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Integrating computationally assembled mouse transcript sequences with the Mouse Genome Informatics (MGI) database

GENOME BIOLOGY 2003年第2期4卷 R16-R16页

作者： Zhu, YX King, BL Parvizi, B Brunk, BP Stoeckert, CJ Quackenbush, J Richardson, J Bult, CJ Jackson Lab Bar Harbor ME 04609 USA Invitrogen Corp Carlsbad CA 92008 USA Univ Penn Ctr Bioinformat Philadelphia PA 19104 USA Inst Genom Res Rockville MD 20850 USA

databases of experimentally generated and computationally derived transcript sequences are valuable resources for genome analysis and annotation. The utility of such databases is enhanced when the sequences they contain are integrated with such biological information as genomic location, gene function, gene expression and phenotypic variation. We present the analysis and results of a semi-automated process of connecting transcript assemblies with highly curated biological information for mouse genes that is available through the Mouse Genome Informatics (MGI) database.

关键词： additional data file Mouse Genome Informatics Component Sequence Tentative Consensus Sequence Electronic Transcript

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Assessing unmodified 70-mer oligonucleotide probe performance on glass-slide microarrays

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GENOME BIOLOGY 2003年第1期4卷 R5-R5页

作者： Wang, HY Malek, RL Kwitek, AE Greene, AS Luu, TV Behbahani, B Frank, B Quackenbush, J Lee, NH Inst Genom Res Rockville MD 20850 USA Med Coll Wisconsin Dept Physiol Milwaukee WI 53226 USA George Washington Univ Med Ctr Dept Pharmacol Washington DC 20037 USA

Background: Long oligonucleotide microarrays are potentially more cost- and management-efficient than cDNA microarrays, but there is little information on the relative performance of these two probe types. The feasibility of using unmodified oligonucleotides to accurately measure changes in gene expression is also unclear. Results: Unmodified sense and antisense 70-mer oligonucleotides representing 75 known rat genes and 10 Arabidopsis control genes were synthesized, printed and UV cross-linked onto glass slides. Printed alongside were PCR-amplified cDNA clones corresponding to the same genes, enabling us to compare the two probe types simultaneously. Our study was designed to evaluate the mRNA profiles of heart and brain, along with Arabidopsis cRNA spiked into the labeling reaction at different relative copy number. Hybridization signal intensity did not correlate with probe type but depended on the extent of UV irradiation. To determine the effect of oligonucleotide concentration on hybridization signal, 70-mers were serially diluted. No significant change in gene-expression ratio or loss in hybridization signal was detected, even at the lowest concentration tested (6.25 muM). In many instances, signal intensity actually increased with decreasing concentration. The correlation coefficient between oligonucleotide and cDNA probes for identifying differentially expressed genes was 0.80, with an average coefficient of variation of 13.4%. Approximately 8% of the genes showed discordant results with the two probe types, and in each case the cDNA results were more accurate, as determined by real-time PCR. Conclusions: Microarrays of UV cross-linked unmodified oligonucleotides provided sensitive and specific measurements for most of the genes studied.

关键词： additional data file Oligonucleotide Probe Probe Type cDNA Array Hybridization Signal Intensity

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Identification of transcripts with enriched expression in the developing and adult pancreas

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GENOME BIOLOGY 2008年第6期9卷 R99-R99页

作者： Hoffman, Brad G. Zavaglia, Bogard Witzsche, Joy de Algara, Teresa Ruiz Beach, Mike Hoodless, Pamela A. Jones, Steven J. M. Marra, Marco A. Helgason, Cheryl D. British Columbia Canc Res Ctr Dept Canc Endocrinol Vancouver BC V5Z 1L3 Canada British Columbia Canc Res Ctr Terry Fox Lab Vancouver BC V5Z 1L3 Canada Univ British Columbia Fac Med Dept Med Genet Vancouver BC V6T 1Z3 Canada Michael Smith Genome Sci Ctr BC Canc Agcy Vancouver BC V5Z 4S6 Canada Univ British Columbia Fac Med Dept Surg Vancouver BC V5Z 4E3 Canada

Background: Despite recent advances, the transcriptional hierarchy driving pancreas organogenesis remains largely unknown, in part due to the paucity of comprehensive analyses. To address this deficit we generated ten SAGE libraries from the developing murine pancreas spanning Theiler stages 17-26, making use of available Pdx1 enhanced green fluorescent protein (EGFP) and Neurog3 EGFP reporter strains, as well as tissue from adult islets and ducts. Results: We used a specificity metric to identify 2,536 tags with pancreas-enriched expression compared to 195 other mouse SAGE libraries. We subsequently grouped co-expressed transcripts with differential expression during pancreas development using K-means clustering. We validated the clusters first using quantitative real time PCR and then by analyzing the Theiler stage 22 pancreas in situ hybridization staining patterns of over 600 of the identified genes using the GenePaint database. These were then categorized into one of the five expression domains within the developing pancreas. Based on these results we identified a cascade of transcriptional regulators expressed in the endocrine pancreas lineage and, from this, we developed a predictive regulatory network describing beta-cell development. Conclusion: Taken together, this work provides evidence that the SAGE libraries generated here are a valuable resource for continuing to elucidate the molecular mechanisms regulating pancreas development. Furthermore, our studies provide a comprehensive analysis of pancreas development, and insights into the regulatory networks driving this process are revealed.

