planted in connection with a reciprocal recurrent selection programme applied to Coffea canephora in Cote d'Ivoire. Narrow sense heritability values, estimated from a breakdown of the analysis of variance, were high for architectural characters (0.22 to 0.78) and medium for tree vigour (0.13 to 0.40) and for bean weight (0.15 to 0.28). Calculated heritability values for the first two harvests were high (h2>0.7), but much lower for the following harvests (less than 0.2). Heritability estimates by parent-offspring regressions gave variable results, though they were often similar to those obtained by variance analysis, especially for bean weight. Phenotypic and genetic correlations between tree vigour traits and productivity were high. Bean weight was not correlated with vigour or production. Canopy diameter of 4 year-old trees was closely correlated with cumulated productivity from 2 to 5 years in the absence of development competition between trees, but the correlation was low if trees were in competition with each other. The consequences of the results for the choice of characters to be selected among intergroup hybrids or the parents of both populations are discussed.

Two breeding populations have been identified with Coffea canephora, based on geographical and genetic differences: the Guinean group from West Africa and the Congolese group from central Africa. A reciprocal recurrent selection programme based on these groups was initiated in Cote d'Ivoire in 1984. Genotypes of both groups available in field collections are either of cultivated or wild origin. Genotypic and phenotypic variability was evaluated within and between the groups for the following characteristics: isozyme patterns, architecture, drought tolerance and vigour, technological and organoleptic traits, pest and disease resistance. Variation between and within groups was found to be large. Based on isozyme analysis and phenotypic observations, two subgroups were identified within the Congolese group. Phenotypic values of parents and testers used in the first cycle of reciprocal recurrent selection are presented. Results of two intergroup hybrid trials indicate high yield and vigour for these hybrids compared to intragroup hybrids, justifying the reciprocal selection approach. Furthermore, the variation observed indicates that recurrent selection would allow progress for selection traits for both populations and intergroup hybrids.

The number of individual branches attacked by the scolytid coffee‐twig borer was recorded in 1990 and 1991 in a Coffea canephora clonal trial planted in Cote d'Ivoire in 1988, in a randomized single‐tree plot design. An ANCOVA described by Papadakis (1937), taking the mean of neighbouring trees as the covariable, was used to compare the log transformed relative numbers of twigs attacked. Of the 21 clones studied, three proved to be significantly more resistant than all the others. These clones were selected from the progeny of the same cross, suggesting simple transmission of the resistance character. Other clones suffered an intermediate level of attack, sometimes varying from one year to the next. In general, all clones involving the Kouilou type showed high resistance. Tree architecture, vigour and yields did not appear to have any effect on the degree of attack. There was a negative binomial pattern of attack over the plot as a whole, but the residual values (compared to the mean for each clo...

La présente étude montre la transférabilité de marqueurs microsatellites développés sur les deux espèces de caféiers cultivées (Coffea arabica L. et C. canephora Pierre ex Froehn.) à 15 espèces représentant les principaux groupes précédemment identifiés du genre Coffea. L'évaluation de la diversité et des ressources génétiques disponibles au sein du genre Coffea et le développement de marqueurs moléculaires transférables d'une espèce à l'autre sont des étapes importantes pour l'amélioration de ces deux espèces. Nous avons travaillé sur 15 espèces avec 60 marqueurs microsatellites développés suivant différentes méthodologies (banques enrichies en microsatellites, banques BAC, séquences de gènes). Nous avons plus particulièrement analysé quatre espèces d'intérêt en commerce ou en amélioration. Nos résultats établissent que les microsatellites sont hautement transférables dans le genre Coffea. Nous mettons en évidence l'important réservoir de diversité pour l'amélioration que constituent les espèces sauvages de ce genre. Enfin nous discutons des implications pour de futures études de cartographie comparée et l'amélioration des deux espèces cultivées.

