Chlorophyll fluorescence-based high-throughput phenotyping facilitates the genetic dissection of photosynthetic heat tolerance in African (Oryza glaberrima) and Asian (Oryza sativa) rice.

Rising temperatures and extreme heat events threaten rice production. Half of the global population relies on rice for basic nutrition, and therefore developing heat-tolerant rice is essential. During vegetative development, reduced photosynthetic rates can limit growth and the capacity to store soluble carbohydrates. The photosystem II (PSII) complex is a particularly heat-labile component of photosynthesis. We have developed a high-throughput chlorophyll fluorescence-based screen for photosynthetic heat tolerance capable of screening hundreds of plants daily. Through measuring the response of maximum PSII efficiency to increasing temperature, this platform generates data for modelling the PSII-temperature relationship in large populations in a small amount of time. Coefficients from these models (photosynthetic heat tolerance traits) demonstrated high heritabilities across African (Oryza glaberrima) and Asian (Oryza sativa, Bengal Assam Aus Panel) rice diversity sets, highlighting valuable genetic variation accessible for breeding. Genome-wide association studies were performed across both species for these traits, representing the first documented attempt to characterize the genetic basis of photosynthetic heat tolerance in any species to date. A total of 133 candidate genes were highlighted. These were significantly enriched with genes whose predicted roles suggested influence on PSII activity and the response to stress. We discuss the most promising candidates for improving photosynthetic heat tolerance in rice.

Ecological and genomic vulnerability to climate change across native populations of Robusta coffee (Coffea canephora).

The assessment of population vulnerability under climate change is crucial for planning conservation as well as for ensuring food security. Coffea canephora is, in its native habitat, an understorey tree that is mainly distributed in the lowland rainforests of tropical Africa. Also known as Robusta, its commercial value constitutes a significant revenue for many human populations in tropical countries. Comparing ecological and genomic vulnerabilities within the species' native range can provide valuable insights about habitat loss and the species' adaptive potential, allowing to identify genotypes that may act as a resource for varietal improvement. By applying species distribution models, we assessed ecological vulnerability as the decrease in climatic suitability under future climatic conditions from 492 occurrences. We then quantified genomic vulnerability (or risk of maladaptation) as the allelic composition change required to keep pace with predicted climate change. Genomic vulnerability was estimated from genomic environmental correlations throughout the native range. Suitable habitat was predicted to diminish to half its size by 2050, with populations near coastlines and around the Congo River being the most vulnerable. Whole-genome sequencing revealed 165 candidate SNPs associated with climatic adaptation in C. canephora, which were located in genes involved in plant response to biotic and abiotic stressors. Genomic vulnerability was higher for populations in West Africa and in the region at the border between DRC and Uganda. Despite an overall low correlation between genomic and ecological vulnerability at broad scale, these two components of vulnerability overlap spatially in ways that may become damaging. Genomic vulnerability was estimated to be 23% higher in populations where habitat will be lost in 2050 compared to regions where habitat will remain suitable. These results highlight how ecological and genomic vulnerabilities are relevant when planning on how to cope with climate change regarding an economically important species.

MooSciTIC: Training of trainers in West African research and higher education.

The MooSciTIC project is a capacity-building initiative targeting West African research scientists and higher education teachers. The project aimed to improve the self-reliance of researchers and upgrade research practices by providing on-site summer schools on trans-disciplinary topics such as scientific writing, communication, and integrity. Here, we explain how this program was designed and implemented and share the positive responses from our trainees, hoping to inspire similar initiatives.

Redox-related gene expression and sugar accumulation patterns are altered in the edible inflorescence produced by the cultivated form of pacaya palm (Chamaedorea tepejilote).