关键词： Enhance Green Fluorescent Protein additional data file Peak Expression Pancreas Development Adult Islet

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A systematic comparative and structural analysis of protein phosphorylation sites based on the mtcPTM database

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GENOME BIOLOGY 2007年第5期8卷 R90-R90页

作者： Jimenez, Jose L. Hegemann, Bjoren Outchins, James R. A. Peters, Jan-Michael Durbin, Richard Wellcome Trust Sanger Inst Cambridge CB10 1SA England Res Inst Mol Pathol IMP A-1030 Vienna Austria

mtcPTM is an online repository of human and mouse phosphosites where the data are hierarchically organized to preserve biologically relevant experimental information, thus allowing straightforward comparisons of phosphorylation patterns found under different conditions. The database also contains the largest available collection of atomic models of phosphorylatable proteins. Detailed analysis of this structural dataset reveals that phosphorylation sites are found in a highly heterogeneous range of structural and sequence contexts. mtcPTM is available at: httpp://***/cgi-bin/mtcPCM/search

关键词： additional data file Solvent Accessibility Phosphorylation Pattern Phosphorylatable Residue Length Coverage

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Rapid transcriptome responses of maize (Zea mays) to UV-B in irradiated and shielded tissues -: art. no. R16

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GENOME BIOLOGY 2004年第3期5卷 1-19页

作者： Casati, P Walbot, V Stanford Univ Dept Biol Sci Stanford CA 94305 USA

Background: Depletion of stratospheric ozone has raised terrestrial levels of ultraviolet-B radiation (UV-B), an environmental change linked to an increased risk of skin cancer and with potentially deleterious consequences for plants. To better understand the processes of UV-B acclimation that result in altered plant morphology and physiology, we investigated gene expression in different organs of maize at several UV-B fluence rates and exposure times. Results: Microarray hybridization was used to assess UV-B responses in directly exposed maize organs and organs shielded by a plastic that absorbs UV-B. After 8 hours of high UV-B, the abundance of 347 transcripts was altered: 285 were increased significantly in at least one organ and 80 were downregulated. More transcript changes occurred in directly exposed than in shielded organs, and the levels of more transcripts were changed in adult compared to seedling tissues. The time course of transcript abundance changes indicated that the response kinetics to UV-B is very rapid, as some transcript levels were altered within 1 hour of exposure. Conclusions: Most of the UV-B regulated genes are organ-specific. Because shielded tissues, including roots, immature ears, and leaves, displayed altered transcriptome profiles after exposure of the plant to UV-B, some signal(s) must be transmitted from irradiated to shielded tissues. These results indicate that there are integrated responses to UV-B radiation above normal levels. As the same total UV-B irradiation dose applied at three intensities elicited different transcript profiles, the transcriptome changes exhibit threshold effects rather than a reciprocal dose-effect response. Transcriptome profiling highlights possible signaling pathways and molecules for future research.

关键词： additional data file Ribosomal Protein Gene Adult Leaf Transcript Type RuBisCO Small Subunit

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Modeling synthetic lethality

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GENOME BIOLOGY 2008年第9期9卷 R135-R135页

作者： Le Meur, Nolwenn Gentleman, Robert Fred Hutchinson Canc Res Ctr Div Publ Hlth Sci Program Computat Biol Seattle WA 98109 USA INSERM IRISA Symbiose F-35042 Rennes France

Background: Synthetic lethality defines a genetic interaction where the combination of mutations in two or more genes leads to cell death. The implications of synthetic lethal screens have been discussed in the context of drug development as synthetic lethal pairs could be used to selectively kill cancer cells, but leave normal cells relatively unharmed. A challenge is to assess genome-wide experimental data and integrate the results to better understand the underlying biological processes. We propose statistical and computational tools that can be used to find relationships between synthetic lethality and cellular organizational units. Results: In Saccharomyces cerevisiae, we identified multi-protein complexes and pairs of multiprotein complexes that share an unusually high number of synthetic genetic interactions. As previously predicted, we found that synthetic lethality can arise from subunits of an essential multiprotein complex or between pairs of multi-protein complexes. Finally, using multi-protein complexes allowed us to take into account the pleiotropic nature of the gene products. Conclusions: Modeling synthetic lethality using current estimates of the yeast interactome is an efficient approach to disentangle some of the complex molecular interactions that drive a cell. Our model in conjunction with applied statistical methods and computational methods provides new tools to better characterize synthetic genetic interactions.

