Search

Background: The advent and ongoing development of next generation sequencing technologies (NGS) has led to a rapid increase in the rate of human genome re-sequencing data, paving the way for personalized genomics and precision medicine. The body of genome resequencing data is progressively increasing underlining the need for accurate and time-effective bioinformatics systems for genotyping - a crucial prerequisite for identification of candidate causal mutations in diagnostic screens., Results: Here we present CoVaCS, a fully automated, highly accurate system with a web based graphical interface for genotyping and variant annotation. Extensive tests on a gold standard benchmark data-set -the NA12878 Illumina platinum genome- confirm that call-sets based on our consensus strategy are completely in line with those attained by similar command line based approaches, and far more accurate than call-sets from any individual tool. Importantly our system exhibits better sensitivity and higher specificity than equivalent commercial software., Conclusions: CoVaCS offers optimized pipelines integrating state of the art tools for variant calling and annotation for whole genome sequencing (WGS), whole-exome sequencing (WES) and target-gene sequencing (TGS) data. The system is currently hosted at Cineca, and offers the speed of a HPC computing facility, a crucial consideration when large numbers of samples must be analysed. Importantly, all the analyses are performed automatically allowing high reproducibility of the results. As such, we believe that CoVaCS can be a valuable tool for the analysis of human genome resequencing studies. CoVaCS is available at: https://bioinformatics.cineca.it/covacs .

Background: Expression screening of environmental DNA (eDNA) libraries is a popular approach for the identification and characterization of novel microbial enzymes with promising biotechnological properties. In such "functional metagenomics" experiments, inserts, selected on the basis of activity assays, are sequenced with high throughput sequencing technologies. Assembly is followed by gene prediction, annotation and identification of candidate genes that are subsequently evaluated for biotechnological applications., Results: Here we present A-GAME (A GAlaxy suite for functional MEtagenomics), a web service incorporating state of the art tools and workflows for the analysis of eDNA sequence data. We illustrate the potential of A-GAME workflows using real functional metagenomics data, showing that they outperform alternative metagenomics assemblers. Dedicated tools available in A-GAME allow efficient analysis of pooled libraries and rapid identification of candidate genes, reducing sequencing costs and saving the need for laborious manual annotation., Conclusion: In conclusion, we believe A-GAME will constitute a valuable resource for the functional metagenomics community. A-GAME is publicly available at http://beaconlab.it/agame.

Background: Recent discoveries have highlighted the fact that alternative splicing and alternative transcripts are the rule, rather than the exception, in metazoan genes. Since multiple transcript and protein variants expressed by the same gene are, by definition, structurally distinct and need not to be functionally equivalent, the concept of gene orthology should be extended to the transcript level in order to describe evolutionary relationships between structurally similar transcript variants. In other words, the identification of true orthology relationships between gene products now should progress beyond primary sequence and "splicing orthology", consisting in ancestrally shared exon-intron structures, is required to define orthologous isoforms at transcript level., Results: As a starting step in this direction, in this work we performed a large scale human- mouse gene comparison with a twofold goal: first, to assess if and to which extent traditional gene annotations such as RefSeq capture genuine splicing orthology; second, to provide a more detailed annotation and quantification of true human-mouse orthologous transcripts defined as transcripts of orthologous genes exhibiting the same splicing patterns., Conclusions: We observed an identical exon/intron structure for 32% of human and mouse orthologous genes. This figure increases to 87% using less stringent criteria for gene structure similarity, thus implying that for about 13% of the human RefSeq annotated genes (and about 25% of the corresponding transcripts) we could not identify any mouse transcript showing sufficient similarity to be confidently assigned as a splicing ortholog. Our data suggest that current gene and transcript data may still be rather incomplete - with several splicing variants still unknown. The observation that alternative splicing produces large numbers of alternative transcripts and proteins, some of them conserved across species and others truly species-specific, suggests that, still maintaining the conventional definition of gene orthology, a new concept of "splicing orthology" can be defined at transcript level.

