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  • 1
    Keywords: COMBINATION ; Germany ; GENOME ; microarray ; COMPLEX ; COMPLEXES ; DNA ; SEQUENCE ; SEQUENCES ; antibodies ; antibody ; ARRANGEMENT ; ASSAY ; DNA microarray ; DNA microarray technology ; microarrays
    Abstract: While the deciphering of basic sequence information on a genomic scale is yielding complete genomic sequences in ever-shorter intervals, experimental procedures for elucidating the cellular effects and consequences of the DNA-encoded information become critical for further analyses. In recent years, DNA microarray technology has emerged as a prime candidate for the performance of many such functional assays. Technically, array technology has come a long way since its conception some 15 years ago, initially designed as a means for large-scale mapping and sequencing. The basic arrangement, however, could be adapted readily to serve eventually as an analytical tool in a large variety of applications. On their own or in combination with other methods, microarrays open up many new avenues of functional analysis. Copyright (C) 2003 John Wiley Sons, Ltd
    Type of Publication: Journal article published
    PubMed ID: 18629015
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  • 2
    Keywords: EXPRESSION ; GROWTH ; CELL ; Germany ; IN-VIVO ; ENZYMES ; GENE ; GENE-EXPRESSION ; PROTEIN ; PROTEINS ; RNA ; DOMAIN ; BINDING ; SEQUENCE ; SEQUENCES ; RECOGNITION ; ENCODES ; gene expression ; ASSAY ; REGION ; REGIONS ; NUCLEUS ; DEGRADATION ; DOUBLE-STRANDED-RNA ; OVEREXPRESSION ; AU-RICH ELEMENTS ; SINGLE ; MOTIF ; INTERFERENCE ; regulation ; TRANSLATION ; LEVEL ; ENZYME ; ASSAYS ; ROLES ; CELL-DIVISION ; 3'-UNTRANSLATED REGION ; USA ; DEPLETION ; microbiology ; NOV ; DIVISION ; CYCLIN ; BRUCEI ; STEADY-STATE ; CYTOPLASM ; TURNOVER ; AFRICAN TRYPANOSOME ; MAJOR FRIEDLIN CHROMOSOME-1 ; PROCYCLIC FORM
    Abstract: In the African trypanosome Trypanosoma brucei nearly all control of gene expression is posttranscriptional; sequences in the 3'-untranslated regions of mRNAs determine the steady-state mRNA levels by regulation of RNA turnover. Here we investigate the roles of two related proteins, TbUBP1 and TbUBP2, containing a single RNA recognition motif, in trypanosome gene expression. TbUBP1 and TbUBP2 are in the cytoplasm and nucleus, comprise ca. 0.1% of the total protein, and are not associated with polysomes or RNA degradation enzymes. Overexpression of TbUBP2 upregulated the levels of several mRNAs potentially involved in cell division, including the CFB1 mRNA, which encodes a protein with a cyclin F-box domain. CFB1 regulation was mediated by the 3'-untranslated region and involved stabilization of the mRNA. Depletion of ThUBP2 and TbUBP1 inhibited growth and downregulated expression of the cyclin F box protein gene CFB2; trans splicing was unaffected. The results of pull-down assays indicated that all tested mRNAs were bound to TbUBP2 or TbUBP1, with some preference for CFBI. We suggest that TbUBP1 and TbUBP2 may be relatively nonspecific RNA-binding proteins and that specific effects of overexpression or depletion could depend on competition between various different proteins for RNA binding
    Type of Publication: Journal article published
    PubMed ID: 17873084
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  • 3
    Keywords: RECEPTOR ; EXPRESSION ; CELL ; Germany ; INHIBITION ; MICROSCOPY ; SITE ; GENE ; GENE-EXPRESSION ; microarray ; METABOLISM ; DIFFERENTIATION ; COMPLEX ; COMPLEXES ; MECHANISM ; MARKER ; mechanisms ; BINDING ; BIOLOGY ; FORM ; GLYCOPROTEIN ; STAGE ; VECTOR ; MEMBRANE ; MARKERS ; SURFACE ; TRANSFORMATION ; PROGRAMMED CELL-DEATH ; REGULATOR ; OXYGEN ; PPAR-GAMMA ; ACTIVATED RECEPTOR-GAMMA ; DEVELOPMENTAL REGULATION ; ORIGIN ; TRANSCRIPTS ; ONCOLOGY ; RE ; VARIANT ; GAMMA ; LIGHT ; INCREASE ; LONG ; USA ; LOSSES ; uptake ; TRANSMISSION ; LIGHT-MICROSCOPY ; host ; peroxisome ; AFRICAN TRYPANOSOMES ; BLOOD-STREAM FORMS ; BRUCEI ; DIHYDROLIPOAMIDE DEHYDROGENASE ; ESAG8 ; monomorphic ; peroxisome proliferator activated receptor ; pleomorphic ; PROCYCLIC-TRYPOMASTIGOTES ; stumpy ; thiazolidinediones ; trypanosomes ; VSG EXPRESSION SITES
