Georgios J Pappas

Insights into the Musa genome: Syntenic relationships to rice and between Musa species

BackgroundMusa species (Zingiberaceae, Zingiberales) including bananas and plantains are collectively the fourth most important crop in developing countries. Knowledge concerning Musa genome structure and the origin of distinct cultivars has greatly increased over the last few years. Until now, however, no large-scale analyses of Musa genomic sequence have been conducted. This study compares genomic sequence in two Musa species with orthologous regions in the rice genome.ResultsWe produced 1.4 Mb of Musa sequence from 13 BAC clones, annotated and analyzed them along with 4 previously sequenced BACs. The 443 predicted genes revealed that Zingiberales genes share GC content and distribution characteristics with eudicot and Poaceae genomes. Comparison with rice revealed microsynteny regions that have persisted since the divergence of the Commelinid orders Poales and Zingiberales at least 117 Mya. The previously hypothesized large-scale duplication event in the common ancestor of major cereal lineages within the Poaceae was verified. The divergence time distributions for Musa-Zingiber (Zingiberaceae, Zingiberales) orthologs and paralogs provide strong evidence for a large-scale duplication event in the Musa lineage after its divergence from the Zingiberaceae approximately 61 Mya. Comparisons of genomic regions from M. acuminata and M. balbisiana revealed highly conserved genome structure, and indicated that these genomes diverged circa 4.6 Mya.ConclusionThese results point to the utility of comparative analyses between distantly-related monocot species such as rice and Musa for improving our understanding of monocot genome evolution. Sequencing the genome of M. acuminata would provide a strong foundation for comparative genomics in the monocots. In addition a genome sequence would aid genomic and genetic analyses of cultivated Musa polyploid genotypes in research aimed at localizing and cloning genes controlling important agronomic traits for breeding purposes.

Diversity of soil fungal communities of Cerrado and its closely surrounding agriculture fields

Cerrado is a savanna-like region that covers a large area of Brazil. Despite its biological importance, the Cerrado has been the focus of few microbial diversity studies. A molecular approach was chosen to characterize the soil fungal communities in four areas of the Cerrado biome: a native Cerrado, a riverbank forest, an area converted to a soybean plantation, and an area converted to pasture. Global diversity of fungal communities in each area was assessed through Ribosomal intergenic spacer analysis which revealed remarkable differences among the areas studied. Sequencing of approximately 200 clones containing 18S rDNA sequences from each library was performed and, according to the genetic distance between sequences, these were assigned to operational taxonomic units (OTUs). A total of 75, 85, 85, and 70 OTUs were identified for the native Cerrado, riverbank forest, pasture, and soybean plantation, respectively. Analysis of sequences using a similarity cutoff value of 1% showed that the number of OTUs for the native Cerrado area was reduced by 35%; for the soybean plantation, a reduction by more than 50% was observed, indicating a reduction in fungal biodiversity associated with anthropogenic activity. This is the first study demonstrating the anthropogenic impact on Cerrado soil fungal diversity.

Characterization of novel microsatellite markers in Musa acuminata subsp. burmannicoides , var. Calcutta 4

BackgroundBanana is a nutritionally important crop across tropical and sub-tropical countries in sub-Saharan Africa, Central and South America and Asia. Although cultivars have evolved from diploid, triploid and tetraploid wild Asian species of Musa acuminata (A genome) and Musa balbisiana (B genome), many of todays commercial cultivars are sterile triploids or diploids, with fruit developing via parthenocarpy. As a result of restricted genetic variation, improvement has been limited, resulting in a crop frequently lacking resistance to pests and disease. Considering the importance of molecular tools to facilitate development of disease resistant genotypes, the objectives of this study were to develop polymorphic microsatellite markers from BAC clone sequences for M. acuminata subsp. burmannicoides, var. Calcutta 4. This wild diploid species is used as a donor cultivar in breeding programs as a source of resistance to diverse biotic stresses.FindingsMicrosatellite sequences were identified from five Calcutta 4 BAC consensi datasets. Specific primers were designed for 41 loci. Isolated di-nucleotide repeat motifs were the most abundant, followed by tri-nucleotides. From 33 tested loci, 20 displayed polymorphism when screened across 21 diploid M. acuminata accessions, contrasting in resistance to Sigatoka diseases. The number of alleles per SSR locus ranged from two to four, with a total of 56. Six repeat classes were identified, with di-nucleotides the most abundant. Expected heterozygosity values for polymorphic markers ranged from 0.31 to 0.75.ConclusionsThis is the first report identifying polymorphic microsatellite markers from M. acuminata subsp. burmannicoides, var. Calcutta 4 across accessions contrasting in resistance to Sigatoka diseases. These BAC-derived polymorphic microsatellite markers are a useful resource for banana, applicable for genetic map development, germplasm characterization, evolutionary studies and marker assisted selection for traits.