BACKGROUND AND AIMS: The pacaya palm is a dioecious neotropical palm species that is exploited in Latin America for its male inflorescence, which is edible when immature. It is cultivated, in a non-intensive manner, in Guatemala, where a morphotype occurs that produces much larger, more highly branched inflorescences compared with wild palms. We sought to identify molecular factors underlying this phenotypic divergence, which is likely to be a product of domestication. METHODS: We performed RNA-seq-based studies on immature pacaya palm male inflorescences in order to identify genes that might be directly or indirectly affected in their expression in relation to domestication. We also measured the accumulation of a range of soluble sugar molecules to provide information on the biochemical status of the two different types of material. KEY RESULTS: A total of 408 genes were found to display significantly different expression levels between the wild and cultivated morphotypes. Three different functional categories were found to be enriched in the gene set that was upregulated in the cultivated morphotype: redox balance; secondary metabolism; and transport. Several sugars were found to accumulate at higher levels in inflorescences of the cultivated morphotype, in particular myo-inositol, fructose and glucose. CONCLUSIONS: The observed upregulation of redox-related genes in the cultivated morphotype is corroborated by the observation of higher myo-inositol accumulation, which has been shown to be associated with enhanced scavenging of reactive oxygen species in other plants and which may affect meristem activity.

Development and evaluation of a genome-wide Coffee 8.5K SNP array and its application for high-density genetic mapping and for investigating the origin of Coffea arabica L.

Coffee species such as Coffea canephora P. (Robusta) and C. arabica L. (Arabica) are important cash crops in tropical regions around the world. C. arabica is an allotetraploid (2n = 4x = 44) originating from a hybridization event of the two diploid species C. canephora and C. eugenioides (2n = 2x = 22). Interestingly, these progenitor species harbour a greater level of genetic variability and are an important source of genes to broaden the narrow Arabica genetic base. Here, we describe the development, evaluation and use of a single-nucleotide polymorphism (SNP) array for coffee trees. A total of 8580 unique and informative SNPs were selected from C. canephora and C. arabica sequencing data, with 40% of the SNP located in annotated genes. In particular, this array contains 227 markers associated to 149 genes and traits of agronomic importance. Among these, 7065 SNPs (~82.3%) were scorable and evenly distributed over the genome with a mean distance of 54.4 Kb between markers. With this array, we improved the Robusta high-density genetic map by adding 1307 SNP markers, whereas 945 SNPs were found segregating in the Arabica mapping progeny. A panel of C. canephora accessions was successfully discriminated and over 70% of the SNP markers were transferable across the three species. Furthermore, the canephora-derived subgenome of C. arabica was shown to be more closely related to C. canephora accessions from northern Uganda than to other current populations. These validated SNP markers and high-density genetic maps will be useful to molecular genetics and for innovative approaches in coffee breeding.

The coffee genome hub: a resource for coffee genomes.

The whole genome sequence of Coffea canephora, the perennial diploid species known as Robusta, has been recently released. In the context of the C. canephora genome sequencing project and to support post-genomics efforts, we developed the Coffee Genome Hub (http://coffee-genome.org/), an integrative genome information system that allows centralized access to genomics and genetics data and analysis tools to facilitate translational and applied research in coffee. We provide the complete genome sequence of C. canephora along with gene structure, gene product information, metabolism, gene families, transcriptomics, syntenic blocks, genetic markers and genetic maps. The hub relies on generic software (e.g. GMOD tools) for easy querying, visualizing and downloading research data. It includes a Genome Browser enhanced by a Community Annotation System, enabling the improvement of automatic gene annotation through an annotation editor. In addition, the hub aims at developing interoperability among other existing South Green tools managing coffee data (phylogenomics resources, SNPs) and/or supporting data analyses with the Galaxy workflow manager.

TOGGLE: toolbox for generic NGS analyses.

