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Sample records for spotted cdna microarrays

  1. Use of cDNA microarray to isolate differentially expressed genes in White Spot Virus infected shrimp (penaeus stylirostris)

    OpenAIRE

    Dhar, Arun K.; Klimpel, Kurt R.; Bullis, Robert A.; McClenaghan, Leroy R.

    2006-01-01

    White spot syndrome virus (WSSV), the etiologic agent of white spot disease, is currently the most important viral pathogen infecting penaeid shrimp worldwide. Since the initial report, white spot disease has caused losses of catastrophic proportion to shrimp aquaculture globally. Although considerable progress has been made in characterizing the WSSV and developing detection methods, information on the host genes involved in the immune response in shrimp due to WSSV infection is not availabl...

  2. Radioactive cDNA microarray in neurospsychiatry

    International Nuclear Information System (INIS)

    Choe, Jae Gol; Shin, Kyung Ho; Lee, Min Soo; Kim, Meyoung Kon

    2003-01-01

    Microarray technology allows the simultaneous analysis of gene expression patterns of thousands of genes, in a systematic fashion, under a similar set of experimental conditions, thus making the data highly comparable. In some cases arrays are used simply as a primary screen leading to downstream molecular characterization of individual gene candidates. In other cases, the goal of expression profiling is to begin to identify complex regulatory networks underlying developmental processes and disease states. Microarrays were originally used with cell lines or other simple model systems. More recently, microarrays have been used in the analysis of more complex biological tissues including neural systems and the brain. The application of cDNA arrays in neuropsychiatry has lagged behind other fields for a number of reasons. These include a requirement for a large amount of input probe RNA in fluorescent-glass based array systems and the cellular complexity introduced by multicellular brain and neural tissues. An additional factor that impacts the general use of microarrays in neuropsychiatry is the lack of availability of sequenced clone sets from model systems. While human cDNA clones have been widely available, high quality rat, mouse, and drosophilae, among others are just becoming widely available. A final factor in the application of cDNA microarrays in neuropsychiatry is cost of commercial arrays. As academic microarray facilitates become more commonplace custom made arrays will become more widely available at a lower cost allowing more widespread applications. In summary, microarray technology is rapidly having an impact on many areas of biomedical research. Radioisotope-nylon based microarrays offer alternatives that may in some cases be more sensitive, flexible, inexpensive, and universal as compared to other array formats, such as fluorescent-glass arrays. In some situations of limited RNA or exotic species, radioactive membrane microarrays may be the most

  3. Radioactive cDNA microarray in neurospsychiatry

    Energy Technology Data Exchange (ETDEWEB)

    Choe, Jae Gol; Shin, Kyung Ho; Lee, Min Soo; Kim, Meyoung Kon [Korea University Medical School, Seoul (Korea, Republic of)

    2003-02-01

    Microarray technology allows the simultaneous analysis of gene expression patterns of thousands of genes, in a systematic fashion, under a similar set of experimental conditions, thus making the data highly comparable. In some cases arrays are used simply as a primary screen leading to downstream molecular characterization of individual gene candidates. In other cases, the goal of expression profiling is to begin to identify complex regulatory networks underlying developmental processes and disease states. Microarrays were originally used with cell lines or other simple model systems. More recently, microarrays have been used in the analysis of more complex biological tissues including neural systems and the brain. The application of cDNA arrays in neuropsychiatry has lagged behind other fields for a number of reasons. These include a requirement for a large amount of input probe RNA in fluorescent-glass based array systems and the cellular complexity introduced by multicellular brain and neural tissues. An additional factor that impacts the general use of microarrays in neuropsychiatry is the lack of availability of sequenced clone sets from model systems. While human cDNA clones have been widely available, high quality rat, mouse, and drosophilae, among others are just becoming widely available. A final factor in the application of cDNA microarrays in neuropsychiatry is cost of commercial arrays. As academic microarray facilitates become more commonplace custom made arrays will become more widely available at a lower cost allowing more widespread applications. In summary, microarray technology is rapidly having an impact on many areas of biomedical research. Radioisotope-nylon based microarrays offer alternatives that may in some cases be more sensitive, flexible, inexpensive, and universal as compared to other array formats, such as fluorescent-glass arrays. In some situations of limited RNA or exotic species, radioactive membrane microarrays may be the most

  4. Observation of intermittency in gene expression on cDNA microarrays

    CERN Document Server

    Peterson, L E

    2002-01-01

    We used scaled factorial moments to search for intermittency in the log expression ratios (LERs) for thousands of genes spotted on cDNA microarrays (gene chips). Results indicate varying levels of intermittency in gene expression. The observation of intermittency in the data analyzed provides a complimentary handle on moderately expressed genes, generally not tackled by conventional techniques.

  5. cDNA microarray screening in food safety

    International Nuclear Information System (INIS)

    Roy, Sashwati; Sen, Chandan K.

    2006-01-01

    The cDNA microarray technology and related bioinformatics tools presents a wide range of novel application opportunities. The technology may be productively applied to address food safety. In this mini-review article, we present an update highlighting the late breaking discoveries that demonstrate the vitality of cDNA microarray technology as a tool to analyze food safety with reference to microbial pathogens and genetically modified foods. In order to bring the microarray technology to mainstream food safety, it is important to develop robust user-friendly tools that may be applied in a field setting. In addition, there needs to be a standardized process for regulatory agencies to interpret and act upon microarray-based data. The cDNA microarray approach is an emergent technology in diagnostics. Its values lie in being able to provide complimentary molecular insight when employed in addition to traditional tests for food safety, as part of a more comprehensive battery of tests

  6. AN IMPROVED FUZZY CLUSTERING ALGORITHM FOR MICROARRAY IMAGE SPOTS SEGMENTATION

    Directory of Open Access Journals (Sweden)

    V.G. Biju

    2015-11-01

    Full Text Available An automatic cDNA microarray image processing using an improved fuzzy clustering algorithm is presented in this paper. The spot segmentation algorithm proposed uses the gridding technique developed by the authors earlier, for finding the co-ordinates of each spot in an image. Automatic cropping of spots from microarray image is done using these co-ordinates. The present paper proposes an improved fuzzy clustering algorithm Possibility fuzzy local information c means (PFLICM to segment the spot foreground (FG from background (BG. The PFLICM improves fuzzy local information c means (FLICM algorithm by incorporating typicality of a pixel along with gray level information and local spatial information. The performance of the algorithm is validated using a set of simulated cDNA microarray images added with different levels of AWGN noise. The strength of the algorithm is tested by computing the parameters such as the Segmentation matching factor (SMF, Probability of error (pe, Discrepancy distance (D and Normal mean square error (NMSE. SMF value obtained for PFLICM algorithm shows an improvement of 0.9 % and 0.7 % for high noise and low noise microarray images respectively compared to FLICM algorithm. The PFLICM algorithm is also applied on real microarray images and gene expression values are computed.

  7. Microarrays in ecological research: A case study of a cDNA microarray for plant-herbivore interactions

    Directory of Open Access Journals (Sweden)

    Gase Klaus

    2004-09-01

    Full Text Available Abstract Background Microarray technology allows researchers to simultaneously monitor changes in the expression ratios (ERs of hundreds of genes and has thereby revolutionized most of biology. Although this technique has the potential of elucidating early stages in an organism's phenotypic response to complex ecological interactions, to date, it has not been fully incorporated into ecological research. This is partially due to a lack of simple procedures of handling and analyzing the expression ratio (ER data produced from microarrays. Results We describe an analysis of the sources of variation in ERs from 73 hybridized cDNA microarrays, each with 234 herbivory-elicited genes from the model ecological expression system, Nicotiana attenuata, using procedures that are commonly used in ecologic research. Each gene is represented by two independently labeled PCR products and each product was arrayed in quadruplicate. We present a robust method of normalizing and analyzing ERs based on arbitrary thresholds and statistical criteria, and characterize a "norm of reaction" of ERs for 6 genes (4 of known function, 2 of unknown with different ERs as determined across all analyzed arrays to provide a biologically-informed alternative to the use of arbitrary expression ratios in determining significance of expression. These gene-specific ERs and their variance (gene CV were used to calculate array-based variances (array CV, which, in turn, were used to study the effects of array age, probe cDNA quantity and quality, and quality of spotted PCR products as estimates of technical variation. Cluster analysis and a Principal Component Analysis (PCA were used to reveal associations among the transcriptional "imprints" of arrays hybridized with cDNA probes derived from mRNA from N. attenuata plants variously elicited and attacked by different herbivore species and from three congeners: N. quadrivalis, N. longiflora and N. clevelandii. Additionally, the PCA

  8. Radioactive cDNA microarray (II): Gene expression profiling of antidepressant treatment by human cDNA microarray

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Ji Hye; Kang, Rhee Hun; Ham, Byung Joo; Lee, Min Su; Shin, Kyung Ho; Choe, Jae Gol; Kim, Meyoung Kon [College of Medicine, Univ. of Korea, Seoul (Korea, Republic of)

    2003-07-01

    Major depressive disorder is a prevalent psychiatric disorder in primary care, associated with impaired patient functioning and well-being. Fluoxetine is a selective serotonin-reuptake inhibitors (SSRIs) and is a commonly prescribed antidepressant compound. Its action is primarily attributed to selective inhibition of the reuptake of serotonin (5-hydroxytryptamine) in the central nervous system. Objectives ; the aims of this study were two-fold: (1) to determine the usefulness for investigation of the transcription profiles in depression patients, and (2) to assess the differences in gene expression profiles between positive response group and negative response groups by fluoxetine treatment. This study included 53 patients with major depression (26 in positive response group with antidepressant treatment, 27 in negative response group with antidepressant treatment), and 53 healthy controls. To examine the difference of gene expression profile in depression patients, radioactive complementary DNA microarrays were used to evaluate changes in the expression of 1,152 genes in total. Using 33p-labeled probes, this method provided highly sensitive gene expression profiles including brain receptors, drug metabolism, and cellular signaling. Gene transcription profiles were classified into several categories in accordance with the antidepressant gene-regulation. The gene profiles were significantly up-(22 genes) and down-(16 genes) regulated in the positive response group when compared to the control group. Also, in the negative response group, 35 genes were up-regulated and 8 genes were down-regulated when compared to the control group. Consequently, we demonstrated that radioactive human cDNA microarray is highly likely to be an efficient technology for evaluating the gene regulation of antidepressants, such as selective serotonin-reuptake inhibitors (SSRIs), by using high-throughput biotechnology.

  9. Radioactive cDNA microarray (II): Gene expression profiling of antidepressant treatment by human cDNA microarray

    International Nuclear Information System (INIS)

    Lee, Ji Hye; Kang, Rhee Hun; Ham, Byung Joo; Lee, Min Su; Shin, Kyung Ho; Choe, Jae Gol; Kim, Meyoung Kon

    2003-01-01

    Major depressive disorder is a prevalent psychiatric disorder in primary care, associated with impaired patient functioning and well-being. Fluoxetine is a selective serotonin-reuptake inhibitors (SSRIs) and is a commonly prescribed antidepressant compound. Its action is primarily attributed to selective inhibition of the reuptake of serotonin (5-hydroxytryptamine) in the central nervous system. Objectives ; the aims of this study were two-fold: (1) to determine the usefulness for investigation of the transcription profiles in depression patients, and (2) to assess the differences in gene expression profiles between positive response group and negative response groups by fluoxetine treatment. This study included 53 patients with major depression (26 in positive response group with antidepressant treatment, 27 in negative response group with antidepressant treatment), and 53 healthy controls. To examine the difference of gene expression profile in depression patients, radioactive complementary DNA microarrays were used to evaluate changes in the expression of 1,152 genes in total. Using 33p-labeled probes, this method provided highly sensitive gene expression profiles including brain receptors, drug metabolism, and cellular signaling. Gene transcription profiles were classified into several categories in accordance with the antidepressant gene-regulation. The gene profiles were significantly up-(22 genes) and down-(16 genes) regulated in the positive response group when compared to the control group. Also, in the negative response group, 35 genes were up-regulated and 8 genes were down-regulated when compared to the control group. Consequently, we demonstrated that radioactive human cDNA microarray is highly likely to be an efficient technology for evaluating the gene regulation of antidepressants, such as selective serotonin-reuptake inhibitors (SSRIs), by using high-throughput biotechnology

  10. [Preparation of the cDNA microarray on the differential expressed cDNA of senescence-accelerated mouse's hippocampus].

    Science.gov (United States)

    Cheng, Xiao-Rui; Zhou, Wen-Xia; Zhang, Yong-Xiang

    2006-05-01

    Alzheimer' s disease (AD) is the most common form of dementia in the elderly. AD is an invariably fatal neurodegenerative disorder with no effective treatment. Senescence-accelerated mouse prone 8 (SAMP8) is a model for studying age-related cognitive impairments and also is a good model to study brain aging and one of mouse model of AD. The technique of cDNA microarray can monitor the expression levels of thousands of genes simultaneously and can be used to study AD with the character of multi-mechanism, multi-targets and multi-pathway. In order to disclose the mechanism of AD and find the drug targets of AD, cDNA microarray containing 3136 cDNAs amplified from the suppression subtracted cDNA library of hippocampus of SAMP8 and SAMR1 was prepared with 16 blocks and 14 x 14 pins, the housekeeping gene beta-actin and G3PDH as inner conference. The background of this microarray was low and unanimous, and dots divided evenly. The conditions of hybridization and washing were optimized during the hybridization of probe and target molecule. After the data of hybridization analysis, the differential expressed cDNAs were sequenced and analyzed by the bioinformatics, and some of genes were quantified by the real time RT-PCR and the reliability of this cDNA microarray were validated. This cDNA microarray may be the good means to select the differential expressed genes and disclose the molecular mechanism of SAMP8's brain aging and AD.

  11. Advanced spot quality analysis in two-colour microarray experiments

    Directory of Open Access Journals (Sweden)

    Vetter Guillaume

    2008-09-01

    Full Text Available Abstract Background Image analysis of microarrays and, in particular, spot quantification and spot quality control, is one of the most important steps in statistical analysis of microarray data. Recent methods of spot quality control are still in early age of development, often leading to underestimation of true positive microarray features and, consequently, to loss of important biological information. Therefore, improving and standardizing the statistical approaches of spot quality control are essential to facilitate the overall analysis of microarray data and subsequent extraction of biological information. Findings We evaluated the performance of two image analysis packages MAIA and GenePix (GP using two complementary experimental approaches with a focus on the statistical analysis of spot quality factors. First, we developed control microarrays with a priori known fluorescence ratios to verify the accuracy and precision of the ratio estimation of signal intensities. Next, we developed advanced semi-automatic protocols of spot quality evaluation in MAIA and GP and compared their performance with available facilities of spot quantitative filtering in GP. We evaluated these algorithms for standardised spot quality analysis in a whole-genome microarray experiment assessing well-characterised transcriptional modifications induced by the transcription regulator SNAI1. Using a set of RT-PCR or qRT-PCR validated microarray data, we found that the semi-automatic protocol of spot quality control we developed with MAIA allowed recovering approximately 13% more spots and 38% more differentially expressed genes (at FDR = 5% than GP with default spot filtering conditions. Conclusion Careful control of spot quality characteristics with advanced spot quality evaluation can significantly increase the amount of confident and accurate data resulting in more meaningful biological conclusions.

  12. Preparation of fluorescent-dye-labeled cDNA from RNA for microarray hybridization.

    Science.gov (United States)

    Ares, Manuel

    2014-01-01

    This protocol describes how to prepare fluorescently labeled cDNA for hybridization to microarrays. It consists of two steps: first, a mixture of anchored oligo(dT) and random hexamers is used to prime amine-modified cDNA synthesis by reverse transcriptase using a modified deoxynucleotide with a reactive amine group (aminoallyl-dUTP) and an RNA sample as a template. Second, the cDNA is purified and exchanged into bicarbonate buffer so that the amine groups in the cDNA react with the dye N-hydroxysuccinimide (NHS) esters, covalently joining the dye to the cDNA. The dye-coupled cDNA is purified again, and the amount of dye incorporated per microgram of cDNA is determined.

  13. Construction of a cDNA microarray derived from the ascidian Ciona intestinalis.

    Science.gov (United States)

    Azumi, Kaoru; Takahashi, Hiroki; Miki, Yasufumi; Fujie, Manabu; Usami, Takeshi; Ishikawa, Hisayoshi; Kitayama, Atsusi; Satou, Yutaka; Ueno, Naoto; Satoh, Nori

    2003-10-01

    A cDNA microarray was constructed from a basal chordate, the ascidian Ciona intestinalis. The draft genome of Ciona has been read and inferred to contain approximately 16,000 protein-coding genes, and cDNAs for transcripts of 13,464 genes have been characterized and compiled as the "Ciona intestinalis Gene Collection Release I". In the present study, we constructed a cDNA microarray of these 13,464 Ciona genes. A preliminary experiment with Cy3- and Cy5-labeled probes showed extensive differential gene expression between fertilized eggs and larvae. In addition, there was a good correlation between results obtained by the present microarray analysis and those from previous EST analyses. This first microarray of a large collection of Ciona intestinalis cDNA clones should facilitate the analysis of global gene expression and gene networks during the embryogenesis of basal chordates.

  14. APPLICATION OF CDNA MICROARRAY TO THE STUDY OF ARSENIC TOXICOLOGY AND CARCINOGENESIS

    Science.gov (United States)

    Arsenic (As) is a common environmental toxicant and known human carcinogen. Epidemiological studies link As exposure to various disorders and cancers. However, the molecular mechanisms for As toxicity and carcinogenicity are not completely known. The cDNA microarray, a high-th...

  15. GENE EXPRESSION IN THE TESTES OF NORMOSPERMIC VERSUS TERATOSPERMIC DOMESTIC CATS USING HUMAN CDNA MICROARRAY ANALYSES

    Science.gov (United States)

    GENE EXPRESSION IN THE TESTES OF NORMOSPERMIC VERSUS TERATOSPERMIC DOMESTIC CATS USING HUMAN cDNA MICROARRAY ANALYSESB.S. Pukazhenthi1, J. C. Rockett2, M. Ouyang3, D.J. Dix2, J.G. Howard1, P. Georgopoulos4, W.J. J. Welsh3 and D. E. Wildt11Department of Reproductiv...

  16. Transcription analysis of apple fruit development using cDNA microarrays

    NARCIS (Netherlands)

    Soglio, V.; Costa, F.; Molthoff, J.W.; Weemen-Hendriks, M.; Schouten, H.J.; Gianfranceschi, L.

    2009-01-01

    The knowledge of the molecular mechanisms underlying fruit quality traits is fundamental to devise efficient marker-assisted selection strategies and to improve apple breeding. In this study, cDNA microarray technology was used to identify genes whose expression changes during fruit development and

  17. ESTs, cDNA microarrays, and gene expression profiling: tools for dissecting plant physiology and development.

    Science.gov (United States)

    Alba, Rob; Fei, Zhangjun; Payton, Paxton; Liu, Yang; Moore, Shanna L; Debbie, Paul; Cohn, Jonathan; D'Ascenzo, Mark; Gordon, Jeffrey S; Rose, Jocelyn K C; Martin, Gregory; Tanksley, Steven D; Bouzayen, Mondher; Jahn, Molly M; Giovannoni, Jim

    2004-09-01

    Gene expression profiling holds tremendous promise for dissecting the regulatory mechanisms and transcriptional networks that underlie biological processes. Here we provide details of approaches used by others and ourselves for gene expression profiling in plants with emphasis on cDNA microarrays and discussion of both experimental design and downstream analysis. We focus on methods and techniques emphasizing fabrication of cDNA microarrays, fluorescent labeling, cDNA hybridization, experimental design, and data processing. We include specific examples that demonstrate how this technology can be used to further our understanding of plant physiology and development (specifically fruit development and ripening) and for comparative genomics by comparing transcriptome activity in tomato and pepper fruit.

  18. Temporal Gene Expression Profiling of the Wheat Leaf Rust Pathosystem Using cDNA Microarray Reveals Differences in Compatible and Incompatible Defence Pathways

    OpenAIRE

    Fofana, Bourlaye; Banks, Travis W.; McCallum, Brent; Strelkov, Stephen E.; Cloutier, Sylvie

    2007-01-01

    In this study, we detail the construction of a custom cDNA spotted microarray containing 7728 wheat ESTs and the use of the array to identify host genes that are differentially expressed upon challenges with leaf rust fungal pathogens. Wheat cultivar RL6003 (Thatcher Lr1) was inoculated with Puccinia triticina virulence phenotypes BBB (incompatible) or TJB (7-2) (compatible) and sampled at four different time points (3, 6, 12, and 24 hours) after inoculation. Transcript expression levels rela...

  19. Chromosomal Localization of DNA Amplifications in Neuroblastoma Tumors Using cDNA Microarray Comparative Genomic Hybridization

    Directory of Open Access Journals (Sweden)

    Ben Beheshti

    2003-01-01

    Full Text Available Conventional comparative genomic hybridization (CGH profiling of neuroblastomas has identified many genomic aberrations, although the limited resolution has precluded a precise localization of sequences of interest within amplicons. To map high copy number genomic gains in clinically matched stage IV neuroblastomas, CGH analysis using a 19,200-feature cDNA microarray was used. A dedicated (freely available algorithm was developed for rapid in silico determination of chromosomal localizations of microarray cDNA targets, and for generation of an ideogram-type profile of copy number changes. Using these methodologies, novel gene amplifications undetectable by chromosome CGH were identified, and larger MYCN amplicon sizes (in one tumor up to 6 Mb than those previously reported in neuroblastoma were identified. The genes HPCAL1, LPIN1/KIAA0188, NAG, and NSE1/LOC151354 were found to be coamplified with MYCN. To determine whether stage IV primary tumors could be further subclassified based on their genomic copy number profiles, hierarchical clustering was performed. Cluster analysis of microarray CGH data identified three groups: 1 no amplifications evident, 2 a small MYCN amplicon as the only detectable imbalance, and 3 a large MYCN amplicon with additional gene amplifications. Application of CGH to cDNA microarray targets will help to determine both the variation of amplicon size and help better define amplification-dependent and independent pathways of progression in neuroblastoma.

  20. Construction and application of a bovine immune-endocrine cDNA microarray.

    Science.gov (United States)

    Tao, Wenjing; Mallard, Bonnie; Karrow, Niel; Bridle, Byram

    2004-09-01

    A variety of commercial DNA arrays specific for humans and rodents are widely available; however, microarrays containing well-characterized genes to study pathway-specific gene expression are not as accessible for domestic animals, such as cattle, sheep and pigs. Therefore, a small-scale application-targeted bovine immune-endocrine cDNA array was developed to evaluate genetic pathways involved in the immune-endocrine axis of cattle during periods of altered homeostasis provoked by physiological or environmental stressors, such as infection, vaccination or disease. For this purpose, 167 cDNA sequences corresponding to immune, endocrine and inflammatory response genes were collected and categorized. Positive controls included 5 housekeeping genes (glyceraldehydes-3-phosphate dehydrogenase, hypoxanthine phosphoribosyltransferase, ribosomal protein L19, beta-actin, beta2-microglobulin) and bovine genomic DNA. Negative controls were a bacterial gene (Rhodococcus equi 17-kDa virulence-associated protein) and a partial sequence of the plasmid pACYC177. In addition, RNA extracted from un-stimulated, as well as superantigen (Staphylococcus aureus enterotoxin-A, S. aureus Cowan Pansorbin Cells) and mitogen-stimulated (LPS, ConA) bovine blood leukocytes was mixed, reverse transcribed and PCR amplified using gene-specific primers. The endocrine-associated genes were amplified from cDNA derived from un-stimulated bovine hypothalamus, pituitary, adrenal and thyroid gland tissues. The array was constructed in 4 repeating grids of 180 duplicated spots by coupling the PCR amplified 213-630 bp gene fragments onto poly-l-lysine coated glass slides. The bovine immune-endocrine arrays were standardized and preliminary gene expression profiles generated using Cy3 and Cy5 labelled cDNA from un-stimulated and ConA (5 microg/ml) stimulated PBMC of 4 healthy Holstein cows (2-4 replicate arrays/cow) in a time course study. Mononuclear cell-derived cytokine and chemokine (IL-2, IL-1alpha

  1. Spot detection and image segmentation in DNA microarray data.

    Science.gov (United States)

    Qin, Li; Rueda, Luis; Ali, Adnan; Ngom, Alioune

    2005-01-01

    Following the invention of microarrays in 1994, the development and applications of this technology have grown exponentially. The numerous applications of microarray technology include clinical diagnosis and treatment, drug design and discovery, tumour detection, and environmental health research. One of the key issues in the experimental approaches utilising microarrays is to extract quantitative information from the spots, which represent genes in a given experiment. For this process, the initial stages are important and they influence future steps in the analysis. Identifying the spots and separating the background from the foreground is a fundamental problem in DNA microarray data analysis. In this review, we present an overview of state-of-the-art methods for microarray image segmentation. We discuss the foundations of the circle-shaped approach, adaptive shape segmentation, histogram-based methods and the recently introduced clustering-based techniques. We analytically show that clustering-based techniques are equivalent to the one-dimensional, standard k-means clustering algorithm that utilises the Euclidean distance.

  2. CDNA Microarray Based Comparative Gene Expression Analysis of Primary Breast Tumors Versus In Vitro Transformed Neoplastic Breast Epithelium

    National Research Council Canada - National Science Library

    Szallasi, Zoltan

    2001-01-01

    .... The first group of clones is being sorted by their ability to form tumors. We are currently performing cDNA microarray analysis quantifying the expression level of about 15,000 genes in these cell lines...

  3. The effects of radiation on p53-mutated glioma cells using cDNA microarray technique

    International Nuclear Information System (INIS)

    Ngo, F.Q.H.; Hsiao, Y.-Y.H.

    2003-01-01

    Full text: In this study, we investigated the effects of 10-Gy irradiation on cell-cycle arrest, apoptosis and clonogenic death in the p53-mutated human U138MG (malignant glioblastoma) cell line. In order to evaluate time-dependent events in cellular responses to radiation, we did a time course study by incubating cells ranging from 0.5 to 48 hours after irradiation. Cell-cycle distribution and apoptosis were evaluated by flow cytometry using propidium iodide (PI) and annexin-V plus PI staining. Cell viability and proliferative capacity were studied by colony formation assay. Dual fluorescence cDNA microarray technique was used to examine the differential expression patterns of the irradiated cells. The cDNA microarray chips used contained DNA sequences corresponding to 12,814 human genes. From the flow cytometry data, it can be observed that radiation induced G2/M phase arrest and that late apoptosis was more evident following G2/M arrest. After 36 hours, some cells underwent senescence and the remains continued on with the cell cycle. Microarray analyses revealed changes in the expression of a small number of cell-cycle-related genes (p21, cyclin B1, etc.) and cell-death genes (tumor necrosis factors, DDB2, etc.) suggesting their involvement in radiation-induced cell-cycle arrest and apoptosis. In silico interpretations of the molecular mechanisms responsible for these radiation effects are in progress

  4. Development of a porcine skeletal muscle cDNA microarray: analysis of differential transcript expression in phenotypically distinct muscles

    Directory of Open Access Journals (Sweden)

    Stear Michael

    2003-03-01

    Full Text Available Abstract Background Microarray profiling has the potential to illuminate the molecular processes that govern the phenotypic characteristics of porcine skeletal muscles, such as hypertrophy or atrophy, and the expression of specific fibre types. This information is not only important for understanding basic muscle biology but also provides underpinning knowledge for enhancing the efficiency of livestock production. Results We report on the de novo development of a composite skeletal muscle cDNA microarray, comprising 5500 clones from two developmentally distinct cDNA libraries (longissimus dorsi of a 50-day porcine foetus and the gastrocnemius of a 3-day-old pig. Clones selected for the microarray assembly were of low to moderate abundance, as indicated by colony hybridisation. We profiled the differential expression of genes between the psoas (red muscle and the longissimus dorsi (white muscle, by co-hybridisation of Cy3 and Cy5 labelled cDNA derived from these two muscles. Results from seven microarray slides (replicates correctly identified genes that were expected to be differentially expressed, as well as a number of novel candidate regulatory genes. Quantitative real-time RT-PCR on selected genes was used to confirm the results from the microarray. Conclusion We have developed a porcine skeletal muscle cDNA microarray and have identified a number of candidate genes that could be involved in muscle phenotype determination, including several members of the casein kinase 2 signalling pathway.

  5. Microarray and cDNA sequence analysis of transcription during nerve-dependent limb regeneration

    Directory of Open Access Journals (Sweden)

    Bryant Susan V

    2009-01-01

    Full Text Available Abstract Background Microarray analysis and 454 cDNA sequencing were used to investigate a centuries-old problem in regenerative biology: the basis of nerve-dependent limb regeneration in salamanders. Innervated (NR and denervated (DL forelimbs of Mexican axolotls were amputated and transcripts were sampled after 0, 5, and 14 days of regeneration. Results Considerable similarity was observed between NR and DL transcriptional programs at 5 and 14 days post amputation (dpa. Genes with extracellular functions that are critical to wound healing were upregulated while muscle-specific genes were downregulated. Thus, many processes that are regulated during early limb regeneration do not depend upon nerve-derived factors. The majority of the transcriptional differences between NR and DL limbs were correlated with blastema formation; cell numbers increased in NR limbs after 5 dpa and this yielded distinct transcriptional signatures of cell proliferation in NR limbs at 14 dpa. These transcriptional signatures were not observed in DL limbs. Instead, gene expression changes within DL limbs suggest more diverse and protracted wound-healing responses. 454 cDNA sequencing complemented the microarray analysis by providing deeper sampling of transcriptional programs and associated biological processes. Assembly of new 454 cDNA sequences with existing expressed sequence tag (EST contigs from the Ambystoma EST database more than doubled (3935 to 9411 the number of non-redundant human-A. mexicanum orthologous sequences. Conclusion Many new candidate gene sequences were discovered for the first time and these will greatly enable future studies of wound healing, epigenetics, genome stability, and nerve-dependent blastema formation and outgrowth using the axolotl model.

  6. Dissection of the inflammatory bowel disease transcriptome using genome-wide cDNA microarrays.

    Directory of Open Access Journals (Sweden)

    Christine M Costello

    2005-08-01

    Full Text Available BACKGROUND: The differential pathophysiologic mechanisms that trigger and maintain the two forms of inflammatory bowel disease (IBD, Crohn disease (CD, and ulcerative colitis (UC are only partially understood. cDNA microarrays can be used to decipher gene regulation events at a genome-wide level and to identify novel unknown genes that might be involved in perpetuating inflammatory disease progression. METHODS AND FINDINGS: High-density cDNA microarrays representing 33,792 UniGene clusters were prepared. Biopsies were taken from the sigmoid colon of normal controls (n = 11, CD patients (n = 10 and UC patients (n = 10. 33P-radiolabeled cDNA from purified poly(A+ RNA extracted from biopsies (unpooled was hybridized to the arrays. We identified 500 and 272 transcripts differentially regulated in CD and UC, respectively. Interesting hits were independently verified by real-time PCR in a second sample of 100 individuals, and immunohistochemistry was used for exemplary localization. The main findings point to novel molecules important in abnormal immune regulation and the highly disturbed cell biology of colonic epithelial cells in IBD pathogenesis, e.g., CYLD (cylindromatosis, turban tumor syndrome and CDH11 (cadherin 11, type 2. By the nature of the array setup, many of the genes identified were to our knowledge previously uncharacterized, and prediction of the putative function of a subsection of these genes indicate that some could be involved in early events in disease pathophysiology. CONCLUSION: A comprehensive set of candidate genes not previously associated with IBD was revealed, which underlines the polygenic and complex nature of the disease. It points out substantial differences in pathophysiology between CD and UC. The multiple unknown genes identified may stimulate new research in the fields of barrier mechanisms and cell signalling in the context of IBD, and ultimately new therapeutic approaches.

  7. Evaluation of normalization methods for cDNA microarray data by k-NN classification

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Wei; Xing, Eric P; Myers, Connie; Mian, Saira; Bissell, Mina J

    2004-12-17

    Non-biological factors give rise to unwanted variations in cDNA microarray data. There are many normalization methods designed to remove such variations. However, to date there have been few published systematic evaluations of these techniques for removing variations arising from dye biases in the context of downstream, higher-order analytical tasks such as classification. Ten location normalization methods that adjust spatial- and/or intensity-dependent dye biases, and three scale methods that adjust scale differences were applied, individually and in combination, to five distinct, published, cancer biology-related cDNA microarray data sets. Leave-one-out cross-validation (LOOCV) classification error was employed as the quantitative end-point for assessing the effectiveness of a normalization method. In particular, a known classifier, k-nearest neighbor (k-NN), was estimated from data normalized using a given technique, and the LOOCV error rate of the ensuing model was computed. We found that k-NN classifiers are sensitive to dye biases in the data. Using NONRM and GMEDIAN as baseline methods, our results show that single-bias-removal techniques which remove either spatial-dependent dye bias (referred later as spatial effect) or intensity-dependent dye bias (referred later as intensity effect) moderately reduce LOOCV classification errors; whereas double-bias-removal techniques which remove both spatial- and intensity effect reduce LOOCV classification errors even further. Of the 41 different strategies examined, three two-step processes, IGLOESS-SLFILTERW7, ISTSPLINE-SLLOESS and IGLOESS-SLLOESS, all of which removed intensity effect globally and spatial effect locally, appear to reduce LOOCV classification errors most consistently and effectively across all data sets. We also found that the investigated scale normalization methods do not reduce LOOCV classification error. Using LOOCV error of k-NNs as the evaluation criterion, three double

  8. Sharpening spots: correcting for bleedover in cDNA array images.

    Science.gov (United States)

    Therneau, Terry; Tschumper, Renee C; Jelinek, Diane

    2002-03-01

    For cDNA array methods that depend on imaging of a radiolabel, we show that bleedover of one spot onto another, due to the gap between the array and the imaging media, can be a major problem. The images can be sharpened, however, using a blind convolution method based on the EM algorithm. The sharpened images look like a set of donuts, which concurs with our knowledge of the spotting process. Oversharpened images are actually useful as well, in locating the centers of each spot.

  9. A cell spot microarray method for production of high density siRNA transfection microarrays

    Directory of Open Access Journals (Sweden)

    Mpindi John-Patrick

    2011-03-01

    Full Text Available Abstract Background High-throughput RNAi screening is widely applied in biological research, but remains expensive, infrastructure-intensive and conversion of many assays to HTS applications in microplate format is not feasible. Results Here, we describe the optimization of a miniaturized cell spot microarray (CSMA method, which facilitates utilization of the transfection microarray technique for disparate RNAi analyses. To promote rapid adaptation of the method, the concept has been tested with a panel of 92 adherent cell types, including primary human cells. We demonstrate the method in the systematic screening of 492 GPCR coding genes for impact on growth and survival of cultured human prostate cancer cells. Conclusions The CSMA method facilitates reproducible preparation of highly parallel cell microarrays for large-scale gene knockdown analyses. This will be critical towards expanding the cell based functional genetic screens to include more RNAi constructs, allow combinatorial RNAi analyses, multi-parametric phenotypic readouts or comparative analysis of many different cell types.

