Sequencing the banana genome (W069)

Bananas (Musa) are the fourth most important crop in developing countries. They are important as an export crop but also play a major role in local food security. Crops of Musa are susceptible to an ever increasing range of diseases requiring massive use of pesticides that have dramatic environmental and health impacts and threaten the sustainability of the crop. There is thus an urgent need for a wider diversity of genetically improved banana cultivars with more robust disease resistance, increased productivity and better adaptability to a large range of growing conditions. The production of export dessert bananas relies on very few related genotypes of the Cavendish subgroup with an AAA genome constitution. However, numerous dessert and cooking types with an AAA, AAB (including plantain) or ABB genome constitution are grown for local consumption. The Musa accession selected for sequencing is a doubled haploid of the accession 'Pahang' (DH Pahang). This accession belongs to the Musa acuminata species (AA genome) malaccensis subspecies. We generated 20x coverage using paired and single 454 reads, complemented by Sanger BESs and ~50 x coverage of Illumina shotgun data. The assembly was performed with Newbler, and the scaffolds were anchored to a genetic map. Genes were predicted using a reconciliation approach taking ESTs, protein sequences and ab initio data as input. A genetic map of the parent accession 'Pahang' was developed with SSR and DArT markers to assemble the scaffolds in pseudo-molecules. (Texte intégral

The Banana (Musa acuminata) genome and the evolution of monocotyledonous plants : W401

Bananas (Musa spp.), including dessert and cooking types, are giant perennial monocotyledonous herbs of the Zingiberales order, a sister group to the well-studied Poales. We sequenced and assembled the 520 Mb genome of a doubled-haploid of the accession 'Pahang'. This accession belongs to the Musa acuminata species (AA genome) malaccensis subspecies. We detected three rounds of whole-genome duplications in the Musa lineage, independently of those previously described in the Poales lineage and the one we detected in the Arecales lineage. This first monocotyledon high-continuity whole-genome sequence reported outside Poales represents an essential bridge for comparative genome analysis in plants. As such, it sheds new light on the monocotyledon lineage, reveals Poaceae specific features and has led to the discovery of conserved noncoding sequences predating monocotyledon-eudicotyledon divergence. The complete list of authors involved in this work can be found in D'Hont et al. Nature. 2012 Aug 9; 488(7410):213-7 (Résumé d'auteur

Fluorescence in situ hybridization in sugarcane or fish-ing in the genomic wilderness. [MO06]

Cytogenetics applied to sugarcane has brought our fundamental understanding of the sugarcane genome to a new level. In the mid-nineties, Genomic in situ Hybridisation (GISH) was first applied to sugarcane to determine the specific composition of the modern cultivar R570. GISH revealed the chromosomal composition of R570 was 80% Saccharum officinarum, 10% S. spontaneum and 10% of recombined chromosomes. The Australian counterpart Q165, revealed a slightly different species composition as 75%, 15% and 10%, respectively. Both R570 and Q165 genetic maps have portrayed a partial coverage of linkage groups (LG) despite the large number of molecular markers invested in the maps. It also shows that S. spontaneum chromosomes seem to have a better vertical coverage than S. officinarum chromosomes as the S. spontaneum genome is more polymorphic. To gain a better understanding of the genome composition in terms of LG number per homology group (HG) and species attribution of the LG, we applied BAC-FISH to sugarcane. Bacterial Artificial Chromosomes (BAC) consist of large chromosome segments (around 100kb). BAC from the Sorghum or Saccharum genomes were used as anchorage points on the sugarcane cultivars to identify homologous/homeologous chromosomes for each HG. We will present some examples of results of BAC-FISH applied to several cultivars for at least 4 different HG. The determination and comparison of the number of chromosomes per HG to the number of LG from the genetic maps will determine the saturation level of the genetic maps. This will help us to obtain critical knowledge of the horizontal chromosome distribution for a particular cultivar and compare its structure to another cultivar. Eventually we will have a better understanding of the distribution of the chromosomes during crossing and this will help breeders to make more informed and targeted choices in their selection programs. (Texte intégral

Characterization of large chromosomal structural variations between Musa acuminata sub-species by NGS re-sequencing