International audience; Coffea canephora Pierre breeding in Cote d'Ivoire is based on indirect reciprocal recurrent selection (RRSi) using the Congolese and Guinean populations. In practice, the genotypes of each population (78 Congolese genotypes and 100 Guinean genotypes) are tested with several testers from the reciprocal population: two Guinean and three Congolese testers. These testers are heterozygous as C. canephora is a strictly outcrossing species. After a first cycle of RRSi, we judged the efficiency of these testers for yield, susceptibility to coffee leaf rust (SCLR) and hundred bean weight (HBW). All the testers discriminated the tested genotypes highly significantly (P < 0.01), even though differences were found in the degrees of discrimination. For yield and SCLR, no interaction between tested genotypes and testers was observed and correlations between test values obtained from different testers were significant or highly significant. Furthermore, test values of tested genotypes could be used to predict the yield and SCLR of between-population (BP) hybrids. Prediction was always more accurate with the best tester than with the mean of several testers. As opposed to yield and SCLR, HBW showed significant interactions between tested genotypes and testers and HBW of hybrids could not be predicted from the test value of the genotypes. We conclude that only one reciprocal tester can be used in the next cycles of RRSi applied to C. canephora. This will allow either a reduced cost of selection or increased selection intensity.

Sucrose is one of the compounds in the raw coffee bean that has been identified as an important precursor of coffee flavour and aroma. In order to increase our knowledge of sucrose metabolism in coffee species, sucrose and reducing sugar content were investigated in the fast fruit-ripening coffee species Coffea racemosa. Fruits were harvested regularly from anthesis, until the point of complete fruit maturation and tissue development, followed by measurement of fruit tissue weight. Pericarp was the most abundant tissue, and always represented around 70?80% of fruit fresh weight. The perisperm present in young fruits was rapidly replaced by the endosperm at around 40 days after flowering (DAF). In the latter tissue, total and reducing sugars decreased during development. However, sucrose content was maintained at a relatively high level throughout fruit development, with a peak at 40 DAF that coincided with the highest level of sucrose synthase (SUS) activity detected in this tissue. For all endosperm developmental stages analysed, very low invertase activity was observed, suggesting a limited role for this enzyme in sucrose metabolism. Northern blot experiments using SUS1 and SUS2 cDNA sequences from Coffea arabica as probes were carried out to study the expression of SUS-encoding genes. The SUS1 transcripts of C. racemosa overlapped with the peak of SUS activity in the endosperm, whereas SUS2 transcripts accumulated mainly during the latest stages of endosperm development. Altogether, these results suggest that the SUS1 isoform of SUS is essential for sucrose accumulation in the fruits of C. racemosa. Introduction Coffea racemosa Lour. (Rubiaceae) originates from East Africa and is characterised by abundant branching, high leaf drop during the dry season and rapid fruit maturation (Guerreiro Filho, 1992). C. racemosa produces a pale coloured aromatic

Soixante-dix pour cent des 103 espèces de Coffea actuellement identifiées sont menacées d'extinction, mais peu d'entre elles sont étudiées. Un jeu de 40 marqueurs microsatellites polymorphes a été développé en utilisant une banque génomique de Coffea canephora enrichie en GA/GT. L'amplification de ces marqueurs a été testée sur des accessions de C. heterocalyx (une espèce déclarée " Gravement Menacée ") et C. pseudozanguebariae (une espèce déclarée " Vulnérable "), appartenant à différents clades géographiques Africains. Tous les microsatellites sont polymorphes chez C. canephora, avec un nombre moyen d'allèles par locus de plus de 3 (sur un minimum de 9 génotypes testés). Les hétérozygoties observées et attendues, calculées pour les espèces C. canephora et C. pseudozanguebariae varient respectivement entre 0,10 et 0,91; et entre 0,20 et 0,77. Au total, 95 % des paires d'amorces ont permis une amplification chez C. heterocalyx et C. pseudozanguebariae, indiquant un fort taux de transfert au sein du genre Coffea. Ce large jeu de marqueurs sera utile pour des études génétiques approfondies des espèces menacées de Coffea.

To understand the importance of ethylene receptor genes in the quality of coVee berries three full-length cDNAs corresponding to a putative ethyl- ene receptor gene (ETR1) were isolated from CoVea canephora cDNA libraries. They diVered by their 3UTR and contained a main ORF and a 5UTR short ORF putatively encoding a small polypeptide. The CcETR1 gene, present as a single copy in the C. cane- phora genome, contained Wve introns in the coding region and one in its 5UTR. Alternative splicing can occur in C. canephora and C. pseudozanguebariae, lead- ing to a truncated polypeptide. C. pseudozanguebariae ETR1 transcripts showed various forms of splicing alterations. This gene was equally expressed at all stages of fruit development. A segregation study on an inter-speciWc progeny showed that ETR1 is related to the fructiWcation time, the caVeine content of the green beans, and seed weight. Arabidopsis transformed etio- lated seedlings, which over-expressed CcETR1, dis- played highly reduced gravitropism, but the triple response was observed in an ethylene enriched envi- ronment. These plants behaved like a low-concentra- tion ethylene-insensitive mutant thus conWrming the receptor function of the encoded protein. This gene showed no induction during the climacteric crisis but some linkage with traits related to quality.