  10. A comparison of parametric and nonparametric methods for normalising cDNA microarray data.

    Science.gov (United States)

    Khondoker, Mizanur R; Glasbey, Chris A; Worton, Bruce J

    2007-12-01

    Normalisation is an essential first step in the analysis of most cDNA microarray data, to correct for effects arising from imperfections in the technology. Loess smoothing is commonly used to correct for trends in log-ratio data. However, parametric models, such as the additive plus multiplicative variance model, have been preferred for scale normalisation, though the variance structure of microarray data may be of a more complex nature than can be accommodated by a parametric model. We propose a new nonparametric approach that incorporates location and scale normalisation simultaneously using a Generalised Additive Model for Location, Scale and Shape (GAMLSS, Rigby and Stasinopoulos, 2005, Applied Statistics, 54, 507-554). We compare its performance in inferring differential expression with Huber et al.'s (2002, Bioinformatics, 18, 96-104) arsinh variance stabilising transformation (AVST) using real and simulated data. We show GAMLSS to be as powerful as AVST when the parametric model is correct, and more powerful when the model is wrong. (c) 2007 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

  11. Normal uniform mixture differential gene expression detection for cDNA microarrays

    Directory of Open Access Journals (Sweden)

    Raftery Adrian E

    2005-07-01

    Full Text Available Abstract Background One of the primary tasks in analysing gene expression data is finding genes that are differentially expressed in different samples. Multiple testing issues due to the thousands of tests run make some of the more popular methods for doing this problematic. Results We propose a simple method, Normal Uniform Differential Gene Expression (NUDGE detection for finding differentially expressed genes in cDNA microarrays. The method uses a simple univariate normal-uniform mixture model, in combination with new normalization methods for spread as well as mean that extend the lowess normalization of Dudoit, Yang, Callow and Speed (2002 1. It takes account of multiple testing, and gives probabilities of differential expression as part of its output. It can be applied to either single-slide or replicated experiments, and it is very fast. Three datasets are analyzed using NUDGE, and the results are compared to those given by other popular methods: unadjusted and Bonferroni-adjusted t tests, Significance Analysis of Microarrays (SAM, and Empirical Bayes for microarrays (EBarrays with both Gamma-Gamma and Lognormal-Normal models. Conclusion The method gives a high probability of differential expression to genes known/suspected a priori to be differentially expressed and a low probability to the others. In terms of known false positives and false negatives, the method outperforms all multiple-replicate methods except for the Gamma-Gamma EBarrays method to which it offers comparable results with the added advantages of greater simplicity, speed, fewer assumptions and applicability to the single replicate case. An R package called nudge to implement the methods in this paper will be made available soon at http://www.bioconductor.org.

  12. An improved K-means clustering method for cDNA microarray image segmentation.

    Science.gov (United States)

    Wang, T N; Li, T J; Shao, G F; Wu, S X

    2015-07-14

    Microarray technology is a powerful tool for human genetic research and other biomedical applications. Numerous improvements to the standard K-means algorithm have been carried out to complete the image segmentation step. However, most of the previous studies classify the image into two clusters. In this paper, we propose a novel K-means algorithm, which first classifies the image into three clusters, and then one of the three clusters is divided as the background region and the other two clusters, as the foreground region. The proposed method was evaluated on six different data sets. The analyses of accuracy, efficiency, expression values, special gene spots, and noise images demonstrate the effectiveness of our method in improving the segmentation quality.

  13. c-DNA of HIV-1 detection on spot of Buffy-Coat of leukocytes (DBCS)

    OpenAIRE

    Marco Rossi de Gasperis; Maria Daniela Caione; Carlo Concato; Ersilia Fiscarelli; Nicola Di Pietro; Vittorio Salotti; Lorenza Putignani; Donato Menichella; Francesco Callea

    2010-01-01

    Introduction:The elective way for the diagnosis of HIV-1-infection in the window period and in children under the age of 16-18 months is to search virus integrated in leukocytes. Aim of the study was to assess the sensitivity and specificity of extraction from Buffy-Dried Coat Spot (DBCS) in leukocyte to detect c-DNA with nested-PCR in HIV-1-infected individuals compared to Dried Blood Spot (DBS) both extracted by automated instrument EZ1 (QIAGEN, Hilden, Germany). Both DBCS and both DBS were...

  14. Gene expression of panaxydol-treated human melanoma cells using radioactive cDNA microarrays

    Energy Technology Data Exchange (ETDEWEB)

    Cho, Joong Youn; Yu, Su Jin; Soh, Jeong Won; Kim, Meyoung Kon [College of Medicine, Korea Univ., Seoul (Korea, Republic of)

    2001-07-01

    Polyacetylenic alcohols derived from Panax ginseng have been studied to be an anticancer reagent previously. One of the Panax ginseng polyacetylenic alcohols, i.e., panaxydol, has been studied to possess an antiproliferative effect on human melanoma cell line (SK-MEL-1). In ths study, radioactive cDNA microarrays enabled an efficient approach to analyze the pattern of gene expression (3.194 genes in a total) simultaneously. The bioinformatics selection of human cDNAs, which is specifically designed for immunology, apoptosis and signal transduction, were arrayed on nylon membranes. Using with {sup 33}P labeled probes, this method provided highly sensitive gene expression profiles of our interest including apoptosis, cell proliferation, cell cycle, and signal transduction. Gene expression profiles were also classified into several categories in accordance with the duration of panaxydol treatment. Consequently, the gene profiles of our interest were significantly up (199 genes, > 2.0 of Z-ratio) or down-(196 genes, < 2.0 of Z-ratio) regulated in panaxydol-treated human melanoma cells.

  15. Gene expression of panaxydol-treated human melanoma cells using radioactive cDNA microarrays

    International Nuclear Information System (INIS)

    Cho, Joong Youn; Yu, Su Jin; Soh, Jeong Won; Kim, Meyoung Kon

    2001-01-01

    Polyacetylenic alcohols derived from Panax ginseng have been studied to be an anticancer reagent previously. One of the Panax ginseng polyacetylenic alcohols, i.e., panaxydol, has been studied to possess an antiproliferative effect on human melanoma cell line (SK-MEL-1). In ths study, radioactive cDNA microarrays enabled an efficient approach to analyze the pattern of gene expression (3.194 genes in a total) simultaneously. The bioinformatics selection of human cDNAs, which is specifically designed for immunology, apoptosis and signal transduction, were arrayed on nylon membranes. Using with 33 P labeled probes, this method provided highly sensitive gene expression profiles of our interest including apoptosis, cell proliferation, cell cycle, and signal transduction. Gene expression profiles were also classified into several categories in accordance with the duration of panaxydol treatment. Consequently, the gene profiles of our interest were significantly up (199 genes, > 2.0 of Z-ratio) or down-(196 genes, < 2.0 of Z-ratio) regulated in panaxydol-treated human melanoma cells

  16. Examination of gene expression in mice exposed to low dose radiation using affymetrix cDNA microarrays

    Energy Technology Data Exchange (ETDEWEB)

    Morris, D.; Knox, D.; Lavoie, J.; Lemon, J.; Boreham, D. [McMaster Univ., Hamilton, Ontario (Canada)

    2005-07-01

    'Full text:' Gamma radiation acts via the indirect effect to damage cells by producing reactive oxygen species (ROS). These ROS are capable damaging macromolecules and, altering signal pathways and gene transcription. Cells have evolved enzymes and mechanisms to scavenge ROS and repair oxidative damage. Microarrays allow the survey of the gene transcription activity of thousands of genes simultaneously. Messenger RNA is extracted from cells, hybridized with the complementary DNA (cDNA) of a microarray chip, and examined with a chip reader. Affymetrix microarray chips have been produced by the CSCHAH in Winnipeg containing 26000 murine genes. Groups of female mice have been exposed to low dose whole body chronic gamma radiation exposures of 0,50,100, and 120 mGy, corresponding to 15,30,60, and 75 weeks, respectively. MRNA from mice brain tissue has been extracted, isolated, converted to cDNA and labeled. Gene expression in each irradiated mouse was compared to the pooled expression of the control mice. Analysis of gene expression levels are performed with microarray analytical software, Array Pro by Media Cybernetics, and powerful statistical software, BRB microarray tools. Differences in gene expressions, focusing on genes for cytokines, DNA repair mechanisms, immuno-modulators, apoptosis pathways, and enzymatic anti-oxidant systems, are being examined and will be reported. (author)

  17. High-throughput screening of suppression subtractive hybridization cDNA libraries using DNA microarray analysis

    CSIR Research Space (South Africa)

    Van den Berg, N

    2004-11-01

    Full Text Available Efficient construction of cDNA libraries enriched for differentially expressed transcripts is an important first step in many biological investigations. We present a quantitative procedure for screening cDNA libraries constructed by suppression...

  18. c-DNA of HIV-1 detection on spot of Buffy-Coat of leukocytes (DBCS

    Directory of Open Access Journals (Sweden)

    Marco Rossi de Gasperis

    2010-03-01

    Full Text Available Introduction:The elective way for the diagnosis of HIV-1-infection in the window period and in children under the age of 16-18 months is to search virus integrated in leukocytes. Aim of the study was to assess the sensitivity and specificity of extraction from Buffy-Dried Coat Spot (DBCS in leukocyte to detect c-DNA with nested-PCR in HIV-1-infected individuals compared to Dried Blood Spot (DBS both extracted by automated instrument EZ1 (QIAGEN, Hilden, Germany. Both DBCS and both DBS were compared with those tests from whole blood by conventional DNA-extraction Methods: Five ml of whole blood from 50 HIV-infected individuals were collected. 40 μl of each sample were spotted on “FTA ELUTE Micro Card” (Whatman, Inc., Clifton, NJ, 200 μl were extracted according to the manual procedure (QIAGEN “QIAamp DNA minikit and the remaining sample was incubated at 37 °C for 120 minutes. Plasma was centrifuged at 1000 rcf/1g for 10 minutes at room temperature. Forty μl of the obtained buffy-coat was spotted. Both DBCS and both DBS were dried at room temperature for 24 hours.Two of 5 punch from each spot were extracted with TISSUE DNA kit (Biorobot EZ1 DSP “Qiagen” and eluted in 50 μl of buffer.The recovery of genomic DNA was measured amplifying the ß-globin gene by Real-Time “SybrGreen I”.The DNA was amplified for the “pol” gene of HIV-1 by nested PCR and revealed in “SYBR-green I”. Eight HIV-antibody-negative samples were used as internal control. Results:The experimental protocol adopted for the DBCS showed high sensitivity and specificity.The extracted DNA from DBS and DBCS was characterized by excellent quality and without any inhibitory agents. The amount of proviral DNA extracted from DBCS is similar to that obtained by conventional extraction, while the DBS test was significantly less sensitive. Conclusion:These preliminary data suggest that the amount of c-DNA obtained with DBS technique is often not enough for the

  19. Application of a cDNA microarray for profiling the gene expression of Echinococcus granulosus protoscoleces treated with albendazole and artemisinin.

    Science.gov (United States)

    Lü, Guodong; Zhang, Wenbao; Wang, Jianhua; Xiao, Yunfeng; Zhao, Jun; Zhao, Jianqin; Sun, Yimin; Zhang, Chuanshan; Wang, Junhua; Lin, Renyong; Liu, Hui; Zhang, Fuchun; Wen, Hao

    2014-12-01

    Cystic echinoccocosis (CE) is a neglected zoonosis that is caused by the dog-tapeworm Echinococcus granulosus. The disease is endemic worldwide. There is an urgent need for searching effective drug for the treatment of the disease. In this study, we sequenced a cDNA library constructed using RNA isolated from oncospheres, protoscoleces, cyst membrane and adult worms of E. granulosus. A total of 9065 non-redundant or unique sequences were obtained and spotted on chips as uniEST probes to profile the gene expression in protoscoleces of E. granulosus treated with the anthelmintic drugs albendazole and artemisinin, respectively. The results showed that 7 genes were up-regulated and 38 genes were down-regulated in the protoscoleces treated with albendazole. Gene analysis showed that these genes are responsible for energy metabolism, cell cycle and assembly of cell structure. We also identified 100 genes up-regulated and 6 genes down-regulated in the protoscoleces treated with artemisinin. These genes play roles in the transduction of environmental signals, and metabolism. Albendazole appeared its drug efficacy in damaging cell structure, while artemisinin was observed to increase the formation of the heterochromatin in protoscolex cells. Our results highlight the utility of using cDNA microarray methods to detect gene expression profiles of E. granulosus and, in particular, to understand the pharmacologic mechanism of anti-echinococcosis drugs. Copyright © 2014 Elsevier B.V. All rights reserved.

  20. cDNA microarray analysis of human keratinocytes cells of patients submitted to chemoradiotherapy and oral photobiomodulation therapy: pilot study.

    Science.gov (United States)

    Antunes, Heliton S; Wajnberg, Gabriel; Pinho, Marcos B; Jorge, Natasha Andressa Nogueira; de Moraes, Joyce Luana Melo; Stefanoff, Claudio Gustavo; Herchenhorn, Daniel; Araújo, Carlos M M; Viégas, Celia Maria Pais; Rampini, Mariana P; Dias, Fernando L; de Araujo-Souza, Patricia Savio; Passetti, Fabio; Ferreira, Carlos G

    2018-01-01

    Oral mucositis is an acute toxicity that occurs in patients submitted to chemoradiotherapy to treat head and neck squamous cell carcinoma. In this study, we evaluated differences in gene expression in the keratinocytes of the oral mucosa of patients treated with photobiomodulation therapy and tried to associate the molecular mechanisms with clinical findings. From June 2009 to December 2010, 27 patients were included in a randomized double-blind pilot study. Buccal smears from 13 patients were obtained at days 1 and 10 of chemoradiotherapy, and overall gene expression of samples from both dates were analyzed by complementary DNA (cDNA) microarray. In addition, samples from other 14 patients were also collected at D1 and D10 of chemoradiotherapy for subsequent validation of cDNA microarray findings by qPCR. The expression array analysis identified 105 upregulated and 60 downregulated genes in our post-treatment samples when compared with controls. Among the upregulated genes with the highest fold change, it was interesting to observe the presence of genes related to keratinocyte differentiation. Among downregulated genes were observed genes related to cytotoxicity and immune response. The results indicate that genes known to be induced during differentiation of human epidermal keratinocytes were upregulated while genes associated with cytotoxicity and immune response were downregulated in the laser group. These results support previous clinical findings indicating that the lower incidence of oral mucositis associated with photobiomodulation therapy might be correlated to the activation of genes involved in keratinocyte differentiation.

  1. cDNA microarrays as a tool for identification of biomineralization proteins in the coccolithophorid Emiliania huxleyi (Haptophyta).

    Science.gov (United States)

    Quinn, Patrick; Bowers, Robert M; Zhang, Xiaoyu; Wahlund, Thomas M; Fanelli, Michael A; Olszova, Daniela; Read, Betsy A

    2006-08-01

    Marine unicellular coccolithophore algae produce species-specific calcite scales otherwise known as coccoliths. While the coccoliths and their elaborate architecture have attracted the attention of investigators from various scientific disciplines, our knowledge of the underpinnings of the process of biomineralization in this alga is still in its infancy. The processes of calcification and coccolithogenesis are highly regulated and likely to be complex, requiring coordinated expression of many genes and pathways. In this study, we have employed cDNA microarrays to investigate changes in gene expression associated with biomineralization in the most abundant coccolithophorid, Emiliania huxleyi. Expression profiling of cultures grown under calcifying and noncalcifying conditions has been carried out using cDNA microarrays corresponding to approximately 2,300 expressed sequence tags. A total of 127 significantly up- or down-regulated transcripts were identified using a P value of 0.01 and a change of >2.0-fold. Real-time reverse transcriptase PCR was used to test the overall validity of the microarray data, as well as the relevance of many of the proteins predicted to be associated with biomineralization, including a novel gamma-class carbonic anhydrase (A. R. Soto, H. Zheng, D. Shoemaker, J. Rodriguez, B. A. Read, and T. M. Wahlund, Appl. Environ. Microbiol. 72:5500-5511, 2006). Differentially regulated genes include those related to cellular metabolism, ion channels, transport proteins, vesicular trafficking, and cell signaling. The putative function of the vast majority of candidate transcripts could not be defined. Nonetheless, the data described herein represent profiles of the transcription changes associated with biomineralization-related pathways in E. huxleyi and have identified novel and potentially useful targets for more detailed analysis.

  2. Use of the cDNA microarray technology in thesafety assessment of GM food plants

    DEFF Research Database (Denmark)

    Pedersen, Jan W.; Knudsen, Ib; Eriksen, Folmer Damsted

    This report focuses on new analytical approaches that might give more insight into possible changes in a genetically modified plant. Primarily the focus is on the new DNA microarray technique but also proteomics and metabolomics are discussed.The report describes the new techniques and evaluates ...

  3. Use of the cDNA microarray technology in the safety assessment of GM food plants

    NARCIS (Netherlands)

    Kok, E.J.; Kleter, G.A.; Dijk, van J.P.

    2003-01-01

    This report focuses on new analytical approaches that might give more insight into possible changes in a genetically modified plant. Primarily the focus is on the new DNA microarray technique but also proteomics and metabolomics are discussed.The report describes the new techniques and evaluates the

  4. Development of a cDNA microarray for the measurement of gene expression in the sheep scab mite Psoroptes ovis

    Directory of Open Access Journals (Sweden)

    Burgess Stewart TG

    2012-02-01

    Full Text Available Abstract Background Sheep scab is caused by the ectoparasitic mite Psoroptes ovis which initiates a profound cutaneous inflammatory response, leading to the development of the skin lesions which are characteristic of the disease. Existing control strategies rely upon injectable endectocides and acaricidal dips but concerns over residues, eco-toxicity and the development of acaricide resistance limit the sustainability of this approach. In order to identify alternative means of disease control, a deeper understanding of both the parasite and its interaction with the host are required. Methods Herein we describe the development and utilisation of an annotated P. ovis cDNA microarray containing 3,456 elements for the measurement of gene expression in this economically important ectoparasite. The array consists of 981 P. ovis EST sequences printed in triplicate along with 513 control elements. Array performance was validated through the analysis of gene expression differences between fed and starved P. ovis mites. Results Sequences represented on the array include homologues of major house dust mite allergens and tick salivary proteins, along with factors potentially involved in mite reproduction and xenobiotic metabolism. In order to validate the performance of this unique resource under biological conditions we used the array to analyse gene expression differences between fed and starved P. ovis mites. These analyses identified a number of house dust mite allergen homologues up-regulated in fed mites and P. ovis transcripts involved in stress responses, autophagy and chemosensory perception up-regulated in starved mites. Conclusion The P. ovis cDNA microarray described here has been shown to be both robust and reproducible and will enable future studies to analyse gene expression in this important ectoparasite.

  5. Identification of Differentially Expressed IGFBP5-Related Genes in Breast Cancer Tumor Tissues Using cDNA Microarray Experiments.

    Science.gov (United States)

    Akkiprik, Mustafa; Peker, İrem; Özmen, Tolga; Amuran, Gökçe Güllü; Güllüoğlu, Bahadır M; Kaya, Handan; Özer, Ayşe

    2015-11-10

    IGFBP5 is an important regulatory protein in breast cancer progression. We tried to identify differentially expressed genes (DEGs) between breast tumor tissues with IGFBP5 overexpression and their adjacent normal tissues. In this study, thirty-eight breast cancer and adjacent normal breast tissue samples were used to determine IGFBP5 expression by qPCR. cDNA microarrays were applied to the highest IGFBP5 overexpressed tumor samples compared to their adjacent normal breast tissue. Microarray analysis revealed that a total of 186 genes were differentially expressed in breast cancer compared with normal breast tissues. Of the 186 genes, 169 genes were downregulated and 17 genes were upregulated in the tumor samples. KEGG pathway analyses showed that protein digestion and absorption, focal adhesion, salivary secretion, drug metabolism-cytochrome P450, and phenylalanine metabolism pathways are involved. Among these DEGs, the prominent top two genes (MMP11 and COL1A1) which potentially correlated with IGFBP5 were selected for validation using real time RT-qPCR. Only COL1A1 expression showed a consistent upregulation with IGFBP5 expression and COL1A1 and MMP11 were significantly positively correlated. We concluded that the discovery of coordinately expressed genes related with IGFBP5 might contribute to understanding of the molecular mechanism of the function of IGFBP5 in breast cancer. Further functional studies on DEGs and association with IGFBP5 may identify novel biomarkers for clinical applications in breast cancer.

  6. Correlating gene expression to physiological parameters and environmental conditons during cold acclimation of Pinus sylvestris, identification of molecular markers using cDNA microarrays

    NARCIS (Netherlands)

    Joosen, R.V.L.; Lammers, M.; Balk, P.A.; Bronnum, P.; Konings, M.C.J.M.; Perks, M.; Stattin, E.; Wordragen, van M.F.; Geest, van der A.H.M.

    2006-01-01

    Scots pine (Pinus sylvestris L.) seedlings were grown under different conditions (three field locations, two seasons and two climate room regimes), and then analyzed for freezing tolerance of shoots and roots and for transcript abundance in apical buds based on a cDNA microarray containing about

  7. Discovery of distinctive gene expression profiles in rheumatoid synovium using cDNA microarray technology: evidence for the existence of multiple pathways of tissue destruction and repair.

    NARCIS (Netherlands)

    Kraan, TC van der Pouw; Gaalen, van FA; Huizinga, T.W.; Pieterman, E; Breedveld, F.C.; Verweij, C.L.

    2003-01-01

    Rheumatoid arthritis (RA) is a heterogeneous disease. We used cDNA microarray technology to subclassify RA patients and disclose disease pathways in rheumatoid synovium. Hierarchical clustering of gene expression data identified two main groups of tissues (RA-I and RA-II). A total of 121 genes were

  8. cDNA Microarray Analysis of Serially Sampled Cervical Cancer Specimens From Patients Treated With Thermochemoradiotherapy

    International Nuclear Information System (INIS)

    Borkamo, Erling Dahl; Schem, Baard-Christian; Fluge, Oystein; Bruland, Ove; Dahl, Olav; Mella, Olav

    2009-01-01

    Purpose: To elucidate changes in gene expression after treatment with regional thermochemoradiotherapy in locally advanced squamous cell cervical cancer. Methods and Materials: Tru-Cut biopsy specimens were serially collected from 16 patients. Microarray gene expression levels before and 24 h after the first and second trimodality treatment sessions were compared. Pathway and network analyses were conducted by use of Ingenuity Pathways Analysis (IPA; Ingenuity Systems, Redwood City, CA). Single gene expressions were analyzed by quantitative real-time reverse transcription-polymerase chain reaction. Results: We detected 53 annotated genes that were differentially expressed after trimodality treatment. Central in the three top networks detected by IPA were interferon alfa, interferon beta, and interferon gamma receptor; nuclear factor κB; and tumor necrosis factor, respectively. These genes encode proteins that are important in regulation cell signaling, proliferation, gene expression, and immune stimulation. Biological processes over-represented among the 53 genes were fibrosis, tumorigenesis, and immune response. Conclusions: Microarrays showed minor changes in gene expression after thermochemoradiotherapy in locally advanced cervical cancer. We detected 53 differentially expressed genes, mainly involved in fibrosis, tumorigenesis, and immune response. A limitation with the use of serial biopsy specimens was low quality of ribonucleic acid from tumors that respond to highly effective therapy. Another 'key limitation' is timing of the post-treatment biopsy, because 24 h may be too late to adequately assess the impact of hyperthermia on gene expression.

  9. Detecting Outlier Microarray Arrays by Correlation and Percentage of Outliers Spots

    Directory of Open Access Journals (Sweden)

    Song Yang

    2006-01-01

    Full Text Available We developed a quality assurance (QA tool, namely microarray outlier filter (MOF, and have applied it to our microarray datasets for the identification of problematic arrays. Our approach is based on the comparison of the arrays using the correlation coefficient and the number of outlier spots generated on each array to reveal outlier arrays. For a human universal reference (HUR dataset, which is used as a technical control in our standard hybridization procedure, 3 outlier arrays were identified out of 35 experiments. For a human blood dataset, 12 outlier arrays were identified from 185 experiments. In general, arrays from human blood samples displayed greater variation in their gene expression profiles than arrays from HUR samples. As a result, MOF identified two distinct patterns in the occurrence of outlier arrays. These results demonstrate that this methodology is a valuable QA practice to identify questionable microarray data prior to downstream analysis.

  10. A cDNA microarray, UniShrimpChip, for identification of genes relevant to testicular development in the black tiger shrimp (Penaeus monodon

    Directory of Open Access Journals (Sweden)

    Klinbunga Sirawut

    2011-04-01

    Full Text Available Abstract Background Poor reproductive maturation in captive male broodstock of the black tiger shrimp (Penaeus monodon is one of the serious problems to the farming industries. Without genome sequence, EST libraries of P. monodon were previously constructed to identify transcripts with important biological functions. In this study, a new version of cDNA microarray, UniShrimpChip, was constructed from the Peneaus monodon EST libraries of 12 tissues, containing 5,568 non-redundant cDNA clones from 10,536 unique cDNA in the P. monodon EST database. UniShrimpChip was used to study testicular development by comparing gene expression levels of wild brooders from the West and East coasts of Thailand and domesticated brooders with different ages (10-, 14-, 18-month-old. Results The overall gene expression patterns from the microarray experiments revealed distinct transcriptomic patterns between the wild and domesticated groups. Moreover, differentially expressed genes from the microarray comparisons were identified, and the expression patterns of eight selected transcripts were subsequently confirmed by reverse-transcriptase quantitative PCR (RT-qPCR. Among these, expression levels of six subunits (CSN2, 4, 5, 6, 7a, and 8 of the COP9 signalosome (CSN gene family in wild and different ages of domesticated brooders were examined by RT-qPCR. Among the six subunits, CSN5 and CSN6 were most highly expressed in wild brooders and least expressed in the 18-month-old domesticated group; therefore, their full-length cDNA sequences were characterized. Conclusions This study is the first report to employ cDNA microarray to study testicular development in the black tiger shrimp. We show that there are obvious differences between the wild and domesticated shrimp at the transcriptomic level. Furthermore, our study is the first to investigate the feasibility that the CSN gene family might have involved in reproduction and development of this economically important

  11. Importance of the efficiency of double-stranded DNA formation in cDNA synthesis for the imprecision of microarray expression analysis.

    Science.gov (United States)

    Thormar, Hans G; Gudmundsson, Bjarki; Eiriksdottir, Freyja; Kil, Siyoen; Gunnarsson, Gudmundur H; Magnusson, Magnus Karl; Hsu, Jason C; Jonsson, Jon J

    2013-04-01

    The causes of imprecision in microarray expression analysis are poorly understood, limiting the use of this technology in molecular diagnostics. Two-dimensional strandness-dependent electrophoresis (2D-SDE) separates nucleic acid molecules on the basis of length and strandness, i.e., double-stranded DNA (dsDNA), single-stranded DNA (ssDNA), and RNA·DNA hybrids. We used 2D-SDE to measure the efficiency of cDNA synthesis and its importance for the imprecision of an in vitro transcription-based microarray expression analysis. The relative amount of double-stranded cDNA formed in replicate experiments that used the same RNA sample template was highly variable, ranging between 0% and 72% of the total DNA. Microarray experiments showed an inverse relationship between the difference between sample pairs in probe variance and the relative amount of dsDNA. Approximately 15% of probes showed between-sample variation (P cDNA synthesized can be an important component of the imprecision in T7 RNA polymerase-based microarray expression analysis. © 2013 American Association for Clinical Chemistry

  12. Understanding the radiosensitivity of hematopoietic stem cells through CDNA micro-arrays profiling

    Energy Technology Data Exchange (ETDEWEB)

    Pawlik, A.; Cebo, Ch.; Vaigot, P.; Tronik-Le Roux, D. [Evry Univ., Lab. de Genomique et Radiobiologie de l' Hematopoiese, Service de Genomique Fonctionnelle, CEA, 91 (France)

    2006-07-01

    Eradication of circulating hematopoietic cells has been long known to be the first noticeable somatic effect following total body ionizing radiation (IR) exposure. Among these hematopoietic cells a marked differences in sensitivity to IR have been documented reflecting the remarkable degree of heterogeneity in cell type, proliferative capacity and cell cycle status within the bone marrow cells. From all the hematopoietic cells, the small lymphocyte has the greatest radiosensitivity. In fact, a decline in absolute lymphocyte count has been used to assess IR dose in the early phase of observation after IR exposure. At moderate doses, bone marrow recovery is triggered by the differentiation of stem/early progenitor cells, which confirms further their differential sensitivity to radiation exposure. Although differences in radiosensitivity of the stem cell pool have also been documented, little is known from a molecular viewpoint. To gain insight into the molecular programs underlying the response o f hematopoietic cells to radiation exposure, we have applied a genome wide analysis strategy based on cDNA micro arrays. This technology offers a unique opportunity to dissect complex biological process by assessing three types of questions, which are, in order of complexity: Which genes are differentially expressed among the samples studied:Which genes are expressed in a coordinated manner and what are the regulators involved,what are the global biological pathways mobilized. To answer these questions transcriptional changes occurring after exposure of mice to whole body irradiation (2 Gy) were monitored in bone marrow and spleen. The time course was established in vivo and encompassed the reversible eradication of cells. For each kinetic point RNA was collected from both, spleen or sorted B.M. populations from irradiated and sham irradiated mice. The sham irradiated mice were used to eliminate stress modifications due to handling.The results highlight numerous

  13. Monitoring expression profiles of rice (Oryza sativa L.) genes under abiotic stresses using cDNA Microarray Analysis (abstract)

    International Nuclear Information System (INIS)

    Rabbani, M.A.

    2005-01-01

    Transcript regulation in response to cold, drought, high salinity and ABA application was investigated in rice (Oryza sativa L., Nipponbare) with microarray analysis including approx. 1700 independent DNA elements derived from three cDNA libraries constructed from 15-day old rice seedlings stressed with drought, cold and high salinity. A total of 141 non-redundant genes were identified, whose expression ratios were more than three-fold compared with the control genes for at least one of stress treatments in microarray analysis. However, after RNA gel blot analysis, a total of 73 genes were identified, among them the transcripts of 36, 62, 57 and 43 genes were found increased after cold, drought, high salinity and ABA application, respectively. Sixteen of these identified genes have been reported previously to be stress inducible in rice, while 57 of which are novel that have not been reported earlier as stress responsive in rice. We observed a strong association in the expression patterns of stress responsive genes and found 15 stress inducible genes that responded to all four treatments. Based on Venn diagram analysis, 56 genes were induced by both drought and high salinity, whereas 22 genes were upregulated by both cold and high salinity stress. Similarly 43 genes were induced by both drought stress and ABA application, while only 17 genes were identified as cold and ABA inducible genes. These results indicated the existence of greater cross talk between drought, ABA and high salinity stress signaling processes than those between cold and ABA, and cold and high salinity stress signaling pathways. The cold, drought, high salinity and ABA inducible genes were classified into four gene groups from their expression profiles. Analysis of data enabled us to identify a number of promoters and possible cis-acting DNA elements of several genes induced by a variety of abiotic stresses by combining expression data with genomic sequence data of rice. Comparative analysis of

  14. Transcriptomic identification of candidate genes involved in sunflower responses to chilling and salt stresses based on cDNA microarray analysis

    Directory of Open Access Journals (Sweden)

    Paniego Norma

    2008-01-01

    Full Text Available Abstract Background Considering that sunflower production is expanding to arid regions, tolerance to abiotic stresses as drought, low temperatures and salinity arises as one of the main constrains nowadays. Differential organ-specific sunflower ESTs (expressed sequence tags were previously generated by a subtractive hybridization method that included a considerable number of putative abiotic stress associated sequences. The objective of this work is to analyze concerted gene expression profiles of organ-specific ESTs by fluorescence microarray assay, in response to high sodium chloride concentration and chilling treatments with the aim to identify and follow up candidate genes for early responses to abiotic stress in sunflower. Results Abiotic-related expressed genes were the target of this characterization through a gene expression analysis using an organ-specific cDNA fluorescence microarray approach in response to high salinity and low temperatures. The experiment included three independent replicates from leaf samples. We analyzed 317 unigenes previously isolated from differential organ-specific cDNA libraries from leaf, stem and flower at R1 and R4 developmental stage. A statistical analysis based on mean comparison by ANOVA and ordination by Principal Component Analysis allowed the detection of 80 candidate genes for either salinity and/or chilling stresses. Out of them, 50 genes were up or down regulated under both stresses, supporting common regulatory mechanisms and general responses to chilling and salinity. Interestingly 15 and 12 sequences were up regulated or down regulated specifically in one stress but not in the other, respectively. These genes are potentially involved in different regulatory mechanisms including transcription/translation/protein degradation/protein folding/ROS production or ROS-scavenging. Differential gene expression patterns were confirmed by qRT-PCR for 12.5% of the microarray candidate sequences. Conclusion

  15. Extending Immunological Profiling in the Gilthead Sea Bream, Sparus aurata, by Enriched cDNA Library Analysis, Microarray Design and Initial Studies upon the Inflammatory Response to PAMPs

    Directory of Open Access Journals (Sweden)

    Sebastian Boltaña

    2017-02-01

    Full Text Available This study describes the development and validation of an enriched oligonucleotide-microarray platform for Sparus aurata (SAQ to provide a platform for transcriptomic studies in this species. A transcriptome database was constructed by assembly of gilthead sea bream sequences derived from public repositories of mRNA together with reads from a large collection of expressed sequence tags (EST from two extensive targeted cDNA libraries characterizing mRNA transcripts regulated by both bacterial and viral challenge. The developed microarray was further validated by analysing monocyte/macrophage activation profiles after challenge with two Gram-negative bacterial pathogen-associated molecular patterns (PAMPs; lipopolysaccharide (LPS and peptidoglycan (PGN. Of the approximately 10,000 EST sequenced, we obtained a total of 6837 EST longer than 100 nt, with 3778 and 3059 EST obtained from the bacterial-primed and from the viral-primed cDNA libraries, respectively. Functional classification of contigs from the bacterial- and viral-primed cDNA libraries by Gene Ontology (GO showed that the top five represented categories were equally represented in the two libraries: metabolism (approximately 24% of the total number of contigs, carrier proteins/membrane transport (approximately 15%, effectors/modulators and cell communication (approximately 11%, nucleoside, nucleotide and nucleic acid metabolism (approximately 7.5% and intracellular transducers/signal transduction (approximately 5%. Transcriptome analyses using this enriched oligonucleotide platform identified differential shifts in the response to PGN and LPS in macrophage-like cells, highlighting responsive gene-cassettes tightly related to PAMP host recognition. As observed in other fish species, PGN is a powerful activator of the inflammatory response in S. aurata macrophage-like cells. We have developed and validated an oligonucleotide microarray (SAQ that provides a platform enriched for the study

  16. Extending Immunological Profiling in the Gilthead Sea Bream, Sparus aurata, by Enriched cDNA Library Analysis, Microarray Design and Initial Studies upon the Inflammatory Response to PAMPs.