关键词： additional data file Genetic Interaction Array Gene Query Gene Synthetic Lethality

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Development and application of versatile high density microarrays for genome-wide analysis of Streptomyces coelicolor: characterization of the HspR regulon

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GENOME BIOLOGY 2009年第1期10卷 R5-R5页

作者： Bucca, Giselda Laing, Emma Mersinias, Vassilis Allenby, Nicholas Hurd, Douglas Holdstock, Jolyon Brenner, Volker Harrison, Marcus Smith, Colin P. Univ Surrey Fac Hlth & Med Sci Microbial Sci Div Guildford GU2 7XH Surrey England Oxford Gene Technol Ltd Oxford OX15 1PF England

Background: DNA microarrays are a key resource for global analysis of genome content, gene expression and the distribution of transcription factor binding sites. We describe the development and application of versatile high density ink-jet in situ-synthesized DNA arrays for the G+C rich bacterium Streptomyces coelicolor. High G+C content DNA probes often perform poorly on arrays, yielding either weak hybridization or non-specific signals. Thus, more than one million 60-mer oligonucleotide probes were experimentally tested for sensitivity and specificity to enable selection of optimal probe sets for the genome microarrays. The heat-shock HspR regulatory system of S. coelicolor, a well-characterized repressor with a small number of known targets, was exploited to test and validate the arrays for use in global chromatin immunoprecipitation-on-chip (ChIP-chip) and gene expression analysis. Results: In addition to confirming dnaK, clpB and lon as in vivo targets of HspR, it was revealed, using a novel ChIP-chip data clustering method, that HspR also apparently interacts with ribosomal RNA (rrnD operon) and specific transfer RNA genes (the tRNA(Gln)/tRNA(Glu) cluster). It is suggested that enhanced synthesis of Glu-tRNA(Glu) may reflect increased demand for tetrapyrrole biosynthesis following heat-shock. Moreover, it was found that heatshock-induced genes are significantly enriched for Gln/Glu codons relative to the whole genome, a finding that would be consistent with HspR-mediated control of the tRNA species. Conclusions: This study suggests that HspR fulfils a broader, unprecedented role in adaptation to stresses than previously recognized - influencing expression of key components of the translational apparatus in addition to molecular chaperone and protease-encoding genes. It is envisaged that these experimentally optimized arrays will provide a key resource for systems level studies of Streptomyces biology.

关键词： Streptomyces additional data file Feature Extraction Software Tetrapyrrole Biosynthesis Dual Label Probe

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Increased cardiac index due to terbutaline treatment aggravates capillary-alveolar macromolecular leakage in oleic acid lung injury in dogs

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CRITICAL CARE 2009年第5期13卷 R166-R166页

作者： Briot, Raphael Bayat, Sam Anglade, Daniel Martiel, Jean-Louis Grimbert, Francis Univ Grenoble 1 Ctr Hosp Univ CNRS Lab TIMCEquipe PRETAUMR 5525 F-38043 Grenoble 09 France

Introduction We assessed the in vivo effects of terbutaline, a beta2-agonist assumed to reduce microvascular permeability in acute lung injury. Methods We used a recently developed broncho-alveolar lavage (BAL) technique to repeatedly measure (every 15 min. for 4 hours) the time-course of capillary-alveolar leakage of a macromolecule (fluorescein-labeled dextran) in 19 oleic acid (OA) lung injured dogs. BAL was performed in a closed lung sampling site, using a bronchoscope fitted with an inflatable cuff. Fluorescein-labeled Dextran (FITC-D70) was continuously infused and its concentration measured in plasma and BAL fluid. A two-compartment model (blood and alveoli) was used to calculate KAB, the transport rate coefficient of FITC-D70 from blood to alveoli. KAB was estimated every 15 minutes over 4 hours. Terbutaline intra-venous perfusion was started 90 min. after the onset of the injury and then continuously infused until the end of the experiment. Results In the non-treated injured group, the capillary-alveolar leakage of FITC-D70 reached a peak within 30 minutes after the OA injury. Thereafter the FITC-D70 leakage decreased gradually until the end of the experiment. Terbutaline infusion, started 90 min after injury, interrupted the recovery with an aggravation in FITC-D70 leakage. Conclusions As cardiac index increased with terbutaline infusion, we speculate that terbutaline recruits leaky capillaries and increases FITC-D70 leakage after OA injury. These findings suggest that therapies inducing an increase in cardiac output and a decrease in pulmonary vascular resistances have the potential to heighten the early increase in protein transport from plasma to alveoli within the acutely injured lung.

