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GeneOrder3.0: software for comparing the order of genes in pairs of small bacterial genomes.

BACKGROUND: An increasing number of whole viral and bacterial genomes are being sequenced and deposited in public databases. In parallel to the mounting interest in whole genomes, the number of whole genome analyses software tools is also increasing. GeneOrder was originally developed to provide an analysis of genes between two genomes, allowing visualization of gene order and synteny comparisons of any small genomes. It was originally developed for comparing virus, mitochondrion and chloroplast genomes. This is now extended to small bacterial genomes of sizes less than 2 Mb. RESULTS: GeneOrder3.0 has been developed and validated successfully on several small bacterial genomes (ca. 580 kb to 1.83 Mb) archived in the NCBI GenBank database. It is an updated web-based "on-the-fly" computational tool allowing gene order and synteny comparisons of any two small bacterial genomes. Analyses of several bacterial genomes show that a large amount of gene and genome re-arrangement occurs, as seen with earlier DNA software tools. This can be displayed at the protein level using GeneOrder3.0. Whole genome alignments of genes are presented in both a table and a dot plot. This allows the detection of evolutionary more distant relationships since protein sequences are more conserved than DNA sequences. CONCLUSIONS: GeneOrder3.0 allows researchers to perform comparative analysis of gene order and synteny in genomes of sizes up to 2 Mb "on-the-fly." AVAILABILITY: http://binf.gmu.edu/genometools.html and http://pasteur.atcc.org:8050/GeneOrder3.0.

Chromosome Mapping↗

Visualization of single copies of the Epstein-Barr virus genome by in situ hybridization.

Conditions have been established for demonstrating small numbers of genes of the Epstein-Barr virus (EBV) in B-lymphoid cells by in situ hybridization using biotinylated EBV-specific DNA from cloned BamHI fragments of the viral genome. Single copies of EBV genomes were successfully visualized with minimal background when the probe concentration was 0.2 micrograms/ml, the DNA denaturation step was performed at 100 degrees C, and the immunochemical detection system employed a three-layer peroxidase protocol with gold-silver amplification of the diaminobenzidine substrate. The minimal target DNA detectable was about 10 kilobase pairs. In the case of sectioned cells fixed overnight with formalin, simulating conditions used in routine tissue fixation, this approach failed to demonstrate EBV DNA present at less than 100 copies per cell, that is, at the level found in Raji cells. However, when denaturation was performed using microwave irradiation with the other optimized conditions maintained, EBV DNA could be visualized in 10-20% of such cells, although not in cells known to contain fewer than 10 copies per cell. Thus, microwave irradiation partially overcomes the limit of DNA target detection imposed by formalin.

Animals↗

PLATCOM: a Platform for Computational Comparative Genomics.

MOTIVATION: As more whole genome sequences become available, comparing multiple genomes at the sequence level can provide insight into new biological discovery. However, there are significant challenges for genome comparison. The challenge includes requirement for computational resources owing to the large volume of genome data. More importantly, since the choice of genomes to be compared is entirely subjective, there are too many choices for genome comparison. For these reasons, there is pressing need for bioinformatics systems for comparing multiple genomes where users can choose genomes to be compared freely. RESULTS: PLATCOM (Platform for Computational Comparative Genomics) is an integrated system for the comparative analysis of multiple genomes. The system is built on several public databases and a suite of genome analysis applications are provided as exemplary genome data mining tools over these internal databases. Researchers are able to visually investigate genomic sequence similarities, conserved gene neighborhoods, conserved metabolic pathways and putative gene fusion events among a set of selected multiple genomes. AVAILABILITY: http://platcom.informatics.indiana.edu/platcom

Chromosome Mapping↗

Informatics issues in large-scale sequence analysis: elucidating the protein kinases of C. elegans.

With the availability of the nearly complete genomic sequence of C. elegans, the first multicellular organism to be sequenced, molecular biology has definitely entered the postgenomic era. Annotation of the genomic sequence, which refers to identifying the genes and other biologically relevant sections of the genome, is an important and nontrivial next step. A first-pass annotation will be necessarily incomplete but will drive further biological experiments, which in turn will help to annotate the genome better. Given the scale of the genome sequence analysis, it is clear that the annotation should be automated as much as possible without sacrificing the quality of analysis. In this work, we outline our approach to identifying the protein kinases of C. elegans from the genomic sequence. We describe new tools we have developed for analysis, management and visualization of genomic data. By developing modular and scalable solutions, this study has provided a framework for future analysis of the Drosophila and human genomes.

Animals↗

Visualization of parental HSV-1 genomes and replication compartments in association with ND10 in live infected cells.