    Science.gov (United States)

    Boltaña, Sebastian; Castellana, Barbara; Goetz, Giles; Tort, Lluis; Teles, Mariana; Mulero, Victor; Novoa, Beatriz; Figueras, Antonio; Goetz, Frederick W; Gallardo-Escarate, Cristian; Planas, Josep V; Mackenzie, Simon

    2017-02-03

    This study describes the development and validation of an enriched oligonucleotide-microarray platform for Sparus aurata (SAQ) to provide a platform for transcriptomic studies in this species. A transcriptome database was constructed by assembly of gilthead sea bream sequences derived from public repositories of mRNA together with reads from a large collection of expressed sequence tags (EST) from two extensive targeted cDNA libraries characterizing mRNA transcripts regulated by both bacterial and viral challenge. The developed microarray was further validated by analysing monocyte/macrophage activation profiles after challenge with two Gram-negative bacterial pathogen-associated molecular patterns (PAMPs; lipopolysaccharide (LPS) and peptidoglycan (PGN)). Of the approximately 10,000 EST sequenced, we obtained a total of 6837 EST longer than 100 nt, with 3778 and 3059 EST obtained from the bacterial-primed and from the viral-primed cDNA libraries, respectively. Functional classification of contigs from the bacterial- and viral-primed cDNA libraries by Gene Ontology (GO) showed that the top five represented categories were equally represented in the two libraries: metabolism (approximately 24% of the total number of contigs), carrier proteins/membrane transport (approximately 15%), effectors/modulators and cell communication (approximately 11%), nucleoside, nucleotide and nucleic acid metabolism (approximately 7.5%) and intracellular transducers/signal transduction (approximately 5%). Transcriptome analyses using this enriched oligonucleotide platform identified differential shifts in the response to PGN and LPS in macrophage-like cells, highlighting responsive gene-cassettes tightly related to PAMP host recognition. As observed in other fish species, PGN is a powerful activator of the inflammatory response in S. aurata macrophage-like cells. We have developed and validated an oligonucleotide microarray (SAQ) that provides a platform enriched for the study of gene

  17. Statistical Analysis of Microarray Data with Replicated Spots: A Case Study with Synechococcus WH8102

    Directory of Open Access Journals (Sweden)

    E. V. Thomas

    2009-01-01

    Full Text Available Until recently microarray experiments often involved relatively few arrays with only a single representation of each gene on each array. A complete genome microarray with multiple spots per gene (spread out spatially across the array was developed in order to compare the gene expression of a marine cyanobacterium and a knockout mutant strain in a defined artificial seawater medium. Statistical methods were developed for analysis in the special situation of this case study where there is gene replication within an array and where relatively few arrays are used, which can be the case with current array technology. Due in part to the replication within an array, it was possible to detect very small changes in the levels of expression between the wild type and mutant strains. One interesting biological outcome of this experiment is the indication of the extent to which the phosphorus regulatory system of this cyanobacterium affects the expression of multiple genes beyond those strictly involved in phosphorus acquisition.

  18. Systematic spatial bias in DNA microarray hybridization is caused by probe spot position-dependent variability in lateral diffusion.

    Science.gov (United States)

    Steger, Doris; Berry, David; Haider, Susanne; Horn, Matthias; Wagner, Michael; Stocker, Roman; Loy, Alexander

    2011-01-01

    The hybridization of nucleic acid targets with surface-immobilized probes is a widely used assay for the parallel detection of multiple targets in medical and biological research. Despite its widespread application, DNA microarray technology still suffers from several biases and lack of reproducibility, stemming in part from an incomplete understanding of the processes governing surface hybridization. In particular, non-random spatial variations within individual microarray hybridizations are often observed, but the mechanisms underpinning this positional bias remain incompletely explained. This study identifies and rationalizes a systematic spatial bias in the intensity of surface hybridization, characterized by markedly increased signal intensity of spots located at the boundaries of the spotted areas of the microarray slide. Combining observations from a simplified single-probe block array format with predictions from a mathematical model, the mechanism responsible for this bias is found to be a position-dependent variation in lateral diffusion of target molecules. Numerical simulations reveal a strong influence of microarray well geometry on the spatial bias. Reciprocal adjustment of the size of the microarray hybridization chamber to the area of surface-bound probes is a simple and effective measure to minimize or eliminate the diffusion-based bias, resulting in increased uniformity and accuracy of quantitative DNA microarray hybridization.

  19. Genome-Wide Screening of Genes Showing Altered Expression in Liver Metastases of Human Colorectal Cancers by cDNA Microarray

    Directory of Open Access Journals (Sweden)

    Rempei Yanagawa

    2001-01-01

    Full Text Available In spite of intensive and increasingly successful attempts to determine the multiple steps involved in colorectal carcinogenesis, the mechanisms responsible for metastasis of colorectal tumors to the liver remain to be clarified. To identify genes that are candidates for involvement in the metastatic process, we analyzed genome-wide expression profiles of 10 primary colorectal cancers and their corresponding metastatic lesions by means of a cDNA microarray consisting of 9121 human genes. This analysis identified 40 genes whose expression was commonly upregulated in metastatic lesions, and 7 that were commonly downregulated. The upregulated genes encoded proteins involved in cell adhesion, or remodeling of the actin cytoskeleton. Investigation of the functions of more of the altered genes should improve our understanding of metastasis and may identify diagnostic markers and/or novel molecular targets for prevention or therapy of metastatic lesions.

  20. USE OF cDNA MICROARRAY TECHNOLOGY FOR IDENTIFICATION OF NOVEL GENES RESPONDING TO ABSCISIC ACID PHYTOHORMONE

    OpenAIRE

    D. Cabezas; Sandra Pérez; R. Huelva; Dong Haitao; Li Debao

    2005-01-01

    Se utilizó el cDNA microarreglo con 4370 unigenes, provenientes de la biblioteca del endospermo del arroz y de los tejidos de las hojas, para detectar los niveles de expresión del mRNA de los tejidos del tallo del arroz tratados con agua y con la hormona ácido abscísico (ABA). Los resultados mostraron que los niveles de expresión de cinco genes fueron reprimidos por la fitohormona ABA. El Reverse Northern confirmó que uno de los cinco genes (H024g06) fue realmente reprimido por el ABA. Los an...

  1. Hematopoietic Lineage Transcriptome Stability and Representation in PAXgeneTM Collected Peripheral Blood Utilising SPIA Single-Stranded cDNA Probes for Microarray

    Directory of Open Access Journals (Sweden)

    Laura Kennedy

    2008-01-01

    Full Text Available Peripheral blood as a surrogate tissue for transcriptome profiling holds great promise for the discovery of diagnostic and prognostic disease biomarkers, particularly when target tissues of disease are not readily available. To maximize the reliability of gene expression data generated from clinical blood samples, both the sample collection and the microarray probe generation methods should be optimized to provide stabilized, reproducible and representative gene expression profiles faithfully representing the transcriptional profiles of the constituent blood cell types present in the circulation. Given the increasing innovation in this field in recent years, we investigated a combination of methodological advances in both RNA stabilisation and microarray probe generation with the goal of achieving robust, reliable and representative transcriptional profiles from whole blood. To assess the whole blood profiles, the transcriptomes of purified blood cell types were measured and compared with the global transcriptomes measured in whole blood. The results demonstrate that a combination of PAXgeneTM RNA stabilising technology and single-stranded cDNA probe generation afforded by the NuGEN Ovation RNA amplification system V2TM enables an approach that yields faithful representation of specific hematopoietic cell lineage transcriptomes in whole blood without the necessity for prior sample fractionation, cell enrichment or globin reduction. Storage stability assessments of the PAXgeneTM blood samples also advocate a short, fixed room temperature storage time for all PAXgeneTM blood samples collected for the purposes of global transcriptional profiling in clinical studies.

  2. Hematopoietic Lineage Transcriptome Stability and Representation in PAXgene Collected Peripheral Blood Utilising SPIA Single-Stranded cDNA Probes for Microarray.

    Science.gov (United States)

    Kennedy, Laura; Vass, J Keith; Haggart, D Ross; Moore, Steve; Burczynski, Michael E; Crowther, Dan; Miele, Gino

    2008-08-25

    Peripheral blood as a surrogate tissue for transcriptome profiling holds great promise for the discovery of diagnostic and prognostic disease biomarkers, particularly when target tissues of disease are not readily available. To maximize the reliability of gene expression data generated from clinical blood samples, both the sample collection and the microarray probe generation methods should be optimized to provide stabilized, reproducible and representative gene expression profiles faithfully representing the transcriptional profiles of the constituent blood cell types present in the circulation. Given the increasing innovation in this field in recent years, we investigated a combination of methodological advances in both RNA stabilisation and microarray probe generation with the goal of achieving robust, reliable and representative transcriptional profiles from whole blood. To assess the whole blood profiles, the transcriptomes of purified blood cell types were measured and compared with the global transcriptomes measured in whole blood. The results demonstrate that a combination of PAXgene() RNA stabilising technology and single-stranded cDNA probe generation afforded by the NuGEN Ovation RNA amplification system V2() enables an approach that yields faithful representation of specific hematopoietic cell lineage transcriptomes in whole blood without the necessity for prior sample fractionation, cell enrichment or globin reduction. Storage stability assessments of the PAXgene() blood samples also advocate a short, fixed room temperature storage time for all PAXgene() blood samples collected for the purposes of global transcriptional profiling in clinical studies.

  3. Hematopoietic Lineage Transcriptome Stability and Representation in PAXgene™ Collected Peripheral Blood Utilising SPIA Single-Stranded cDNA Probes for Microarray

    Science.gov (United States)

    Kennedy, Laura; Vass, J. Keith; Haggart, D. Ross; Moore, Steve; Burczynski, Michael E.; Crowther, Dan; Miele, Gino

    2008-01-01

    Peripheral blood as a surrogate tissue for transcriptome profiling holds great promise for the discovery of diagnostic and prognostic disease biomarkers, particularly when target tissues of disease are not readily available. To maximize the reliability of gene expression data generated from clinical blood samples, both the sample collection and the microarray probe generation methods should be optimized to provide stabilized, reproducible and representative gene expression profiles faithfully representing the transcriptional profiles of the constituent blood cell types present in the circulation. Given the increasing innovation in this field in recent years, we investigated a combination of methodological advances in both RNA stabilisation and microarray probe generation with the goal of achieving robust, reliable and representative transcriptional profiles from whole blood. To assess the whole blood profiles, the transcriptomes of purified blood cell types were measured and compared with the global transcriptomes measured in whole blood. The results demonstrate that a combination of PAXgene™ RNA stabilising technology and single-stranded cDNA probe generation afforded by the NuGEN Ovation RNA amplification system V2™ enables an approach that yields faithful representation of specific hematopoietic cell lineage transcriptomes in whole blood without the necessity for prior sample fractionation, cell enrichment or globin reduction. Storage stability assessments of the PAXgene™ blood samples also advocate a short, fixed room temperature storage time for all PAXgene™ blood samples collected for the purposes of global transcriptional profiling in clinical studies. PMID:19578521

  4. APPLICATION OF CDNA MICROARRAY TECHNOLOGY TO IN VITRO TOXICOLOGY AND THE SELECTION OF GENES FOR A REAL TIME RT-PCR-BASED SCREEN FOR OXIDATIVE STRESS IN HEP-G2 CELLS

    Science.gov (United States)

    Large-scale analysis of gene expression using cDNA microarrays promises therapid detection of the mode of toxicity for drugs and other chemicals. cDNAmicroarrays were used to examine chemically-induced alterations of geneexpression in HepG2 cells exposed to oxidative ...

  5. Identification of late O{sub 3}-responsive genes in Arabidopsis thaliana by cDNA microarray analysis

    Energy Technology Data Exchange (ETDEWEB)

    D' Haese, D. [Univ. of Antwerp, Dept. of Biology, Antwerp (BE) and Univ. of Newcastle, School of Biology and Psychology, Div. of Biology, Newcastle-Upon-Tyne (United Kingdom); Horemans, N.; Coen, W. De; Guisez, Y. [Univ. of Antwerp, Dept. of Biology, Antwerp (Belgium)

    2006-09-15

    To better understand the response of a plant to 0{sub 3} stress, an integrated microarray analysis was performed on Arabidopsis plants exposed during 2 days to purified air or 150 nl l{sup -1} O{sub 3}, 8 h day-l. Agilent Arabidopsis 2 Oligo Microarrays were used of which the reliability was confirmed by quantitative real-time PCR of nine randomly selected genes. We confirmed the O{sub 3} responsiveness of heat shock proteins (HSPs), glutathione-S-tranferases and genes involved in cell wall stiffening and microbial defence. Whereas, a previous study revealed that during an early stage of the O{sub 3} stress response, gene expression was strongly dependent on jasmonic acid and ethylene, we report that at a later stage (48 h) synthesis of jasrnonic acid and ethylene was downregulated. In addition, we observed the simultaneous induction of salicylic acid synthesis and genes involved in programmed cell death and senescence. Also typically, the later stage of the response to O{sub 3} appeared to be the induction of the complete pathway leading to the biosynthesis of anthocyanin diglucosides and the induction of thioredoxin-based redox control. Surprisingly absent in the list of induced genes were genes involved in ASC-dependent antioxidation, few of which were found to be induced after 12 h of 0{sub 3} exposure in another study. We discuss these and other particular results of the microarray analysis and provide a map depicting significantly affected genes and their pathways highlighting their interrelationships and subcellular localization. (au)

  6. Gene expression profiling in gill tissues of White spot syndrome virus infected black tiger shrimp Penaeus monodon by DNA microarray.

    Science.gov (United States)

    Shekhar, M S; Gomathi, A; Gopikrishna, G; Ponniah, A G

    2015-06-01

    White spot syndrome virus (WSSV) continues to be the most devastating viral pathogen infecting penaeid shrimp the world over. The genome of WSSV has been deciphered and characterized from three geographical isolates and significant progress has been made in developing various molecular diagnostic methods to detect the virus. However, the information on host immune gene response to WSSV pathogenesis is limited. Microarray analysis was carried out as an approach to analyse the gene expression in black tiger shrimp Penaeus monodon in response to WSSV infection. Gill tissues collected from the WSSV infected shrimp at 6, 24, 48 h and moribund stage were analysed for differential gene expression. Shrimp cDNAs of 40,059 unique sequences were considered for designing the microarray chip. The Cy3-labeled cRNA derived from healthy and WSSV-infected shrimp was subjected to hybridization with all the DNA spots in the microarray which revealed 8,633 and 11,147 as up- and down-regulated genes respectively at different time intervals post infection. The altered expression of these numerous genes represented diverse functions such as immune response, osmoregulation, apoptosis, nucleic acid binding, energy and metabolism, signal transduction, stress response and molting. The changes in gene expression profiles observed by microarray analysis provides molecular insights and framework of genes which are up- and down-regulated at different time intervals during WSSV infection in shrimp. The microarray data was validated by Real Time analysis of four differentially expressed genes involved in apoptosis (translationally controlled tumor protein, inhibitor of apoptosis protein, ubiquitin conjugated enzyme E2 and caspase) for gene expression levels. The role of apoptosis related genes in WSSV infected shrimp is discussed herein.

  7. Molecular characterisation of the early response in pigs to experimental infection with Actinobacillus pleuropneumoniae using cDNA microarrays

    DEFF Research Database (Denmark)

    Hedegaard, Jakob; Skovgaard, Kerstin; Mortensen, Shila

    2007-01-01

    Background: The bacterium Actinobacillus pleuropneumoniae is responsible for porcine pleuropneumonia, a widespread, highly contagious and often fatal respiratory disease of pigs. The general porcine innate immune response after A. pleuropneumoniae infection is still not clarified. The objective...... lymph node tissue were hybridised to an expanded version of the porcine microarray with 26879 unique PCR products. Results: A total of 357 genes differed significantly in expression between infected and non-infected lung tissue, 713 genes differed in expression in liver tissue from infected versus non-infected...... animals and 130 genes differed in expression in tracheobronchial lymph node tissue from infected versus non-infected animals. Among these genes, several have previously been described to be part of a general host response to infections encoding immune response related proteins. In inflamed lung tissue...

  8. Analysis of the effects of sex hormone background on the rat choroid plexus transcriptome by cDNA microarrays.

    Directory of Open Access Journals (Sweden)

    Telma Quintela

    Full Text Available The choroid plexus (CP are highly vascularized branched structures that protrude into the ventricles of the brain, and form a unique interface between the blood and the cerebrospinal fluid (CSF, the blood-CSF barrier, that are the main site of production and secretion of CSF. Sex hormones are widely recognized as neuroprotective agents against several neurodegenerative diseases, and the presence of sex hormones cognate receptors suggest that it may be a target for these hormones. In an effort to provide further insight into the neuroprotective mechanisms triggered by sex hormones we analyzed gene expression differences in the CP of female and male rats subjected to gonadectomy, using microarray technology. In gonadectomized female and male animals, 3045 genes were differentially expressed by 1.5-fold change, compared to sham controls. Analysis of the CP transcriptome showed that the top-five pathways significantly regulated by the sex hormone background are olfactory transduction, taste transduction, metabolism, steroid hormone biosynthesis and circadian rhythm pathways. These results represent the first overview of global expression changes in CP of female and male rats induced by gonadectomy and suggest that sex hormones are implicated in pathways with central roles in CP functions and CSF homeostasis.

  9. Gene organization in rice revealed by full-length cDNA mapping and gene expression analysis through microarray.

    Directory of Open Access Journals (Sweden)

    Kouji Satoh

    Full Text Available Rice (Oryza sativa L. is a model organism for the functional genomics of monocotyledonous plants since the genome size is considerably smaller than those of other monocotyledonous plants. Although highly accurate genome sequences of indica and japonica rice are available, additional resources such as full-length complementary DNA (FL-cDNA sequences are also indispensable for comprehensive analyses of gene structure and function. We cross-referenced 28.5K individual loci in the rice genome defined by mapping of 578K FL-cDNA clones with the 56K loci predicted in the TIGR genome assembly. Based on the annotation status and the presence of corresponding cDNA clones, genes were classified into 23K annotated expressed (AE genes, 33K annotated non-expressed (ANE genes, and 5.5K non-annotated expressed (NAE genes. We developed a 60mer oligo-array for analysis of gene expression from each locus. Analysis of gene structures and expression levels revealed that the general features of gene structure and expression of NAE and ANE genes were considerably different from those of AE genes. The results also suggested that the cloning efficiency of rice FL-cDNA is associated with the transcription activity of the corresponding genetic locus, although other factors may also have an effect. Comparison of the coverage of FL-cDNA among gene families suggested that FL-cDNA from genes encoding rice- or eukaryote-specific domains, and those involved in regulatory functions were difficult to produce in bacterial cells. Collectively, these results indicate that rice genes can be divided into distinct groups based on transcription activity and gene structure, and that the coverage bias of FL-cDNA clones exists due to the incompatibility of certain eukaryotic genes in bacteria.

  10. Differential gene expression in a DNA double-strand-break repair mutant XRS-5 defective in Ku80. Analysis by cDNA microarray

    Energy Technology Data Exchange (ETDEWEB)

    Chan, John Y.H.; Chen, Lung-Kun; Chang, Jui-Feng [National Yang Ming Univ., Taipei, Taiwan (China). Inst. of Radiological Sciences] (and others)

    2001-12-01

    The ability of cells to rejoin DNA double-strand breaks (DSBs) usually correlates with their radiosensitivity. This correlation has been demonstrated in radiosensitive cells, including the Chinese hamster ovary mutant XRS-5. XRS-5 is defective in a DNA end-binding protein, Ku80, which is a component of a DNA-dependent protein kinase complex used for joining strand breaks. However, Ku80-deficient cells are known to be retarded in cell proliferation and growth as well as other yet to be identified defects. Using custom-made 600-gene cDNA microarray filters, we found differential gene expressions between the wild-type and XRS-5 cells. Defective Ku80 apparently affects the expression of several repair genes, including topoisomerase-I and -IIA, ERCC5, MLH1, and ATM. In contrast, other DNA repair-associated genes, such as GADD45A, EGR1 MDM2 and p53, were not affected. In addition, for large numbers of growth-associated genes, such as cyclins and clks, the growth factors and cytokines were also affected. Down-regulated expression was also found in several categories of seemingly unrelated genes, including apoptosis, angiogenesis, kinase and signaling, phosphatase, stress protein, proto-oncogenes and tumor suppressors, transcription and translation factors. A RT-PCR analysis confirmed that the XRS-5 cells used were defective in Ku80 expression. The diversified groups of genes being affected could mean that Ku80, a multi-functional DNA-binding protein, not only affects DNA repair, but is also involved in transcription regulation. Our data, taken together, indicate that there are specific genes being modulated in Ku80- deficient cells, and that some of the DNA repair pathways and other biological functions are apparently linked, suggesting that a defect in one gene could have global effects on many other processes. (author)

  11. Differential gene expression in a DNA double-strand-break repair mutant XRS-5 defective in Ku80. Analysis by cDNA microarray

    International Nuclear Information System (INIS)

    Chan, John Y.H.; Chen, Lung-Kun; Chang, Jui-Feng

    2001-01-01

    The ability of cells to rejoin DNA double-strand breaks (DSBs) usually correlates with their radiosensitivity. This correlation has been demonstrated in radiosensitive cells, including the Chinese hamster ovary mutant XRS-5. XRS-5 is defective in a DNA end-binding protein, Ku80, which is a component of a DNA-dependent protein kinase complex used for joining strand breaks. However, Ku80-deficient cells are known to be retarded in cell proliferation and growth as well as other yet to be identified defects. Using custom-made 600-gene cDNA microarray filters, we found differential gene expressions between the wild-type and XRS-5 cells. Defective Ku80 apparently affects the expression of several repair genes, including topoisomerase-I and -IIA, ERCC5, MLH1, and ATM. In contrast, other DNA repair-associated genes, such as GADD45A, EGR1 MDM2 and p53, were not affected. In addition, for large numbers of growth-associated genes, such as cyclins and clks, the growth factors and cytokines were also affected. Down-regulated expression was also found in several categories of seemingly unrelated genes, including apoptosis, angiogenesis, kinase and signaling, phosphatase, stress protein, proto-oncogenes and tumor suppressors, transcription and translation factors. A RT-PCR analysis confirmed that the XRS-5 cells used were defective in Ku80 expression. The diversified groups of genes being affected could mean that Ku80, a multi-functional DNA-binding protein, not only affects DNA repair, but is also involved in transcription regulation. Our data, taken together, indicate that there are specific genes being modulated in Ku80- deficient cells, and that some of the DNA repair pathways and other biological functions are apparently linked, suggesting that a defect in one gene could have global effects on many other processes. (author)

  12. Gene expression analysis of the rat testis after treatment with di(2-ethylhexyl) phthalate using cDNA microarray and real-time RT-PCR

    International Nuclear Information System (INIS)

    Kijima, Kazuyasu; Toyosawa, Kaoru; Yasuba, Masashi; Matsuoka, Nobuo; Adachi, Tetsuya; Komiyama, Masatoshi; Mori, Chisato

    2004-01-01

    To investigate the effects of di(2-ethylhexyl) phthalate (DEHP) on gene expression in rat testis, 6-week-old male Sprague-Dawley rats were given a single oral dose of 20 or 2000 mg/kg and euthanized 3, 6, 24, or 72 h thereafter. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL)-positive cells were significantly increased in the testis at 24 and 72 h after the exposure to 2000 mg/kg of DEHP. On cDNA microarray analysis, in addition to apoptosis-related genes, genes associated with atrophy, APEX nuclease, MutS homologue (E. coli), testosterone-repressed-prostatic-message-2 (TRPM-2), connective tissue growth factor, collagen alpha 2 type V, and cell adhesion kinase were differentially expressed. To investigate the relationship between histopathological alteration and gene expression, we selected genes associated with apoptosis and analyzed their expression by real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR). With 20 mg/kg of DEHP treatment, bcl-2, key gene related to apoptosis, was increased. Up-regulation of bcl-2, inhibitor of Apaf-1/caspase-9/caspase-2 cascade of apoptosis, may be related to the fact that no morphological apoptotic change was induced after dosing of 20 mg/kg DEHP. With 2000 mg/kg of DEHP treatment, the apoptotic activator cascade, Fas/FasL, FADD/caspase-8/caspase-3 cascade, and Apaf-1/caspase-9/caspase-2 cascade were increased and bcl-2 was decreased. Thus, these gene regulations might lead the cells into apoptosis in the case of high exposure to DEHP. In contrast, FADD/caspase-10/caspase-6 cascade and caspase-11/caspase-3 cascade were not increased. These results indicate that the cascades of FADD/caspase-10/caspase-6 and caspase-11/caspase-3 are not related to apoptosis with DEHP treatment

  13. Analysis of cellular responses to aflatoxin B1 in yeast expressing human cytochrome P450 1A2 using cDNA microarrays

    International Nuclear Information System (INIS)

    Guo Yingying; Breeden, Linda L.; Fan, Wenhong; Zhao Lueping; Eaton, David L.; Zarbl, Helmut

    2006-01-01

    Aflatoxin B1 (AFB 1 ) is a potent human hepatotoxin and hepatocarcinogen produced by the mold Aspergillus flavus. In human, AFB 1 is bioactivated by cytochrome P450 (CYP450) enzymes, primarily CYP1A2, to the genotoxic epoxide that forms N 7 -guanine DNA adducts. To characterize the transcriptional responses to genotoxic insults from AFB 1 , a strain of Saccharomyces cerevisiae engineered to express human CYP1A2 was exposed to doses of AFB 1 that resulted in minimal lethality, but substantial genotoxicity. Flow cytometric analysis demonstrated a dose and time dependent S phase delay under the same treatment conditions, indicating a checkpoint response to DNA damage. Replicate cDNA microarray analyses of AFB 1 treated cells showed that about 200 genes were significantly affected by the exposure. The genes activated by AFB 1 -treatment included RAD51, DUN1 and other members of the DNA damage response signature reported in a previous study with methylmethane sulfonate and ionizing radiation [A.P. Gasch, M. Huang, S. Metzner, D. Botstein, S.J. Elledge, P.O. Brown, Genomic expression responses to DNA-damaging agents and the regulatory role of the yeast ATR homolog Mec1p, Mol. Biol. Cell 12 (2001) 2987-3003]. However, unlike previous studies using highly cytotoxic doses, environmental stress response genes [A.P. Gasch, P.T. Spellman, C.M. Kao, O. Carmel-Harel, M.B. Eisen, G. Storz, D. Botstein, P.O. Brown, Genomic expression programs in the response of yeast cells to environmental changes, Mol. Biol. Cell 11 (2000) 4241-4257] were largely unaffected by our dosing regimen. About half of the transcripts affected are also known to be cell cycle regulated. The most strongly repressed transcripts were those encoding the histone genes and a group of genes that are cell cycle regulated and peak in M phase and early G1. These include most of the known daughter-specific genes. The rapid and coordinated repression of histones and M/G1-specific transcripts cannot be explained by

  14. Analysis of cellular responses to aflatoxin B{sub 1} in yeast expressing human cytochrome P450 1A2 using cDNA microarrays

    Energy Technology Data Exchange (ETDEWEB)

    Guo Yingying [Departmental of Environmental and Occupational Health Sciences, University of Washington, Seattle, WA (United States); Fred Hutchinson Cancer Research Center, Seattle, WA (United States); Breeden, Linda L. [Fred Hutchinson Cancer Research Center, Seattle, WA (United States); Fan, Wenhong [Fred Hutchinson Cancer Research Center, Seattle, WA (United States); Zhao Lueping [Fred Hutchinson Cancer Research Center, Seattle, WA (United States); Eaton, David L. [Departmental of Environmental and Occupational Health Sciences, University of Washington, Seattle, WA (United States); Fred Hutchinson Cancer Research Center, Seattle, WA (United States); Zarbl, Helmut [Departmental of Environmental and Occupational Health Sciences, University of Washington, Seattle, WA (United States) and Fred Hutchinson Cancer Research Center, Seattle, WA (United States)]. E-mail: hzarbl@fhcrc.org

    2006-01-29

    Aflatoxin B1 (AFB{sub 1}) is a potent human hepatotoxin and hepatocarcinogen produced by the mold Aspergillus flavus. In human, AFB{sub 1} is bioactivated by cytochrome P450 (CYP450) enzymes, primarily CYP1A2, to the genotoxic epoxide that forms N{sup 7}-guanine DNA adducts. To characterize the transcriptional responses to genotoxic insults from AFB{sub 1}, a strain of Saccharomyces cerevisiae engineered to express human CYP1A2 was exposed to doses of AFB{sub 1} that resulted in minimal lethality, but substantial genotoxicity. Flow cytometric analysis demonstrated a dose and time dependent S phase delay under the same treatment conditions, indicating a checkpoint response to DNA damage. Replicate cDNA microarray analyses of AFB{sub 1} treated cells showed that about 200 genes were significantly affected by the exposure. The genes activated by AFB{sub 1}-treatment included RAD51, DUN1 and other members of the DNA damage response signature reported in a previous study with methylmethane sulfonate and ionizing radiation [A.P. Gasch, M. Huang, S. Metzner, D. Botstein, S.J. Elledge, P.O. Brown, Genomic expression responses to DNA-damaging agents and the regulatory role of the yeast ATR homolog Mec1p, Mol. Biol. Cell 12 (2001) 2987-3003]. However, unlike previous studies using highly cytotoxic doses, environmental stress response genes [A.P. Gasch, P.T. Spellman, C.M. Kao, O. Carmel-Harel, M.B. Eisen, G. Storz, D. Botstein, P.O. Brown, Genomic expression programs in the response of yeast cells to environmental changes, Mol. Biol. Cell 11 (2000) 4241-4257] were largely unaffected by our dosing regimen. About half of the transcripts affected are also known to be cell cycle regulated. The most strongly repressed transcripts were those encoding the histone genes and a group of genes that are cell cycle regulated and peak in M phase and early G1. These include most of the known daughter-specific genes. The rapid and coordinated repression of histones and M/G1-specific

  15. Microarray and RT-PCR screening for white spot syndrome virus immediate-early genes in cycloheximide-treated shrimp

    International Nuclear Information System (INIS)

    Liu Wangjing; Chang Yunshiang; Wang Chunghsiung; Kou, Guang-Hsiung; Lo Chufang

    2005-01-01

    Here, we report for the first time the successful use of cycloheximide (CHX) as an inhibitor to block de novo viral protein synthesis during WSSV (white spot syndrome virus) infection. Sixty candidate IE (immediate-early) genes were identified using a global analysis microarray technique. RT-PCR showed that the genes corresponding to ORF126, ORF242 and ORF418 in the Taiwan isolate were consistently CHX-insensitive, and these genes were designated ie1, ie2 and ie3, respectively. The sequences for these IE genes also appear in the two other WSSV isolates that have been sequenced. Three corresponding ORFs were identified in the China WSSV isolate, but only an ORF corresponding to ie1 was predicted in the Thailand isolate. In a promoter activity assay in Sf9 insect cells using EGFP (enhanced green fluorescence protein) as a reporter, ie1 showed very strong promoter activity, producing higher EGFP signals than the insect Orgyia pseudotsugata multicapsid nuclear polyhedrosis virus (OpMNPV) ie2 promoter

  16. Application of four dyes in gene expression analyses by microarrays

    Directory of Open Access Journals (Sweden)

    van Schooten Frederik J

    2005-07-01

    Full Text Available Abstract Background DNA microarrays are widely used in gene expression analyses. To increase throughput and minimize costs without reducing gene expression data obtained, we investigated whether four mRNA samples can be analyzed simultaneously by applying four different fluorescent dyes. Results Following tests for cross-talk of fluorescence signals, Alexa 488, Alexa 594, Cyanine 3 and Cyanine 5 were selected for hybridizations. For self-hybridizations, a single RNA sample was labelled with all dyes and hybridized on commercial cDNA arrays or on in-house spotted oligonucleotide arrays. Correlation coefficients for all combinations of dyes were above 0.9 on the cDNA array. On the oligonucleotide array they were above 0.8, except combinations with Alexa 488, which were approximately 0.5. Standard deviation of expression differences for replicate spots were similar on the cDNA array for all dye combinations, but on the oligonucleotide array combinations with Alexa 488 showed a higher variation. Conclusion In conclusion, the four dyes can be used simultaneously for gene expression experiments on the tested cDNA array, but only three dyes can be used on the tested oligonucleotide array. This was confirmed by hybridizations of control with test samples, as all combinations returned similar numbers of differentially expressed genes with comparable effects on gene expression.

  17. [Screening of phosphoprotein associated with glycosphingolipid microdomains 1 (PAG1) by cDNA microarray and influence of overexpression of PAG1 on biologic behavior of human metastatic prostatic cancer cell line in vitro].