关键词： Acute Respiratory Distress Syndrome additional data file Pulmonary Vascular Resistance Terbutaline Injured Lung

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Cell-specific microarray profiling experiments reveal a comprehensive picture of gene expression in the C-elegans nervous system

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GENOME BIOLOGY 2007年第7期8卷 R135-R135页

作者： Von Stetina, Stephen E. Watson, Joseph D. Fox, Rebecca M. Olszewski, Kellen L. Spencer, W. Clay Roy, Peter J. Miller, David M., III Vanderbilt Univ Dept Cell & Dev Biol Nashville TN 37232 USA Vanderbilt Univ Ctr Mol Neurosci Grad Program Neurosci Nashville TN 37232 USA Johns Hopkins Univ Sch Med Dept Cell Biol Baltimore MD 21205 USA Princeton Univ Lewis Sigler Inst Integrat Genom Dept Mol Biol Princeton NJ 08544 USA Univ Toronto Donnelly Ctr Cellular & Biomol Res Dept Med Genet & Microbiol Toronto ON M5S 1A Canada

Background: With its fully sequenced genome and simple, well-defined nervous system, the nematode Caenorhabditis elegans offers a unique opportunity to correlate gene expression with neuronal differentiation. The lineal origin, cellular morphology and synaptic connectivity of each of the 302 neurons are known. In many instances, specific behaviors can be attributed to particular neurons or circuits. Here we describe microarray-based methods that monitor gene expression in C. elegans neurons and, thereby, link comprehensive profiles of neuronal transcription to key developmental and functional properties of the nervous system. Results: We employed complementary microarray-based strategies to profile gene expression in the embryonic and larval nervous systems. In the MAPCeL ( Microarray Profiling C. elegans cells) method, we used fluorescence activated cell sorting ( FACS) to isolate GFP-tagged embryonic neurons for microarray analysis. To profile the larval nervous system, we used the mRNA-tagging technique in which an epitope-labeled mRNA binding protein ( FLAG-PAB-1) was transgenically expressed in neurons for immunoprecipitation of cell-specific transcripts. These combined approaches identified approximately 2,500 mRNAs that are highly enriched in either the embryonic or larval C. elegans nervous system. These data are validated in part by the detection of gene classes ( for example, transcription factors, ion channels, synaptic vesicle components) with established roles in neuronal development or function. Of particular interest are 19 conserved transcripts of unknown function that are also expressed in the mammalian brain. In addition to utilizing these profiling approaches to define stage-specific gene expression, we also applied the mRNA-tagging method to fingerprint a specific neuron type, the A-class group of cholinergic motor neurons, during early larval development. A comparison of these data to a MAPCeL profile of embryonic A-class motor neurons identifie

关键词： Motor Neuron Synaptic Vesicle additional data file Fluorescence Activate Cell Sorting Ventral Nerve Cord

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Development of a 950-gene DNA array for examining gene expression patterns in mouse testis

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GENOME BIOLOGY 2001年第4期2卷 research0014.1-research0014.9页

作者： Rockett, John C. Luft, J. Christopher Garges, J. Brian Krawetz, Stephen A. Hughes, Mark R. Kim, Kwan Hee Oudes, Asa J. Dix, David J. US EPA Natl Hlth & Environm Effects Res Lab Reprod Toxciol Div Res Triangle Pk NC 27711 USA Wayne State Univ Sch Med Ctr Mol Med & Genet Detroit MI 48201 USA Wayne State Univ Sch Med Dept Obstet & Gynecol Detroit MI 48201 USA Washington State Univ Sch Mol Biosci Pullman WA 99164 USA

Background: Over the past five years, interest in and use of DNA array technology has increased dramatically, and there has been a surge in demand for different types of arrays. Although manufacturers offer a number of pre-made arrays, these are generally of utilitarian design and often cannot accommodate the specific requirements of focused research, such as a particular set of genes from a particular tissue. We found that suppliers did not provide an array to suit our particular interest in testicular toxicology, and therefore elected to design and produce our own. Results: We describe the procedures used by members of the US Environmental Protection Agency MicroArray Consortium (EPAMAC) to produce a mouse testis expression array on both filter and glass-slide formats. The approaches used in the selection and assembly of a pertinent, nonredundant list of testis-expressed genes are detailed. Hybridization of the filter arrays with normal and bromochloroacetic acid-treated mouse testicular RNAs demonstrated that all the selected genes on the array were expressed in mouse testes. Conclusion: We have assembled two lists of mouse (950) and human (960) genes expressed in the mouse and/or human adult testis, essentially all of which are available as sequence-verified clones from public sources. Of these, 764 are homologous and will therefore enable close comparison of gene expression between murine models and human clinical testicular samples.

关键词： additional data file Mouse Testis Membrane Array Mouse Gene Expression Transcriptome Gene

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