    Abstract: Trypanosoma brucei, a protozoan parasite causing sleeping sickness, is transmitted by the tsetse fly and undergoes a complex lifecycle including several defined stages within the insect vector and its mammalian host. In the latter, differentiation from the long slender to the short stumpy form is induced by a yet unknown factor of trypanosomal origin. Here we describe that some thiazolidinediones are also able to induce differentiation. In higher eukaryotes, thiazolidinediones are involved in metabolism and differentiation processes mainly by binding to the intracellular receptor peroxisome proliferator activated receptor gamma. Our studies focus on the effects of troglitazone on bloodstream form trypanosomes. Differentiation was monitored using mitochondrial markers (membrane potential, succinate dehydrogenase activity, inhibition of oxygen uptake by KCN, amount of cytochrome transcripts), morphological changes (Transmission EM and light microscopy), and transformation experiments (loss of the Variant Surface Glycoprotein coat and increase of dihydroliponamide dehydrogenase activity). To further investigate the mechanisms responsible for these changes, microarray analyses were performed, showing an upregulation of expression site associated gene 8 (ESAG8), a potential differentiation regulator. (c) 2007 Elsevier Inc. All rights reserved
    Type of Publication: Journal article published
    PubMed ID: 17428467
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  • 4
    Keywords: CELLS ; EXPRESSION ; Germany ; SYSTEM ; GENE ; GENES ; GENOME ; HYBRIDIZATION ; microarray ; transcription ; DIFFERENTIATION ; validation ; TIME ; MACROPHAGES ; TRANSCRIPTION FACTOR ; INDUCTION ; NERVOUS-SYSTEM ; TRANSCRIPTION FACTORS ; IN-SITU ; ENCODES ; microarrays ; DESIGN ; Drosophila ; NUMBER ; genetics ; CENTRAL-NERVOUS-SYSTEM ; specificity ; expression profiling ; CELL-DIFFERENTIATION ; in situ hybridization ; SUBSET ; DEPENDENCE ; TARGET GENES ; function ; glial cells ; BARRIER FORMATION ; Drosophila embryogenesis ; EMBRYONIC NERVOUS-SYSTEM ; FATE ; gcm ; glial development ; glial genes ; IDENTIFIED NEUROBLASTS ; macrophage ; MOLECULAR MARKERS ; PROMOTING FACTOR ; SEPTATE JUNCTION
    Abstract: In the central nervous system of Drosophila, the induction of the glial cell fate is dependent on the transcription factor glial cells missing (gcm). Though a considerable number of other genes have been shown to be expressed in all or in subsets of glial cells, the course of glial cell differentiation and subtype specification is only poorly understood. This prompted us to design a whole genome microarray approach comparing gem gain-of-function and, for the first time, gem loss-of-function genetics to wildtype in time course experiments along embryogenesis. The microarray data were analyzed with special emphasis on the temporal profile of differential regulation. A comparison of both experiments enabled us to identify more than 300 potential gcm target genes. Validation by in situ hybridization revealed expression in glial cells, macrophages, and tendon cells (all three cell types depend on gem) for 70 genes, of which more than 50 had been unknown to be under gem control. Eighteen genes are exclusively expressed in glial cells, and their dependence on gem was confirmed in situ. Initial considerations regarding the role of the newly discovered glial genes are discussed based on gene ontology and the temporal profile and subtype specificity of their expression. This collection of glial genes provides an important basis for the clarification of the genetic network controlling various aspects of glial development and function. (c) 2006 Elsevier Inc. All rights reserved
    Type of Publication: Journal article published
    PubMed ID: 16762338
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  • 5