    Science.gov (United States)

    Yu, Wen-juan; Wang, Yue-wei; Xie, Zhi-gang; You, Jiang-feng; Wang, Jie-liang; Cui, Xiang-lin; Pei, Fei; Zheng, Jie

    2010-02-01

    To screen for novel gene(s) associated with tumor metastasis, and to investigate the effect of overexpression of phosphoprotein associated with glycosphingolipid microdomains 1 (PAG1) on the biological behaviors of human prostatic cancer cell line PC-3M-1E8 in vitro. Four cDNA microarrays were constructed using cDNA library of prostatic cancer cells PC-3M-1E8 (high metastatic potential), PC-3M-2B4 (low metastatic potential), lung cancer cells PG-BE1 (high metastatic potential)and PG-LH7 (low metastatic potential)to screen genes which were differentially expressed according to their different metastatic properties. From a battery of differentially expressed genes, PAG1, which was markedly downregulated in both high metastatic sublines of PC-3M and PG was chosen for further investigation. Real-time PCR and Western blot were used to confirm the gene expression of PAG1 at mRNA and protein levels. Full-length coding sequence of human PAG1 was subcloned into plasmid pcDNA3.0 and the recombinant plasmids were stably transfected into PC-3M-1E8. The cell proliferation ability, anchorage-independent growth, cell cycle distribution, apoptosis rates and invasive ability were detected by MTT, and in addition, soft agar colony formation, flow cytometry analysis and matrigel invasion assay using Boyden chamber were also carried out respectively. All experiments contained pcDNA3.0-PAG1-transfected clones, vector transfected clones and non-transfected parental cells. A total of 327 differentially expressed genes were obtained between the high and low metastatic sublines of PC-3M cells, including 123 upregulated and 204 downregulated genes in PC-3M-1E8. A total of 281 genes, including 167 upregulated and 114 downregulated genes were obtained in PG-BE1 cells. Nine genes were simultaneously downregulated and 8 genes were upregulated in both high metastatic cell lines of PC-3M and PG. The expression of PAG1 at mRNA and protein level were decreased in the high metastatic subline PC-3M-1

  18. Plant-pathogen interactions: what microarray tells about it?

    Science.gov (United States)

    Lodha, T D; Basak, J

    2012-01-01

    Plant defense responses are mediated by elementary regulatory proteins that affect expression of thousands of genes. Over the last decade, microarray technology has played a key role in deciphering the underlying networks of gene regulation in plants that lead to a wide variety of defence responses. Microarray is an important tool to quantify and profile the expression of thousands of genes simultaneously, with two main aims: (1) gene discovery and (2) global expression profiling. Several microarray technologies are currently in use; most include a glass slide platform with spotted cDNA or oligonucleotides. Till date, microarray technology has been used in the identification of regulatory genes, end-point defence genes, to understand the signal transduction processes underlying disease resistance and its intimate links to other physiological pathways. Microarray technology can be used for in-depth, simultaneous profiling of host/pathogen genes as the disease progresses from infection to resistance/susceptibility at different developmental stages of the host, which can be done in different environments, for clearer understanding of the processes involved. A thorough knowledge of plant disease resistance using successful combination of microarray and other high throughput techniques, as well as biochemical, genetic, and cell biological experiments is needed for practical application to secure and stabilize yield of many crop plants. This review starts with a brief introduction to microarray technology, followed by the basics of plant-pathogen interaction, the use of DNA microarrays over the last decade to unravel the mysteries of plant-pathogen interaction, and ends with the future prospects of this technology.

  19. A probabilistic framework for microarray data analysis: fundamental probability models and statistical inference.

    Science.gov (United States)

    Ogunnaike, Babatunde A; Gelmi, Claudio A; Edwards, Jeremy S

    2010-05-21

    Gene expression studies generate large quantities of data with the defining characteristic that the number of genes (whose expression profiles are to be determined) exceed the number of available replicates by several orders of magnitude. Standard spot-by-spot analysis still seeks to extract useful information for each gene on the basis of the number of available replicates, and thus plays to the weakness of microarrays. On the other hand, because of the data volume, treating the entire data set as an ensemble, and developing theoretical distributions for these ensembles provides a framework that plays instead to the strength of microarrays. We present theoretical results that under reasonable assumptions, the distribution of microarray intensities follows the Gamma model, with the biological interpretations of the model parameters emerging naturally. We subsequently establish that for each microarray data set, the fractional intensities can be represented as a mixture of Beta densities, and develop a procedure for using these results to draw statistical inference regarding differential gene expression. We illustrate the results with experimental data from gene expression studies on Deinococcus radiodurans following DNA damage using cDNA microarrays. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

  20. Carbohydrate microarrays

    DEFF Research Database (Denmark)

    Park, Sungjin; Gildersleeve, Jeffrey C; Blixt, Klas Ola

    2012-01-01

    In the last decade, carbohydrate microarrays have been core technologies for analyzing carbohydrate-mediated recognition events in a high-throughput fashion. A number of methods have been exploited for immobilizing glycans on the solid surface in a microarray format. This microarray...... of substrate specificities of glycosyltransferases. This review covers the construction of carbohydrate microarrays, detection methods of carbohydrate microarrays and their applications in biological and biomedical research....

  1. Advanced microarray technologies for clinical diagnostics

    NARCIS (Netherlands)

    Pierik, Anke

    2011-01-01

    DNA microarrays become increasingly important in the field of clinical diagnostics. These microarrays, also called DNA chips, are small solid substrates, typically having a maximum surface area of a few cm2, onto which many spots are arrayed in a pre-determined pattern. Each of these spots contains

  2. Microarray BASICA: Background Adjustment, Segmentation, Image Compression and Analysis of Microarray Images

    Directory of Open Access Journals (Sweden)

    Jianping Hua

    2004-01-01

    Full Text Available This paper presents microarray BASICA: an integrated image processing tool for background adjustment, segmentation, image compression, and analysis of cDNA microarray images. BASICA uses a fast Mann-Whitney test-based algorithm to segment cDNA microarray images, and performs postprocessing to eliminate the segmentation irregularities. The segmentation results, along with the foreground and background intensities obtained with the background adjustment, are then used for independent compression of the foreground and background. We introduce a new distortion measurement for cDNA microarray image compression and devise a coding scheme by modifying the embedded block coding with optimized truncation (EBCOT algorithm (Taubman, 2000 to achieve optimal rate-distortion performance in lossy coding while still maintaining outstanding lossless compression performance. Experimental results show that the bit rate required to ensure sufficiently accurate gene expression measurement varies and depends on the quality of cDNA microarray images. For homogeneously hybridized cDNA microarray images, BASICA is able to provide from a bit rate as low as 5 bpp the gene expression data that are 99% in agreement with those of the original 32 bpp images.

  3. Direct calibration of PICKY-designed microarrays

    Directory of Open Access Journals (Sweden)

    Ronald Pamela C

    2009-10-01

    Full Text Available Abstract Background Few microarrays have been quantitatively calibrated to identify optimal hybridization conditions because it is difficult to precisely determine the hybridization characteristics of a microarray using biologically variable cDNA samples. Results Using synthesized samples with known concentrations of specific oligonucleotides, a series of microarray experiments was conducted to evaluate microarrays designed by PICKY, an oligo microarray design software tool, and to test a direct microarray calibration method based on the PICKY-predicted, thermodynamically closest nontarget information. The complete set of microarray experiment results is archived in the GEO database with series accession number GSE14717. Additional data files and Perl programs described in this paper can be obtained from the website http://www.complex.iastate.edu under the PICKY Download area. Conclusion PICKY-designed microarray probes are highly reliable over a wide range of hybridization temperatures and sample concentrations. The microarray calibration method reported here allows researchers to experimentally optimize their hybridization conditions. Because this method is straightforward, uses existing microarrays and relatively inexpensive synthesized samples, it can be used by any lab that uses microarrays designed by PICKY. In addition, other microarrays can be reanalyzed by PICKY to obtain the thermodynamically closest nontarget information for calibration.

  4. Gene Expression Analysis Using Agilent DNA Microarrays

    DEFF Research Database (Denmark)

    Stangegaard, Michael

    2009-01-01

    Hybridization of labeled cDNA to microarrays is an intuitively simple and a vastly underestimated process. If it is not performed, optimized, and standardized with the same attention to detail as e.g., RNA amplification, information may be overlooked or even lost. Careful balancing of the amount ...

  5. Normalization and gene p-value estimation: issues in microarray data processing.

    Science.gov (United States)

    Fundel, Katrin; Küffner, Robert; Aigner, Thomas; Zimmer, Ralf

    2008-05-28

    Numerous methods exist for basic processing, e.g. normalization, of microarray gene expression data. These methods have an important effect on the final analysis outcome. Therefore, it is crucial to select methods appropriate for a given dataset in order to assure the validity and reliability of expression data analysis. Furthermore, biological interpretation requires expression values for genes, which are often represented by several spots or probe sets on a microarray. How to best integrate spot/probe set values into gene values has so far been a somewhat neglected problem. We present a case study comparing different between-array normalization methods with respect to the identification of differentially expressed genes. Our results show that it is feasible and necessary to use prior knowledge on gene expression measurements to select an adequate normalization method for the given data. Furthermore, we provide evidence that combining spot/probe set p-values into gene p-values for detecting differentially expressed genes has advantages compared to combining expression values for spots/probe sets into gene expression values. The comparison of different methods suggests to use Stouffer's method for this purpose. The study has been conducted on gene expression experiments investigating human joint cartilage samples of osteoarthritis related groups: a cDNA microarray (83 samples, four groups) and an Affymetrix (26 samples, two groups) data set. The apparently straight forward steps of gene expression data analysis, e.g. between-array normalization and detection of differentially regulated genes, can be accomplished by numerous different methods. We analyzed multiple methods and the possible effects and thereby demonstrate the importance of the single decisions taken during data processing. We give guidelines for evaluating normalization outcomes. An overview of these effects via appropriate measures and plots compared to prior knowledge is essential for the biological

  6. Improved intra-array and interarray normalization of peptide microarray phosphorylation for phosphorylome and kinome profiling by rational selection of relevant spots

    Science.gov (United States)

    Scholma, Jetse; Fuhler, Gwenny M.; Joore, Jos; Hulsman, Marc; Schivo, Stefano; List, Alan F.; Reinders, Marcel J. T.; Peppelenbosch, Maikel P.; Post, Janine N.

    2016-01-01

    Massive parallel analysis using array technology has become the mainstay for analysis of genomes and transcriptomes. Analogously, the predominance of phosphorylation as a regulator of cellular metabolism has fostered the development of peptide arrays of kinase consensus substrates that allow the charting of cellular phosphorylation events (often called kinome profiling). However, whereas the bioinformatical framework for expression array analysis is well-developed, no advanced analysis tools are yet available for kinome profiling. Especially intra-array and interarray normalization of peptide array phosphorylation remain problematic, due to the absence of “housekeeping” kinases and the obvious fallacy of the assumption that different experimental conditions should exhibit equal amounts of kinase activity. Here we describe the development of analysis tools that reliably quantify phosphorylation of peptide arrays and that allow normalization of the signals obtained. We provide a method for intraslide gradient correction and spot quality control. We describe a novel interarray normalization procedure, named repetitive signal enhancement, RSE, which provides a mathematical approach to limit the false negative results occuring with the use of other normalization procedures. Using in silico and biological experiments we show that employing such protocols yields superior insight into cellular physiology as compared to classical analysis tools for kinome profiling. PMID:27225531

  7. MICROARRAY IMAGE GRIDDING USING GRID LINE REFINEMENT TECHNIQUE

    Directory of Open Access Journals (Sweden)

    V.G. Biju

    2015-05-01

    Full Text Available An important stage in microarray image analysis is gridding. Microarray image gridding is done to locate sub arrays in a microarray image and find co-ordinates of spots within each sub array. For accurate identification of spots, most of the proposed gridding methods require human intervention. In this paper a fully automatic gridding method which enhances spot intensity in the preprocessing step as per a histogram based threshold method is used. The gridding step finds co-ordinates of spots from horizontal and vertical profile of the image. To correct errors due to the grid line placement, a grid line refinement technique is proposed. The algorithm is applied on different image databases and results are compared based on spot detection accuracy and time. An average spot detection accuracy of 95.06% depicts the proposed method’s flexibility and accuracy in finding the spot co-ordinates for different database images.

  8. The EADGENE Microarray Data Analysis Workshop

    DEFF Research Database (Denmark)

    de Koning, Dirk-Jan; Jaffrézic, Florence; Lund, Mogens Sandø

    2007-01-01

    Microarray analyses have become an important tool in animal genomics. While their use is becoming widespread, there is still a lot of ongoing research regarding the analysis of microarray data. In the context of a European Network of Excellence, 31 researchers representing 14 research groups from...... 10 countries performed and discussed the statistical analyses of real and simulated 2-colour microarray data that were distributed among participants. The real data consisted of 48 microarrays from a disease challenge experiment in dairy cattle, while the simulated data consisted of 10 microarrays...... statistical weights, to omitting a large number of spots or omitting entire slides. Surprisingly, these very different approaches gave quite similar results when applied to the simulated data, although not all participating groups analysed both real and simulated data. The workshop was very successful...

  9. Microarrays for global expression constructed with a low redundancy set of 27,500 sequenced cDNAs representing an array of developmental stages and physiological conditions of the soybean plant

    Directory of Open Access Journals (Sweden)

    Retzel Ernest

    2004-09-01

    Full Text Available Abstract Background Microarrays are an important tool with which to examine coordinated gene expression. Soybean (Glycine max is one of the most economically valuable crop species in the world food supply. In order to accelerate both gene discovery as well as hypothesis-driven research in soybean, global expression resources needed to be developed. The applications of microarray for determining patterns of expression in different tissues or during conditional treatments by dual labeling of the mRNAs are unlimited. In addition, discovery of the molecular basis of traits through examination of naturally occurring variation in hundreds of mutant lines could be enhanced by the construction and use of soybean cDNA microarrays. Results We report the construction and analysis of a low redundancy 'unigene' set of 27,513 clones that represent a variety of soybean cDNA libraries made from a wide array of source tissue and organ systems, developmental stages, and stress or pathogen-challenged plants. The set was assembled from the 5' sequence data of the cDNA clones using cluster analysis programs. The selected clones were then physically reracked and sequenced at the 3' end. In order to increase gene discovery from immature cotyledon libraries that contain abundant mRNAs representing storage protein gene families, we utilized a high density filter normalization approach to preferentially select more weakly expressed cDNAs. All 27,513 cDNA inserts were amplified by polymerase chain reaction. The amplified products, along with some repetitively spotted control or 'choice' clones, were used to produce three 9,728-element microarrays that have been used to examine tissue specific gene expression and global expression in mutant isolines. Conclusions Global expression studies will be greatly aided by the availability of the sequence-validated and low redundancy cDNA sets described in this report. These cDNAs and ESTs represent a wide array of developmental

  10. Extended -Regular Sequence for Automated Analysis of Microarray Images

    Directory of Open Access Journals (Sweden)

    Jin Hee-Jeong

    2006-01-01

    Full Text Available Microarray study enables us to obtain hundreds of thousands of expressions of genes or genotypes at once, and it is an indispensable technology for genome research. The first step is the analysis of scanned microarray images. This is the most important procedure for obtaining biologically reliable data. Currently most microarray image processing systems require burdensome manual block/spot indexing work. Since the amount of experimental data is increasing very quickly, automated microarray image analysis software becomes important. In this paper, we propose two automated methods for analyzing microarray images. First, we propose the extended -regular sequence to index blocks and spots, which enables a novel automatic gridding procedure. Second, we provide a methodology, hierarchical metagrid alignment, to allow reliable and efficient batch processing for a set of microarray images. Experimental results show that the proposed methods are more reliable and convenient than the commercial tools.

  11. Liver spots

    Science.gov (United States)

    ... skin changes - liver spots; Senile or solar lentigines; Skin spots - aging; Age spots ... Liver spots are changes in skin color that occur in older skin. The coloring may be due to aging, exposure to the sun ...

  12. Multiplex cDNA quantification method that facilitates the standardization of gene expression data

    Science.gov (United States)

    Gotoh, Osamu; Murakami, Yasufumi; Suyama, Akira

    2011-01-01

    Microarray-based gene expression measurement is one of the major methods for transcriptome analysis. However, current microarray data are substantially affected by microarray platforms and RNA references because of the microarray method can provide merely the relative amounts of gene expression levels. Therefore, valid comparisons of the microarray data require standardized platforms, internal and/or external controls and complicated normalizations. These requirements impose limitations on the extensive comparison of gene expression data. Here, we report an effective approach to removing the unfavorable limitations by measuring the absolute amounts of gene expression levels on common DNA microarrays. We have developed a multiplex cDNA quantification method called GEP-DEAN (Gene expression profiling by DCN-encoding-based analysis). The method was validated by using chemically synthesized DNA strands of known quantities and cDNA samples prepared from mouse liver, demonstrating that the absolute amounts of cDNA strands were successfully measured with a sensitivity of 18 zmol in a highly multiplexed manner in 7 h. PMID:21415008

  13. Development and application of a microarray meter tool to optimize microarray experiments

    Directory of Open Access Journals (Sweden)

    Rouse Richard JD

    2008-07-01

    Full Text Available Abstract Background Successful microarray experimentation requires a complex interplay between the slide chemistry, the printing pins, the nucleic acid probes and targets, and the hybridization milieu. Optimization of these parameters and a careful evaluation of emerging slide chemistries are a prerequisite to any large scale array fabrication effort. We have developed a 'microarray meter' tool which assesses the inherent variations associated with microarray measurement prior to embarking on large scale projects. Findings The microarray meter consists of nucleic acid targets (reference and dynamic range control and probe components. Different plate designs containing identical probe material were formulated to accommodate different robotic and pin designs. We examined the variability in probe quality and quantity (as judged by the amount of DNA printed and remaining post-hybridization using three robots equipped with capillary printing pins. Discussion The generation of microarray data with minimal variation requires consistent quality control of the (DNA microarray manufacturing and experimental processes. Spot reproducibility is a measure primarily of the variations associated with printing. The microarray meter assesses array quality by measuring the DNA content for every feature. It provides a post-hybridization analysis of array quality by scoring probe performance using three metrics, a a measure of variability in the signal intensities, b a measure of the signal dynamic range and c a measure of variability of the spot morphologies.

  14. Spot table - RPD | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available switchLanguage; BLAST Search Image Search Home About Archive Update History Data ...d_spot.zip File URL: ftp://ftp.biosciencedbc.jp/archive/rpd/LATEST/rpd_spot.zip F... cDNA. (multiple entries) About This Database Database Description Download License Update History of This Database Site Policy | Contact Us Spot table - RPD | LSDB Archive ...

  15. Normalized cDNA libraries

    Science.gov (United States)

    Soares, Marcelo B.; Efstratiadis, Argiris

    1997-01-01

    This invention provides a method to normalize a directional cDNA library constructed in a vector that allows propagation in single-stranded circle form comprising: (a) propagating the directional cDNA library in single-stranded circles; (b) generating fragments complementary to the 3' noncoding sequence of the single-stranded circles in the library to produce partial duplexes; (c) purifying the partial duplexes; (d) melting and reassociating the purified partial duplexes to moderate Cot; and (e) purifying the unassociated single-stranded circles, thereby generating a normalized cDNA library.

  16. Segmentation and intensity estimation of microarray images using a gamma-t mixture model.

    Science.gov (United States)

    Baek, Jangsun; Son, Young Sook; McLachlan, Geoffrey J

    2007-02-15

    use of the bivariate t distribution for the foreground intensity provides a model that is less sensitive to extreme observations; (5) as a consequence of the aforementioned properties, it allows segmentation to be undertaken for a wide range of spot shapes, including doughnut, sickle shape and artifacts. We apply our method for gridding, segmentation and estimation to cDNA microarray real images and artificial data. Our method provides better segmentation results in spot shapes as well as intensity estimation than Spot and spotSegmentation R language softwares. It detected blank spots as well as bright artifact for the real data, and estimated spot intensities with high-accuracy for the synthetic data. The algorithms were implemented in Matlab. The Matlab codes implementing both the gridding and segmentation/estimation are available upon request. Supplementary material is available at Bioinformatics online.

  17. Simulation of microarray data with realistic characteristics

    Directory of Open Access Journals (Sweden)

    Lehmussola Antti

    2006-07-01

    Full Text Available Abstract Background Microarray technologies have become common tools in biological research. As a result, a need for effective computational methods for data analysis has emerged. Numerous different algorithms have been proposed for analyzing the data. However, an objective evaluation of the proposed algorithms is not possible due to the lack of biological ground truth information. To overcome this fundamental problem, the use of simulated microarray data for algorithm validation has been proposed. Results We present a microarray simulation model which can be used to validate different kinds of data analysis algorithms. The proposed model is unique in the sense that it includes all the steps that affect the quality of real microarray data. These steps include the simulation of biological ground truth data, applying biological and measurement technology specific error models, and finally simulating the microarray slide manufacturing and hybridization. After all these steps are taken into account, the simulated data has realistic biological and statistical characteristics. The applicability of the proposed model is demonstrated by several examples. Conclusion The proposed microarray simulation model is modular and can be used in different kinds of applications. It includes several error models that have been proposed earlier and it can be used with different types of input data. The model can be used to simulate both spotted two-channel and oligonucleotide based single-channel microarrays. All this makes the model a valuable tool for example in validation of data analysis algorithms.

  18. Automation of cDNA Synthesis and Labelling Improves Reproducibility

    Directory of Open Access Journals (Sweden)

    Daniel Klevebring

    2009-01-01

    Full Text Available Background. Several technologies, such as in-depth sequencing and microarrays, enable large-scale interrogation of genomes and transcriptomes. In this study, we asses reproducibility and throughput by moving all laboratory procedures to a robotic workstation, capable of handling superparamagnetic beads. Here, we describe a fully automated procedure for cDNA synthesis and labelling for microarrays, where the purification steps prior to and after labelling are based on precipitation of DNA on carboxylic acid-coated paramagnetic beads. Results. The fully automated procedure allows for samples arrayed on a microtiter plate to be processed in parallel without manual intervention and ensuring high reproducibility. We compare our results to a manual sample preparation procedure and, in addition, use a comprehensive reference dataset to show that the protocol described performs better than similar manual procedures. Conclusions. We demonstrate, in an automated gene expression microarray experiment, a reduced variance between replicates, resulting in an increase in the statistical power to detect differentially expressed genes, thus allowing smaller differences between samples to be identified. This protocol can with minor modifications be used to create cDNA libraries for other applications such as in-depth analysis using next-generation sequencing technologies.

  19. Evaluation of Sex-Specific Gene Expression in Archived Dried Blood Spots (DBS

    Directory of Open Access Journals (Sweden)

    Scott Jewell

    2012-08-01

    Full Text Available Screening newborns for treatable serious conditions is mandated in all US states and many other countries. After screening, Guthrie cards with residual blood (whole spots or portions of spots are typically stored at ambient temperature in many facilities. The potential of archived dried blood spots (DBS for at-birth molecular studies in epidemiological and clinical research is substantial. However, it is also challenging as analytes from DBS may be degraded due to preparation and storage conditions. We previously reported an improved assay for obtaining global RNA gene expression from blood spots. Here, we evaluated sex-specific gene expression and its preservation in DBS using oligonucleotide microarray technology. We found X inactivation-specific transcript (XIST, lysine-specific demethylase 5D (KDM5D (also known as selected cDNA on Y, homolog of mouse (SMCY, uncharacterized LOC729444 (LOC729444, and testis-specific transcript, Y-linked 21 (TTTY21 to be differentially-expressed by sex of the newborn. Our finding that trait-specific RNA gene expression is preserved in unfrozen DBS, demonstrates the technical feasibility of performing molecular genetic profiling using such samples. With millions of DBS potentially available for research, we see new opportunities in using newborn molecular gene expression to better understand molecular pathogenesis of perinatal diseases.

  20. DNA Microarray Technologies: A Novel Approach to Geonomic Research

    Energy Technology Data Exchange (ETDEWEB)

    Hinman, R.; Thrall, B.; Wong, K,

    2002-01-01

    A cDNA microarray allows biologists to examine the expression of thousands of genes simultaneously. Researchers may analyze the complete transcriptional program of an organism in response to specific physiological or developmental conditions. By design, a cDNA microarray is an experiment with many variables and few controls. One question that inevitably arises when working with a cDNA microarray is data reproducibility. How easy is it to confirm mRNA expression patterns? In this paper, a case study involving the treatment of a murine macrophage RAW 264.7 cell line with tumor necrosis factor alpha (TNF) was used to obtain a rough estimate of data reproducibility. Two trials were examined and a list of genes displaying either a > 2-fold or > 4-fold increase in gene expression was compiled. Variations in signal mean ratios between the two slides were observed. We can assume that erring in reproducibility may be compensated by greater inductive levels of similar genes. Steps taken to obtain results included serum starvation of cells before treatment, tests of mRNA for quality/consistency, and data normalization.

  1. Bier spots

    OpenAIRE

    Ahu Yorulmaz,; Seray Kulcu Cakmak; Esra Ar?; Ferda Artuz

    2015-01-01

    Also called as physiologic anemic macules, Bier spots are small, hypopigmented irregularly shaped macules against a background of diffuse erythema, which creates an appearance of speckled vascular mottling of the skin. Bier spots most commonly appear on distal portions of the limbs though there are case reports describing diffuse involvement, which also affect trunk and mucous membranes of the patient. Although the exact pathophysiological mechanisms underlying Bier spots still need to be elu...

  2. Universal Reference RNA as a standard for microarray experiments

    Directory of Open Access Journals (Sweden)

    Fero Michael

    2004-03-01

    Full Text Available Abstract Background Obtaining reliable and reproducible two-color microarray gene expression data is critically important for understanding the biological significance of perturbations made on a cellular system. Microarray design, RNA preparation and labeling, hybridization conditions and data acquisition and analysis are variables difficult to simultaneously control. A useful tool for monitoring and controlling intra- and inter-experimental variation is Universal Reference RNA (URR, developed with the goal of providing hybridization signal at each microarray probe location (spot. Measuring signal at each spot as the ratio of experimental RNA to reference RNA targets, rather than relying on absolute signal intensity, decreases variability by normalizing signal output in any two-color hybridization experiment. Results Human, mouse and rat URR (UHRR, UMRR and URRR, respectively were prepared from pools of RNA derived from individual cell lines representing different tissues. A variety of microarrays were used to determine percentage of spots hybridizing with URR and producing signal above a user defined threshold (microarray coverage. Microarray coverage was consistently greater than 80% for all arrays tested. We confirmed that individual cell lines contribute their own unique set of genes to URR, arguing for a pool of RNA from several cell lines as a better configuration for URR as opposed to a single cell line source for URR. Microarray coverage comparing two separately prepared batches each of UHRR, UMRR and URRR were highly correlated (Pearson's correlation coefficients of 0.97. Conclusion Results of this study demonstrate that large quantities of pooled RNA from individual cell lines are reproducibly prepared and possess diverse gene representation. This type of reference provides a standard for reducing variation in microarray experiments and allows more reliable comparison of gene expression data within and between experiments and

  3. Fibre optic microarrays.

    Science.gov (United States)

    Walt, David R

    2010-01-01

    This tutorial review describes how fibre optic microarrays can be used to create a variety of sensing and measurement systems. This review covers the basics of optical fibres and arrays, the different microarray architectures, and describes a multitude of applications. Such arrays enable multiplexed sensing for a variety of analytes including nucleic acids, vapours, and biomolecules. Polymer-coated fibre arrays can be used for measuring microscopic chemical phenomena, such as corrosion and localized release of biochemicals from cells. In addition, these microarrays can serve as a substrate for fundamental studies of single molecules and single cells. The review covers topics of interest to chemists, biologists, materials scientists, and engineers.

  4. Fluorescent microarray for multiplexed quantification of environmental contaminants in seawater samples

    Science.gov (United States)

    The development of a fluorescent multiplexed microarray platform able to detect and quantify a wide variety of pollutants in seawater is reported. The microarray platform has been manufactured by spotting 6 different bioconjugate competitors and it uses a cocktail of 6 monoclonal and polyclonal anti...

  5. DNA Microarray Technology

    Science.gov (United States)

    Skip to main content DNA Microarray Technology Enter Search Term(s): Español Research Funding An Overview Bioinformatics Current Grants Education and Training Funding Extramural Research News Features Funding Divisions Funding ...

  6. DNA Microarray Technology; TOPICAL

    International Nuclear Information System (INIS)

    WERNER-WASHBURNE, MARGARET; DAVIDSON, GEORGE S.

    2002-01-01

    Collaboration between Sandia National Laboratories and the University of New Mexico Biology Department resulted in the capability to train students in microarray techniques and the interpretation of data from microarray experiments. These studies provide for a better understanding of the role of stationary phase and the gene regulation involved in exit from stationary phase, which may eventually have important clinical implications. Importantly, this research trained numerous students and is the basis for three new Ph.D. projects

  7. Protein microarray: sensitive and effective immunodetection for drug residues

    Directory of Open Access Journals (Sweden)

    Zer Cindy

    2010-02-01

    Full Text Available Abstract Background Veterinary drugs such as clenbuterol (CL and sulfamethazine (SM2 are low molecular weight ( Results The artificial antigens were spotted on microarray slides. Standard concentrations of the compounds were added to compete with the spotted antigens for binding to the antisera to determine the IC50. Our microarray assay showed the IC50 were 39.6 ng/ml for CL and 48.8 ng/ml for SM2, while the traditional competitive indirect-ELISA (ci-ELISA showed the IC50 were 190.7 ng/ml for CL and 156.7 ng/ml for SM2. We further validated the two methods with CL fortified chicken muscle tissues, and the protein microarray assay showed 90% recovery while the ci-ELISA had 76% recovery rate. When tested with CL-fed chicken muscle tissues, the protein microarray assay had higher sensitivity (0.9 ng/g than the ci-ELISA (0.1 ng/g for detection of CL residues. Conclusions The protein microarrays showed 4.5 and 3.5 times lower IC50 than the ci-ELISA detection for CL and SM2, respectively, suggesting that immunodetection of small molecules with protein microarray is a better approach than the traditional ELISA technique.

  8. Plasmonically amplified fluorescence bioassay with microarray format

    Science.gov (United States)

    Gogalic, S.; Hageneder, S.; Ctortecka, C.; Bauch, M.; Khan, I.; Preininger, Claudia; Sauer, U.; Dostalek, J.

    2015-05-01

    Plasmonic amplification of fluorescence signal in bioassays with microarray detection format is reported. A crossed relief diffraction grating was designed to couple an excitation laser beam to surface plasmons at the wavelength overlapping with the absorption and emission bands of fluorophore Dy647 that was used as a label. The surface of periodically corrugated sensor chip was coated with surface plasmon-supporting gold layer and a thin SU8 polymer film carrying epoxy groups. These groups were employed for the covalent immobilization of capture antibodies at arrays of spots. The plasmonic amplification of fluorescence signal on the developed microarray chip was tested by using interleukin 8 sandwich immunoassay. The readout was performed ex situ after drying the chip by using a commercial scanner with high numerical aperture collecting lens. Obtained results reveal the enhancement of fluorescence signal by a factor of 5 when compared to a regular glass chip.

  9. Bier spots

    Directory of Open Access Journals (Sweden)

    Ahu Yorulmaz,

    2015-10-01

    Full Text Available Also called as physiologic anemic macules, Bier spots are small, hypopigmented irregularly shaped macules against a background of diffuse erythema, which creates an appearance of speckled vascular mottling of the skin. Bier spots most commonly appear on distal portions of the limbs though there are case reports describing diffuse involvement, which also affect trunk and mucous membranes of the patient. Although the exact pathophysiological mechanisms underlying Bier spots still need to be elucidated, Bier spots have been suggested to be a vascular anomaly caused by vasoconstriction of small vessels. In addition, several diseases have been proposed to be associated with Bier spots, including scleroderma renal crisis, cryoglobulinemia, Peutz-Jeghers syndrome, alopecia areata and hypoplasia of the aorta, although it has not been shown whether these associations are casual or coincidental. The clinical presentation of Bier spots is quite typical. These tiny whitish macules easily become prominent when the affected limb is placed in a dependent position and fade away when the limb is raised. Here we report a case of Bier spots in a 32-year-old male patient with characteristical clinical manifestations.

  10. Microarray analysis identified Puccinia striiformis f. sp. tritici genes involved in infection and sporulation.

    Science.gov (United States)

    Puccinia striiformis f. sp. tritici (Pst) causes stripe rust, one of the most important diseases of wheat worldwide. To identify Pst genes involved in infection and sporulation, a custom oligonucleotide Genechip was made using sequences of 442 genes selected from Pst cDNA libraries. Microarray analy...

  11. Age Spots

    Science.gov (United States)

    ... for Every Season How to Choose the Best Skin Care Products In This Section Dermatologic Surgery What is dermatologic ... for Every Season How to Choose the Best Skin Care Products Age Spots Treatment Options Learn more about treatment ...

  12. Spotted inflation

    International Nuclear Information System (INIS)

    Matsuda, Tomohiro

    2010-01-01

    We describe new scenarios for generating curvature perturbations when inflaton (curvaton) has significant interactions. We consider a ''spot'', which arises from interactions associated with an enhanced symmetric point (ESP) on the trajectory. Our first example uses the spot to induce a gap in the field equation. We observe that the gap in the field equation may cause generation of curvature perturbation if it does not appear simultaneous in space. The mechanism is similar to the scenario of inhomogeneous phase transition. Then we observe that the spot interactions may initiate warm inflation in the cold Universe. Creation of cosmological perturbation is discussed in relation to the inflaton dynamics and the modulation associated with the spot interactions

  13. Emerging use of gene expression microarrays in plant physiology.

    Science.gov (United States)

    Wullschleger, Stan D; Difazio, Stephen P

    2003-01-01

    Microarrays have become an important technology for the global analysis of gene expression in humans, animals, plants, and microbes. Implemented in the context of a well-designed experiment, cDNA and oligonucleotide arrays can provide highthroughput, simultaneous analysis of transcript abundance for hundreds, if not thousands, of genes. However, despite widespread acceptance, the use of microarrays as a tool to better understand processes of interest to the plant physiologist is still being explored. To help illustrate current uses of microarrays in the plant sciences, several case studies that we believe demonstrate the emerging application of gene expression arrays in plant physiology were selected from among the many posters and presentations at the 2003 Plant and Animal Genome XI Conference. Based on this survey, microarrays are being used to assess gene expression in plants exposed to the experimental manipulation of air temperature, soil water content and aluminium concentration in the root zone. Analysis often includes characterizing transcript profiles for multiple post-treatment sampling periods and categorizing genes with common patterns of response using hierarchical clustering techniques. In addition, microarrays are also providing insights into developmental changes in gene expression associated with fibre and root elongation in cotton and maize, respectively. Technical and analytical limitations of microarrays are discussed and projects attempting to advance areas of microarray design and data analysis are highlighted. Finally, although much work remains, we conclude that microarrays are a valuable tool for the plant physiologist interested in the characterization and identification of individual genes and gene families with potential application in the fields of agriculture, horticulture and forestry.

  14. Emerging Use of Gene Expression Microarrays in Plant Physiology

    Directory of Open Access Journals (Sweden)

    Stephen P. Difazio

    2006-04-01

    Full Text Available Microarrays have become an important technology for the global analysis of gene expression in humans, animals, plants, and microbes. Implemented in the context of a well-designed experiment, cDNA and oligonucleotide arrays can provide highthroughput, simultaneous analysis of transcript abundance for hundreds, if not thousands, of genes. However, despite widespread acceptance, the use of microarrays as a tool to better understand processes of interest to the plant physiologist is still being explored. To help illustrate current uses of microarrays in the plant sciences, several case studies that we believe demonstrate the emerging application of gene expression arrays in plant physiology were selected from among the many posters and presentations at the 2003 Plant and Animal Genome XI Conference. Based on this survey, microarrays are being used to assess gene expression in plants exposed to the experimental manipulation of air temperature, soil water content and aluminium concentration in the root zone. Analysis often includes characterizing transcript profiles for multiple post-treatment sampling periods and categorizing genes with common patterns of response using hierarchical clustering techniques. In addition, microarrays are also providing insights into developmental changes in gene expression associated with fibre and root elongation in cotton and maize, respectively. Technical and analytical limitations of microarrays are discussed and projects attempting to advance areas of microarray design and data analysis are highlighted. Finally, although much work remains, we conclude that microarrays are a valuable tool for the plant physiologist interested in the characterization and identification of individual genes and gene families with potential application in the fields of agriculture, horticulture and forestry.