Genome-wide SNP discovery from a pooled sample of accessions of the biofuel plant Jatropha curcas based on whole-transcriptome Illumina resequencing

Background Jatropha curcas(JC) is an oil-rich, drought-tolerant perennial shrub of the Euphorbiaceae family widely dispersed throughout the world. Thought to be native to Central America, it has been the object of an increasing number of studies in recent years for it exhibits a number of appealing attributes as a promising source of biodiesel. Although its undomesticated nature and preferential outcrossed mating system would suggest a high degree of genetic variation to be exploited in breeding, studies have shown limited genetic diversity in the existing germplasm collections [1]. In spite of the increased interest in this bioenergy plant, challenges still exist to turn this species into a genuine crop and improved varieties that consolidate desirable traits are not yet available, making JC large scale plantation an uncertain business [2]. Genomic studies to potentially assist JC breeding efforts have started in the last few years. JC is diploid (2n=22), with a haploid genome size estimated at 416 Mbp [3]. EST databases focusing on gene discovery were constructed [4]and a draft genome sequence was recently published covering 285 Mbp (~68%) of the genome in 120,586 contigs with 40,929 predicted gene models [5]. The focus of our work with JC is to provide effective tools to accelerate breeding through Genomic Selection (GS) [6] and to help assess the levels, organization and enrichment strategies of genetic diversity in germplasm banks and breeding populations. To this end we have started the development of SNP markers. Available EST databases built from single individual plants do not provide the necessary sequence diversity for SNP discovery.In this work we report on the discovery of a set of SNPs for JC derived from a pool of genetically diverse accessions using Illumina sequencing and a SNP selection pipeline recently described [7].

Genetic variation and recombination of RdRp and HSP 70h genes of Citrus tristeza virus isolates from orange trees showing symptoms of citrus sudden death disease.

BackgroundCitrus sudden death (CSD), a disease that rapidly kills orange trees, is an emerging threat to the Brazilian citrus industry. Although the causal agent of CSD has not been definitively determined, based on the diseases distribution and symptomatology it is suspected that the agent may be a new strain of Citrus tristeza virus (CTV). CTV genetic variation was therefore assessed in two Brazilian orange trees displaying CSD symptoms and a third with more conventional CTV symptoms.ResultsA total of 286 RNA-dependent-RNA polymerase (RdRp) and 284 heat shock protein 70 homolog (HSP70h) gene fragments were determined for CTV variants infecting the three trees. It was discovered that, despite differences in symptomatology, the trees were all apparently coinfected with similar populations of divergent CTV variants. While mixed CTV infections are common, the genetic distance between the most divergent population members observed (24.1% for RdRp and 11.0% for HSP70h) was far greater than that in previously described mixed infections. Recombinants of five distinct RdRp lineages and three distinct HSP70h lineages were easily detectable but respectively accounted for only 5.9 and 11.9% of the RdRp and HSP70h gene fragments analysed and there was no evidence of an association between particular recombinant mosaics and CSD. Also, comparisons of CTV population structures indicated that the two most similar CTV populations were those of one of the trees with CSD and the tree without CSD.ConclusionWe suggest that if CTV is the causal agent of CSD, it is most likely a subtle feature of population structures within mixed infections and not merely the presence (or absence) of a single CTV variant within these populations that triggers the disease.