Unlabelled: The version of this article published in BMC Genomics 2009, 10:558, contains data in Table 1 which are now known to be unreliable, and an illustration, in Figure 1, of unusual miRNA processing events predicted by these unreliable data. In this full-length correction, new data replace those found to be unreliable, leading to a more straightforward interpretation without altering the principle conclusions of the study. Table 1 and associated methods have been corrected, Figure 1 deleted, supplementary file 1 added, and modifications made to the sections "Deep sequencing of small RNAs from grapevine leaf tissue" and "Microarray analysis of miRNA expression". The editors and authors regret the inconvenience caused to readers by premature publication of the original paper., Background: MicroRNAs are short (~21 base) single stranded RNAs that, in plants, are generally coded by specific genes and cleaved specifically from hairpin precursors. MicroRNAs are critical for the regulation of multiple developmental, stress related and other physiological processes in plants. The recent annotation of the genome of the grapevine (Vitis vinifera L.) allowed the identification of many putative conserved microRNA precursors, grouped into multiple gene families., Results: Here we use oligonucleotide arrays to provide the first indication that many of these microRNAs show differential expression patterns between tissues and during the maturation of fruit in the grapevine. Furthermore we demonstrate that whole transcriptome sequencing and deep-sequencing of small RNA fractions can be used both to identify which microRNA precursors are expressed in different tissues and to estimate genomic coordinates and patterns of splicing and alternative splicing for many primary miRNA transcripts., Conclusions: Our results show that many microRNAs are differentially expressed in different tissues and during fruit maturation in the grapevine. Furthermore, the demonstration that whole transcriptome sequencing can be used to identify candidate splicing events and approximate primary microRNA transcript coordinates represents a significant step towards the large-scale elucidation of mechanisms regulating the expression of microRNAs at the transcriptional and post-transcriptional levels.

Background: MicroRNAs are short (approximately 21 base) single stranded RNAs that, in plants, are generally coded by specific genes and cleaved specifically from hairpin precursors. MicroRNAs are critical for the regulation of multiple developmental, stress related and other physiological processes in plants. The recent annotation of the genome of the grapevine (Vitis vinifera L.) allowed the identification of many putative conserved microRNA precursors, grouped into multiple gene families., Results: Here we use oligonucleotide arrays to provide the first indication that many of these microRNAs show differential expression patterns between tissues and during the maturation of fruit in the grapevine. Furthermore we demonstrate that whole transcriptome sequencing and deep-sequencing of small RNA fractions can be used both to identify which microRNA precursors are expressed in different tissues and to estimate genomic coordinates and patterns of splicing and alternative splicing for many primary miRNA transcripts., Conclusion: Our results show that many microRNAs are differentially expressed in different tissues and during fruit maturation in the grapevine. Furthermore, the demonstration that whole transcriptome sequencing can be used to identify candidate splicing events and approximate primary microRNA transcript coordinates represents a significant step towards the large-scale elucidation of mechanisms regulating the expression of microRNAs at the transcriptional and post-transcriptional levels.

Background: Comparative genomic studies of the mitochondrion-lacking protist group Diplomonadida (diplomonads) has been lacking, although Giardia lamblia has been intensively studied. We have performed a sequence survey project resulting in 2341 expressed sequence tags (EST) corresponding to 853 unique clones, 5275 genome survey sequences (GSS), and eleven finished contigs from the diplomonad fish parasite Spironucleus salmonicida (previously described as S. barkhanus)., Results: The analyses revealed a compact genome with few, if any, introns and very short 3' untranslated regions. Strikingly different patterns of codon usage were observed in genes corresponding to frequently sampled ESTs versus genes poorly sampled, indicating that translational selection is influencing the codon usage of highly expressed genes. Rigorous phylogenomic analyses identified 84 genes--mostly encoding metabolic proteins--that have been acquired by diplomonads or their relatively close ancestors via lateral gene transfer (LGT). Although most acquisitions were from prokaryotes, more than a dozen represent likely transfers of genes between eukaryotic lineages. Many genes that provide novel insights into the genetic basis of the biology and pathogenicity of this parasitic protist were identified including 149 that putatively encode variant-surface cysteine-rich proteins which are candidate virulence factors. A number of genomic properties that distinguish S. salmonicida from its human parasitic relative G. lamblia were identified such as nineteen putative lineage-specific gene acquisitions, distinct mutational biases and codon usage and distinct polyadenylation signals., Conclusion: Our results highlight the power of comparative genomic studies to yield insights into the biology of parasitic protists and the evolution of their genomes, and suggest that genetic exchange between distantly-related protist lineages may be occurring at an appreciable rate in eukaryote genome evolution.