  15. SPOT Program

    Science.gov (United States)

    Smith, Jason T.; Welsh, Sam J.; Farinetti, Antonio L.; Wegner, Tim; Blakeslee, James; Deboeck, Toni F.; Dyer, Daniel; Corley, Bryan M.; Ollivierre, Jarmaine; Kramer, Leonard; hide

    2010-01-01

    A Spacecraft Position Optimal Tracking (SPOT) program was developed to process Global Positioning System (GPS) data, sent via telemetry from a spacecraft, to generate accurate navigation estimates of the vehicle position and velocity (state vector) using a Kalman filter. This program uses the GPS onboard receiver measurements to sequentially calculate the vehicle state vectors and provide this information to ground flight controllers. It is the first real-time ground-based shuttle navigation application using onboard sensors. The program is compact, portable, self-contained, and can run on a variety of UNIX or Linux computers. The program has a modular objec-toriented design that supports application-specific plugins such as data corruption remediation pre-processing and remote graphics display. The Kalman filter is extensible to additional sensor types or force models. The Kalman filter design is also strong against data dropouts because it uses physical models from state and covariance propagation in the absence of data. The design of this program separates the functionalities of SPOT into six different executable processes. This allows for the individual processes to be connected in an a la carte manner, making the feature set and executable complexity of SPOT adaptable to the needs of the user. Also, these processes need not be executed on the same workstation. This allows for communications between SPOT processes executing on the same Local Area Network (LAN). Thus, SPOT can be executed in a distributed sense with the capability for a team of flight controllers to efficiently share the same trajectory information currently being computed by the program. SPOT is used in the Mission Control Center (MCC) for Space Shuttle Program (SSP) and International Space Station Program (ISSP) operations, and can also be used as a post -flight analysis tool. It is primarily used for situational awareness, and for contingency situations.

  16. Multisegment one-step RT-PCR fluorescent labeling of influenza A virus genome for use in diagnostic microarray applications

    Energy Technology Data Exchange (ETDEWEB)

    Vasin, A V; Plotnikova, M A; Klotchenko, S A; Elpaeva, E A; Komissarov, A B; Egorov, V V; Kiselev, O I [Research Institute of Influenza of the Ministry of Health and Social Development of the Russian Federation, 15/17 Prof. Popova St., St. Petersburg (Russian Federation); Sandybaev, N T; Chervyakova, O V; Strochkov, V M; Taylakova, E T; Koshemetov, J K; Mamadaliev, S M, E-mail: vasin@influenza.spb.ru [Research Institute for Biological Safety Problems of the RK NBC/SC ME and S RK, Gvardeiskiy (Kazakhstan)

    2011-04-01

    Microarray technology is one of the most challenging methods of influenza A virus subtyping, which is based on the antigenic properties of viral surface glycoproteins - hemagglutinin and neuraminidase. On the example of biochip for detection of influenza A/H5N1 virus we showed the possibility of using multisegment RTPCR method for amplification of fluorescently labeled cDNA of all possible influenza A virus subtypes with a single pair of primers in influenza diagnostic microarrays.

  17. Radioactive cDNA microarrys for gene expression profiles in antidepressant therapy

    International Nuclear Information System (INIS)

    Lee, M. S.; Han, B. J.; Cha, J. H.; Ryu, Y. M.; Shin, E. K.; Park, J. H.; Park, Y. H.; Kim, M. K.

    2002-01-01

    Using radioactive cDNA microarray, we investigated a pattern of gene regulation under treatment of antidepressant on patients of depressive disoder. Basic microarray technology was performed as previously described in our research. The bioinformatic selection of human cDNAs, which is specifically designed for psychiatry, neurology, and signal transduction, were arrayed on nylon membranes. Using with 33P-labeled probes, this method provided highly sensitive gene expression profiles of our interest including brain receptors, drug metabolism, and cellular signalings. Gene expression profiles were also classified into several categories in accordance with the gene-regulation of antidepressant. The gene profiles of our interest were significantly up- (16 genes, >2.0 of Z-ratio) or down- (24 genes, <-2.0 of Z ratio) regulated when compared the good responsed group with the bad-responsed one. Consequently, we demonstrated that radioactive human cDNA microarray is highly likely to be an efficient technology for evaluating the gene regulation of antidepressants, such as selective serotonin-reuptake inhibitors (SSRIs), by using high-throughput biotechnology

  18. A Customized DNA Microarray for Microbial Source Tracking ...

    Science.gov (United States)

    It is estimated that more than 160, 000 miles of rivers and streams in the United States are impaired due to the presence of waterborne pathogens. These pathogens typically originate from human and other animal fecal pollution sources; therefore, a rapid microbial source tracking (MST) method is needed to facilitate water quality assessment and impaired water remediation. We report a novel qualitative DNA microarray technology consisting of 453 probes for the detection of general fecal and host-associated bacteria, viruses, antibiotic resistance, and other environmentally relevant genetic indicators. A novel data normalization and reduction approach is also presented to help alleviate false positives often associated with high-density microarray applications. To evaluate the performance of the approach, DNA and cDNA was isolated from swine, cattle, duck, goose and gull fecal reference samples, as well as soiled poultry liter and raw municipal sewage. Based on nonmetric multidimensional scaling analysis of results, findings suggest that the novel microarray approach may be useful for pathogen detection and identification of fecal contamination in recreational waters. The ability to simultaneously detect a large collection of environmentally important genetic indicators in a single test has the potential to provide water quality managers with a wide range of information in a short period of time. Future research is warranted to measure microarray performance i

  19. Cross-platform analysis of cancer microarray data improves gene expression based classification of phenotypes

    Directory of Open Access Journals (Sweden)

    Eils Roland

    2005-11-01

    Full Text Available Abstract Background The extensive use of DNA microarray technology in the characterization of the cell transcriptome is leading to an ever increasing amount of microarray data from cancer studies. Although similar questions for the same type of cancer are addressed in these different studies, a comparative analysis of their results is hampered by the use of heterogeneous microarray platforms and analysis methods. Results In contrast to a meta-analysis approach where results of different studies are combined on an interpretative level, we investigate here how to directly integrate raw microarray data from different studies for the purpose of supervised classification analysis. We use median rank scores and quantile discretization to derive numerically comparable measures of gene expression from different platforms. These transformed data are then used for training of classifiers based on support vector machines. We apply this approach to six publicly available cancer microarray gene expression data sets, which consist of three pairs of studies, each examining the same type of cancer, i.e. breast cancer, prostate cancer or acute myeloid leukemia. For each pair, one study was performed by means of cDNA microarrays and the other by means of oligonucleotide microarrays. In each pair, high classification accuracies (> 85% were achieved with training and testing on data instances randomly chosen from both data sets in a cross-validation analysis. To exemplify the potential of this cross-platform classification analysis, we use two leukemia microarray data sets to show that important genes with regard to the biology of leukemia are selected in an integrated analysis, which are missed in either single-set analysis. Conclusion Cross-platform classification of multiple cancer microarray data sets yields discriminative gene expression signatures that are found and validated on a large number of microarray samples, generated by different laboratories and

  20. The IronChip evaluation package: a package of perl modules for robust analysis of custom microarrays

    Directory of Open Access Journals (Sweden)

    Brazma Alvis

    2010-03-01

    Full Text Available Abstract Background Gene expression studies greatly contribute to our understanding of complex relationships in gene regulatory networks. However, the complexity of array design, production and manipulations are limiting factors, affecting data quality. The use of customized DNA microarrays improves overall data quality in many situations, however, only if for these specifically designed microarrays analysis tools are available. Results The IronChip Evaluation Package (ICEP is a collection of Perl utilities and an easy to use data evaluation pipeline for the analysis of microarray data with a focus on data quality of custom-designed microarrays. The package has been developed for the statistical and bioinformatical analysis of the custom cDNA microarray IronChip but can be easily adapted for other cDNA or oligonucleotide-based designed microarray platforms. ICEP uses decision tree-based algorithms to assign quality flags and performs robust analysis based on chip design properties regarding multiple repetitions, ratio cut-off, background and negative controls. Conclusions ICEP is a stand-alone Windows application to obtain optimal data quality from custom-designed microarrays and is freely available here (see "Additional Files" section and at: http://www.alice-dsl.net/evgeniy.vainshtein/ICEP/

  1. PCR-based cDNA library construction: general cDNA libraries at the level of a few cells.

    OpenAIRE

    Belyavsky, A; Vinogradova, T; Rajewsky, K

    1989-01-01

    A procedure for the construction of general cDNA libraries is described which is based on the amplification of total cDNA in vitro. The first cDNA strand is synthesized from total RNA using an oligo(dT)-containing primer. After oligo(dG) tailing the total cDNA is amplified by PCR using two primers complementary to oligo(dA) and oligo(dG) ends of the cDNA. For insertion of the cDNA into a vector a controlled trimming of the 3' ends of the cDNA by Klenow enzyme was used. Starting from 10 J558L ...

  2. Insights into the iron and sulfur energetic metabolism of Acidithiobacillus ferrooxidans by microarray transcriptome profiling

    Energy Technology Data Exchange (ETDEWEB)

    R. Quatrini; C. Appia-Ayme; Y. Denis; J. Ratouchniak; F. Veloso; J. Valdes; C. Lefimil; S. Silver; F. Roberto; O. Orellana; F. Denizot; E. Jedlicki; D. Holmes; V. Bonnefoy

    2006-09-01

    Acidithiobacillus ferrooxidans is a well known acidophilic, chemolithoautotrophic, Gram negative, bacterium involved in bioleaching and acid mine drainage. In aerobic conditions, it gains energy mainly from the oxidation of ferrous iron and/or reduced sulfur compounds present in ores. After initial oxidation of the substrate, electrons from ferrous iron or sulfur enter respiratory chains and are transported through several redox proteins to oxygen. However, the oxidation of ferrous iron and reduced sulfur compounds has also to provide electrons for the reduction of NAD(P) that is subsequently required for many metabolic processes including CO2 fixation. To help to unravel the enzymatic pathways and the electron transfer chains involved in these processes, a genome-wide microarray transcript profiling analysis was carried out. Oligonucleotides corresponding to approximately 3000 genes of the A. ferrooxidans type strain ATCC23270 were spotted onto glass-slides and hybridized with cDNA retrotranscribed from RNA extracted from ferrous iron and sulfur grown cells. The genes which are preferentially transcribed in ferrous iron conditions and those preferentially transcribed in sulfur conditions were analyzed. The expression of a substantial number of these genes has been validated by real-time PCR, Northern blot hybridization and/or immunodetection analysis. Our results support and extend certain models of iron and sulfur oxidation and highlight previous observations regarding the possible presence of alternate electron pathways. Our findings also suggest ways in which iron and sulfur oxidation may be co-ordinately regulated. An accompanying paper (Appia-Ayme et al.) describes results pertaining to other metabolic functions.

  3. DNA microarrays : a molecular cloning manual

    National Research Council Canada - National Science Library

    Sambrook, Joseph; Bowtell, David

    2002-01-01

    .... DNA Microarrays provides authoritative, detailed instruction on the design, construction, and applications of microarrays, as well as comprehensive descriptions of the software tools and strategies...

  4. Rocky Mountain spotted fever

    Science.gov (United States)

    ... spotted fever on the foot Rocky Mountain spotted fever, petechial rash Antibodies Deer and dog tick References McElligott SC, Kihiczak GG, Schwartz RA. Rocky Mountain spotted fever and other rickettsial infections. In: Lebwohl MG, Heymann ...

  5. Gene targeting associated with the radiation sensitivity in squamous cell carcinoma by using microarray analysis

    International Nuclear Information System (INIS)

    Nimura, Yoshinori; Kumagai, Ken; Kouzu, Yoshinao; Higo, Morihiro; Kato, Yoshikuni; Seki, Naohiko; Yamada, Shigeru

    2005-01-01

    In order to identify a set of genes related to radiation sensitivity of squamous cell carcinoma (SCC) and establish a predictive method, we compared expression profiles of radio-sensitive/radio-resistant SCC cell lines, using the in-house cDNA microarray consisting of 2,201 human genes derived from full-length enriched SCC cDNA libraries and the Human oligo chip 30 K (Hitachi Software Engineering). Surviving fractions (SF) after irradiation of heavy iron were calculated by colony formation assay. Three pairs (TE2-TE13, YES5-YES6, and HSC3-HSC2), sensitive (SF1 0.6), were selected for the microarray analysis. The results of cDNA microarray analysis showed that 20 genes in resistant cell lines and 5 genes in sensitive cell lines were up regulated more than 1.5-fold compared with sensitive and resistant cell lines respectively. Fourteen out of 25 genes were confirmed the gene expression profiles by real-time polymerase chain reaction (PCR). Twenty-seven genes identified by Human oligo chip 30 K are candidate for the markers to distinguish radio-sensitive from radio-resistant. These results suggest that the isolated 27 genes are the candidates that might be used as specific molecular markers to predict radiation sensitivity. (author)

  6. Lectin cDNA and transgenic plants derived therefrom

    Science.gov (United States)

    Raikhel, Natasha V.

    2000-10-03

    Transgenic plants containing cDNA encoding Gramineae lectin are described. The plants preferably contain cDNA coding for barley lectin and store the lectin in the leaves. The transgenic plants, particularly the leaves exhibit insecticidal and fungicidal properties.

  7. cDNA sequence quality data - Budding yeast cDNA sequencing project | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available List Contact us Budding yeast cDNA sequencing project cDNA sequence quality data Data detail Data name cDNA sequence quality... data DOI 10.18908/lsdba.nbdc00838-003 Description of data contents Phred's quality score. P...tion Download License Update History of This Database Site Policy | Contact Us cDNA sequence quality

  8. Nonlinear matching measure for the analysis of on-off type DNA microarray images

    Science.gov (United States)

    Kim, Jong D.; Park, Misun; Kim, Jongwon

    2003-07-01

    In this paper, we propose a new nonlinear matching measure for automatic analysis of the on-off type DNA microarray images in which the hybridized spots are detected by the template matching method. The targeting spots of HPV DNA chips are designed for genotyping the human papilloma virus(HPV). The proposed measure is obtained by binarythresholding over the whole template region and taking the number of white pixels inside the spotted area. This measure is evaluated in terms of the accuracy of the estimated marker location to show better performance than the normalized covariance.

  9. Printing Proteins as Microarrays for High-Throughput Function Determination

    Science.gov (United States)

    MacBeath, Gavin; Schreiber, Stuart L.

    2000-09-01

    Systematic efforts are currently under way to construct defined sets of cloned genes for high-throughput expression and purification of recombinant proteins. To facilitate subsequent studies of protein function, we have developed miniaturized assays that accommodate extremely low sample volumes and enable the rapid, simultaneous processing of thousands of proteins. A high-precision robot designed to manufacture complementary DNA microarrays was used to spot proteins onto chemically derivatized glass slides at extremely high spatial densities. The proteins attached covalently to the slide surface yet retained their ability to interact specifically with other proteins, or with small molecules, in solution. Three applications for protein microarrays were demonstrated: screening for protein-protein interactions, identifying the substrates of protein kinases, and identifying the protein targets of small molecules.

  10. Nové možnosti v diagnostice kolorektálního karcinomu s využitím technologie cDNA mikročipů

    Czech Academy of Sciences Publication Activity Database

    Jansová, E.; Krontorád, P.; Svoboda, Z.; Pavlík, T.; Koutná, I.; Kozubek, Michal; Žaloudík, J.; Kozubek, Stanislav

    2004-01-01

    Roč. 6, č. 17 (2004), s. 203-207 ISSN 0862-495X R&D Projects: GA MZd(CZ) NC6987 Institutional research plan: CEZ:AV0Z5004920 Keywords : cDNA microarray technology - cluster analysis - colorectal cancer Subject RIV: BO - Biophysics

  11. The Porcelain Crab Transcriptome and PCAD, the Porcelain Crab Microarray and Sequence Database

    Energy Technology Data Exchange (ETDEWEB)

    Tagmount, Abderrahmane; Wang, Mei; Lindquist, Erika; Tanaka, Yoshihiro; Teranishi, Kristen S.; Sunagawa, Shinichi; Wong, Mike; Stillman, Jonathon H.

    2010-01-27

    Background: With the emergence of a completed genome sequence of the freshwater crustacean Daphnia pulex, construction of genomic-scale sequence databases for additional crustacean sequences are important for comparative genomics and annotation. Porcelain crabs, genus Petrolisthes, have been powerful crustacean models for environmental and evolutionary physiology with respect to thermal adaptation and understanding responses of marine organisms to climate change. Here, we present a large-scale EST sequencing and cDNA microarray database project for the porcelain crab Petrolisthes cinctipes. Methodology/Principal Findings: A set of ~;;30K unique sequences (UniSeqs) representing ~;;19K clusters were generated from ~;;98K high quality ESTs from a set of tissue specific non-normalized and mixed-tissue normalized cDNA libraries from the porcelain crab Petrolisthes cinctipes. Homology for each UniSeq was assessed using BLAST, InterProScan, GO and KEGG database searches. Approximately 66percent of the UniSeqs had homology in at least one of the databases. All EST and UniSeq sequences along with annotation results and coordinated cDNA microarray datasets have been made publicly accessible at the Porcelain Crab Array Database (PCAD), a feature-enriched version of the Stanford and Longhorn Array Databases.Conclusions/Significance: The EST project presented here represents the third largest sequencing effort for any crustacean, and the largest effort for any crab species. Our assembly and clustering results suggest that our porcelain crab EST data set is equally diverse to the much larger EST set generated in the Daphnia pulex genome sequencing project, and thus will be an important resource to the Daphnia research community. Our homology results support the pancrustacea hypothesis and suggest that Malacostraca may be ancestral to Branchiopoda and Hexapoda. Our results also suggest that our cDNA microarrays cover as much of the transcriptome as can reasonably be captured in

  12. Improvement in the amine glass platform by bubbling method for a DNA microarray.

    Science.gov (United States)

    Jee, Seung Hyun; Kim, Jong Won; Lee, Ji Hyeong; Yoon, Young Soo

    2015-01-01

    A glass platform with high sensitivity for sexually transmitted diseases microarray is described here. An amino-silane-based self-assembled monolayer was coated on the surface of a glass platform using a novel bubbling method. The optimized surface of the glass platform had highly uniform surface modifications using this method, as well as improved hybridization properties with capture probes in the DNA microarray. On the basis of these results, the improved glass platform serves as a highly reliable and optimal material for the DNA microarray. Moreover, in this study, we demonstrated that our glass platform, manufactured by utilizing the bubbling method, had higher uniformity, shorter processing time, lower background signal, and higher spot signal than the platforms manufactured by the general dipping method. The DNA microarray manufactured with a glass platform prepared using bubbling method can be used as a clinical diagnostic tool.

  13. Molecular and histological characterization of age spots

    Science.gov (United States)

    Choi, Wonseon; Yin, Lanlan; Smuda, Christoph; Batzer, Jan; Hearing, Vincent J.; Kolbe, Ludger

    2016-01-01

    Age spots, also called solar lentigines and lentigo senilis, are light brown to black pigmented lesions of various sizes that typically develop in chronically sun-exposed skin. It is well known that age spots are strongly related to chronic sun exposure and are associated with photodamage and an increased risk for skin cancer, however, the mechanism(s) underlying their development remain poorly understood. We used immunohistochemical analysis and microarray analysis to investigate the processes involved in their formation, focusing on specific markers associated with the functions and proliferation of melanocytes and keratinocytes. A total of 193 genes were differentially expressed in age spots but melanocyte pigment genes were not among them. The increased expression of keratins 5 and 10, markers of basal and suprabasal keratinocytes, respectively, in age spots suggests that the increased proliferation of basal keratinocytes combined with the decreased turnover of suprabasal keratinocytes leads to the exaggerated formation of rete ridges in lesional epidermis which in turn disrupts the normal processing of melanin upwards from the basal layer. Based on our results, we propose a model for the development of age spots that explains the accumulation of melanin and the development of extensive rete ridges in those hyperpigmented lesions. PMID:27621222

  14. Comparison of small n statistical tests of differential expression applied to microarrays

    Directory of Open Access Journals (Sweden)

    Lee Anna Y

    2009-02-01

    Full Text Available Abstract Background DNA microarrays provide data for genome wide patterns of expression between observation classes. Microarray studies often have small samples sizes, however, due to cost constraints or specimen availability. This can lead to poor random error estimates and inaccurate statistical tests of differential expression. We compare the performance of the standard t-test, fold change, and four small n statistical test methods designed to circumvent these problems. We report results of various normalization methods for empirical microarray data and of various random error models for simulated data. Results Three Empirical Bayes methods (CyberT, BRB, and limma t-statistics were the most effective statistical tests across simulated and both 2-colour cDNA and Affymetrix experimental data. The CyberT regularized t-statistic in particular was able to maintain expected false positive rates with simulated data showing high variances at low gene intensities, although at the cost of low true positive rates. The Local Pooled Error (LPE test introduced a bias that lowered false positive rates below theoretically expected values and had lower power relative to the top performers. The standard two-sample t-test and fold change were also found to be sub-optimal for detecting differentially expressed genes. The generalized log transformation was shown to be beneficial in improving results with certain data sets, in particular high variance cDNA data. Conclusion Pre-processing of data influences performance and the proper combination of pre-processing and statistical testing is necessary for obtaining the best results. All three Empirical Bayes methods assessed in our study are good choices for statistical tests for small n microarray studies for both Affymetrix and cDNA data. Choice of method for a particular study will depend on software and normalization preferences.

  15. MicroArray Facility: a laboratory information management system with extended support for Nylon based technologies

    Directory of Open Access Journals (Sweden)

    Beaudoing Emmanuel

    2006-09-01

    Full Text Available Abstract Background High throughput gene expression profiling (GEP is becoming a routine technique in life science laboratories. With experimental designs that repeatedly span thousands of genes and hundreds of samples, relying on a dedicated database infrastructure is no longer an option. GEP technology is a fast moving target, with new approaches constantly broadening the field diversity. This technology heterogeneity, compounded by the informatics complexity of GEP databases, means that software developments have so far focused on mainstream techniques, leaving less typical yet established techniques such as Nylon microarrays at best partially supported. Results MAF (MicroArray Facility is the laboratory database system we have developed for managing the design, production and hybridization of spotted microarrays. Although it can support the widely used glass microarrays and oligo-chips, MAF was designed with the specific idiosyncrasies of Nylon based microarrays in mind. Notably single channel radioactive probes, microarray stripping and reuse, vector control hybridizations and spike-in controls are all natively supported by the software suite. MicroArray Facility is MIAME supportive and dynamically provides feedback on missing annotations to help users estimate effective MIAME compliance. Genomic data such as clone identifiers and gene symbols are also directly annotated by MAF software using standard public resources. The MAGE-ML data format is implemented for full data export. Journalized database operations (audit tracking, data anonymization, material traceability and user/project level confidentiality policies are also managed by MAF. Conclusion MicroArray Facility is a complete data management system for microarray producers and end-users. Particular care has been devoted to adequately model Nylon based microarrays. The MAF system, developed and implemented in both private and academic environments, has proved a robust solution for

  16. SpotADAPT

    DEFF Research Database (Denmark)

    Kaulakiene, Dalia; Thomsen, Christian; Pedersen, Torben Bach

    2015-01-01

    by Amazon Web Services (AWS). The users aiming for the spot market are presented with many instance types placed in multiple datacenters in the world, and thus it is difficult to choose the optimal deployment. In this paper, we propose the framework SpotADAPT (Spot-Aware (re-)Deployment of Analytical...... of typical analytical workloads and real spot price traces. SpotADAPT's suggested deployments are comparable to the theoretically optimal ones, and in particular, it shows good cost benefits for the budget optimization -- on average SpotADAPT is at most 0.3% more expensive than the theoretically optimal...

  17. Polyadenylation state microarray (PASTA) analysis.

    Science.gov (United States)

    Beilharz, Traude H; Preiss, Thomas

    2011-01-01

    Nearly all eukaryotic mRNAs terminate in a poly(A) tail that serves important roles in mRNA utilization. In the cytoplasm, the poly(A) tail promotes both mRNA stability and translation, and these functions are frequently regulated through changes in tail length. To identify the scope of poly(A) tail length control in a transcriptome, we developed the polyadenylation state microarray (PASTA) method. It involves the purification of mRNA based on poly(A) tail length using thermal elution from poly(U) sepharose, followed by microarray analysis of the resulting fractions. In this chapter we detail our PASTA approach and describe some methods for bulk and mRNA-specific poly(A) tail length measurements of use to monitor the procedure and independently verify the microarray data.

  18. Rocky Mountain Spotted Fever

    Science.gov (United States)

    ... with facebook share with twitter share with linkedin Rocky Mountain Spotted Fever Credit: CDC A male cayenne tick, Amblyomma cajennense, ... and New Mexico. Why Is the Study of Rocky Mountain Spotted Fever a Priority for NIAID? Tickborne diseases are becoming ...

  19. An efficient algorithm for the stochastic simulation of the hybridization of DNA to microarrays

    Directory of Open Access Journals (Sweden)

    Laurenzi Ian J

    2009-12-01

    Full Text Available Abstract Background Although oligonucleotide microarray technology is ubiquitous in genomic research, reproducibility and standardization of expression measurements still concern many researchers. Cross-hybridization between microarray probes and non-target ssDNA has been implicated as a primary factor in sensitivity and selectivity loss. Since hybridization is a chemical process, it may be modeled at a population-level using a combination of material balance equations and thermodynamics. However, the hybridization reaction network may be exceptionally large for commercial arrays, which often possess at least one reporter per transcript. Quantification of the kinetics and equilibrium of exceptionally large chemical systems of this type is numerically infeasible with customary approaches. Results In this paper, we present a robust and computationally efficient algorithm for the simulation of hybridization processes underlying microarray assays. Our method may be utilized to identify the extent to which nucleic acid targets (e.g. cDNA will cross-hybridize with probes, and by extension, characterize probe robustnessusing the information specified by MAGE-TAB. Using this algorithm, we characterize cross-hybridization in a modified commercial microarray assay. Conclusions By integrating stochastic simulation with thermodynamic prediction tools for DNA hybridization, one may robustly and rapidly characterize of the selectivity of a proposed microarray design at the probe and "system" levels. Our code is available at http://www.laurenzi.net.

  20. Transcriptional profiling of endocrine cerebro-osteodysplasia using microarray and next-generation sequencing.

    Directory of Open Access Journals (Sweden)

    Piya Lahiry

    Full Text Available BACKGROUND: Transcriptome profiling of patterns of RNA expression is a powerful approach to identify networks of genes that play a role in disease. To date, most mRNA profiling of tissues has been accomplished using microarrays, but next-generation sequencing can offer a richer and more comprehensive picture. METHODOLOGY/PRINCIPAL FINDINGS: ECO is a rare multi-system developmental disorder caused by a homozygous mutation in ICK encoding intestinal cell kinase. We performed gene expression profiling using both cDNA microarrays and next-generation mRNA sequencing (mRNA-seq of skin fibroblasts from ECO-affected subjects. We then validated a subset of differentially expressed transcripts identified by each method using quantitative reverse transcription-polymerase chain reaction (qRT-PCR. Finally, we used gene ontology (GO to identify critical pathways and processes that were abnormal according to each technical platform. Methodologically, mRNA-seq identifies a much larger number of differentially expressed genes with much better correlation to qRT-PCR results than the microarray (r² = 0.794 and 0.137, respectively. Biologically, cDNA microarray identified functional pathways focused on anatomical structure and development, while the mRNA-seq platform identified a higher proportion of genes involved in cell division and DNA replication pathways. CONCLUSIONS/SIGNIFICANCE: Transcriptome profiling with mRNA-seq had greater sensitivity, range and accuracy than the microarray. The two platforms generated different but complementary hypotheses for further evaluation.

  1. Detecting imbalanced expression of SNP alleles by minisequencing on microarrays

    Directory of Open Access Journals (Sweden)

    Dahlgren Andreas

    2004-10-01

    Full Text Available Abstract Background Each of the human genes or transcriptional units is likely to contain single nucleotide polymorphisms that may give rise to sequence variation between individuals and tissues on the level of RNA. Based on recent studies, differential expression of the two alleles of heterozygous coding single nucleotide polymorphisms (SNPs may be frequent for human genes. Methods with high accuracy to be used in a high throughput setting are needed for systematic surveys of expressed sequence variation. In this study we evaluated two formats of multiplexed, microarray based minisequencing for quantitative detection of imbalanced expression of SNP alleles. We used a panel of ten SNPs located in five genes known to be expressed in two endothelial cell lines as our model system. Results The accuracy and sensitivity of quantitative detection of allelic imbalance was assessed for each SNP by constructing regression lines using a dilution series of mixed samples from individuals of different genotype. Accurate quantification of SNP alleles by both assay formats was evidenced for by R2 values > 0.95 for the majority of the regression lines. According to a two sample t-test, we were able to distinguish 1–9% of a minority SNP allele from a homozygous genotype, with larger variation between SNPs than between assay formats. Six of the SNPs, heterozygous in either of the two cell lines, were genotyped in RNA extracted from the endothelial cells. The coefficient of variation between the fluorescent signals from five parallel reactions was similar for cDNA and genomic DNA. The fluorescence signal intensity ratios measured in the cDNA samples were compared to those in genomic DNA to determine the relative expression levels of the two alleles of each SNP. Four of the six SNPs tested displayed a higher than 1.4-fold difference in allelic ratios between cDNA and genomic DNA. The results were verified by allele-specific oligonucleotide hybridisation and

  2. Current Knowledge on Microarray Technology - An Overview

    African Journals Online (AJOL)

    Erah

    This paper reviews basics and updates of each microarray technology and serves to .... through protein microarrays. Protein microarrays also known as protein chips are nothing but grids that ... conditioned media, patient sera, plasma and urine. Clontech ... based antibody arrays) is similar to membrane-based antibody ...

  3. Diagnostic and analytical applications of protein microarrays

    DEFF Research Database (Denmark)

    Dufva, Hans Martin; Christensen, C.B.V.

    2005-01-01

    DNA microarrays have changed the field of biomedical sciences over the past 10 years. For several reasons, antibody and other protein microarrays have not developed at the same rate. However, protein and antibody arrays have emerged as a powerful tool to complement DNA microarrays during the post...

  4. A salmonid EST genomic study: genes, duplications, phylogeny and microarrays

    Directory of Open Access Journals (Sweden)

    Brahmbhatt Sonal

    2008-11-01

    Full Text Available Abstract Background Salmonids are of interest because of their relatively recent genome duplication, and their extensive use in wild fisheries and aquaculture. A comprehensive gene list and a comparison of genes in some of the different species provide valuable genomic information for one of the most widely studied groups of fish. Results 298,304 expressed sequence tags (ESTs from Atlantic salmon (69% of the total, 11,664 chinook, 10,813 sockeye, 10,051 brook trout, 10,975 grayling, 8,630 lake whitefish, and 3,624 northern pike ESTs were obtained in this study and have been deposited into the public databases. Contigs were built and putative full-length Atlantic salmon clones have been identified. A database containing ESTs, assemblies, consensus sequences, open reading frames, gene predictions and putative annotation is available. The overall similarity between Atlantic salmon ESTs and those of rainbow trout, chinook, sockeye, brook trout, grayling, lake whitefish, northern pike and rainbow smelt is 93.4, 94.2, 94.6, 94.4, 92.5, 91.7, 89.6, and 86.2% respectively. An analysis of 78 transcript sets show Salmo as a sister group to Oncorhynchus and Salvelinus within Salmoninae, and Thymallinae as a sister group to Salmoninae and Coregoninae within Salmonidae. Extensive gene duplication is consistent with a genome duplication in the common ancestor of salmonids. Using all of the available EST data, a new expanded salmonid cDNA microarray of 32,000 features was created. Cross-species hybridizations to this cDNA microarray indicate that this resource will be useful for studies of all 68 salmonid species. Conclusion An extensive collection and analysis of salmonid RNA putative transcripts indicate that Pacific salmon, Atlantic salmon and charr are 94–96% similar while the more distant whitefish, grayling, pike and smelt are 93, 92, 89 and 86% similar to salmon. The salmonid transcriptome reveals a complex history of gene duplication that is

  5. Single-cell multiple gene expression analysis based on single-molecule-detection microarray assay for multi-DNA determination

    Energy Technology Data Exchange (ETDEWEB)

    Li, Lu [School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China); Wang, Xianwei [School of Life Sciences, Shandong University, Jinan 250100 (China); Zhang, Xiaoli [School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China); Wang, Jinxing [School of Life Sciences, Shandong University, Jinan 250100 (China); Jin, Wenrui, E-mail: jwr@sdu.edu.cn [School of Chemistry and Chemical Engineering, Shandong University, Jinan 250100 (China)

    2015-01-07

    Highlights: • A single-molecule-detection (SMD) microarray for 10 samples is fabricated. • The based-SMD microarray assay (SMA) can determine 8 DNAs for each sample. • The limit of detection of SMA is as low as 1.3 × 10{sup −16} mol L{sup −1}. • The SMA can be applied in single-cell multiple gene expression analysis. - Abstract: We report a novel ultra-sensitive and high-selective single-molecule-detection microarray assay (SMA) for multiple DNA determination. In the SMA, a capture DNA (DNAc) microarray consisting of 10 subarrays with 9 spots for each subarray is fabricated on a silanized glass coverslip as the substrate. On the subarrays, the spot-to-spot spacing is 500 μm and each spot has a diameter of ∼300 μm. The sequence of the DNAcs on the 9 spots of a subarray is different, to determine 8 types of target DNAs (DNAts). Thus, 8 types of DNAts are captured to their complementary DNAcs at 8 spots of a subarray, respectively, and then labeled with quantum dots (QDs) attached to 8 types of detection DNAs (DNAds) with different sequences. The ninth spot is used to detect the blank value. In order to determine the same 8 types of DNAts in 10 samples, the 10 DNAc-modified subarrays on the microarray are identical. Fluorescence single-molecule images of the QD-labeled DNAts on each spot of the subarray are acquired using a home-made single-molecule microarray reader. The amounts of the DNAts are quantified by counting the bright dots from the QDs. For a microarray, 8 types of DNAts in 10 samples can be quantified in parallel. The limit of detection of the SMA for DNA determination is as low as 1.3 × 10{sup −16} mol L{sup −1}. The SMA for multi-DNA determination can also be applied in single-cell multiple gene expression analysis through quantification of complementary DNAs (cDNAs) corresponding to multiple messenger RNAs (mRNAs) in single cells. To do so, total RNA in single cells is extracted and reversely transcribed into their cDNAs. Three

  6. cDNA structure, genomic organization and expression patterns of ...