SisGen: A CORBA---Based Data Management Program for DNA Sequencing Projects

Biological data deluge has challenged researchers over the last decade. Expressed sequence tag (EST) analyzes provide a rapid and economical means to identify candidate genes, gene expression profiles in different cell conditions, as well as functional annotation of putative gene products. Although EST analysis tools are publicly available there is still a lack of comprehensive data analysis and management programs. This work presents SisGen , an integrated software system capable of efficiently managing multi---user genomic projects. SisGen is a Java client---server application that uses CORBA as a middleware in a multi---layer architecture. The software integrates data management an annotation pipeline in a rich graphical visualization environment. The architectural design is presented and highlights the advantages in terms of portability, interconnectivity, modularity and user interface that can be achieved with this concept.

ConSSeq: a web-based application for analysis of amino acid conservation based on HSSP database and within context of structure

SUMMARY A web-based application to analyze protein amino acids conservation-Consensus Sequence (ConSSeq) is presented. ConSSeq graphically represents information about amino acid conservation based on sequence alignments reported in homology-derived structures of proteins. Beyond the relative entropy for each position in the alignment, ConSSeq also presents the consensus sequence and information about the amino acids, which are predominant at each position of the alignment. ConSSeq is part of the STING Millennium Suite and is implemented as a Java Applet. AVAILABILITY http://sms.cbi.cnptia.embrapa.br/SMS/STINGm/consseq/, http://trantor.bioc.columbia.edu/SMS/STINGm/consseq/, http://mirrors.rcsb.org//SMS/STINGm/consseq/, http://www.es.embnet.org/SMS/STINGm/consseq/ and http://www.ar.embnet.org/SMS/STINGm/consseq/

A new set of 182 microsatellites for Eucalyptus: characterization and mapping in a four-species consensus linkage map

Background Eucalyptus is the most widely planted hardwood crop in the tropical and subtropical world. Plantations of Eucalyptus species supply high-quality wood for industrial applications and are important sources of carbon neutral renewable energy in Brazil. E. grandis and E. urophylla and their hybrids are the most widely planted species in fast growing commercial forests in Brazil. E. globulus is the preferred raw material by the mills generating a pulp that is considered superior by the market. However as a pure species it does not grow adequately in Brazil but performs well in hybrid combinations. Breeding programs have increasingly incorporated E. globulus germplasm in fast moving elite populations. Molecular breeding in such populations will require information on markers, comparative mapping and QTL validation across pedigrees involving these different species. Highly multiallelic and transferable microsatellites not only are excellent tools for individual identification, but also provide robust and efficient framework genetic maps that serve well for mapping thousands of bialellic higher throughput markers such as Single Nucleotide Polymorphisms (SNPs) and Diversity Array Technology (DArT). Furthermore microsatellites provide a powerful way for QTL validation across species. We describe the development and characterization of 182 new microsatellites, most of them derived from ESTs and some from a genomic shotgun library. These markers, together with other previously developed ones were used to build a consensus map involving three different pedigrees derived from intercrossing four species of Eucalyptus.

Analysis of the effects of multiple sequence alignments in protein secondary structure prediction

Secondary structure prediction methods are widely used bioinformatics algorithms providing initial insights about protein structure from sequence information. Significant efforts to improve the prediction accuracy over the past years were made, specially the incorporation of information from multiple sequence alignments. This motivated the search for the factors contributing for this improvement. We show that in two of the highly ranked secondary structure prediction methods, DSC and PREDATOR, the use of multiple alignments consistently improves the prediction accuracy as compared to the use of single sequences. This is validated by using different measures of accuracy, which also permit to identify that helical regions benefit the most from alignments, whereas β-strands seem to have reached a plateau in terms of predictability. Also, the origins of this improvement is explored in terms of sequence specificity, secondary structure composition and the extent of sequence similarity which provides the optimal performance. It is found that divergent sequences, in the identity range of 25–55% provide the largest accuracy gain and that above 65% identity there is almost no advantage in using multiple alignments.