The relative location of active and repressed genes within the nucleus is becoming recognized as a significant factor in the control of gene expression. We have developed systems to visualize parental and replicated herpes simplex virus type 1 (HSV-1) amplicon genomes in association with PML nuclear bodies (ND10) in live cells. Plasmids containing viral replication and packaging signals, a gene expressing enhanced yellow fluorescent protein linked to the tetracycline repressor DNA binding domain and 14 copies of the tetracycline operator sequence were used to produce amplicon genomes packaged into normal viral particles. The frequency of the juxtaposition of viral genomes and ND10 was substantially increased by inclusion of an active HSV-1 Early gene transcription unit, indicating that the association is neither random nor passive. Furthermore, the ND10-associated genomes preferentially progressed to form viral replication compartments. Thus, active viral transcription contributes to the efficiency of viral genome association with ND10, and this in turn increases the probability that the genome will engage in active DNA replication. These studies in live cells provide a novel insight into virus-ND10 interactions and provide compelling visualization of their functional relevance.

Animals↗

Molecular cloning and structural analysis of the human gene encoding cellular retinaldehyde-binding protein.

Cellular retinaldehyde-binding protein (CRALBP) appears to play a role in the vertebrate visual process as a substrate-routing protein, influencing the enzymatic partitioning of 11-cis-retinol at a key branch point in the visual cycle. Genomic clones spanning 29 kilobases and encompassing the human CRALBP gene have been isolated by screening two human genomic libraries and from polymerase chain reaction amplification of human leukocyte DNA. The sequence of 13,647 contiguous nucleotides has been determined, including 3130 and 516 bases from the 5'- and 3'-flanking regions, respectively. The human CRALBP gene exists as a single copy in the genome based on Southern analyses and localization to a single site on human chromosome 15 (Sparkes, R. S., Heinzmann, C., Goldflam, S., Kojis, T., Saari, J. C., Mohandes, T., Klisak, I., Bateman, J. B., and Crabb, J. W. (1992) Genomics 12, 58-62). The gene is composed of eight exons and seven introns with average lengths of 198 base pairs and 1.2 kilobases, respectively, and which exhibit conventional vertebrate splicing. Alu repetitive sequences exist in introns 4 and 5 as well as in the 5'- and 3'-flanking regions of the gene. RNase protection and primer extension analyses indicate that the human CRALBP gene transcription start site is 922 bases upstream of the initiation codon. The first exon is entirely untranslated and both exon 2 and exon 8 contain untranslated regions. The proximal 5'-flanking region lacks GC boxes and consensus TATA and CCAAT boxes at the usual positions. The 3'-untranslated region of CRALBP exon 8 is essentially identical to a partial cDNA clone reportedly isolated from a human hippocampus cDNA library, suggesting that the protein may be expressed in a wider spectrum of tissues than previously recognized. The human CRALBP genomic clones and structure provide valuable tools for studying the physiological role of the protein in vision and visual disorders.

Animals↗

Interactive gene-order comparison for multiple small genomes.

UNLABELLED: The Genome Organization Analysis Tool (GOAT) is a program that performs comparative sequence analysis on ordered gene lists from annotated genomes, provides visual and tabular output, and provides means of accessing and analyzing related gene sequence data, for the purpose of comparing genome organization at the gene-order level. GOAT can be used to compare any two or more genomes or chromosomes on demand, or configured to provide access to precomputed comparisons of a specific group of genome sequences. AVAILABILITY: Demonstration web server and software download, subject to the Virginia Tech Noncommercial License are available at http://gaia.biotech.vt.edu/goat/. SUPPLEMENTARY INFORMATION: Updates, installation and configuration information are available at http://gaia.biotech.vt.edu/goat.

Algorithms↗

Relative prevalence of typical and atypical strains among rotaviruses from diarrheic pigs in conventional swine herds.

Polyacrylamide gel electrophoresis was conducted on genomic RNA extracted from rotaviruses detected in diarrheic pigs from conventional swine herds. Ninety samples contained sufficient virus for RNA band visualization and genome classification. Genome profiles were characteristic of typical group A rotaviruses in 67.8% of the 90 samples, of group B rotaviruses in 10.0%, and of group C rotaviruses in 11.1%. In 11.1% of the samples, the presence of more than 11 bands suggested concurrent infection with more than 1 strain of rotavirus. In infections among nursing pigs, 76.4% were group A rotaviruses, 7.4% were group B, 7.4% were group C, and 8.8% were coinfections. In infections among weaned pigs, 40.9% were group A, 18.2% were group B, 22.7% were group C, and 18.2% were coinfections. Coelectrophoresis with prototype OSU and Gottfried strains revealed a great diversity in electropherotype among field strains of rotavirus.

Animals↗

Blast2GO: a universal tool for annotation, visualization and analysis in functional genomics research.