    African Journals Online (AJOL)

    Visfatin was a newly identified adipocytokine, which was involved in various physiologic and pathologic processes of organisms. The cDNA structure, genomic organization and expression patterns of silver Prussian carp visfatin were described in this report. The silver Prussian carp visfatin cDNA cloned from the liver was ...

  7. Characterization of the porcine carboxypeptidase E cDNA

    DEFF Research Database (Denmark)

    Hreidarsdôttir, G.E.; Cirera, Susanna; Fredholm, Merete

    2007-01-01

    the sequence of the cDNA for the porcine CPE gene including all the coding region and the 3'-UTR region was generated. Comparisons with bovine, human, mouse, and rat CPE cDNA sequences showed that the coding regions of the gene are highly conserved both at the nucleotide and at the amino acid level. A very low...

  8. Infectious Maize rayado fino virus from cloned cDNA

    Science.gov (United States)

    Maize rayado fino virus (MRFV) is the type member of the marafiviruses within the family Tymoviridae. A cDNA clone from which infectious RNA can be transcribed was produced from a US isolate of MRFV (MRFV-US). Infectivity of transcripts derived from cDNA clones was demonstrated by infection of mai...

  9. cDNA encoding a polypeptide including a hevein sequence

    Energy Technology Data Exchange (ETDEWEB)

    Raikhel, N.V.; Broekaert, W.F.; Namhai Chua; Kush, A.

    1993-02-16

    A cDNA clone (HEV1) encoding hevein was isolated via polymerase chain reaction (PCR) using mixed oligonucleotides corresponding to two regions of hevein as primers and a Hevea brasiliensis latex cDNA library as a template. HEV1 is 1,018 nucleotides long and includes an open reading frame of 204 amino acids.

  10. Molecular cloning of lupin leghemoglobin cDNA

    DEFF Research Database (Denmark)

    Konieczny, A; Jensen, E O; Marcker, K A

    1987-01-01

    Poly(A)+ RNA isolated from root nodules of yellow lupin (Lupinus luteus, var. Ventus) has been used as a template for the construction of a cDNA library. The ds cDNA was synthesized and inserted into the Hind III site of plasmid pBR 322 using synthetic Hind III linkers. Clones containing sequences...... specific for nodules were selected by differential colony hybridization using 32P-labeled cDNA synthesized either from nodule poly(A)+ RNA or from poly(A)+ RNA of uninfected root as probes. Among the recombinant plasmids, the cDNA gene for leghemoglobin was identified. The protein structure derived from...... its nucleotide sequence was consistent with known amino acid sequence of lupin Lb II. The cloned lupin Lb cDNA hybridized to poly(A)+ RNA from nodules only, which is in accordance with the general concept, that leghemoglobin is expressed exclusively in nodules. Udgivelsesdato: 1987-null...

  11. Universal ligation-detection-reaction microarray applied for compost microbes

    Directory of Open Access Journals (Sweden)

    Romantschuk Martin

    2008-12-01

    Full Text Available Abstract Background Composting is one of the methods utilised in recycling organic communal waste. The composting process is dependent on aerobic microbial activity and proceeds through a succession of different phases each dominated by certain microorganisms. In this study, a ligation-detection-reaction (LDR based microarray method was adapted for species-level detection of compost microbes characteristic of each stage of the composting process. LDR utilises the specificity of the ligase enzyme to covalently join two adjacently hybridised probes. A zip-oligo is attached to the 3'-end of one probe and fluorescent label to the 5'-end of the other probe. Upon ligation, the probes are combined in the same molecule and can be detected in a specific location on a universal microarray with complementary zip-oligos enabling equivalent hybridisation conditions for all probes. The method was applied to samples from Nordic composting facilities after testing and optimisation with fungal pure cultures and environmental clones. Results Probes targeted for fungi were able to detect 0.1 fmol of target ribosomal PCR product in an artificial reaction mixture containing 100 ng competing fungal ribosomal internal transcribed spacer (ITS area or herring sperm DNA. The detection level was therefore approximately 0.04% of total DNA. Clone libraries were constructed from eight compost samples. The LDR microarray results were in concordance with the clone library sequencing results. In addition a control probe was used to monitor the per-spot hybridisation efficiency on the array. Conclusion This study demonstrates that the LDR microarray method is capable of sensitive and accurate species-level detection from a complex microbial community. The method can detect key species from compost samples, making it a basis for a tool for compost process monitoring in industrial facilities.

  12. DNA microarray technique for detecting food-borne pathogens

    Directory of Open Access Journals (Sweden)

    Xing GAO

    2012-08-01

    Full Text Available Objective To study the application of DNA microarray technique for screening and identifying multiple food-borne pathogens. Methods The oligonucleotide probes were designed by Clustal X and Oligo 6.0 at the conserved regions of specific genes of multiple food-borne pathogens, and then were validated by bioinformatic analyses. The 5' end of each probe was modified by amino-group and 10 Poly-T, and the optimized probes were synthesized and spotted on aldehyde-coated slides. The bacteria DNA template incubated with Klenow enzyme was amplified by arbitrarily primed PCR, and PCR products incorporated into Aminoallyl-dUTP were coupled with fluorescent dye. After hybridization of the purified PCR products with DNA microarray, the hybridization image and fluorescence intensity analysis was acquired by ScanArray and GenePix Pro 5.1 software. A series of detection conditions such as arbitrarily primed PCR and microarray hybridization were optimized. The specificity of this approach was evaluated by 16 different bacteria DNA, and the sensitivity and reproducibility were verified by 4 food-borne pathogens DNA. The samples of multiple bacteria DNA and simulated water samples of Shigella dysenteriae were detected. Results Nine different food-borne bacteria were successfully discriminated under the same condition. The sensitivity of genomic DNA was 102 -103pg/ μl, and the coefficient of variation (CV of the reproducibility of assay was less than 15%. The corresponding specific hybridization maps of the multiple bacteria DNA samples were obtained, and the detection limit of simulated water sample of Shigella dysenteriae was 3.54×105cfu/ml. Conclusions The DNA microarray detection system based on arbitrarily primed PCR can be employed for effective detection of multiple food-borne pathogens, and this assay may offer a new method for high-throughput platform for detecting bacteria.

  13. cDNA library Table - KAIKOcDNA | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available c00951-005 Description of data contents List of Bombyx mori cDNA libraries. Data file File name: kaiko_cdna_...library.zip File URL: ftp://ftp.biosciencedbc.jp/archive/kaiko-cdna/LATEST/kaiko_cdna_library.zip File size:... 4.8 KB Simple search URL http://togodb.biosciencedbc.jp/togodb/view/kaiko_cdna_l

  14. Fiscal 1998 achievement report. Industrial technology research and development project. (Strategic human cDNA genome application technology development); 1998 nendo senryakuteki hito cDNA genome oyo gijutsu kaihatsu seika hokokusho

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    2000-03-01

    A human genome related project named above was started, and studies were conducted for base sequence determination and function analysis for approximately 10,000 kinds of full-length or long-chain human cDNA clones owned by research organizations in this country. The Institute of Medical Science of University of Tokyo and Helix Research Institute dealt with a full-length human cDNA library constructed by oligo-capping, and determined the base sequences of all specimens in the library. The Kazusa DNA Research Institute determined partial sequences for long-chain clones which are not shorter than 4-5kbp, and determined entire sequences for some bases. The obtained base sequence data were subjected to homology analysis, the base sequences were converted into amino acid sequences, and functions of proteins were predicted. In the analysis of gene functions, ATAC-PCR (adaptor tagged competitive-polymerase chain reaction) was applied to the clones covered by this project, and a database was prepared by use of the results of analyses of frequency-related information. For the preparation of a comprehensive gene expression profile, technologies for cDNA microarray construction were established. (NEDO)

  15. Transcriptome sequencing of the Microarray Quality Control (MAQC RNA reference samples using next generation sequencing

    Directory of Open Access Journals (Sweden)

    Thierry-Mieg Danielle

    2009-06-01

    Full Text Available Abstract Background Transcriptome sequencing using next-generation sequencing platforms will soon be competing with DNA microarray technologies for global gene expression analysis. As a preliminary evaluation of these promising technologies, we performed deep sequencing of cDNA synthesized from the Microarray Quality Control (MAQC reference RNA samples using Roche's 454 Genome Sequencer FLX. Results We generated more that 3.6 million sequence reads of average length 250 bp for the MAQC A and B samples and introduced a data analysis pipeline for translating cDNA read counts into gene expression levels. Using BLAST, 90% of the reads mapped to the human genome and 64% of the reads mapped to the RefSeq database of well annotated genes with e-values ≤ 10-20. We measured gene expression levels in the A and B samples by counting the numbers of reads that mapped to individual RefSeq genes in multiple sequencing runs to evaluate the MAQC quality metrics for reproducibility, sensitivity, specificity, and accuracy and compared the results with DNA microarrays and Quantitative RT-PCR (QRTPCR from the MAQC studies. In addition, 88% of the reads were successfully aligned directly to the human genome using the AceView alignment programs with an average 90% sequence similarity to identify 137,899 unique exon junctions, including 22,193 new exon junctions not yet contained in the RefSeq database. Conclusion Using the MAQC metrics for evaluating the performance of gene expression platforms, the ExpressSeq results for gene expression levels showed excellent reproducibility, sensitivity, and specificity that improved systematically with increasing shotgun sequencing depth, and quantitative accuracy that was comparable to DNA microarrays and QRTPCR. In addition, a careful mapping of the reads to the genome using the AceView alignment programs shed new light on the complexity of the human transcriptome including the discovery of thousands of new splice variants.

  16. Cloning of the human androgen receptor cDNA

    International Nuclear Information System (INIS)

    Govindan, M.V.; Burelle, M.; Cantin, C.; Kabrie, C.; Labrie, F.; Lachance, Y.; Leblanc, G.; Lefebvre, C.; Patel, P.; Simard, J.

    1988-01-01

    The authors discuss how in order to define the functional domains of the human androgen receptor, complementary DNA (cDNA) clones encoding the human androgen receptor (hAR) have been isolated from a human testis λgtll cDNA library using synthetic oligonnucleotide probes, homologous to segments of the human glucocorticoid, estradiol and progesterone receptors. The cDNA clones corresponding to the human glucocorticoid, estradiol and progesterone receptors were eliminated after cross-hybridization with their respective cDNA probes and/or after restriction mapping of the cDNA clones. The remaining cDNA clones were classified into different groups after analysis by restriction digestion and cross-hybridization. Two of the largest cDNA clones from each group were inserted into an expression vector in both orientations. The linearized plasmids were used as templates in in vitro transcription with T7 RNA polymerase. Subsequent in vitro translation of the purified transcripts in rabbit reticulocyte lysate followed by sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE) permitted the characterization of the encoded polyeptides. The expressed proteins larger than 30,000 Da were analyzed for their ability to bind tritium-labelled dihydrotestosterone ([ 3 H] DHT) with high affinity and specificity

  17. In silico design and performance of peptide microarrays for breast cancer tumour-auto-antibody testing

    Directory of Open Access Journals (Sweden)

    Andreas Weinhäusel

    2012-06-01

    Full Text Available The simplicity and potential of minimally invasive testing using sera from patients makes auto-antibody based biomarkers a very promising tool for use in cancer diagnostics. Protein microarrays have been used for the identification of such auto-antibody signatures. Because high throughput protein expression and purification is laborious, synthetic peptides might be a good alternative for microarray generation and multiplexed analyses. In this study, we designed 1185 antigenic peptides, deduced from proteins expressed by 642 cDNA expression clones found to be sero-reactive in both breast tumour patients and controls. The sero-reactive proteins and the corresponding peptides were used for the production of protein and peptide microarrays. Serum samples from females with benign and malignant breast tumours and healthy control sera (n=16 per group were then analysed. Correct classification of the serum samples on peptide microarrays were 78% for discrimination of ‘malignant versus healthy controls’, 72% for ‘benign versus malignant’ and 94% for ‘benign versus controls’. On protein arrays, correct classification for these contrasts was 69%, 59% and 59%, respectively. The over-representation analysis of the classifiers derived from class prediction showed enrichment of genes associated with ribosomes, spliceosomes, endocytosis and the pentose phosphate pathway. Sequence analyses of the peptides with the highest sero-reactivity demonstrated enrichment of the zinc-finger domain. Peptides’ sero-reactivities were found negatively correlated with hydrophobicity and positively correlated with positive charge, high inter-residue protein contact energies and a secondary structure propensity bias. This study hints at the possibility of using in silico designed antigenic peptide microarrays as an alternative to protein microarrays for the improvement of tumour auto-antibody based diagnostics.

  18. Sequence of human protamine 2 cDNA

    Energy Technology Data Exchange (ETDEWEB)

    Domenjoud, L; Fronia, C; Uhde, F; Engel, W [Universitaet Goettingen (West Germany)

    1988-08-11

    The authors report the cloning and sequencing of a cDNA clone for human protamine 2 (hp2), isolated from a human testis cDNA library cloned in the vector {lambda}-gt11. A 66mer oligonucleotide, that corresponds to an amino acid sequence which is highly conserved between hp2 and mouse protamine 2 (mp2) served as hybridization probe. The homology between the amino acid sequence deduced from our cDNA and the published amino acid sequence for hp2 is 100%.

  19. Spot market for uranium

    International Nuclear Information System (INIS)

    Colhoun, C.

    1982-01-01

    The spot market is always quoted for the price of uranium because little information is available about long-term contracts. A review of the development of spot market prices shows the same price curve swings that occur with all raw materials. Future long-term contracts will probably be lower to reflect spot market prices, which are currently in the real-value range of $30-$35. An upswing in the price of uranium could come in the next few months as utilities begin making purchases and trading from stockpiles. The US, unlike Europe and Japan, has already reached a supply and demand point where the spot market share is increasing. Forecasters cannot project the market price, they can only predict the presence of an oscillating spot or a secondary market. 5 figures

  20. Method for construction of normalized cDNA libraries

    Science.gov (United States)

    Soares, Marcelo B.; Efstratiadis, Argiris

    1998-01-01

    This invention provides a method to normalize a directional cDNA library constructed in a vector that allows propagation in single-stranded circle form comprising: (a) propagating the directional cDNA library in single-stranded circles; (b) generating fragments complementary to the 3' noncoding sequence of the single-stranded circles in the library to produce partial duplexes; (c) purifying the partial duplexes; (d) melting and reassociating the purified partial duplexes to appropriate Cot; and (e) purifying the unassociated single-stranded circles, thereby generating a normalized cDNA library. This invention also provides normalized cDNA libraries generated by the above-described method and uses of the generated libraries.

  1. Cloning, sequencing and expression of cDNA encoding growth ...

    Indian Academy of Sciences (India)

    Unknown

    of medicine, animal husbandry, fish farming and animal ..... northern pike (Esox lucius) growth hormone; Mol. Mar. Biol. ... prolactin 1-luciferase fusion gene in African catfish and ... 1988 Cloning and sequencing of cDNA that encodes goat.

  2. "Harshlighting" small blemishes on microarrays

    Directory of Open Access Journals (Sweden)

    Wittkowski Knut M

    2005-03-01

    Full Text Available Abstract Background Microscopists are familiar with many blemishes that fluorescence images can have due to dust and debris, glass flaws, uneven distribution of fluids or surface coatings, etc. Microarray scans show similar artefacts, which affect the analysis, particularly when one tries to detect subtle changes. However, most blemishes are hard to find by the unaided eye, particularly in high-density oligonucleotide arrays (HDONAs. Results We present a method that harnesses the statistical power provided by having several HDONAs available, which are obtained under similar conditions except for the experimental factor. This method "harshlights" blemishes and renders them evident. We find empirically that about 25% of our chips are blemished, and we analyze the impact of masking them on screening for differentially expressed genes. Conclusion Experiments attempting to assess subtle expression changes should be carefully screened for blemishes on the chips. The proposed method provides investigators with a novel robust approach to improve the sensitivity of microarray analyses. By utilizing topological information to identify and mask blemishes prior to model based analyses, the method prevents artefacts from confounding the process of background correction, normalization, and summarization.

  3. Drop drying on surfaces determines chemical reactivity - the specific case of immobilization of oligonucleotides on microarrays

    Science.gov (United States)

    2013-01-01

    Background Drop drying is a key factor in a wide range of technical applications, including spotted microarrays. The applied nL liquid volume provides specific reaction conditions for the immobilization of probe molecules to a chemically modified surface. Results We investigated the influence of nL and μL liquid drop volumes on the process of probe immobilization and compare the results obtained to the situation in liquid solution. In our data, we observe a strong relationship between drop drying effects on immobilization and surface chemistry. In this work, we present results on the immobilization of dye labeled 20mer oligonucleotides with and without an activating 5′-aminoheptyl linker onto a 2D epoxysilane and a 3D NHS activated hydrogel surface. Conclusions Our experiments identified two basic processes determining immobilization. First, the rate of drop drying that depends on the drop volume and the ambient relative humidity. Oligonucleotides in a dried spot react unspecifically with the surface and long reaction times are needed. 3D hydrogel surfaces allow for immobilization in a liquid environment under diffusive conditions. Here, oligonucleotide immobilization is much faster and a specific reaction with the reactive linker group is observed. Second, the effect of increasing probe concentration as a result of drop drying. On a 3D hydrogel, the increasing concentration of probe molecules in nL spotting volumes accelerates immobilization dramatically. In case of μL volumes, immobilization depends on whether the drop is allowed to dry completely. At non-drying conditions, very limited immobilization is observed due to the low oligonucleotide concentration used in microarray spotting solutions. The results of our study provide a general guideline for microarray assay development. They allow for the initial definition and further optimization of reaction conditions for the immobilization of oligonucleotides and other probe molecule classes to different

  4. Development and evaluation of a high-throughput, low-cost genotyping platform based on oligonucleotide microarrays in rice

    Directory of Open Access Journals (Sweden)

    Liu Bin

    2008-05-01

    Full Text Available Abstract Background We report the development of a microarray platform for rapid and cost-effective genetic mapping, and its evaluation using rice as a model. In contrast to methods employing whole-genome tiling microarrays for genotyping, our method is based on low-cost spotted microarray production, focusing only on known polymorphic features. Results We have produced a genotyping microarray for rice, comprising 880 single feature polymorphism (SFP elements derived from insertions/deletions identified by aligning genomic sequences of the japonica cultivar Nipponbare and the indica cultivar 93-11. The SFPs were experimentally verified by hybridization with labeled genomic DNA prepared from the two cultivars. Using the genotyping microarrays, we found high levels of polymorphism across diverse rice accessions, and were able to classify all five subpopulations of rice with high bootstrap support. The microarrays were used for mapping of a gene conferring resistance to Magnaporthe grisea, the causative organism of rice blast disease, by quantitative genotyping of samples from a recombinant inbred line population pooled by phenotype. Conclusion We anticipate this microarray-based genotyping platform, based on its low cost-per-sample, to be particularly useful in applications requiring whole-genome molecular marker coverage across large numbers of individuals.

  5. Constructing and detecting a cDNA library for mites.

    Science.gov (United States)

    Hu, Li; Zhao, YaE; Cheng, Juan; Yang, YuanJun; Li, Chen; Lu, ZhaoHui

    2015-10-01

    RNA extraction and construction of complementary DNA (cDNA) library for mites have been quite challenging due to difficulties in acquiring tiny living mites and breaking their hard chitin. The present study is to explore a better method to construct cDNA library for mites that will lay the foundation on transcriptome and molecular pathogenesis research. We selected Psoroptes cuniculi as an experimental subject and took the following steps to construct and verify cDNA library. First, we combined liquid nitrogen grinding with TRIzol for total RNA extraction. Then, switching mechanism at 5' end of the RNA transcript (SMART) technique was used to construct full-length cDNA library. To evaluate the quality of cDNA library, the library titer and recombination rate were calculated. The reliability of cDNA library was detected by sequencing and analyzing positive clones and genes amplified by specific primers. The results showed that the RNA concentration was 836 ng/μl and the absorbance ratio at 260/280 nm was 1.82. The library titer was 5.31 × 10(5) plaque-forming unit (PFU)/ml and the recombination rate was 98.21%, indicating that the library was of good quality. In the 33 expressed sequence tags (ESTs) of P. cuniculi, two clones of 1656 and 1658 bp were almost identical with only three variable sites detected, which had an identity of 99.63% with that of Psoroptes ovis, indicating that the cDNA library was reliable. Further detection by specific primers demonstrated that the 553-bp Pso c II gene sequences of P. cuniculi had an identity of 98.56% with those of P. ovis, confirming that the cDNA library was not only reliable but also feasible.

  6. Cloning of the cDNA for human 12-lipoxygenase

    International Nuclear Information System (INIS)

    Izumi, T.; Hoshiko, S.; Radmark, O.; Samuelsson, B.

    1990-01-01

    A full-length cDNA clone encoding 12-lipoxygenase was isolated from a human platelet cDNA library by using a cDNA for human reticulocyte 15-lipoxygenase as probe for the initial screening. The cDNA had an open reading frame encoding 662 amino acid residues with a calculated molecular weight of 75,590. Three independent clones revealed minor heterogeneities in their DNA sequences. Thus, in three positions of the deduced amino acid sequence, there is a choice between two different amino acids. The deduced sequence from the clone plT3 showed 65% identity with human reticulocyte 15-lipoxygenase and 42% identity with human leukocyte 5-lipoxygenase. The 12-lipoxygenase cDNA recognized a 3.0-kilobase mRNA species in platelets and human erythroleukemia cells (HEL cells). Phorbol 12-tetradecanoyl 13-acetate induced megakaryocytic differentiation of HEL cells and 12-lipoxygenase activity and increased mRNA for 12-lipoxygenase. The identity of the cloned 12-lipoxygenase was assured by expression in a mammalian cell line (COS cells). Human platelet 12-lipoxygenase has been difficult to purify to homogeneity. The cloning of this cDNA will increase the possibilities to elucidate the structure and function of this enzyme

  7. SSHscreen and SSHdb, generic software for microarray based gene discovery: application to the stress response in cowpea

    Directory of Open Access Journals (Sweden)

    Oelofse Dean

    2010-04-01

    Full Text Available Abstract Background Suppression subtractive hybridization is a popular technique for gene discovery from non-model organisms without an annotated genome sequence, such as cowpea (Vigna unguiculata (L. Walp. We aimed to use this method to enrich for genes expressed during drought stress in a drought tolerant cowpea line. However, current methods were inefficient in screening libraries and management of the sequence data, and thus there was a need to develop software tools to facilitate the process. Results Forward and reverse cDNA libraries enriched for cowpea drought response genes were screened on microarrays, and the R software package SSHscreen 2.0.1 was developed (i to normalize the data effectively using spike-in control spot normalization, and (ii to select clones for sequencing based on the calculation of enrichment ratios with associated statistics. Enrichment ratio 3 values for each clone showed that 62% of the forward library and 34% of the reverse library clones were significantly differentially expressed by drought stress (adjusted p value 88% of the clones in both libraries were derived from rare transcripts in the original tester samples, thus supporting the notion that suppression subtractive hybridization enriches for rare transcripts. A set of 118 clones were chosen for sequencing, and drought-induced cowpea genes were identified, the most interesting encoding a late embryogenesis abundant Lea5 protein, a glutathione S-transferase, a thaumatin, a universal stress protein, and a wound induced protein. A lipid transfer protein and several components of photosynthesis were down-regulated by the drought stress. Reverse transcriptase quantitative PCR confirmed the enrichment ratio values for the selected cowpea genes. SSHdb, a web-accessible database, was developed to manage the clone sequences and combine the SSHscreen data with sequence annotations derived from BLAST and Blast2GO. The self-BLAST function within SSHdb grouped

  8. Fabrication of protein microarrays for alpha fetoprotein detection by using a rapid photo-immobilization process

    Directory of Open Access Journals (Sweden)

    Sirasa Yodmongkol

    2016-03-01

    Full Text Available In this study, protein microarrays based on sandwich immunoassays are generated to quantify the amount of alpha fetoprotein (AFP in blood serum. For chip generation a mixture of capture antibody and a photoactive copolymer consisting of N,N-dimethylacrylamide (DMAA, methacryloyloxy benzophenone (MaBP, and Na-4-styrenesulfonate (SSNa was spotted onto unmodified polymethyl methacrylate (PMMA substrates. Subsequently to printing of the microarray, the polymer and protein were photochemically cross-linked and the forming, biofunctionalized hydrogels simultaneously bound to the chip surface by short UV- irradiation. The obtained biochip was incubated with AFP antigen, followed by biotinylated AFP antibody and streptavidin-Cy5 and the fluorescence signal read-out. The developed microarray biochip covers the range of AFP in serum samples such as maternal serum in the range of 5 and 100 ng/ml. The chip production process is based on a fast and simple immobilization process, which can be applied to conventional plastic surfaces. Therefore, this protein microarray production process is a promising method to fabricate biochips for AFP screening processes. Keywords: Photo-immobilization, Protein microarray, Alpha fetoprotein, Hydrogel, 3D surface, Down syndrome

  9. Improvement in the amine glass platform by bubbling method for a DNA microarray

    Directory of Open Access Journals (Sweden)

    Jee SH

    2015-10-01

    Full Text Available Seung Hyun Jee,1 Jong Won Kim,2 Ji Hyeong Lee,2 Young Soo Yoon11Department of Chemical and Biological Engineering, Gachon University, Seongnam, Gyeonggi, Republic of Korea; 2Genomics Clinical Research Institute, LabGenomics Co., Ltd., Bundang-gu, Seongnam-si, Gyeonggi-do, Republic of KoreaAbstract: A glass platform with high sensitivity for sexually transmitted diseases microarray is described here. An amino-silane-based self-assembled monolayer was coated on the surface of a glass platform using a novel bubbling method. The optimized surface of the glass platform had highly uniform surface modifications using this method, as well as improved hybridization properties with capture probes in the DNA microarray. On the basis of these results, the improved glass platform serves as a highly reliable and optimal material for the DNA microarray. Moreover, in this study, we demonstrated that our glass platform, manufactured by utilizing the bubbling method, had higher uniformity, shorter processing time, lower background signal, and higher spot signal than the platforms manufactured by the general dipping method. The DNA microarray manufactured with a glass platform prepared using bubbling method can be used as a clinical diagnostic tool. Keywords: DNA microarray, glass platform, bubbling method, self-assambled monolayer

  10. Microarray analysis of thioacetamide-treated type 1 diabetic rats

    International Nuclear Information System (INIS)

    Devi, Sachin S.; Mehendale, Harihara M.

    2006-01-01

    It is well known that diabetes imparts high sensitivity to numerous hepatotoxicants. Previously, we have shown that a normally non-lethal dose of thioacetamide (TA, 300 mg/kg) causes 90% mortality in type 1 diabetic (DB) rats due to inhibited tissue repair allowing progression of liver injury. On the other hand, DB rats exposed to 30 mg TA/kg exhibit delayed tissue repair and delayed recovery from injury. The objective of this study was to investigate the mechanism of impaired tissue repair and progression of liver injury in TA-treated DB rats by using cDNA microarray. Gene expression pattern was examined at 0, 6, and 12 h after TA challenge, and selected mechanistic leads from microarray experiments were confirmed by real-time RT-PCR and further investigated at protein level over the time course of 0 to 36 h after TA treatment. Diabetic condition itself increased gene expression of proteases and decreased gene expression of protease inhibitors. Administration of 300 mg TA/kg to DB rats further elevated gene expression of proteases and suppressed gene expression of protease inhibitors, explaining progression of liver injury in DB rats after TA treatment. Inhibited expression of genes involved in cell division cycle (cyclin D1, IGFBP-1, ras, E2F) was observed after exposure of DB rats to 300 mg TA/kg, explaining inhibited tissue repair in these rats. On the other hand, DB rats receiving 30 mg TA/kg exhibit delayed expression of genes involved in cell division cycle, explaining delayed tissue repair in these rats. In conclusion, impaired cyclin D1 signaling along with increased proteases and decreased protease inhibitors may explain impaired tissue repair that leads to progression of liver injury initiated by TA in DB rats

  11. Microarrays for the evaluation of cell-biomaterial surface interactions

    Science.gov (United States)

    Thissen, H.; Johnson, G.; McFarland, G.; Verbiest, B. C. H.; Gengenbach, T.; Voelcker, N. H.

    2007-01-01

    The evaluation of cell-material surface interactions is important for the design of novel biomaterials which are used in a variety of biomedical applications. While traditional in vitro test methods have routinely used samples of relatively large size, microarrays representing different biomaterials offer many advantages, including high throughput and reduced sample handling. Here, we describe the simultaneous cell-based testing of matrices of polymeric biomaterials, arrayed on glass slides with a low cell-attachment background coating. Arrays were constructed using a microarray robot at 6 fold redundancy with solid pins having a diameter of 375 μm. Printed solutions contained at least one monomer, an initiator and a bifunctional crosslinker. After subsequent UV polymerisation, the arrays were washed and characterised by X-ray photoelectron spectroscopy. Cell culture experiments were carried out over 24 hours using HeLa cells. After labelling with CellTracker ® Green for the final hour of incubation and subsequent fixation, the arrays were scanned. In addition, individual spots were also viewed by fluorescence microscopy. The evaluation of cell-surface interactions in high-throughput assays as demonstrated here is a key enabling technology for the effective development of future biomaterials.

  12. Screening for C3 deficiency in newborns using microarrays.

    Directory of Open Access Journals (Sweden)

    Magdalena Janzi

    Full Text Available BACKGROUND: Dried blood spot samples (DBSS from newborns are widely used in neonatal screening for selected metabolic diseases and diagnostic possibilities for additional disorders are continuously being evaluated. Primary immunodeficiency disorders comprise a group of more than one hundred diseases, several of which are fatal early in life. Yet, a majority of the patients are not diagnosed due to lack of high-throughput screening methods. METHODOLOGY/PRINCIPAL FINDINGS: We have previously developed a system using reverse phase protein microarrays for analysis of IgA levels in serum samples. In this study, we extended the applicability of the method to include determination of complement component C3 levels in eluates from DBSS collected at birth. Normal levels of C3 were readily detected in 269 DBSS from healthy newborns, while no C3 was detected in sera and DBSS from C3 deficient patients. CONCLUSIONS/SIGNIFICANCE: The findings suggest that patients with deficiencies of specific serum proteins can be identified by analysis of DBSS using reverse phase protein microarrays.

  13. Biocompatible Hydrogels for Microarray Cell Printing and Encapsulation

    Directory of Open Access Journals (Sweden)

    Akshata Datar

    2015-10-01

    Full Text Available Conventional drug screening processes are a time-consuming and expensive endeavor, but highly rewarding when they are successful. To identify promising lead compounds, millions of compounds are traditionally screened against therapeutic targets on human cells grown on the surface of 96-wells. These two-dimensional (2D cell monolayers are physiologically irrelevant, thus, often providing false-positive or false-negative results, when compared to cells grown in three-dimensional (3D structures such as hydrogel droplets. However, 3D cell culture systems are not easily amenable to high-throughput screening (HTS, thus inherently low throughput, and requiring relatively large volume for cell-based assays. In addition, it is difficult to control cellular microenvironments and hard to obtain reliable cell images due to focus position and transparency issues. To overcome these problems, miniaturized 3D cell cultures in hydrogels were developed via cell printing techniques where cell spots in hydrogels can be arrayed on the surface of glass slides or plastic chips by microarray spotters and cultured in growth media to form cells encapsulated 3D droplets for various cell-based assays. These approaches can dramatically reduce assay volume, provide accurate control over cellular microenvironments, and allow us to obtain clear 3D cell images for high-content imaging (HCI. In this review, several hydrogels that are compatible to microarray printing robots are discussed for miniaturized 3D cell cultures.

  14. Mononucleosis spot test

    Science.gov (United States)

    Monospot test; Heterophile antibody test; Heterophile agglutination test; Paul-Bunnell test; Forssman antibody test ... The mononucleosis spot test is done when symptoms of mononucleosis are ... Fatigue Fever Large spleen (possibly) Sore throat Tender ...

  15. Carbohydrate Microarrays in Plant Science

    DEFF Research Database (Denmark)

    Fangel, Jonatan Ulrik; Pedersen, H.L.; Vidal-Melgosa, S.

    2012-01-01

    Almost all plant cells are surrounded by glycan-rich cell walls, which form much of the plant body and collectively are the largest source of biomass on earth. Plants use polysaccharides for support, defense, signaling, cell adhesion, and as energy storage, and many plant glycans are also important...... industrially and nutritionally. Understanding the biological roles of plant glycans and the effective exploitation of their useful properties requires a detailed understanding of their structures, occurrence, and molecular interactions. Microarray technology has revolutionized the massively high...... for plant research and can be used to map glycan populations across large numbers of samples to screen antibodies, carbohydrate binding proteins, and carbohydrate binding modules and to investigate enzyme activities....

  16. Arc cathode spots

    International Nuclear Information System (INIS)

    Schrade, H.O.

    1989-01-01

    Arc spots are usually highly unstable and jump statistically over the cathode surface. In a magnetic field parallel to the surface, preferably they move in the retrograde direction; i.e., opposite to the Lorentzian rule. If the field is inclined with respect to the surface, the spots drift away at a certain angle with respect to the proper retrograde direction (Robson drift motion). These well-known phenomena are explained by one stability theory

  17. The Use of cDNA Microarray to Study Gene Expression in Wnt-1 Induced Mammary Tumors

    National Research Council Canada - National Science Library

    Huang, Shixia

    2002-01-01

    .... Specifically, we have collected tissue samples from virgin mammary glands, hyperplastic mammary glands, Wnt- 1 mammary tumors, and tumors metastasized to the lung, and compared their gene expression patterns...