BACKGROUND: The explosion of NGS (Next Generation Sequencing) sequence data requires a huge effort in Bioinformatics methods and analyses. The creation of dedicated, robust and reliable pipelines able to handle dozens of samples from raw FASTQ data to relevant biological data is a time-consuming task in all projects relying on NGS. To address this, we created a generic and modular toolbox for developing such pipelines. RESULTS: TOGGLE (TOolbox for Generic nGs anaLysEs) is a suite of tools able to design pipelines that manage large sets of NGS softwares and utilities. Moreover, TOGGLE offers an easy way to manipulate the various options of the different softwares through the pipelines in using a single basic configuration file, which can be changed for each assay without having to change the code itself. We also describe one implementation of TOGGLE in a complete analysis pipeline designed for SNP discovery for large sets of genomic data, ready to use in different environments (from a single machine to HPC clusters). CONCLUSION: TOGGLE speeds up the creation of robust pipelines with reliable log tracking and data flow, for a large range of analyses. Moreover, it enables Biologists to concentrate on the biological relevance of results, and change the experimental conditions easily. The whole code and test data are available at https://github.com/SouthGreenPlatform/TOGGLE .

Haitian coffee agroforestry systems harbor complex arabica variety mixtures and under-recognized genetic diversity.

Though facing significant challenges, coffee (Coffea arabica) grown in Haitian agroforestry systems are important contributors to rural livelihoods and provide several ecosystem services. However, little is known about their genetic diversity and the variety mixtures used. In light of this, there is a need to characterize Haitian coffee diversity to help inform revitalization of this sector. We sampled 28 diverse farms in historically important coffee growing regions of northern and southern Haiti. We performed KASP-genotyping of SNP markers and HiPlex multiplex amplicon sequencing for haplotype calling on our samples, as well as several Ethiopian and commercial accessions from international collections. This allowed us to assign Haitian samples to varietal groups. Our analyses revealed considerable genetic diversity in Haitian farms, higher in fact than many farmers realized. Notably, genetic structure analyses revealed the presence of clusters related to Typica, Bourbon, and Catimor groups, another group that was not represented in our reference accession panel, and several admixed individuals. Across the study areas, we found both mixed-variety farms and monovarietal farms with the historical and traditional Typica variety. This study is, to our knowledge, the first to genetically characterize Haitian C. arabica variety mixtures, and report the limited cultivation of C. canephora (Robusta coffee) in the study area. Our results show that some coffee farms are repositories of historical, widely-abandoned varieties while others are generators of new diversity through genetic mixing.

Glutaredoxin regulation of primary root growth is associated with early drought stress tolerance in pearl millet.

Seedling root traits impact plant establishment under challenging environments. Pearl millet is one of the most heat and drought tolerant cereal crops that provides a vital food source across the sub-Saharan Sahel region. Pearl millet's early root system features a single fast-growing primary root which we hypothesize is an adaptation to the Sahelian climate. Using crop modeling, we demonstrate that early drought stress is an important constraint in agrosystems in the Sahel where pearl millet was domesticated. Furthermore, we show that increased pearl millet primary root growth is correlated with increased early water stress tolerance in field conditions. Genetics including genome-wide association study and quantitative trait loci (QTL) approaches identify genomic regions controlling this key root trait. Combining gene expression data, re-sequencing and re-annotation of one of these genomic regions identified a glutaredoxin-encoding gene PgGRXC9 as the candidate stress resilience root growth regulator. Functional characterization of its closest Arabidopsis homolog AtROXY19 revealed a novel role for this glutaredoxin (GRX) gene clade in regulating cell elongation. In summary, our study suggests a conserved function for GRX genes in conferring root cell elongation and enhancing resilience of pearl millet to its Sahelian environment.

