52 research outputs found
A survey of research in the application of tolerance analysis to the design of mechanical assemblies
Volatilização de N-NH3 na cultura de milho:: II. avaliação de fontes sólidas e fluidas em sistema de plantio direto e convencional
Making sense of EST sequences by CLOBBing them
Background
Expressed sequence tags (ESTs) are single pass reads from randomly selected cDNA clones. They provide a highly cost-effective method to access and identify expressed genes. However, they are often prone to sequencing errors and typically define incomplete transcripts. To increase the amount of information obtainable from ESTs and reduce sequencing errors, it is necessary to cluster ESTs into groups sharing significant sequence similarity.
Results
As part of our ongoing EST programs investigating 'orphan' genomes, we have developed a clustering algorithm, CLOBB (Cl uster o n the b asis of B LAST similarity) to identify and cluster ESTs. CLOBB may be used incrementally, preserving original cluster designations. It tracks cluster-specific events such as merging, identifies 'superclusters' of related clusters and avoids the expansion of chimeric clusters. Based on the Perl scripting language, CLOBB is highly portable relying only on a local installation of NCBI's freely available BLAST executable and can be usefully applied to > 95 % of the current EST datasets. Analysis of the Danio rerio EST dataset demonstrates that CLOBB compares favourably with two less portable systems, UniGene and TIGR Gene Indices.
Conclusions
CLOBB provides a highly portable EST clustering solution and is freely downloaded from: http://www.nematodes.org/CLOB
200 000 nematode expressed sequence tags on the Net
Expressed sequence tags (ESTs) are single-pass sequence reads made from randomly selected cDNA clones, which represent the expressed genes of an organism. EST analysis is an efficient and cost-effective method for sampling the genes expressed by an organism or tissue. ESTs have been a focus of eukaryotic parasite genome initiatives for several years 1 (see the Parasite Genome web server at http://www.ebi.ac.uk/parasites/parasite-genome.html), and parasitic organism ESTs make up a significant portion of the dbEST subsection of GenBank 2 , 3 . EST data sets can be mined for useful or interesting content using standard similarity-based search tools such as BLAST. In the field of molecular parasitology, such approaches have led to the discovery of many new potential drug targets and vaccine candidates. However, EST data sets also contain important additional types of information
IL-4 dependent alternatively-activated macrophages have a distinctive in vivo gene expression phenotype
Background
"Alternatively-activated" macrophages are found in Th2-mediated inflammatory settings such as nematode infection and allergic pulmonary inflammation. Due in part to a lack of markers, these cells have not been well characterized in vivo and their function remains unknown.
Results
We have used murine macrophages elicited by nematode infection (NeMφ) as a source of in vivo derived alternatively activated macrophages. Using three distinct yet complementary molecular approaches we have established a gene expression profile of alternatively activated macrophages and identified macrophage genes that are regulated in vivo by IL-4. First, genes abundantly expressed were identified by an expressed sequence tag strategy. Second, an array of 1176 known mouse genes was screened for differential expression between NeMφ from wild type or IL-4 deficient mice. Third, a subtractive library was screened to identify novel IL-4 dependent macrophage genes. Differential expression was confirmed by real time RT-PCR analysis.
Conclusions
Our data demonstrate that alternatively activated macrophages generated in vivo have a gene expression profile distinct from any macrophage population described to date. Several of the genes we identified, including those most abundantly expressed, have not previously been associated with macrophages and thus this study provides unique new information regarding the phenotype of macrophages found in Th2-mediated, chronic inflammatory settings. Our data also provide additional in vivo evidence for parallels between the inflammatory processes involved in nematode infection and allergy
The Brugia malayi genome project: expressed sequence tags and gene discovery
To advance and facilitate molecular studies of Brugia malayi, one of the causative agents of human lymphatic filariasis, an expressed sequence tag (EST)-based gene discovery programme has been carried out. Over 22 000 ESTs have been produced and deposited in the public databases by a consortium of laboratories from endemic and non-endemic countries. The ESTs have been analysed using custom informatic tools to reveal patterns of individual gene expression that may point to potential targets for future research on anti-filarial drugs and vaccines. Many genes first discovered as ESTs are now being analysed by researchers for immunodiagnostic, vaccine and drug target potential. Building on the success of the B. malayi EST programme, significant EST datasets are being generated for a number of other major parasites of humans and domesticated animals, and model parasitic species
Leveling the playing field: Bringing development of biomarkers and molecular diagnostics up to the standards for drug development
Molecular diagnostics are becoming increasingly important in clinical research to stratify or identify molecularly profiled patient cohorts for targeted therapies, to modify the dose of a therapeutic, and to assess early response to therapy or monitor patients. Molecular diagnostics can also be used to identify the pharmacogenetic risk of adverse drug reactions. The articles in this CCR Focus section on molecular diagnosis describe the development and use of markers to guide medical decisions regarding cancer patients. They define sources of preanalytic variability that need to be minimized, as well as the regulatory and financial challenges involved in developing diagnostics and integrating them into clinical practice. They also outline a National Cancer Institute program to assist diagnostic development. Molecular diagnostic clinical tests require rigor in their development and clinical validation, with sensitivity, specificity, and validity comparable to those required for the development of therapeutics. These diagnostics must be offered at a realistic cost that reflects both their clinical value and the costs associated with their development. When genome-sequencing technologies move into the clinic, they must be integrated with and traceable to current technology because they may identify more efficient and accurate approaches to drug development. In addition, regulators may define progressive drug approval for companion diagnostics that requires further evidence regarding efficacy and safety before full approval can be achieved. One way to accomplish this is to emphasize phase IV postmarketing, hypothesis-driven clinical trials with biological characterization that would permit an accurate definition of the association of low-prevalence gene alterations with toxicity or response in large cohorts
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