  18. cDNA table - RPD | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available of data contents Results of homology search to cDNA clones in the KOME. Data file File name: rpd_cdna.zip F...ile URL: ftp://ftp.biosciencedbc.jp/archive/rpd/LATEST/rpd_cdna.zip File size: 15 KB Simple search URL http:...//togodb.biosciencedbc.jp/togodb/view/rpd_cdna#en Data acquisition method - Data

  19. Gene ARMADA: an integrated multi-analysis platform for microarray data implemented in MATLAB.

    Science.gov (United States)

    Chatziioannou, Aristotelis; Moulos, Panagiotis; Kolisis, Fragiskos N

    2009-10-27

    The microarray data analysis realm is ever growing through the development of various tools, open source and commercial. However there is absence of predefined rational algorithmic analysis workflows or batch standardized processing to incorporate all steps, from raw data import up to the derivation of significantly differentially expressed gene lists. This absence obfuscates the analytical procedure and obstructs the massive comparative processing of genomic microarray datasets. Moreover, the solutions provided, heavily depend on the programming skills of the user, whereas in the case of GUI embedded solutions, they do not provide direct support of various raw image analysis formats or a versatile and simultaneously flexible combination of signal processing methods. We describe here Gene ARMADA (Automated Robust MicroArray Data Analysis), a MATLAB implemented platform with a Graphical User Interface. This suite integrates all steps of microarray data analysis including automated data import, noise correction and filtering, normalization, statistical selection of differentially expressed genes, clustering, classification and annotation. In its current version, Gene ARMADA fully supports 2 coloured cDNA and Affymetrix oligonucleotide arrays, plus custom arrays for which experimental details are given in tabular form (Excel spreadsheet, comma separated values, tab-delimited text formats). It also supports the analysis of already processed results through its versatile import editor. Besides being fully automated, Gene ARMADA incorporates numerous functionalities of the Statistics and Bioinformatics Toolboxes of MATLAB. In addition, it provides numerous visualization and exploration tools plus customizable export data formats for seamless integration by other analysis tools or MATLAB, for further processing. Gene ARMADA requires MATLAB 7.4 (R2007a) or higher and is also distributed as a stand-alone application with MATLAB Component Runtime. Gene ARMADA provides a

  20. Microarray mRNA expression analysis of Fanconi anemia fibroblasts.

    Science.gov (United States)

    Galetzka, D; Weis, E; Rittner, G; Schindler, D; Haaf, T

    2008-01-01

    Fanconi anemia (FA) cells are generally hypersensitive to DNA cross-linking agents, implying that mutations in the different FANC genes cause a similar DNA repair defect(s). By using a customized cDNA microarray chip for DNA repair- and cell cycle-associated genes, we identified three genes, cathepsin B (CTSB), glutaredoxin (GLRX), and polo-like kinase 2 (PLK2), that were misregulated in untreated primary fibroblasts from three unrelated FA-D2 patients, compared to six controls. Quantitative real-time RT PCR was used to validate these results and to study possible molecular links between FA-D2 and other FA subtypes. GLRX was misregulated to opposite directions in a variety of different FA subtypes. Increased CTSB and decreased PLK2 expression was found in all or almost all of the analyzed complementation groups and, therefore, may be related to the defective FA pathway. Transcriptional upregulation of the CTSB proteinase appears to be a secondary phenomenon due to proliferation differences between FA and normal fibroblast cultures. In contrast, PLK2 is known to play a pivotal role in processes that are linked to FA defects and may contribute in multiple ways to the FA phenotype: PLK2 is a target gene for TP53, is likely to function as a tumor suppressor gene in hematologic neoplasia, and Plk2(-/-) mice are small because of defective embryonal development. (c) 2008 S. Karger AG, Basel.

  1. Use of a multi-thermal washer for DNA microarrays simplifies probe design and gives robust genotyping assays

    DEFF Research Database (Denmark)

    Petersen, J.; Poulsen, Lena; Petronis, S.

    2008-01-01

    is called a multi-thermal array washer (MTAW), and it has eight individually controlled heating zones, each of which corresponds to the location of a subarray on a slide. Allele-specific oligonucleotide probes for nine mutations in the beta-globin gene were spotted in eight identical subarrays at positions......DNA microarrays are generally operated at a single condition, which severely limits the freedom of designing probes for allele-specific hybridization assays. Here, we demonstrate a fluidic device for multi-stringency posthybridization washing of microarrays on microscope slides. This device...

  2. Second-strand cDNA synthesis: classical method

    International Nuclear Information System (INIS)

    Gubler, U.

    1987-01-01

    The classical scheme for the synthesis of double-stranded cDNA as it was reported in 1976 is described. Reverse transcription of mRNA with oligo(dT) as the primer generates first strands with a small loop at the 3' end of the cDNA (the end that corresponds to the 5' end of the mRNA). Subsequent removal of the mRNA by alkaline hydrolysis leaves single-stranded cDNA molecules again with a small 3' loop. This loop can be used by either reverse transcriptase or Klenow fragment of DNA polymerase I as a primer for second-strand synthesis. The resulting products are double-stranded cDNA molecules that are covalently closed at the end corresponding to the 5' end of the original mRNA. Subsequent cleavage of the short piece of single-stranded cDNA within the loop with the single-strand-specific S 1 nuclease generate open double-stranded molecules that can be used for molecular cloning in plasmids or in phage. Useful variations of this scheme have been described

  3. cDNA library construction of two human Demodexspecies.

    Science.gov (United States)

    Niu, DongLing; Wang, RuiLing; Zhao, YaE; Yang, Rui; Hu, Li; Lei, YuYang; Dan, WeiChao

    2017-06-01

    The research of Demodex, a type of pathogen causing various dermatoses in animals and human beings, is lacking at RNA level. This study aims at extracting RNA and constructing cDNA library for Demodex. First, P. cuniculiand D. farinaewere mixed to establish homogenization method for RNA extraction. Second, D. folliculorumand D. breviswere collected and preserved in Trizol, which were mixed with D. farinaerespectively to extract RNA. Finally, cDNA library was constructed and its quality was assessed. The results indicated that for D. folliculorum& D. farinae, the recombination rate of cDNA library was 90.67% and the library titer was 7.50 × 104 pfu/ml. 17 of the 59 positive clones were predicted to be of D. folliculorum; For D. brevis& D. farinae, the recombination rate was 90.96% and the library titer was 7.85 x104 pfu/ml. 40 of the 59 positive clones were predicted to be of D. brevis. Further detection by specific primers demonstrated that mtDNA cox1, cox3and ATP6 detected from cDNA libraries had 96.52%-99.73% identities with the corresponding sequences in GenBank. In conclusion, the cDNA libraries constructed for Demodexmixed with D. farinaewere successful and could satisfy the requirements for functional genes detection.

  4. Cloning and characterization of the human colipase cDNA

    International Nuclear Information System (INIS)

    Lowe, M.E.; Rosenblum, J.L.; McEwen, P.; Strauss, A.W.

    1990-01-01

    Pancreatic lipase hydrolyzes dietary triglycerides to monoglycerides and fatty acids. In the presence of bile salts, the activity of pancreatic lipase is markedly decreased. The activity can be restored by the addition of colipase, a low molecular weight protein secreted by the pancreas. The action of pancreatic lipase in the gut lumen is dependent upon its interaction with colipase. As a first step in elucidating the molecular events governing the interaction of lipase and colipase with each other and with fatty acids, a cDNA encoding human colipase was isolated from a λgt11 cDNA library with a rabbit polyclonal anti-human colipase antibody. The full-length 525 bp cDNA contained an open reading frame encoding 112 amino acids, including a 17 amino acid signal peptide. The predicted sequence contains 100% of the published protein sequence for human colipase determined by chemical methods, but predicts the presence of five additional NH 2 -terminal amino acids and four additional COOH-terminal amino acids. Comparison of the predicted protein sequence with the known sequences of colipase from other species reveals regions of extensive identity. The authors report, for the first time, a cDNA for colipase. The cDNA predicts a human procolipase an suggests that there may also be processing at the COOH-terminus. The regions of identity with colipase from other species will aid in defining the interaction with lipase and lipids through site-specific mutagenesis

  5. MARS: Microarray analysis, retrieval, and storage system

    Directory of Open Access Journals (Sweden)

    Scheideler Marcel

    2005-04-01

    Full Text Available Abstract Background Microarray analysis has become a widely used technique for the study of gene-expression patterns on a genomic scale. As more and more laboratories are adopting microarray technology, there is a need for powerful and easy to use microarray databases facilitating array fabrication, labeling, hybridization, and data analysis. The wealth of data generated by this high throughput approach renders adequate database and analysis tools crucial for the pursuit of insights into the transcriptomic behavior of cells. Results MARS (Microarray Analysis and Retrieval System provides a comprehensive MIAME supportive suite for storing, retrieving, and analyzing multi color microarray data. The system comprises a laboratory information management system (LIMS, a quality control management, as well as a sophisticated user management system. MARS is fully integrated into an analytical pipeline of microarray image analysis, normalization, gene expression clustering, and mapping of gene expression data onto biological pathways. The incorporation of ontologies and the use of MAGE-ML enables an export of studies stored in MARS to public repositories and other databases accepting these documents. Conclusion We have developed an integrated system tailored to serve the specific needs of microarray based research projects using a unique fusion of Web based and standalone applications connected to the latest J2EE application server technology. The presented system is freely available for academic and non-profit institutions. More information can be found at http://genome.tugraz.at.

  6. Annotating breast cancer microarray samples using ontologies

    Science.gov (United States)

    Liu, Hongfang; Li, Xin; Yoon, Victoria; Clarke, Robert

    2008-01-01

    As the most common cancer among women, breast cancer results from the accumulation of mutations in essential genes. Recent advance in high-throughput gene expression microarray technology has inspired researchers to use the technology to assist breast cancer diagnosis, prognosis, and treatment prediction. However, the high dimensionality of microarray experiments and public access of data from many experiments have caused inconsistencies which initiated the development of controlled terminologies and ontologies for annotating microarray experiments, such as the standard microarray Gene Expression Data (MGED) ontology (MO). In this paper, we developed BCM-CO, an ontology tailored specifically for indexing clinical annotations of breast cancer microarray samples from the NCI Thesaurus. Our research showed that the coverage of NCI Thesaurus is very limited with respect to i) terms used by researchers to describe breast cancer histology (covering 22 out of 48 histology terms); ii) breast cancer cell lines (covering one out of 12 cell lines); and iii) classes corresponding to the breast cancer grading and staging. By incorporating a wider range of those terms into BCM-CO, we were able to indexed breast cancer microarray samples from GEO using BCM-CO and MGED ontology and developed a prototype system with web interface that allows the retrieval of microarray data based on the ontology annotations. PMID:18999108

  7. Procedure for normalization of cDNA libraries

    Science.gov (United States)

    Bonaldo, Maria DeFatima; Soares, Marcelo Bento

    1997-01-01

    This invention provides a method to normalize a cDNA library constructed in a vector capable of being converted to single-stranded circles and capable of producing complementary nucleic acid molecules to the single-stranded circles comprising: (a) converting the cDNA library in single-stranded circles; (b) generating complementary nucleic acid molecules to the single-stranded circles; (c) hybridizing the single-stranded circles converted in step (a) with complementary nucleic acid molecules of step (b) to produce partial duplexes to an appropriate Cot; (e) separating the unhybridized single-stranded circles from the hybridized single-stranded circles, thereby generating a normalized cDNA library.

  8. Cloning and expression of cDNA coding for bouganin.

    Science.gov (United States)

    den Hartog, Marcel T; Lubelli, Chiara; Boon, Louis; Heerkens, Sijmie; Ortiz Buijsse, Antonio P; de Boer, Mark; Stirpe, Fiorenzo

    2002-03-01

    Bouganin is a ribosome-inactivating protein that recently was isolated from Bougainvillea spectabilis Willd. In this work, the cloning and expression of the cDNA encoding for bouganin is described. From the cDNA, the amino-acid sequence was deduced, which correlated with the primary sequence data obtained by amino-acid sequencing on the native protein. Bouganin is synthesized as a pro-peptide consisting of 305 amino acids, the first 26 of which act as a leader signal while the 29 C-terminal amino acids are cleaved during processing of the molecule. The mature protein consists of 250 amino acids. Using the cDNA sequence encoding the mature protein of 250 amino acids, a recombinant protein was expressed, purified and characterized. The recombinant molecule had similar activity in a cell-free protein synthesis assay and had comparable toxicity on living cells as compared to the isolated native bouganin.

  9. Metric learning for DNA microarray data analysis

    International Nuclear Information System (INIS)

    Takeuchi, Ichiro; Nakagawa, Masao; Seto, Masao

    2009-01-01

    In many microarray studies, gene set selection is an important preliminary step for subsequent main task such as tumor classification, cancer subtype identification, etc. In this paper, we investigate the possibility of using metric learning as an alternative to gene set selection. We develop a simple metric learning algorithm aiming to use it for microarray data analysis. Exploiting a property of the algorithm, we introduce a novel approach for extending the metric learning to be adaptive. We apply the algorithm to previously studied microarray data on malignant lymphoma subtype identification.

  10. cDNA encoding a polypeptide including a hevein sequence

    Energy Technology Data Exchange (ETDEWEB)

    Raikhel, Natasha V. (Okemos, MI); Broekaert, Willem F. (Dilbeek, BE); Chua, Nam-Hai (Scarsdale, NY); Kush, Anil (New York, NY)

    1993-02-16

    A cDNA clone (HEV1) encoding hevein was isolated via polymerase chain reaction (PCR) using mixed oligonucleotides corresponding to two regions of hevein as primers and a Hevea brasiliensis latex cDNA library as a template. HEV1 is 1018 nucleotides long and includes an open reading frame of 204 amino acids. The deduced amino acid sequence contains a pu GOVERNMENT RIGHTS This application was funded under Department of Energy Contract DE-AC02-76ER01338. The U.S. Government has certain rights under this application and any patent issuing thereon.

  11. Focused Screening of ECM-Selective Adhesion Peptides on Cellulose-Bound Peptide Microarrays.

    Science.gov (United States)

    Kanie, Kei; Kondo, Yuto; Owaki, Junki; Ikeda, Yurika; Narita, Yuji; Kato, Ryuji; Honda, Hiroyuki

    2016-11-19

    The coating of surfaces with bio-functional proteins is a promising strategy for the creation of highly biocompatible medical implants. Bio-functional proteins from the extracellular matrix (ECM) provide effective surface functions for controlling cellular behavior. We have previously screened bio-functional tripeptides for feasibility of mass production with the aim of identifying those that are medically useful, such as cell-selective peptides. In this work, we focused on the screening of tripeptides that selectively accumulate collagen type IV (Col IV), an ECM protein that accelerates the re-endothelialization of medical implants. A SPOT peptide microarray was selected for screening owing to its unique cellulose membrane platform, which can mimic fibrous scaffolds used in regenerative medicine. However, since the library size on the SPOT microarray was limited, physicochemical clustering was used to provide broader variation than that of random peptide selection. Using the custom focused microarray of 500 selected peptides, we assayed the relative binding rates of tripeptides to Col IV, collagen type I (Col I), and albumin. We discovered a cluster of Col IV-selective adhesion peptides that exhibit bio-safety with endothelial cells. The results from this study can be used to improve the screening of regeneration-enhancing peptides.

  12. Focused Screening of ECM-Selective Adhesion Peptides on Cellulose-Bound Peptide Microarrays

    Directory of Open Access Journals (Sweden)

    Kei Kanie

    2016-11-01

    Full Text Available The coating of surfaces with bio-functional proteins is a promising strategy for the creation of highly biocompatible medical implants. Bio-functional proteins from the extracellular matrix (ECM provide effective surface functions for controlling cellular behavior. We have previously screened bio-functional tripeptides for feasibility of mass production with the aim of identifying those that are medically useful, such as cell-selective peptides. In this work, we focused on the screening of tripeptides that selectively accumulate collagen type IV (Col IV, an ECM protein that accelerates the re-endothelialization of medical implants. A SPOT peptide microarray was selected for screening owing to its unique cellulose membrane platform, which can mimic fibrous scaffolds used in regenerative medicine. However, since the library size on the SPOT microarray was limited, physicochemical clustering was used to provide broader variation than that of random peptide selection. Using the custom focused microarray of 500 selected peptides, we assayed the relative binding rates of tripeptides to Col IV, collagen type I (Col I, and albumin. We discovered a cluster of Col IV-selective adhesion peptides that exhibit bio-safety with endothelial cells. The results from this study can be used to improve the screening of regeneration-enhancing peptides.

  13. Gene Expression and Microarray Investigation of Dendrobium ...

    African Journals Online (AJOL)

    blood glucose > 16.7 mmol/L were used as the model group and treated with Dendrobium mixture. (DEN ... Keywords: Diabetes, Gene expression, Dendrobium mixture, Microarray testing ..... homeostasis in airway smooth muscle. Am J.

  14. SLIMarray: Lightweight software for microarray facility management

    Directory of Open Access Journals (Sweden)

    Marzolf Bruz

    2006-10-01

    Full Text Available Abstract Background Microarray core facilities are commonplace in biological research organizations, and need systems for accurately tracking various logistical aspects of their operation. Although these different needs could be handled separately, an integrated management system provides benefits in organization, automation and reduction in errors. Results We present SLIMarray (System for Lab Information Management of Microarrays, an open source, modular database web application capable of managing microarray inventories, sample processing and usage charges. The software allows modular configuration and is well suited for further development, providing users the flexibility to adapt it to their needs. SLIMarray Lite, a version of the software that is especially easy to install and run, is also available. Conclusion SLIMarray addresses the previously unmet need for free and open source software for managing the logistics of a microarray core facility.

  15. cDNA library information - Dicty_cDB | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available List Contact us Dicty_cDB cDNA library information Data detail Data name cDNA library information DOI 10.189...s Data item Description cDNA library name Names of cDNA libraries (AF, AH, CF, CH, FC, FC-IC, FCL, SF, SH, S...(C) 5) sexually fusion-competent KAX3 cells (Gamete phase) (F) cDNA library construction method How to construct cDNA library...dir) 2) Full-length cDNA libraries (oligocapped method)(fl) 3) Gamete-specific subtraction library (sub) cDNA library... construction protocol Link to the webpage describing the protocol for generating cDNA library Size

  16. PATMA: parser of archival tissue microarray

    Directory of Open Access Journals (Sweden)

    Lukasz Roszkowiak

    2016-12-01

    Full Text Available Tissue microarrays are commonly used in modern pathology for cancer tissue evaluation, as it is a very potent technique. Tissue microarray slides are often scanned to perform computer-aided histopathological analysis of the tissue cores. For processing the image, splitting the whole virtual slide into images of individual cores is required. The only way to distinguish cores corresponding to specimens in the tissue microarray is through their arrangement. Unfortunately, distinguishing the correct order of cores is not a trivial task as they are not labelled directly on the slide. The main aim of this study was to create a procedure capable of automatically finding and extracting cores from archival images of the tissue microarrays. This software supports the work of scientists who want to perform further image processing on single cores. The proposed method is an efficient and fast procedure, working in fully automatic or semi-automatic mode. A total of 89% of punches were correctly extracted with automatic selection. With an addition of manual correction, it is possible to fully prepare the whole slide image for extraction in 2 min per tissue microarray. The proposed technique requires minimum skill and time to parse big array of cores from tissue microarray whole slide image into individual core images.

  17. Automatic Identification and Quantification of Extra-Well Fluorescence in Microarray Images.

    Science.gov (United States)

    Rivera, Robert; Wang, Jie; Yu, Xiaobo; Demirkan, Gokhan; Hopper, Marika; Bian, Xiaofang; Tahsin, Tasnia; Magee, D Mitchell; Qiu, Ji; LaBaer, Joshua; Wallstrom, Garrick

    2017-11-03

    In recent studies involving NAPPA microarrays, extra-well fluorescence is used as a key measure for identifying disease biomarkers because there is evidence to support that it is better correlated with strong antibody responses than statistical analysis involving intraspot intensity. Because this feature is not well quantified by traditional image analysis software, identification and quantification of extra-well fluorescence is performed manually, which is both time-consuming and highly susceptible to variation between raters. A system that could automate this task efficiently and effectively would greatly improve the process of data acquisition in microarray studies, thereby accelerating the discovery of disease biomarkers. In this study, we experimented with different machine learning methods, as well as novel heuristics, for identifying spots exhibiting extra-well fluorescence (rings) in microarray images and assigning each ring a grade of 1-5 based on its intensity and morphology. The sensitivity of our final system for identifying rings was found to be 72% at 99% specificity and 98% at 92% specificity. Our system performs this task significantly faster than a human, while maintaining high performance, and therefore represents a valuable tool for microarray image analysis.

  18. Cloning and expression of human deoxycytidine kinase cDNA

    International Nuclear Information System (INIS)

    Chottiner, E.G.; Shewach, D.S.; Datta, N.S.; Ashcraft, E.; Gribbin, D.; Ginsburg, D.; Fox, I.H.; Mitchell, B.S.

    1991-01-01

    Deoxycytidine (dCyd) kinase is required for the phosphorylation of several deoxyribonucleosides and certain nucleoside analogs widely employed as antiviral and chemotherapeutic agents. Detailed analysis of this enzyme has been limited, however, by its low abundance and instability. Using oligonucleotides based on primary amino acid sequence derived from purified dCyd kinase, the authors have screened T-lymphoblast cDNA libraries and identified a cDNA sequence that encodes a 30.5-kDa protein corresponding to the subunit molecular mass of the purified protein. Expression of the cDNA in Escherichia coli results in a 40-fold increase in dCyd kinase activity over control levels. Northern blot analysis reveals a single 2.8-kilobase mRNA expressed in T lymphoblasts at 5- to 10-fold higher levels than in B lymphoblasts, and decreased dCyd kinase mRNA levels are present in T-lymphoblast cell lines resistant to arabinofuranosylcytosine and dideoxycytidine. These findings document that this cDNA encodes the T-lymphoblast dCyd kinase responsible for the phosphorylation of dAdo and dGuo as well as dCyd and arabinofuranosylcytosine

  19. MOLECULAR CHARACTERIZATION OF ENDOCRINE DISRUPTION IN FISH USING CDNA ARRAYS.

    Science.gov (United States)

    We are developing cDNA macroarrays to measure the induction of gene expression in sheepshead minnows and largemouth bass exposed to anthropogenic chemicals that can mimic the action of endogenous hormones. For sheepshead minnows exposed in aqua, we observed similar genetic profil...

  20. A BIOINFORMATIC STRATEGY TO RAPIDLY CHARACTERIZE CDNA LIBRARIES

    Science.gov (United States)

    A Bioinformatic Strategy to Rapidly Characterize cDNA LibrariesG. Charles Ostermeier1, David J. Dix2 and Stephen A. Krawetz1.1Departments of Obstetrics and Gynecology, Center for Molecular Medicine and Genetics, & Institute for Scientific Computing, Wayne State Univer...

  1. NORMAL NASAL GENE EXPRESSION LEVELS USING CDNA ARRAY TECHNOLOGY

    Science.gov (United States)

    Normal Nasal Gene Expression Levels Using cDNA Array Technology. The nasal epithelium is a target site for chemically-induced toxicity and carcinogenicity. To detect and analyze genetic events which contribute to nasal tumor development, we first defined the gene expressi...

  2. Complete cDNA sequence coding for human docking protein

    Energy Technology Data Exchange (ETDEWEB)

    Hortsch, M; Labeit, S; Meyer, D I

    1988-01-11

    Docking protein (DP, or SRP receptor) is a rough endoplasmic reticulum (ER)-associated protein essential for the targeting and translocation of nascent polypeptides across this membrane. It specifically interacts with a cytoplasmic ribonucleoprotein complex, the signal recognition particle (SRP). The nucleotide sequence of cDNA encoding the entire human DP and its deduced amino acid sequence are given.

  3. cDNA, genomic sequence cloning and overexpression of ribosomal ...

    African Journals Online (AJOL)

    RPS16 of eukaryote is a component of the 40S small ribosomal subunit encoded by RPS16 gene and is also a homolog of prokaryotic RPS9. The cDNA and genomic sequence of RPS16 was cloned successfully for the first time from the Giant Panda (Ailuropoda melanoleuca) using reverse transcription-polymerase chain ...

  4. TV spots' impact.

    Science.gov (United States)

    El-bakly, S

    1994-09-01

    The Information, Education and Communication (IEC) Center of the State Information Service was established in 1979 for the purpose of providing information to the people on the population issue. The Ministry of Information has accorded the State Information Service free TV and radio air time for family planning dramas and spots. In the early years information campaigns were organized to make people aware of the population problem by slogans, songs, and cartoons. Around 1984 misconceptions about family planning and contraceptives were attacked through a number of TV and radio spots. A few years later 21 spots on specific contraceptive methods were broadcast which were aired for three years over 3000 times. They were extremely successful. The impact of these TV spots was one of the major reasons why the contraceptive prevalence rate increased from 30% in 1984 to 38% in 1988 and 47% in 1992. Spots were also broadcast about the social implications of large families. The TV soap opera "And The Nile Flows On", with the family planning message interwoven into it, was very well received by the target audience. A program entitled "Wedding of the Month" features couples who know family planning well. The most successful radio program is a 15-20 minute long quiz show for residents of the villages where the Select Villages Project is being implemented. The State Information Service has 60 local information centers in the 26 governorates of Egypt that make plans for the family planning campaign. In 1992 the Minya Initiative, a family planning project was implemented in the Minya Governorate. As a result, the contraceptive prevalence rate rose from 22% to 30% over 18 months. A new project, the Select Village Project, was developed in 1993 that replicates the Minya Initiative on the village level in other governorates. This new project that was implemented in sixteen governorates.

  5. The function analysis of full-length cDNA sequence from IRM-2 mouse cDNA library

    International Nuclear Information System (INIS)

    Wang Qin; Liu Xiaoqiu; Xu Chang; Du Liqing; Sun Zhijuan; Wang Yan; Liu Qiang; Song Li; Li Jin; Fan Feiyue

    2013-01-01

    Objective: To identify the function of full-length cDNA sequence from IRM-2 mouse cDNA library. Methods: Full-length cDNA products were amplified by PCR from IRM-2 mouse cDNA library according to twenty-one pieces of expressed sequence tag. The expression of full-length cDNAs were detected after mouse embryonic fibroblasts were exposed to 6.5 Gy γ-ray radiation. And the effect on the growth of radiosensitivity cells AT5B1VA transfected with full-length cDNAs was investigated. Results: The expression of No.4, 5 and 2 full-length cDNAs from IRM-2 mouse were higher than that of parental ICR and 615 mouse after mouse embryonic fibroblasts irradiated with γ-ray radiation. And the survival rate of AT5B1VA cells transfected with No.4, 5 and 2 full-length cDNAs was high. Conclusion: No.4, 5 and 2 full-length cDNAs of IRM-2 mouse are of high radioresistance. (authors)

  6. Roth spots in pernicious anaemia

    OpenAIRE

    Macauley, Mavin; Nag, Satyajit

    2011-01-01

    Roth spots are white-centred retinal haemorrhages, previously thought to be pathognomonic for subacute bacterial endocarditis. A number of other conditions can be associated with Roth spots. In this case, the authors describe the association of Roth spots and pernicious anaemia. This association has been rarely described in the medical literature. Correct diagnosis and treatment with intramuscular vitamin B12 injections resulted in complete resolution of the anaemia and Roth spots. The author...

  7. Generalization of DNA microarray dispersion properties: microarray equivalent of t-distribution

    DEFF Research Database (Denmark)

    Novak, Jaroslav P; Kim, Seon-Young; Xu, Jun

    2006-01-01

    BACKGROUND: DNA microarrays are a powerful technology that can provide a wealth of gene expression data for disease studies, drug development, and a wide scope of other investigations. Because of the large volume and inherent variability of DNA microarray data, many new statistical methods have...

  8. Poisson Spot with Magnetic Levitation

    Science.gov (United States)

    Hoover, Matthew; Everhart, Michael; D'Arruda, Jose

    2010-01-01

    In this paper we describe a unique method for obtaining the famous Poisson spot without adding obstacles to the light path, which could interfere with the effect. A Poisson spot is the interference effect from parallel rays of light diffracting around a solid spherical object, creating a bright spot in the center of the shadow.

  9. Nanotechnology: moving from microarrays toward nanoarrays.

    Science.gov (United States)

    Chen, Hua; Li, Jun

    2007-01-01

    Microarrays are important tools for high-throughput analysis of biomolecules. The use of microarrays for parallel screening of nucleic acid and protein profiles has become an industry standard. A few limitations of microarrays are the requirement for relatively large sample volumes and elongated incubation time, as well as the limit of detection. In addition, traditional microarrays make use of bulky instrumentation for the detection, and sample amplification and labeling are quite laborious, which increase analysis cost and delays the time for obtaining results. These problems limit microarray techniques from point-of-care and field applications. One strategy for overcoming these problems is to develop nanoarrays, particularly electronics-based nanoarrays. With further miniaturization, higher sensitivity, and simplified sample preparation, nanoarrays could potentially be employed for biomolecular analysis in personal healthcare and monitoring of trace pathogens. In this chapter, it is intended to introduce the concept and advantage of nanotechnology and then describe current methods and protocols for novel nanoarrays in three aspects: (1) label-free nucleic acids analysis using nanoarrays, (2) nanoarrays for protein detection by conventional optical fluorescence microscopy as well as by novel label-free methods such as atomic force microscopy, and (3) nanoarray for enzymatic-based assay. These nanoarrays will have significant applications in drug discovery, medical diagnosis, genetic testing, environmental monitoring, and food safety inspection.

  10. Integrative missing value estimation for microarray data.

    Science.gov (United States)

    Hu, Jianjun; Li, Haifeng; Waterman, Michael S; Zhou, Xianghong Jasmine

    2006-10-12

    Missing value estimation is an important preprocessing step in microarray analysis. Although several methods have been developed to solve this problem, their performance is unsatisfactory for datasets with high rates of missing data, high measurement noise, or limited numbers of samples. In fact, more than 80% of the time-series datasets in Stanford Microarray Database contain less than eight samples. We present the integrative Missing Value Estimation method (iMISS) by incorporating information from multiple reference microarray datasets to improve missing value estimation. For each gene with missing data, we derive a consistent neighbor-gene list by taking reference data sets into consideration. To determine whether the given reference data sets are sufficiently informative for integration, we use a submatrix imputation approach. Our experiments showed that iMISS can significantly and consistently improve the accuracy of the state-of-the-art Local Least Square (LLS) imputation algorithm by up to 15% improvement in our benchmark tests. We demonstrated that the order-statistics-based integrative imputation algorithms can achieve significant improvements over the state-of-the-art missing value estimation approaches such as LLS and is especially good for imputing microarray datasets with a limited number of samples, high rates of missing data, or very noisy measurements. With the rapid accumulation of microarray datasets, the performance of our approach can be further improved by incorporating larger and more appropriate reference datasets.

  11. Integrative missing value estimation for microarray data

    Directory of Open Access Journals (Sweden)

    Zhou Xianghong

    2006-10-01

    Full Text Available Abstract Background Missing value estimation is an important preprocessing step in microarray analysis. Although several methods have been developed to solve this problem, their performance is unsatisfactory for datasets with high rates of missing data, high measurement noise, or limited numbers of samples. In fact, more than 80% of the time-series datasets in Stanford Microarray Database contain less than eight samples. Results We present the integrative Missing Value Estimation method (iMISS by incorporating information from multiple reference microarray datasets to improve missing value estimation. For each gene with missing data, we derive a consistent neighbor-gene list by taking reference data sets into consideration. To determine whether the given reference data sets are sufficiently informative for integration, we use a submatrix imputation approach. Our experiments showed that iMISS can significantly and consistently improve the accuracy of the state-of-the-art Local Least Square (LLS imputation algorithm by up to 15% improvement in our benchmark tests. Conclusion We demonstrated that the order-statistics-based integrative imputation algorithms can achieve significant improvements over the state-of-the-art missing value estimation approaches such as LLS and is especially good for imputing microarray datasets with a limited number of samples, high rates of missing data, or very noisy measurements. With the rapid accumulation of microarray datasets, the performance of our approach can be further improved by incorporating larger and more appropriate reference datasets.

  12. On the Spot: Oceans

    OpenAIRE

    Male, Alan; Butterfield, Moira

    2000-01-01

    This a children's non-fiction, knowledge bearing picture book that is part of a Reader's Digest series called 'On the Spot'. The series deals with a range of topics related to the natural world and this one introduces its young audience to the ecosystems of the oceans. \\ud The publication was illustrated and designed by the author (Alan Male) and is technically described as a board book with interactive 'pop up' features, specifically conceived to engage children's discovery and learning thro...

  13. El spot electoral negativo

    Directory of Open Access Journals (Sweden)

    Palma Peña-Jiménez

    2011-01-01

    Full Text Available l spot político tiene durante la campaña un objetivo final inequívoco: la consecución del voto favorable. Se dirige al cuerpo electoral a través de la televisión y de Internet, y presenta, en muchos casos, un planteamiento negativo, albergando mensajes destinados a la crítica frontal contra el adversario, más que a la exposición de propuestas propias. Este artículo se centra en el análisis del spot electoral negativo, en aquellas producciones audiovisuales construidas sin más causa que la reprobación del contrincante. Se trata de vídeos que, lejos de emplearse en difundir las potencialidades de la organización y las virtudes de su candidato –además de su programa electoral–, consumen su tiempo en descalificar al oponente mediante la transmisión de mensajes, muchas veces, ad hominem. Repasamos el planteamiento negativo del spot electoral desde su primera manifestación, que en España data de 1996, año de emisión del conocido como vídeo del dóberman, sin olvidar otros ejemplos que completan el objeto de estudio.

  14. Roth spots in pernicious anaemia.

    Science.gov (United States)

    Macauley, Mavin; Nag, Satyajit

    2011-04-19

    Roth spots are white-centred retinal haemorrhages, previously thought to be pathognomonic for subacute bacterial endocarditis. A number of other conditions can be associated with Roth spots. In this case, the authors describe the association of Roth spots and pernicious anaemia. This association has been rarely described in the medical literature. Correct diagnosis and treatment with intramuscular vitamin B(12) injections resulted in complete resolution of the anaemia and Roth spots. The authors hope to alert clinicians to think of various differentials of Roth spots, and initiate prompt investigation and management.

  15. cDNA - ASTRA | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available ontents List of cDNA in locus Data file File name: astra_cdna.zip File URL: ftp://ftp.biosciencedbc.jp/archive/astra/LATEST/astra_cdn...a.zip File size: 3.3 MB Simple search URL http://togodb.biosciencedbc.jp/togodb/view/astra_cdna...n, Department of Molecular Genetics, National Institute of Agrobiological Sciences (Kikuchi et al., 2003; ftp://cdna

  16. Molecular cloning of growth hormone encoding cDNA of Indian

    Indian Academy of Sciences (India)

    A modified rapid amplification of cDNA ends (RACE) strategy has been developed for cloning highly conserved cDNA sequences. Using this modified method, the growth hormone (GH) encoding cDNA sequences of Labeo rohita, Cirrhina mrigala and Catla catla have been cloned, characterized and overexpressed in ...

  17. EST and microarray analysis of horn development in Onthophagus beetles

    Directory of Open Access Journals (Sweden)

    Tang Zuojian

    2009-10-01

    Full Text Available Abstract Background The origin of novel traits and their subsequent diversification represent central themes in evo-devo and evolutionary ecology. Here we explore the genetic and genomic basis of a class of traits that is both novel and highly diverse, in a group of organisms that is ecologically complex and experimentally tractable: horned beetles. Results We developed two high quality, normalized cDNA libraries for larval and pupal Onthophagus taurus and sequenced 3,488 ESTs that assembled into 451 contigs and 2,330 singletons. We present the annotation and a comparative analysis of the conservation of the sequences. Microarrays developed from the combined libraries were then used to contrast the transcriptome of developing primordia of head horns, prothoracic horns, and legs. Our experiments identify a first comprehensive list of candidate genes for the evolution and diversification of beetle horns. We find that developing horns and legs show many similarities as well as important differences in their transcription profiles, suggesting that the origin of horns was mediated partly, but not entirely, by the recruitment of genes involved in the formation of more traditional appendages such as legs. Furthermore, we find that horns developing from the head and prothorax differ in their transcription profiles to a degree that suggests that head and prothoracic horns are not serial homologs, but instead may have evolved independently from each other. Conclusion We have laid the foundation for a systematic analysis of the genetic basis of horned beetle development and diversification with the potential to contribute significantly to several major frontiers in evolutionary developmental biology.