SUMMARY: We present here Blast2GO (B2G), a research tool designed with the main purpose of enabling Gene Ontology (GO) based data mining on sequence data for which no GO annotation is yet available. B2G joints in one application GO annotation based on similarity searches with statistical analysis and highlighted visualization on directed acyclic graphs. This tool offers a suitable platform for functional genomics research in non-model species. B2G is an intuitive and interactive desktop application that allows monitoring and comprehension of the whole annotation and analysis process. AVAILABILITY: Blast2GO is freely available via Java Web Start at http://www.blast2go.de. SUPPLEMENTARY MATERIAL: http://www.blast2go.de -> Evaluation.

Algorithms↗

GenAlyzer: interactive visualization of sequence similarities between entire genomes.

SUMMARY: Genalyzer is a software tool designed for the interactive visualization of sequence matches between DNA or protein sequences. It provides visualizations on different levels of granularity, from complete overviews via zoomed regions to alignments of particular matching substrings. Genalyzer can efficiently handle very large datasets, allowing to display tens of thousands of matches between sequences of tens of millions of bases. AVAILABILITY: Genalyzer is available free of charge for non-commercial research institutions. For more details, see http://www.genalyzer.de

Algorithms↗

GeneViTo: visualizing gene-product functional and structural features in genomic datasets.

BACKGROUND: The availability of increasing amounts of sequence data from completely sequenced genomes boosts the development of new computational methods for automated genome annotation and comparative genomics. Therefore, there is a need for tools that facilitate the visualization of raw data and results produced by bioinformatics analysis, providing new means for interactive genome exploration. Visual inspection can be used as a basis to assess the quality of various analysis algorithms and to aid in-depth genomic studies. RESULTS: GeneViTo is a JAVA-based computer application that serves as a workbench for genome-wide analysis through visual interaction. The application deals with various experimental information concerning both DNA and protein sequences (derived from public sequence databases or proprietary data sources) and meta-data obtained by various prediction algorithms, classification schemes or user-defined features. Interaction with a Graphical User Interface (GUI) allows easy extraction of genomic and proteomic data referring to the sequence itself, sequence features, or general structural and functional features. Emphasis is laid on the potential comparison between annotation and prediction data in order to offer a supplement to the provided information, especially in cases of "poor" annotation, or an evaluation of available predictions. Moreover, desired information can be output in high quality JPEG image files for further elaboration and scientific use. A compilation of properly formatted GeneViTo input data for demonstration is available to interested readers for two completely sequenced prokaryotes, Chlamydia trachomatis and Methanococcus jannaschii. CONCLUSIONS: GeneViTo offers an inspectional view of genomic functional elements, concerning data stemming both from database annotation and analysis tools for an overall analysis of existing genomes. The application is compatible with Linux or Windows ME-2000-XP operating systems, provided that the appropriate Java Runtime Environment is already installed in the system.

Bacterial Proton-Translocating ATPases↗

M-GCAT: interactively and efficiently constructing large-scale multiple genome comparison frameworks in closely related species.

BACKGROUND: Due to recent advances in whole genome shotgun sequencing and assembly technologies, the financial cost of decoding an organism's DNA has been drastically reduced, resulting in a recent explosion of genomic sequencing projects. This increase in related genomic data will allow for in depth studies of evolution in closely related species through multiple whole genome comparisons. RESULTS: To facilitate such comparisons, we present an interactive multiple genome comparison and alignment tool, M-GCAT, that can efficiently construct multiple genome comparison frameworks in closely related species. M-GCAT is able to compare and identify highly conserved regions in up to 20 closely related bacterial species in minutes on a standard computer, and as many as 90 (containing 75 cloned genomes from a set of 15 published enterobacterial genomes) in an hour. M-GCAT also incorporates a novel comparative genomics data visualization interface allowing the user to globally and locally examine and inspect the conserved regions and gene annotations. CONCLUSION: M-GCAT is an interactive comparative genomics tool well suited for quickly generating multiple genome comparisons frameworks and alignments among closely related species. M-GCAT is freely available for download for academic and non-commercial use at: http://alggen.lsi.upc.es/recerca/align/mgcat/intro-mgcat.html.

Algorithms↗

Analysis of oncornavirus RNA subunits by electron microscopy.

Subunits of oncornavirus (avian myeloblastosis virus) RNA were isolated from purified 60--70S viral RNA by heat dissociation. Molecules sedimenting at 35 S, assumed to be the major component of the viral genome, were visualized in the electron microscope and their lengths were statistically analyzed. The results indicate a rather heterogeneous population of molecules with five distinct, reproducible size groups, an observation that excludes the assumption of random degradation of the genome. In addition, molecules of 28 and 18S RNA, always present in oncornavirus RNA preparations, were examined with the same methods. Some of these molecules possess secondary-structure regions similar to those characteristic for ribosomal RNA.