Pearl millet is a staple food for over 90 million people living in regions of Africa and India that typically experience high temperatures and little rainfall. It was domesticated about 4,500 years ago in the Sahel region of West Africa and is one of the most heat and drought tolerant cereal crops worldwide. In most plants, organs known as roots absorb water and essential nutrients from the soil. Young pearl millet plants develop a fast-growing primary root, but it is unclear how this unique feature helps the crop to grow in hot and dry conditions. Using weather data collected from the Sahel over a 20-year period, Fuente, Grondin et al. predicted by modelling that early drought stress is the major factor limiting pearl millet growth and yield in this region. Field experiments found that plants with primary roots that grow faster within soil were better at tolerating early drought than those with slower growing roots. Further work using genetic approaches revealed that a gene known as PgGRXC9 promotes the growth of the primary root. To better understand how this gene works, the team examined a very similar gene in a well-studied model plant known as Arabidopsis. This suggested that PgGRXC9 helps the primary root to grow by stimulating cell elongation within the root. Since it is well adapted to dry conditions, pearl millet is expected to play an important role in helping agriculture adjust to climate change. The findings of Fuente, Grondin et al. may be used by plant breeders to create more resilient and productive varieties of pearl millet.

The genome and population genomics of allopolyploid Coffea arabica reveal the diversification history of modern coffee cultivars.

Coffea arabica, an allotetraploid hybrid of Coffea eugenioides and Coffea canephora, is the source of approximately 60% of coffee products worldwide, and its cultivated accessions have undergone several population bottlenecks. We present chromosome-level assemblies of a di-haploid C. arabica accession and modern representatives of its diploid progenitors, C. eugenioides and C. canephora. The three species exhibit largely conserved genome structures between diploid parents and descendant subgenomes, with no obvious global subgenome dominance. We find evidence for a founding polyploidy event 350,000-610,000 years ago, followed by several pre-domestication bottlenecks, resulting in narrow genetic variation. A split between wild accessions and cultivar progenitors occurred ~30.5 thousand years ago, followed by a period of migration between the two populations. Analysis of modern varieties, including lines historically introgressed with C. canephora, highlights their breeding histories and loci that may contribute to pathogen resistance, laying the groundwork for future genomics-based breeding of C. arabica.

BAC-end sequences analysis provides first insights into coffee (Coffea canephora P.) genome composition and evolution.

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.

Phylogeography and niche modelling of the relict plant Amborella trichopoda (Amborellaceae) reveal multiple Pleistocene refugia in New Caledonia.

Amborella trichopoda Baill. (Amborellaceae, Amborellales), the sole living member of the sister group to all other extant angiosperms, is endemic to New Caledonia. We addressed the intraspecific phylogeography of Amborella by investigating whether its present population genetic structure could be related to its current and past habitats. We found moderate range-wide genetic diversity based on nuclear microsatellite data and detected four well-differentiated, geographically distinct genetic groups using Bayesian clustering analyses. We modelled the ecological niche of Amborella based on the current climatic and environmental conditions. The predictive ability of the model was very good throughout the Central East mainland zone, but Amborella was predicted in the northern part of the island where this plant has not been reported. Furthermore, no significant barrier was detected based on habitat suitability that could explain the genetic differentiation across the area. Conversely, we found that the main genetic clusters could be related to the distribution of the suitable habitat at the last glacial maximum (LGM, c. 21,000 years BP), when Amborella experienced a dramatic 96.5% reduction in suitable area. At least two lineages survived in distinct putative refugia located in the Massif des Lèvres and in the vicinity of Mount Aoupinié. Our findings finally confirmed the importance of LGM rainforest refugia in shaping the current intra- and interspecific diversity in New Caledonian plants and revealed the possibility of an as yet unreported refugium. The combination of niche modelling and population genetics thereby offered novel insight into the biogeographical history of an emblematic taxon.

Genome-wide association analysis identifies natural allelic variants associated with panicle architecture variation in African rice, Oryza glaberrima Steud.