    Keywords: AB-INITIO ; EXPRESSION ; COMBINATION ; evaluation ; Germany ; human ; GENE ; GENES ; GENOME ; GENOME SEQUENCE ; HYBRIDIZATION ; microarray ; PROTEIN ; transcription ; validation ; DNA ; FAMILY ; PROTEIN FAMILIES ; PROTEIN FAMILY ; BIOLOGY ; SEQUENCE ; SEQUENCES ; DISCOVERY ; IDENTIFICATION ; IN-SITU ; DESIGN ; Drosophila ; DROSOPHILA-MELANOGASTER ; MELANOGASTER ; NUMBER ; DATABASE ; HUMAN GENOME ; RT-PCR ; PREDICTION ; MICROARRAY ANALYSIS ; PROJECT ; max ; RESOURCE ; ANOPHELES-GAMBIAE ; SYSTERS
    Abstract: Background: While the genome sequences for a variety of organisms are now available, the precise number of the genes encoded is still a matter of debate. For the human genome several stringent annotation approaches have resulted in the same number of potential genes, but a careful comparison revealed only limited overlap. This indicates that only the combination of different computational prediction methods and experimental evaluation of such in silico data will provide more complete genome annotations. In order to get a more complete gene content of the Drosophila melanogaster genome, we based our new D. melanogaster whole-transcriptome microarray, the Heidelberg FlyArray, on the combination of the Berkeley Drosophila Genome Project (BDGP) annotation and a novel ab initio gene prediction of lower stringency using the Fgenesh software.Results: Here we provide evidence for the transcription of approximately 2,600 additional genes predicted by Fgenesh. Validation of the developmental profiling data by RT-PCR and in situ hybridization indicates a lower limit of 2,000 novel annotations, thus substantially raising the number of genes that make a fly.Conclusions: The successful design and application of this novel Drosophila microarray on the basis of our integrated in silico/wet biology approach confirms our expectation that in silico approaches alone will always tend to be incomplete. The identification of at least 2,000 novel genes highlights the importance of gathering experimental evidence to discover all genes within a genome. Moreover, as such an approach is independent of homology criteria, it will allow the discovery of novel genes unrelated to known protein families or those that have not been strictly conserved between species
    Type of Publication: Journal article published
    PubMed ID: 14709175
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  • 6
    Keywords: EXPRESSION ; Germany ; GENE ; GENE-EXPRESSION ; GENES ; PROTEIN ; PROTEINS ; DIFFERENTIATION ; FAMILY ; SEQUENCE ; SEQUENCES ; PATTERNS ; PROBES ; ARRAY ; TRANSPORTER ; regulation ; LIFE-CYCLE ; transcript ; EXPRESSION PATTERNS ; EXPRESSION SITES ; FLAGELLUM ; LEISHMANIA ; Trypanosoma ; VARIANT SURFACE GLYCOPROTEIN
    Abstract: We describe developmentally regulated genes in two strains of Trypanosoma brucei: the monomorphic strain Lister 427 and the pleomorphic strain TREU927. Expression patterns were obtained using an array of 24,567 genomic fragments. Probes were prepared from bloodstream-form or procyclic-form trypanosomes. Fourteen procyclic-specific and 77 bloodstream-specific signals were obtained from sequences matching variant surface glycoprotein or associated genes, and a further 17 regulated sequences were repetitive or transposable-element-related. Two hundred and eighty-six regulated spots corresponded to mRNAs from other protein-coding genes; these spots represent 191 different proteins. Regulation of 113 different genes (79 from procyclic forms, 34 from bloodstream-forms) was supported by at least two independent experiments or criteria: of these, about 60 were novel. Only two genes - encoding HSP83 and an importin-related protein - appeared to be regulated in the TREU927 strain only. Our results confirmed previous estimates that 2% of trypanosome genes show developmental regulation at the mRNA level. (C) 2004 Elsevier B.V. All rights reserved
    Type of Publication: Journal article published
    PubMed ID: 15664651
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