Avian Leukosis Virus↗

The ERGO genome analysis and discovery system.

The ERGO (http://ergo.integratedgenomics.com/ERGO/) genome analysis and discovery suite is an integration of biological data from genomics, biochemistry, high-throughput expression profiling, genetics and peer-reviewed journals to achieve a comprehensive analysis of genes and genomes. Far beyond any conventional systems that facilitate functional assignments, ERGO combines pattern-based analysis with comparative genomics by visualizing genes within the context of regulation, expression profiling, phylogenetic clusters, fusion events, networked cellular pathways and chromosomal neighborhoods of other functionally related genes. The result of this multifaceted approach is to provide an extensively curated database of the largest available integration of genomes, with a vast collection of reconstructed cellular pathways spanning all domains of life. Although access to ERGO is provided only under subscription, it is already widely used by the academic community. The current version of the system integrates 500 genomes from all domains of life in various levels of completion, 403 of which are available for subscription.

Animals↗

The electroretinogram and visual evoked potential of freely moving rats.

The vascularised rat retina could be one of the most useful experimental objects in visual neuroscience to understand human visual physiological and pathological processes. We report here on a new method of implantation for studying the visual system of freely moving rats that provides a rat model for simultaneous recording at corneal and cortical level and is stable enough to record for months. We implanted light emitting diodes onto the skull behind the eyeball to stimulate the eye with flashes and to light adapt the retina with constant light levels. A multistrand, stainless steel, flexible fine wire electrode placed on the eyeball was used for electroretinogram recording and screw electrodes (left/right visual and parietal cortical) were used to record the visual evoked potential and the electroencephalogram. In the present report we focus on the new method of implantation for recording the corneal flash electroretinogram of normal, freely moving rats simultaneously with the visual evoked cortical potential showing examples in various visual experiments. We also introduce a program for retinogram and visual evoked potential analysis, which defines various measures (latencies, areas, amplitudes, and durations) and draw attention to the benefits of this method for those involved in visual, functional genomic, pharmacological, and human ophthalmologic research.

Animals↗

High resolution microscopic mapping of DNA using multi-color fluorescent hybridization.

We describe a procedure for microscopically mapping the relative positions of DNA probes along extended strands of DNA. The procedure referred to as direct visual hybridization (DIRVISH) DNA mapping involves the simultaneous hybridization of multiple probes and the fluorescent colors, red green and blue to produce images that convey high-resolution mapping information. The images appear as long strings of fluorescent signals positioned as they are in the genome. A visual multi-color map is generated within 2 days. Cosmid probes span a distance of 10 microms or more and have been observed to contain patterns within the strings of signals. We have developed computer imaging programs to scan through the strings of signals and plot the intensities. Scans through multiple signal strings for one cosmid probe revealed consistent patterns. We have interpreted the patterns as the result of suppression of repetitive DNA sequence hybridization. These patterns may prove useful as fingerprints for regions of DNA.

Animals↗

ProGenExpress: visualization of quantitative data on prokaryotic genomes.

BACKGROUND: The integration of genomic information with quantitative experimental data is a key component of systems biology. An increasing number of microbial genomes are being sequenced, leading to an increasing amount of data from post-genomics technologies. The genomes of prokaryotes contain many structures of interest, such as operons, pathogenicity islands and prophage sequences, whose behaviour is of interest during infection and disease. There is a need for simple and novel tools to display and analyse data from these integrated datasets, and we have developed ProGenExpress as a tool for visualising arbitrarily complex numerical data in the context of prokaryotic genomes. RESULTS: Here we describe ProGenExpress, an R package that allows researchers to easily and quickly visualize quantitative measurements, such as those produced by microarray experiments, in the context of the genome organization of sequenced prokaryotes. Data from microarrays, proteomics or other whole-genome technologies can be accurately displayed on the genome. ProGenExpress can also search for novel regions of interest that consist of groups of adjacent genes that show similar patterns across the experimental data set. We demonstrate ProGenExpress with microarray data from a time-course experiment involving Salmonella typhimurium. CONCLUSION: ProGenExpress can be used to visualize quantitative data from complex experiments in the context of the genome of sequenced prokaryotes, and to find novel regions of interest.

Computational Biology↗

Genome-wide pathway analysis and visualization using gene expression data.

Visualization of results for high performance computing pose special problems due to the complexity and the volume of data these systems manipulate. We present an approach for visualization of c-DNA microarray gene expression data in metabolic and regulatory pathways using multi-resolution animation at different levels of detail. We describe three scoring functions to characterize pathways at the transcriptional level based on gene expression, coregulation and cascade effect. We also assess the significance of each pathway score on the basis of their biological connotation.

Animals↗