Coffee (Coffea sp.) is a perennial plant widely cultivated in many tropical countries. It is a cash crop for millions of small farmers in these areas. As compared with other tree species, coffee has long breeding cycles that make conventional breeding programs time consuming. For that matter, genetic transformation can be an effective technique to introduce a desired trait in an already "elite" variety, or to study a gene function and expression. In this chapter, we describe two Agrobacterium-mediated transformation techniques; the first with A. tumefaciens to introduce an insect resistance gene and the second with A. rhizogenes to study candidate gene expression for nematode resistance in transformed roots.

Coffee (Coffea sp.) is a perennial plant widely cultivated in many tropical countries. It is a cash crop for millions of small farmers in these areas. As compared with other tree species, coffee has long breeding cycles that make conventional breeding programs time consuming. For that matter, genetic transformation can be an effective technique to introduce a desired trait in an already "elite" variety, or to study a gene function and expression. In this chapter, we describe two Agrobacterium-mediated transformation techniques; the first with A. tumefaciens to introduce an insect resistance gene and the second with A. rhizogenes to study candidate gene expression for nematode resistance in transformed roots.

Coffee quality, in the present context of overproduction worldwide, has to be considered as a main selection criterion for coffee improvement. After a definition of quality, and an overview of the non genetic factors affecting its variation, this review focuses on the genetic factors involved in the control of coffee quality variation. Regarding the complexity of this trait, the different types of quality are first presented. Then, the great variation within and between coffee species is underlined, mainly for biochemical compounds related to quality (caffeine, sugars, chlorogenic acids, lipids). The ways for breeding quality traits for cultivated species, Coffea arabica and Coffea canephora are discussed, with specific challenges for each species. For C. arabica, maintaining a good quality in F1 intraspecific hybrids, introgressed lines from Timor hybrid, and grafted varieties are the main challenges. For C. canephora, the improvement is mainly based on intraspecific and interspecific hybrids, using the whole genetic variability available within this species. An improvement is obtained for bean size, with significant genetic gains in current breeding programmes. The content in biochemical compounds related to cup quality is another way to improve Robusta quality. Finally, ongoing programmes towards the understanding of the molecular determinism of coffee quality, particularly using coffee ESTs, are presented, Genética da qualidade do café: No contexto do excedente de produção mundial, a qualidade do café tem sido considerada o principal critério de seleção no melhoramento dessa cultura. Após definir qualidade e fazer considerações sobre os fatores não genéticos afetando sua variação, esta revisão se concentra sobre os fatores genéticos envolvidos no controle da variação da qualidade do café. Em relação à complexidade desta característica, os diferentes tipos de qualidade são apresentados. Então, a grande variação dentro e entre as espécies de café é discutida, rincipalmente em relação aos caracteres bioquímicos relacionados com qualidade (cafeína, açúcares, ácidos clorogênicos, lipídeos). As maneiras para melhorar estes caracteres relacionados à qualidade nas espécies cultivadas Coffea arabica e Coffea canephora são discutidas, assim como os desafios específicos a cada espécie. Para C. arabica, manutenção da boa qualidade em híbridos interespecíficos F1, linhagens geradas por introgressão a partir do Híbrido do Timor, e enxertia de variedades são os principais desafios. Para C. canephora, o melhoramento é principalmente baseado em híbridos intra e interespecíficos, usando a variabilidade genética disponível nesta espécie. Um avanço é obtido com o tamanho da semente, com significante ganho genético em programas de melhoramento. O conteúdo de compostos bioquímicos relacionados com a qualidade da bebida é uma outra maneira de melhorar a qualidade do café Robusta. Finalmente, são comentados programas em andamento, direcionados para a compreensão do determinismo molecular da qualidade do café, particularmente usando ESTs de café