M. acuminata has evolved in several sub-species that have been geographically isolated on various archipelagos, and have accumulated large structural variations. Domestication of banana involving hybridization between these subspecies has been made possible by human migration and selection of diploid and triploid inter-sub-specific hybrids with seedless parthenocarpic fruits. Large structural variations within M. acuminata have been hypothesized based on chromosome pairing analysis and genetic mapping. These structural variations are suspected to be at least partially responsible for M. acuminata hybrids sterility. This sterility allows the production of seedless fruits but complicates breeding programs by limiting crossing possibilities. These structural variations also impact chromosomal segregation and recombination, complicating the transmission of agronomical traits of interest and genetic analyses. There is thus a strong need to characterize these structural variations within the Musa acuminata sub-species. A re-sequencing approach to characterize these structural variations is currently being tested. lt is based on paired-end sequencing of genomic fragments of known sizes from banana accessions, and comparison to the Musa acuminata reference genome to detect discrepancies in paired-read mapping. A specific bioinformatics pipeline has been developed to detect different types of structural variations, and validated on simulated data. The pipeline is currently being tested on diploid banana accessions using various DNA fragment sizes (5kb to 15kb) in order to optimize the detection of large structural variations. (Résumé d'auteur

Molecular breeding of sugarcane using linkage disequilibrium maps and quantitative trait alleles : [Abstract W247]

Sugarcane breeding generally involves forms of recurrent selection using overlapping generations of parent genotypes, with the number of parents used in the order of 100 to 400, depending on the size of the breeding program. Genetic maps of one or a few genotypes are thus not very useful in routine breeding applications, as they provide no information on the allelic variation at important loci within the breeding population. In order to integrate mapping, marker discovery and conventional breeding we have developed methods (i) to create population-level maps of haplotypes in linkage disequilibrium within the breeding population, (ii) to identify potentially useful quantitative trait alleles (QTAs) through association analysis, and (iii) to predict to performance of progeny of bi-parental crosses from the marker (QTA) profile of the parents. The usefulness of the approach has been empirically verified in an experiment which demonstrated that the performance of progeny is better predicted by parental QTAs than by parent phenotype (h2 = 0.82, versus 0.57). By examining the population-level haplotype map, cryptic population structure caused by the complex linkage arrangements that can exist in polyploids could be detected, and accounting for this structure/linkage can further improve the effectiveness of molecular breeding. Additional uses of the map, such as identifying the ancestral origins of haplotypes, and detecting signatures of selection and recombination over several generations of breeding will be demonstrated. (Texte intégral

GNPAnnot community annotation system applied to sugarcane bac clone sequences (W572)

A large amount of data is being produced by current genome sequencing projects. Sequence annotations and analyses need to be organized into databases and widely accessible. Like other species, sugarcane would benefit from centralized and innovative systems to study its genome. The GNPAnnot community annotation system (CAS) could be particularly relevant to the SUGESI sequencing project. It consists in a system for structural and functional annotations supported by comparative genomics allowing both automatic predictions and manual curations of genes and transposable elements. The core of the GNPAnnot CAS dedicated to tropical plants is made of GMOD components.The Chado database can be browsed using the Generic Genome Browser (GBrowse) which provides links to genome editors (ie. Artemis and Apollo). We developed the Chado controller in order to manage public and private annotation projects. It also provides an annotation history page for each gene or transposable element and an annotation inspector that automates several tasks and reports annotation mistakes. GNPAnnot CAS has already been used to annotate sugarcane BAC clones sequences and could be useful to facilitate the annotation of novel sugarcane sequences. (Résumé d'auteur

Detailed analyses of 12 hom(oe)ologous chromosome segments in the highly polyploid sugarcane genome

Modern sugarcane cultivars (Saccharum spp.) are recognized as the crop with the most complex genome studied to date, mainly due to the very high level of vertical redundancy (2n = ca 12x = ca 120), together with an interspecific origin. They are derived from hybridization, performed by breeders a century ago, between two autopolyploid species, namely S. officinarum (domesticated) and S. spontaneum (wild species, 2n=5x=40 to 16x=128). To investigate the impact of polyploidization on its genome organization and more widely on its performance and plasticity, we finely analyzed the structural organization of hom(oe)ologous chromosomes. Thirty-three homoeologous BAC clones from four regions of the sugarcane R570 genome were identified, sequenced, finely annotated and compared, representing more than 3 Mb of sugarcane DNA sequence. For all four regions, almost perfect gene colinearity and high gene structure and sequence conservation were observed, confirming previous preliminary analyses on two of these regions. Moreover, the vast majority of the homoeologous genes were predicted, based on their structure, to be functional and showed signs of evolving under purifying selection. For one of the region carrying the Adh1 gene, we extended the homoeologous series to 13 hom(oe)ologous chromosome segments. Gene similarity and patterns of transposable element insertions are currently being analyzed in order to determine the origin (S. officinarum vs S. spontaneum) and the evolutionary dynamics of these hom(oe)ologous regions. (Résumé d'auteur

The Sugarcane genome sequencing effort: An overview of the strategy, goals and existing data : [Abstract W538]