African rice (Oryza glaberrima Steud), a short-day cereal crop closely related to Asian rice (Oryza sativa L.), has been cultivated in Sub-Saharan Africa for ∼ 3,000 years. Although less cultivated globally, it is a valuable genetic resource in creating high-yielding cultivars that are better adapted to diverse biotic and abiotic stresses. While inflorescence architecture, a key trait for rice grain yield improvement, has been extensively studied in Asian rice, the morphological and genetic determinants of this complex trait are less understood in African rice. In this study, using a previously developed association panel of 162 O. glaberrima accessions and new SNP variants characterized through mapping to a new version of the O. glaberrima reference genome, we conducted a genome-wide association study of four major morphological panicle traits. We have found a total of 41 stable genomic regions that are significantly associated with these traits, of which 13 co-localized with previously identified QTLs in O. sativa populations and 28 were unique for this association panel. Additionally, we found a genomic region of interest on chromosome 3 that was associated with the number of spikelets and primary and secondary branches. Within this region was localized the O. sativa ortholog of the PHYTOCHROME B gene (Oglab_006903/OgPHYB). Haplotype analysis revealed the occurrence of natural sequence variants at the OgPHYB locus associated with panicle architecture variation through modulation of the flowering time phenotype, whereas no equivalent alleles were found in O. sativa. The identification in this study of genomic regions specific to O. glaberrima indicates panicle-related intra-specific genetic variation in this species, increasing our understanding of the underlying molecular processes governing panicle architecture. Identified candidate genes and major haplotypes may facilitate the breeding of new African rice cultivars with preferred panicle traits.

An improved assembly of the pearl millet reference genome using Oxford Nanopore long reads and optical mapping.

Pearl millet (Pennisetum glaucum (L.)) R. Br. syn. Cenchrus americanus (L.) Morrone) is an important crop in South Asia and sub-Saharan Africa which contributes to ensuring food security. Its genome has an estimated size of 1.76 Gb and displays a high level of repetitiveness above 80%. A first assembly was previously obtained for the Tift 23D2B1-P1-P5 cultivar genotype using short-read sequencing technologies. This assembly is, however, incomplete and fragmented with around 200 Mb unplaced on chromosomes. We report here an improved quality assembly of the pearl millet Tift 23D2B1-P1-P5 cultivar genotype obtained with an approach combining Oxford Nanopore long reads and Bionano Genomics optical maps. This strategy allowed us to add around 200 Mb at the chromosome-level assembly. Moreover, we strongly improved continuity in the order of the contigs and scaffolds within the chromosomes, particularly in the centromeric regions. Notably, we added more than 100 Mb around the centromeric region on chromosome 7. This new assembly also displayed a higher gene completeness with a complete BUSCO score of 98.4% using the Poales database. This more complete and higher quality assembly of the Tift 23D2B1-P1-P5 genotype now available to the community will help in the development of research on the role of structural variants and more broadly in genomics studies and the breeding of pearl millet.

Ancestral synteny shared between distantly-related plant species from the asterid (Coffea canephora and Solanum Sp.) and rosid (Vitis vinifera) clades.

BACKGROUND: Coffee trees (Rubiaceae) and tomato (Solanaceae) belong to the Asterid clade, while grapevine (Vitaceae) belongs to the Rosid clade. Coffee and tomato separated from grapevine 125 million years ago, while coffee and tomato diverged 83-89 million years ago. These long periods of divergent evolution should have permitted the genomes to reorganize significantly. So far, very few comparative mappings have been performed between very distantly related species belonging to different clades. We report the first multiple comparison between species from Asterid and Rosid clades, to examine both macro-and microsynteny relationships. RESULTS: Thanks to a set of 867 COSII markers, macrosynteny was detected between coffee, tomato and grapevine. While coffee and tomato genomes share 318 orthologous markers and 27 conserved syntenic segments (CSSs), coffee and grapevine also share a similar number of syntenic markers and CSSs: 299 and 29 respectively. Despite large genome macrostructure reorganization, several large chromosome segments showed outstanding macrosynteny shedding new insights into chromosome evolution between Asterids and Rosids. We also analyzed a sequence of 174 kb containing the ovate gene, conserved in a syntenic block between coffee, tomato and grapevine that showed a high-level of microstructure conservation. A higher level of conservation was observed between coffee and grapevine, both woody and long life-cycle plants, than between coffee and tomato. Out of 16 coffee genes of this syntenic segment, 7 and 14 showed complete synteny between coffee and tomato or grapevine, respectively. CONCLUSIONS: These results show that significant conservation is found between distantly related species from the Asterid (Coffea canephora and Solanum sp.) and Rosid (Vitis vinifera) clades, at the genome macrostructure and microstructure levels. At the ovate locus, conservation did not decline in relation to increasing phylogenetic distance, suggesting that the time factor alone does not explain divergences. Our results are considerably useful for syntenic studies between supposedly remote species for the isolation of important genes for agronomy.

Microsatellite markers for Amborella (Amborellaceae), a monotypic genus endemic to New Caledonia.

PREMISE OF THE STUDY: Informative markers are required for assessing the diversity of Amborella trichopoda, the only species of its order, endemic to New Caledonia and considered to be the sister species to all flowering plants. Therefore, expressed sequence tag (EST)-based microsatellite markers were developed. • METHODS AND RESULTS: Fifty-five microsatellite loci were characterized in 14896 putative unigenes, which were generated by assembling A. trichopoda ESTs from the public sequence database. Seventeen markers revealed polymorphism in 80 adult shrubs from three populations. The number of alleles per locus ranged from two to 12, with a total of 132 alleles scored. The mean expected heterozygosity per population ranged from 0.336 to 0.567. • CONCLUSIONS: These markers offer an appropriate amount of variation to investigate genetic diversity structure, gene flow, and other conservation issues.

Aquaporins are main contributors to root hydraulic conductivity in pearl millet [Pennisetum glaucum (L) R. Br.].

Pearl millet is a key cereal for food security in arid and semi-arid regions but its yield is increasingly threatened by water stress. Physiological mechanisms relating to conservation of soil water or increased water use efficiency can alleviate that stress. Aquaporins (AQP) are water channels that mediate root water transport, thereby influencing plant hydraulics, transpiration and soil water conservation. However, AQP remain largely uncharacterized in pearl millet. Here, we studied AQP function in root water transport in two pearl millet lines contrasting for water use efficiency (WUE). We observed that these lines also contrasted for root hydraulic conductivity (Lpr) and AQP contribution to Lpr. The line with lower WUE showed significantly higher AQP contribution to Lpr. To investigate AQP isoforms contributing to Lpr, we developed genomic approaches to first identify the entire AQP family in pearl millet and secondly, characterize the plasma membrane intrinsic proteins (PIP) gene expression profile. We identified and annotated 33 AQP genes in pearl millet, among which ten encoded PIP isoforms. PgPIP1-3 and PgPIP1-4 were significantly more expressed in the line showing lower WUE, higher Lpr and higher AQP contribution to Lpr. Overall, our study suggests that the PIP1 AQP family are the main regulators of Lpr in pearl millet and may possibly be associated with mechanisms associated to whole plant water use. This study paves the way for further investigations on AQP functions in pearl millet hydraulics and adaptation to environmental stresses.

Genome Wide Association Study Pinpoints Key Agronomic QTLs in African Rice Oryza glaberrima.

BACKGROUND: African rice, Oryza glaberrima, is an invaluable resource for rice cultivation and for the improvement of biotic and abiotic resistance properties. Since its domestication in the inner Niger delta ca. 2500 years BP, African rice has colonized a variety of ecologically and climatically diverse regions. However, little is known about the genetic basis of quantitative traits and adaptive variation of agricultural interest for this species. RESULTS: Using a reference set of 163 fully re-sequenced accessions, we report the results of a Genome Wide Association Study carried out for African rice. We investigated a diverse panel of traits, including flowering date, panicle architecture and resistance to Rice yellow mottle virus. For this, we devised a pipeline using complementary statistical association methods. First, using flowering time as a target trait, we found several association peaks, one of which co-localised with a well described gene in the Asian rice flowering pathway, OsGi, and identified new genomic regions that would deserve more study. Then we applied our pipeline to panicle- and resistance-related traits, highlighting some interesting genomic regions and candidate genes. Lastly, using a high-resolution climate database, we performed an association analysis based on climatic variables, searching for genomic regions that might be involved in adaptation to climatic variations. CONCLUSION: Our results collectively provide insights into the extent to which adaptive variation is governed by sequence diversity within the O. glaberrima genome, paving the way for in-depth studies of the genetic basis of traits of interest that might be useful to the rice breeding community.