Sugarcane is a major feedstock used for biofuel production worldwide. Sugarcane cultivars (Saccharum spp) are derived from interspecific hybridization obtained a century ago by crossing Saccharum officinarum (2n=8x=80) and S. spontaneum (2n=5x=40 to 2n=16x=128). The challenge in a sugarcane genome sequencing project is the size (10 Gb) and complexity of its genome structure that is highly polyploid and aneuploid (2n= ca 110 to 120) with a complete set of homo(eo)logous genes predicted to range from 10 to 12 copies (alleles). A initial strategy is to capture much of the gene-rich recombinationally-active euchromatin. The Sugarcane Genome Sequencing Initiative (SUGESI) was envisaged to join efforts to produce a reference sequence of one sugarcane cultivar using a combination of approaches, including BAC sequencing and whole genome shot-gun approaches. Cultivar R570 was chosen since it is the most intensively characterized to date. We expect that around 4-5 thousand BAC sequences can cover the monoploid euchromatic genome of this cultivar. BAC selection is underway using overgo and EST hybridization data. A next step is to sequence cultivars of interest to breeding programs. Under the shot-gun approach gene rich regions are being targeted for genotypes that are parents of mapping populations. This should allow the identification of very large numbers of polymorphic markers that are expected to assist genome assembly. Pilot experiments are underway to define the best technologies for gene-rich region or promoter identification. A database is under construction (http://sugarcanegenome.org). The initiative is led by researchers in Australia, Brazil, China, France, South Africa and United States. (Texte intégral

Comparison of hom(oe)ologous chromosome segments in the highly polyploid interspecific genome of sugarcane

Modern sugarcane cultivars (Saccharum spp.) present one of the most complex crop genome studied to date, mainly due to a very high degree of polyploidy (2n = 12x = 120), and an interspecific origin from two autopolyploid species, namely S. officinarum and S. spontaneum. To investigate the impact of polyploidization on the sugarcane genome organization and more widely on its performance and plasticity, we finely analyzed the structural organization of hom(oe)ologous chromosome segments. Twenty-seven homoeologous BAC clones from three distinct regions, carrying the genes Adh1 (13 hom(oe)ologous chromosome segments), PST2a (10 hom(oe)ologous chromosome segments) and CAD2 (4 hom(oe)ologous chromosome segments), were identified, sequenced, finely annotated and compared, representing more than 2.5 Mb of sugarcane DNA sequence. A very high gene colinearity, gene structure and sequence conservation (98.1% of average nucleotide sequence identity for the coding sequence, and 93.3% for the aligned part of the introns) was observed among all hom(oe)ologous chromosome segments, confirming preliminary observations. Based on their structure, the homoeologous genes were predicted to be functional and the vast majority of them showed signs of evolving under purifying selection. Colinearity between hom(oe)ologous chromosomes was also extended to many intergenic regions and transposable elements. Divergence between hom(oe)ologous genes and patterns of transposable element insertions are currently being analyzed in order to infer the origin (S. officinarum vs S. spontaneum) of the chromosome segments. The high level of gene colinearity and structure conservation has implication regarding whole genome sequencing strategy of this complex genome, since it suggests that one chromosome segment could serve as reference for the other hom(oe)ologous chromosome segments regarding gene content. The maintenance of a broad set of functional alleles, that we described, may be involved in the high phenotypic plasticity and wide adaptation of sugarcane. (Résumé d'auteur

Paleoploidization events in the Musa (banana) lineage

Bananas (Musa spp.) are giant perennial monocotyledonous herbs of the order Zingiberales, a sister group to the well-studied Poales. Cultivars are mainly triploid, from inter(sub)specific origin and clonally propagated. We sequenced the genome of a Musa acuminata doubled-haploid genotype (Pahang-HD) providing the first monocotyledon high-continuity whole-genome sequence reported outside Poales. The analysis of the genome revealed three rounds of whole genome duplications (WGD), denoted as alpha, beta and gamma (from the most recent event to the oldest). Based on Ks analyses and synteny relationships, twelve beta Musa ancestral blocks were constructed, representing the ancestral genome before alpha and beta duplications. Comparative genomics and phylogenetic approaches revealed that these three WGDs occurred in the Musa lineage independently of those described in the Poales lineage and the one that we detected in the Arecales lineage. Following WGDs, that are particularly frequent in the flowering plant lineages, most duplicated genes are deleted by intrachromosomal recombination, a process referred to as fractionation. We are currently analyzing the fractionation pattern following the Musa polyploidization events. Finally, this Musa reference sequence represents an invaluable reference for studying monocot evolution and associated genomic changes. (Résumé d'auteur