Transcriptome analysis during somatic embryogenesis of the tropical monocot Elaeis guineensis: evidence for conserved gene functions in early development.

With the aim of understanding the molecular mechanisms underlying somatic embryogenesis (SE) in oil palm, we examined transcriptome changes that occur when embryogenic suspension cells are initiated to develop somatic embryos. Two reciprocal suppression subtractive hybridization (SSH) libraries were constructed from oil palm embryogenic cell suspensions: one in which embryo development was blocked by the presence of the synthetic auxin analogue 2,4-dichlorophenoxyacetic acid (2,4-D: ) in the medium (proliferation library); and another in which cells were stimulated to form embryos by the removal of 2,4-D: from the medium (initiation library). A total of 1867 Expressed Sequence Tags (ESTs) consisting of 1567 potential unigenes were assembled from the two libraries. Functional annotation indicated that 928 of the ESTs correspond to proteins that have either no similarity to sequences in public databases or are of unknown function. Gene Ontology (GO) terms assigned to the two EST populations give clues to the underlying molecular functions, biological processes and cellular components involved in the initiation of embryo development. Macroarrays were used for transcript profiling the ESTs during SE. Hierarchical cluster analysis of differential transcript accumulation revealed 4 distinct profiles containing a total of 192 statistically significant developmentally regulated transcripts. Similarities and differences between the global results obtained with in vitro systems from dicots, monocots and gymnosperms will be discussed.

MoccaDB - an integrative database for functional, comparative and diversity studies in the Rubiaceae family.

BACKGROUND: In the past few years, functional genomics information has been rapidly accumulating on Rubiaceae species and especially on those belonging to the Coffea genus (coffee trees). An increasing number of expressed sequence tag (EST) data and EST- or genomic-derived microsatellite markers have been generated, together with Conserved Ortholog Set (COS) markers. This considerably facilitates comparative genomics or map-based genetic studies through the common use of orthologous loci across different species. Similar genomic information is available for e.g. tomato or potato, members of the Solanaceae family. Since both Rubiaceae and Solanaceae belong to the Euasterids I (lamiids) integration of information on genetic markers would be possible and lead to more efficient analyses and discovery of key loci involved in important traits such as fruit development, quality, and maturation, or adaptation. Our goal was to develop a comprehensive web data source for integrated information on validated orthologous markers in Rubiaceae. DESCRIPTION: MoccaDB is an online MySQL-PHP driven relational database that houses annotated and/or mapped microsatellite markers in Rubiaceae. In its current release, the database stores 638 markers that have been defined on 259 ESTs and 379 genomic sequences. Marker information was retrieved from 11 published works, and completed with original data on 132 microsatellite markers validated in our laboratory. DNA sequences were derived from three Coffea species/hybrids. Microsatellite markers were checked for similarity, in vitro tested for cross-amplification and diversity/polymorphism status in up to 38 Rubiaceae species belonging to the Cinchonoideae and Rubioideae subfamilies. Functional annotation was provided and some markers associated with described metabolic pathways were also integrated. Users can search the database for marker, sequence, map or diversity information through multi-option query forms. The retrieved data can be browsed and downloaded, along with protocols used, using a standard web browser. MoccaDB also integrates bioinformatics tools (CMap viewer and local BLAST) and hyperlinks to related external data sources (NCBI GenBank and PubMed, SOL Genomic Network database). CONCLUSION: We believe that MoccaDB will be extremely useful for all researchers working in the areas of comparative and functional genomics and molecular evolution, in general, and population analysis and association mapping of Rubiaceae and Solanaceae species, in particular.