  18. Discovering biological progression underlying microarray samples.

    Directory of Open Access Journals (Sweden)

    Peng Qiu

    2011-04-01

    Full Text Available In biological systems that undergo processes such as differentiation, a clear concept of progression exists. We present a novel computational approach, called Sample Progression Discovery (SPD, to discover patterns of biological progression underlying microarray gene expression data. SPD assumes that individual samples of a microarray dataset are related by an unknown biological process (i.e., differentiation, development, cell cycle, disease progression, and that each sample represents one unknown point along the progression of that process. SPD aims to organize the samples in a manner that reveals the underlying progression and to simultaneously identify subsets of genes that are responsible for that progression. We demonstrate the performance of SPD on a variety of microarray datasets that were generated by sampling a biological process at different points along its progression, without providing SPD any information of the underlying process. When applied to a cell cycle time series microarray dataset, SPD was not provided any prior knowledge of samples' time order or of which genes are cell-cycle regulated, yet SPD recovered the correct time order and identified many genes that have been associated with the cell cycle. When applied to B-cell differentiation data, SPD recovered the correct order of stages of normal B-cell differentiation and the linkage between preB-ALL tumor cells with their cell origin preB. When applied to mouse embryonic stem cell differentiation data, SPD uncovered a landscape of ESC differentiation into various lineages and genes that represent both generic and lineage specific processes. When applied to a prostate cancer microarray dataset, SPD identified gene modules that reflect a progression consistent with disease stages. SPD may be best viewed as a novel tool for synthesizing biological hypotheses because it provides a likely biological progression underlying a microarray dataset and, perhaps more importantly, the

  19. The cDNA sequence of a neutral horseradish peroxidase.

    Science.gov (United States)

    Bartonek-Roxå, E; Eriksson, H; Mattiasson, B

    1991-02-16

    A cDNA clone encoding a horseradish (Armoracia rusticana) peroxidase has been isolated and characterized. The cDNA contains 1378 nucleotides excluding the poly(A) tail and the deduced protein contains 327 amino acids which includes a 28 amino acid leader sequence. The predicted amino acid sequence is nine amino acids shorter than the major isoenzyme belonging to the horseradish peroxidase C group (HRP-C) and the sequence shows 53.7% identity with this isoenzyme. The described clone encodes nine cysteines of which eight correspond well with the cysteines found in HRP-C. Five potential N-glycosylation sites with the general sequence Asn-X-Thr/Ser are present in the deduced sequence. Compared to the earlier described HRP-C this is three glycosylation sites less. The shorter sequence and fewer N-glycosylation sites give the native isoenzyme a molecular weight of several thousands less than the horseradish peroxidase C isoenzymes. Comparison with the net charge value of HRP-C indicates that the described cDNA clone encodes a peroxidase which has either the same or a slightly less basic pI value, depending on whether the encoded protein is N-terminally blocked or not. This excludes the possibility that HRP-n could belong to either the HRP-A, -D or -E groups. The low sequence identity (53.7%) with HRP-C indicates that the described clone does not belong to the HRP-C isoenzyme group and comparison of the total amino acid composition with the HRP-B group does not place the described clone within this isoenzyme group. Our conclusion is that the described cDNA clone encodes a neutral horseradish peroxidase which belongs to a new, not earlier described, horseradish peroxidase group.

  20. [Research on the relevance between the virulent genes differential expression and pathogenecity of Leptospira with microarray].

    Science.gov (United States)

    Yu, De-li; Bao, Lang

    2015-01-01

    To find the change of virulent gene expression and to analyze the relevance between the virulent change and the gene expression. Grouped guinea pigs were inoculated with 1 mL Leptospira cultured in vivo, Leptospira cultured in vitro and the Leptospira culture medium through abdominal subcutaneous respectively. The survival rate, body mass and temperature change of guinea pigs in different groups were measured within 15 d after the inoculation, then the survived guinea pigs were scarified, and the organ coefficient was also measured to know the virulence of Leptospira cultured in different environment. The amplified gene segments from Leptospira were used as probes and wrote the microarray. The total RNA was extracted from Leptospira standard strain cultured in culture medium and guinea pigs. After reverse transcription to cDNA, they were labeled with Cy3 and Cy5 respectively. Labeled cDNA was mixed and hybridized with the microarray. The hybridized mircroarray was scanned and analysed. The survival rate of inoculated guinea pig was different from group to group (in vivo group: 0%; in vitro group: 88.9%; culture medium group: 100%). The guinea pigs in vivo group had a higher temperature (PLeptospira: LA1027, LA1029, LA4004, LA3050, LA3540, LA0327, LA0378, LA1650, LA3937, LA2089, LA2144, LA3576, LA0011 and gene of Loa22 were up regulation after continuously cultured in guinea pigs. The pathogenic ability of Leptospira cultured in different environment is different and the gene expression of Leptospira is different between in vivo and in vitro as well. The understanding of the meaning of this change might help to know the pathogenecity of Leptospira.

  1. The use of microarrays in microbial ecology

    Energy Technology Data Exchange (ETDEWEB)

    Andersen, G.L.; He, Z.; DeSantis, T.Z.; Brodie, E.L.; Zhou, J.

    2009-09-15

    Microarrays have proven to be a useful and high-throughput method to provide targeted DNA sequence information for up to many thousands of specific genetic regions in a single test. A microarray consists of multiple DNA oligonucleotide probes that, under high stringency conditions, hybridize only to specific complementary nucleic acid sequences (targets). A fluorescent signal indicates the presence and, in many cases, the abundance of genetic regions of interest. In this chapter we will look at how microarrays are used in microbial ecology, especially with the recent increase in microbial community DNA sequence data. Of particular interest to microbial ecologists, phylogenetic microarrays are used for the analysis of phylotypes in a community and functional gene arrays are used for the analysis of functional genes, and, by inference, phylotypes in environmental samples. A phylogenetic microarray that has been developed by the Andersen laboratory, the PhyloChip, will be discussed as an example of a microarray that targets the known diversity within the 16S rRNA gene to determine microbial community composition. Using multiple, confirmatory probes to increase the confidence of detection and a mismatch probe for every perfect match probe to minimize the effect of cross-hybridization by non-target regions, the PhyloChip is able to simultaneously identify any of thousands of taxa present in an environmental sample. The PhyloChip is shown to reveal greater diversity within a community than rRNA gene sequencing due to the placement of the entire gene product on the microarray compared with the analysis of up to thousands of individual molecules by traditional sequencing methods. A functional gene array that has been developed by the Zhou laboratory, the GeoChip, will be discussed as an example of a microarray that dynamically identifies functional activities of multiple members within a community. The recent version of GeoChip contains more than 24,000 50mer

  2. 3D Biomaterial Microarrays for Regenerative Medicine

    DEFF Research Database (Denmark)

    Gaharwar, Akhilesh K.; Arpanaei, Ayyoob; Andresen, Thomas Lars

    2015-01-01

    Three dimensional (3D) biomaterial microarrays hold enormous promise for regenerative medicine because of their ability to accelerate the design and fabrication of biomimetic materials. Such tissue-like biomaterials can provide an appropriate microenvironment for stimulating and controlling stem...... for tissue engineering and drug screening applications....... cell differentiation into tissue-specifi c lineages. The use of 3D biomaterial microarrays can, if optimized correctly, result in a more than 1000-fold reduction in biomaterials and cells consumption when engineering optimal materials combinations, which makes these miniaturized systems very attractive...

  3. CDNA encoding a polypeptide including a hevein sequence

    Science.gov (United States)

    Raikhel, Natasha V.; Broekaert, Willem F.; Chua, Nam-Hai; Kush, Anil

    1995-03-21

    A cDNA clone (HEV1) encoding hevein was isolated via polymerase chain reaction (PCR) using mixed oligonucleotides corresponding to two regions of hevein as primers and a Hevea brasiliensis latex cDNA library as a template. HEV1 is 1018 nucleotides long and includes an open reading frame of 204 amino acids. The deduced amino acid sequence contains a putative signal sequence of 17 amino acid residues followed by a 187 amino acid polypeptide. The amino-terminal region (43 amino acids) is identical to hevein and shows homology to several chitin-binding proteins and to the amino-termini of wound-induced genes in potato and poplar. The carboxyl-terminal portion of the polypeptide (144 amino acids) is 74-79% homologous to the carboxyl-terminal region of wound-inducible genes of potato. Wounding, as well as application of the plant hormones abscisic acid and ethylene, resulted in accumulation of hevein transcripts in leaves, stems and latex, but not in roots, as shown by using the cDNA as a probe. A fusion protein was produced in E. coli from the protein of the present invention and maltose binding protein produced by the E. coli.

  4. cDNA encoding a polypeptide including a hevein sequence

    Energy Technology Data Exchange (ETDEWEB)

    Raikhel, N.V.; Broekaert, W.F.; Chua, N.H.; Kush, A.

    2000-07-04

    A cDNA clone (HEV1) encoding hevein was isolated via polymerase chain reaction (PCR) using mixed oligonucleotides corresponding to two regions of hevein as primers and a Hevea brasiliensis latex cDNA library as a template. HEV1 is 1018 nucleotides long and includes an open reading frame of 204 amino acids. The deduced amino acid sequence contains a putative signal sequence of 17 amino acid residues followed by a 187 amino acid polypeptide. The amino-terminal region (43 amino acids) is identical to hevein and shows homology to several chitin-binding proteins and to the amino-termini of wound-induced genes in potato and poplar. The carboxyl-terminal portion of the polypeptide (144 amino acids) is 74--79% homologous to the carboxyl-terminal region of wound-inducible genes of potato. Wounding, as well as application of the plant hormones abscisic acid and ethylene, resulted in accumulation of hevein transcripts in leaves, stems and latex, but not in roots, as shown by using the cDNA as a probe. A fusion protein was produced in E. coli from the protein of the present invention and maltose binding protein produced by the E. coli.

  5. Brain cDNA clone for human cholinesterase

    International Nuclear Information System (INIS)

    McTiernan, C.; Adkins, S.; Chatonnet, A.; Vaughan, T.A.; Bartels, C.F.; Kott, M.; Rosenberry, T.L.; La Du, B.N.; Lockridge, O.

    1987-01-01

    A cDNA library from human basal ganglia was screened with oligonucleotide probes corresponding to portions of the amino acid sequence of human serum cholinesterase. Five overlapping clones, representing 2.4 kilobases, were isolated. The sequenced cDNA contained 207 base pairs of coding sequence 5' to the amino terminus of the mature protein in which there were four ATG translation start sites in the same reading frame as the protein. Only the ATG coding for Met-(-28) lay within a favorable consensus sequence for functional initiators. There were 1722 base pairs of coding sequence corresponding to the protein found circulating in human serum. The amino acid sequence deduced from the cDNA exactly matched the 574 amino acid sequence of human serum cholinesterase, as previously determined by Edman degradation. Therefore, our clones represented cholinesterase rather than acetylcholinesterase. It was concluded that the amino acid sequences of cholinesterase from two different tissues, human brain and human serum, were identical. Hybridization of genomic DNA blots suggested that a single gene, or very few genes coded for cholinesterase

  6. cDNA encoding a polypeptide including a hevein sequence

    Energy Technology Data Exchange (ETDEWEB)

    Raikhel, N.V.; Broekaert, W.F.; Chua, N.H.; Kush, A.

    1999-05-04

    A cDNA clone (HEV1) encoding hevein was isolated via polymerase chain reaction (PCR) using mixed oligonucleotides corresponding to two regions of hevein as primers and a Hevea brasiliensis latex cDNA library as a template. HEV1 is 1018 nucleotides long and includes an open reading frame of 204 amino acids. The deduced amino acid sequence contains a putative signal sequence of 17 amino acid residues followed by a 187 amino acid polypeptide. The amino-terminal region (43 amino acids) is identical to hevein and shows homology to several chitin-binding proteins and to the amino-termini of wound-induced genes in potato and poplar. The carboxyl-terminal portion of the polypeptide (144 amino acids) is 74--79% homologous to the carboxyl-terminal region of wound-inducible genes of potato. Wounding, as well as application of the plant hormones abscisic acid and ethylene, resulted in accumulation of hevein transcripts in leaves, stems and latex, but not in roots, as shown by using the cDNA as a probe. A fusion protein was produced in E. coli from the protein of the present invention and maltose binding protein produced by the E. coli. 12 figs.

  7. cDNA encoding a polypeptide including a hevein sequence

    Energy Technology Data Exchange (ETDEWEB)

    Raikhel, Natasha V. (Okemos, MI); Broekaert, Willem F. (Dilbeek, BE); Chua, Nam-Hai (Scarsdale, NY); Kush, Anil (New York, NY)

    1999-05-04

    A cDNA clone (HEV1) encoding hevein was isolated via polymerase chain reaction (PCR) using mixed oligonucleotides corresponding to two regions of hevein as primers and a Hevea brasiliensis latex cDNA library as a template. HEV1 is 1018 nucleotides long and includes an open reading frame of 204 amino acids. The deduced amino acid sequence contains a putative signal sequence of 17 amino acid residues followed by a 187 amino acid polypeptide. The amino-terminal region (43 amino acids) is identical to hevein and shows homology to several chitin-binding proteins and to the amino-termini of wound-induced genes in potato and poplar. The carboxyl-terminal portion of the polypeptide (144 amino acids) is 74-79% homologous to the carboxyl-terminal region of wound-inducible genes of potato. Wounding, as well as application of the plant hormones abscisic acid and ethylene, resulted in accumulation of hevein transcripts in leaves, stems and latex, but not in roots, as shown by using the cDNA as a probe. A fusion protein was produced in E. coli from the protein of the present invention and maltose binding protein produced by the E. coli.

  8. cDNA encoding a polypeptide including a hevein sequence

    Energy Technology Data Exchange (ETDEWEB)

    Raikhel, N.V.; Broekaert, W.F.; Chua, N.H.; Kush, A.

    1995-03-21

    A cDNA clone (HEV1) encoding hevein was isolated via polymerase chain reaction (PCR) using mixed oligonucleotides corresponding to two regions of hevein as primers and a Hevea brasiliensis latex cDNA library as a template. HEV1 is 1,018 nucleotides long and includes an open reading frame of 204 amino acids. The deduced amino acid sequence contains a putative signal sequence of 17 amino acid residues followed by a 187 amino acid polypeptide. The amino-terminal region (43 amino acids) is identical to hevein and shows homology to several chitin-binding proteins and to the amino-termini of wound-induced genes in potato and poplar. The carboxyl-terminal portion of the polypeptide (144 amino acids) is 74--79% homologous to the carboxyl-terminal region of wound-inducible genes of potato. Wounding, as well as application of the plant hormones abscisic acid and ethylene, resulted in accumulation of hevein transcripts in leaves, stems and latex, but not in roots, as shown by using the cDNA as a probe. A fusion protein was produced in E. coli from the protein of the present invention and maltose binding protein produced by the E. coli. 11 figures.

  9. Radioactive and enzymatic cloned cDNA probes for bovine enteric coronavirus detection by molecular hybridization

    International Nuclear Information System (INIS)

    Collomb, J.; Finance, C.; Alabouch, S.; Laporte, J.

    1992-01-01

    Genomic RNA of F15 strain bovine enteric coronavirus (BECV) was cloned in E. coli. Three clones (174, 160, PG 78), selected in the cDNA library, including a large portion of the nucleocapsid (N), matrix (M) and peplomeric (S) protein genes , were used as probes for a slot blot hybridization assay. Two probe labelling techniques were compared, radiolabelling with 32 P and enzymatic labelling through covalent linkage to peroxidase and chemiluminescence detection. The radioactive probe 174 detected as little as 1 to 3 pg of viral RNA, while the less sensitive enzymatic probe could not reveal more than 100 pg of RNA. No significant detection amplification was achieved when a mixture of the three probes was used. Probe 174 allowed specific identification for BECV. No hybridization was noticed either with rotaviruses or even with other antigenically unrelated members of the family Coronaviridae such as transmissible gastroenteritis virus. The test proved valid for detection of BECV in the supernatant of infected HRT-18 cells: genomic RNA could be detected after direct spotting of samples, but prior nucleic acid extraction after proteinase K treatment improved virus detection. BECV diagnosis in faecal samples using enzymatic probe was compared with conventional diagnostic methods. (authors)

  10. Radioactive and enzymatic cloned cDNA probes for bovine enteric coronavirus detection by molecular hybridization

    Energy Technology Data Exchange (ETDEWEB)

    Collomb, J; Finance, C; Alabouch, S [Lab. de Microbiologie Moleculaire, Faculte des Sciences Pharmaceutiques et Biologiques, Univ. de Nancy I, Nancy (France); Laporte, J [Station de Virologie et d' Immunologie Moleculaires, INRA, Jouy-en-Josas (France)

    1992-01-01

    Genomic RNA of F15 strain bovine enteric coronavirus (BECV) was cloned in E. coli. Three clones (174, 160, PG 78), selected in the cDNA library, including a large portion of the nucleocapsid (N), matrix (M) and peplomeric (S) protein genes , were used as probes for a slot blot hybridization assay. Two probe labelling techniques were compared, radiolabeled with [sup 32]P and enzymatic labeled through covalent linkage to peroxidase for chemiluminescence detection. The radioactive probe 174 detected as little as 1-3 pg of viral RNA, while the less sensitive enzymatic probe could not reveal more than 100 pg of RNA. No significant detection amplification was achieved when a mixture of the three probes was used. Probe 174 allowed specific identification for BECV. No hybridization was noticed either with rotaviruses or even with other antigenically unrelated members of the family Coronaviridae such as transmissible gastroenteritis virus. The test proved valid for detection of BECV in the supernatant of infected HRT-18 cells: genomic RNA could be detected after direct spotting of samples, but prior nucleic acid extraction after proteinase K treatment improved virus detection. BECV diagnosis in fecal samples using enzymatic probe was compared with conventional diagnostic methods. (authors).

  11. Microarray Я US: a user-friendly graphical interface to Bioconductor tools that enables accurate microarray data analysis and expedites comprehensive functional analysis of microarray results.

    Science.gov (United States)

    Dai, Yilin; Guo, Ling; Li, Meng; Chen, Yi-Bu

    2012-06-08

    Microarray data analysis presents a significant challenge to researchers who are unable to use the powerful Bioconductor and its numerous tools due to their lack of knowledge of R language. Among the few existing software programs that offer a graphic user interface to Bioconductor packages, none have implemented a comprehensive strategy to address the accuracy and reliability issue of microarray data analysis due to the well known probe design problems associated with many widely used microarray chips. There is also a lack of tools that would expedite the functional analysis of microarray results. We present Microarray Я US, an R-based graphical user interface that implements over a dozen popular Bioconductor packages to offer researchers a streamlined workflow for routine differential microarray expression data analysis without the need to learn R language. In order to enable a more accurate analysis and interpretation of microarray data, we incorporated the latest custom probe re-definition and re-annotation for Affymetrix and Illumina chips. A versatile microarray results output utility tool was also implemented for easy and fast generation of input files for over 20 of the most widely used functional analysis software programs. Coupled with a well-designed user interface, Microarray Я US leverages cutting edge Bioconductor packages for researchers with no knowledge in R language. It also enables a more reliable and accurate microarray data analysis and expedites downstream functional analysis of microarray results.

  12. White-spot Lesions and Gingivitis Microbiotas in Orthodontic Patients

    Science.gov (United States)

    Tanner, A.C.R.; Sonis, A.L.; Lif Holgerson, P.; Starr, J.R.; Nunez, Y.; Kressirer, C.A.; Paster, B.J.; Johansson, I.

    2012-01-01

    White-spot lesions (WSL) associated with orthodontic appliances are a cosmetic problem and increase risk for cavities. We characterized the microbiota of WSL, accounting for confounding due to gingivitis. Participants were 60 children with fixed appliances, aged between 10 and 19 yrs, half with WSL. Plaque samples were assayed by a 16S rRNA-based microarray (HOMIM) and by PCR. Mean gingival index was positively associated with WSL (p = 0.018). Taxa associated with WSL by microarray included Granulicatella elegans (p = 0.01), Veillonellaceae sp. HOT 155 (p gingivitis by microarray included: Gemella sanguinis (p = 0.002), Actinomyces sp. HOT 448 (p = 0.003), Prevotella cluster IV (p = 0.021), and Streptococcus sp. HOT 071/070 (p = 0.023); and levels of S. mutans (p = 0.02) and Bifidobacteriaceae (p = 0.012) by qPCR. Species’ associations with WSL were minimally changed with adjustment for gingivitis level. Partial least-squares discriminant analysis yielded good discrimination between children with and those without WSL. Granulicatella, Veillonellaceae and Bifidobacteriaceae, in addition to S. mutans and S. wiggsiae, were associated with the presence of WSL in adolescents undergoing orthodontic treatment. Many taxa showed a stronger association with gingivitis than with WSL. PMID:22837552

  13. The Spotting Distribution of Wildfires

    Directory of Open Access Journals (Sweden)

    Jonathan Martin

    2016-06-01

    Full Text Available In wildfire science, spotting refers to non-local creation of new fires, due to downwind ignition of brands launched from a primary fire. Spotting is often mentioned as being one of the most difficult problems for wildfire management, because of its unpredictable nature. Since spotting is a stochastic process, it makes sense to talk about a probability distribution for spotting, which we call the spotting distribution. Given a location ahead of the fire front, we would like to know how likely is it to observe a spot fire at that location in the next few minutes. The aim of this paper is to introduce a detailed procedure to find the spotting distribution. Most prior modelling has focused on the maximum spotting distance, or on physical subprocesses. We will use mathematical modelling, which is based on detailed physical processes, to derive a spotting distribution. We discuss the use and measurement of this spotting distribution in fire spread, fire management and fire breaching. The appendix of this paper contains a comprehensive review of the relevant underlying physical sub-processes of fire plumes, launching fire brands, wind transport, falling and terminal velocity, combustion during transport, and ignition upon landing.

  14. Emerging hot spot analysis

    DEFF Research Database (Denmark)

    Reinau, Kristian Hegner

    Traditionally, focus in the transport field, both politically and scientifically, has been on private cars and public transport. Freight transport has been a neglected topic. Recent years has seen an increased focus upon congestion as a core issue across Europe, resulting in a great need for know...... speed data for freight. Secondly, the analytical methods used, space-time cubes and emerging hot spot analysis, are also new in the freight transport field. The analysis thus estimates precisely how fast freight moves on the roads in Northern Jutland and how this has evolved over time....

  15. Principles of gene microarray data analysis.

    Science.gov (United States)

    Mocellin, Simone; Rossi, Carlo Riccardo

    2007-01-01

    The development of several gene expression profiling methods, such as comparative genomic hybridization (CGH), differential display, serial analysis of gene expression (SAGE), and gene microarray, together with the sequencing of the human genome, has provided an opportunity to monitor and investigate the complex cascade of molecular events leading to tumor development and progression. The availability of such large amounts of information has shifted the attention of scientists towards a nonreductionist approach to biological phenomena. High throughput technologies can be used to follow changing patterns of gene expression over time. Among them, gene microarray has become prominent because it is easier to use, does not require large-scale DNA sequencing, and allows for the parallel quantification of thousands of genes from multiple samples. Gene microarray technology is rapidly spreading worldwide and has the potential to drastically change the therapeutic approach to patients affected with tumor. Therefore, it is of paramount importance for both researchers and clinicians to know the principles underlying the analysis of the huge amount of data generated with microarray technology.

  16. Detection of selected plant viruses by microarrays

    OpenAIRE

    HRABÁKOVÁ, Lenka

    2013-01-01

    The main aim of this master thesis was the simultaneous detection of four selected plant viruses ? Apple mosaic virus, Plum pox virus, Prunus necrotic ringspot virus and Prune harf virus, by microarrays. The intermediate step in the process of the detection was optimizing of multiplex polymerase chain reaction (PCR).

  17. LNA-modified isothermal oligonucleotide microarray for ...

    Indian Academy of Sciences (India)

    2014-10-20

    Oct 20, 2014 ... the advent of DNA microarray techniques (Lee et al. 2007). ... atoms of ribose to form a bicyclic ribosyl structure. It is the .... 532 nm and emission at 570 nm. The signal ..... sis and validation using real-time PCR. Nucleic Acids ...

  18. Microarrays (DNA Chips) for the Classroom Laboratory

    Science.gov (United States)

    Barnard, Betsy; Sussman, Michael; BonDurant, Sandra Splinter; Nienhuis, James; Krysan, Patrick

    2006-01-01

    We have developed and optimized the necessary laboratory materials to make DNA microarray technology accessible to all high school students at a fraction of both cost and data size. The primary component is a DNA chip/array that students "print" by hand and then analyze using research tools that have been adapted for classroom use. The…

  19. Comparing transformation methods for DNA microarray data

    NARCIS (Netherlands)

    Thygesen, Helene H.; Zwinderman, Aeilko H.

    2004-01-01

    Background: When DNA microarray data are used for gene clustering, genotype/phenotype correlation studies, or tissue classification the signal intensities are usually transformed and normalized in several steps in order to improve comparability and signal/noise ratio. These steps may include

  20. [cDNA library construction from panicle meristem of finger millet].

    Science.gov (United States)

    Radchuk, V; Pirko, Ia V; Isaenkov, S V; Emets, A I; Blium, Ia B

    2014-01-01

    The protocol for production of full-size cDNA using SuperScript Full-Length cDNA Library Construction Kit II (Invitrogen) was tested and high quality cDNA library from meristematic tissue of finger millet panicle (Eleusine coracana (L.) Gaertn) was created. The titer of obtained cDNA library comprised 3.01 x 10(5) CFU/ml in avarage. In average the length of cDNA insertion consisted about 1070 base pairs, the effectivity of cDNA fragment insertions--99.5%. The selective sequencing of cDNA clones from created library was performed. The sequences of cDNA clones were identified with usage of BLAST-search. The results of cDNA library analysis and selective sequencing represents prove good functionality and full length character of inserted cDNA clones. Obtained cDNA library from meristematic tissue of finger millet panicle represents good and valuable source for isolation and identification of key genes regulating metabolism and meristematic development and for mining of new molecular markers to conduct out high quality genetic investigations and molecular breeding as well.

  1. Spotting and validation of a genome wide oligonucleotide chip with duplicate measurement of each gene

    International Nuclear Information System (INIS)

    Thomassen, Mads; Skov, Vibe; Eiriksdottir, Freyja; Tan, Qihua; Jochumsen, Kirsten; Fritzner, Niels; Brusgaard, Klaus; Dahlgaard, Jesper; Kruse, Torben A.

    2006-01-01

    The quality of DNA microarray based gene expression data relies on the reproducibility of several steps in a microarray experiment. We have developed a spotted genome wide microarray chip with oligonucleotides printed in duplicate in order to minimise undesirable biases, thereby optimising detection of true differential expression. The validation study design consisted of an assessment of the microarray chip performance using the MessageAmp and FairPlay labelling kits. Intraclass correlation coefficient (ICC) was used to demonstrate that MessageAmp was significantly more reproducible than FairPlay. Further examinations with MessageAmp revealed the applicability of the system. The linear range of the chips was three orders of magnitude, the precision was high, as 95% of measurements deviated less than 1.24-fold from the expected value, and the coefficient of variation for relative expression was 13.6%. Relative quantitation was more reproducible than absolute quantitation and substantial reduction of variance was attained with duplicate spotting. An analysis of variance (ANOVA) demonstrated no significant day-to-day variation

  2. Development of a high-throughput microfluidic integrated microarray for the detection of chimeric bioweapons.

    Energy Technology Data Exchange (ETDEWEB)

    Sheppod, Timothy; Satterfield, Brent; Hukari, Kyle W.; West, Jason A. A.; Hux, Gary A.

    2006-10-01

    The advancement of DNA cloning has significantly augmented the potential threat of a focused bioweapon assault, such as a terrorist attack. With current DNA cloning techniques, toxin genes from the most dangerous (but environmentally labile) bacterial or viral organism can now be selected and inserted into robust organism to produce an infinite number of deadly chimeric bioweapons. In order to neutralize such a threat, accurate detection of the expressed toxin genes, rather than classification on strain or genealogical decent of these organisms, is critical. The development of a high-throughput microarray approach will enable the detection of unknowns chimeric bioweapons. The development of a high-throughput microarray approach will enable the detection of unknown bioweapons. We have developed a unique microfluidic approach to capture and concentrate these threat genes (mRNA's) upto a 30 fold concentration. These captured oligonucleotides can then be used to synthesize in situ oligonucleotide copies (cDNA probes) of the captured genes. An integrated microfluidic architecture will enable us to control flows of reagents, perform clean-up steps and finally elute nanoliter volumes of synthesized oligonucleotides probes. The integrated approach has enabled a process where chimeric or conventional bioweapons can rapidly be identified based on their toxic function, rather than being restricted to information that may not identify the critical nature of the threat.

  3. Cross-species microarray hybridization to identify developmentally regulated genes in the filamentous fungus Sordaria macrospora.

    Science.gov (United States)

    Nowrousian, Minou; Ringelberg, Carol; Dunlap, Jay C; Loros, Jennifer J; Kück, Ulrich

    2005-04-01

    The filamentous fungus Sordaria macrospora forms complex three-dimensional fruiting bodies that protect the developing ascospores and ensure their proper discharge. Several regulatory genes essential for fruiting body development were previously isolated by complementation of the sterile mutants pro1, pro11 and pro22. To establish the genetic relationships between these genes and to identify downstream targets, we have conducted cross-species microarray hybridizations using cDNA arrays derived from the closely related fungus Neurospora crassa and RNA probes prepared from wild-type S. macrospora and the three developmental mutants. Of the 1,420 genes which gave a signal with the probes from all the strains used, 172 (12%) were regulated differently in at least one of the three mutants compared to the wild type, and 17 (1.2%) were regulated differently in all three mutant strains. Microarray data were verified by Northern analysis or quantitative real time PCR. Among the genes that are up- or down-regulated in the mutant strains are genes encoding the pheromone precursors, enzymes involved in melanin biosynthesis and a lectin-like protein. Analysis of gene expression in double mutants revealed a complex network of interaction between the pro gene products.

  4. arrayCGHbase: an analysis platform for comparative genomic hybridization microarrays

    Directory of Open Access Journals (Sweden)

    Moreau Yves

    2005-05-01

    Full Text Available Abstract Background The availability of the human genome sequence as well as the large number of physically accessible oligonucleotides, cDNA, and BAC clones across the entire genome has triggered and accelerated the use of several platforms for analysis of DNA copy number changes, amongst others microarray comparative genomic hybridization (arrayCGH. One of the challenges inherent to this new technology is the management and analysis of large numbers of data points generated in each individual experiment. Results We have developed arrayCGHbase, a comprehensive analysis platform for arrayCGH experiments consisting of a MIAME (Minimal Information About a Microarray Experiment supportive database using MySQL underlying a data mining web tool, to store, analyze, interpret, compare, and visualize arrayCGH results in a uniform and user-friendly format. Following its flexible design, arrayCGHbase is compatible with all existing and forthcoming arrayCGH platforms. Data can be exported in a multitude of formats, including BED files to map copy number information on the genome using the Ensembl or UCSC genome browser. Conclusion ArrayCGHbase is a web based and platform independent arrayCGH data analysis tool, that allows users to access the analysis suite through the internet or a local intranet after installation on a private server. ArrayCGHbase is available at http://medgen.ugent.be/arrayCGHbase/.

  5. Expression microarray reproducibility is improved by optimising purification steps in RNA amplification and labelling

    Directory of Open Access Journals (Sweden)

    Brenton James D

    2004-01-01

    Full Text Available Abstract Background Expression microarrays have evolved into a powerful tool with great potential for clinical application and therefore reliability of data is essential. RNA amplification is used when the amount of starting material is scarce, as is frequently the case with clinical samples. Purification steps are critical in RNA amplification and labelling protocols, and there is a lack of sufficient data to validate and optimise the process. Results Here the purification steps involved in the protocol for indirect labelling of amplified RNA are evaluated and the experimentally determined best method for each step with respect to yield, purity, size distribution of the transcripts, and dye coupling is used to generate targets tested in replicate hybridisations. DNase treatment of diluted total RNA samples followed by phenol extraction is the optimal way to remove genomic DNA contamination. Purification of double-stranded cDNA is best achieved by phenol extraction followed by isopropanol precipitation at room temperature. Extraction with guanidinium-phenol and Lithium Chloride precipitation are the optimal methods for purification of amplified RNA and labelled aRNA respectively. Conclusion This protocol provides targets that generate highly reproducible microarray data with good representation of transcripts across the size spectrum and a coefficient of repeatability significantly better than that reported previously.

  6. Identifying Fishes through DNA Barcodes and Microarrays.

    Directory of Open Access Journals (Sweden)

    Marc Kochzius

    2010-09-01

    Full Text Available International fish trade reached an import value of 62.8 billion Euro in 2006, of which 44.6% are covered by the European Union. Species identification is a key problem throughout the life cycle of fishes: from eggs and larvae to adults in fisheries research and control, as well as processed fish products in consumer protection.This study aims to evaluate the applicability of the three mitochondrial genes 16S rRNA (16S, cytochrome b (cyt b, and cytochrome oxidase subunit I (COI for the identification of 50 European marine fish species by combining techniques of "DNA barcoding" and microarrays. In a DNA barcoding approach, neighbour Joining (NJ phylogenetic trees of 369 16S, 212 cyt b, and 447 COI sequences indicated that cyt b and COI are suitable for unambiguous identification, whereas 16S failed to discriminate closely related flatfish and gurnard species. In course of probe design for DNA microarray development, each of the markers yielded a high number of potentially species-specific probes in silico, although many of them were rejected based on microarray hybridisation experiments. None of the markers provided probes to discriminate the sibling flatfish and gurnard species. However, since 16S-probes were less negatively influenced by the "position of label" effect and showed the lowest rejection rate and the highest mean signal intensity, 16S is more suitable for DNA microarray probe design than cty b and COI. The large portion of rejected COI-probes after hybridisation experiments (>90% renders the DNA barcoding marker as rather unsuitable for this high-throughput technology.Based on these data, a DNA microarray containing 64 functional oligonucleotide probes for the identification of 30 out of the 50 fish species investigated was developed. It represents the next step towards an automated and easy-to-handle method to identify fish, ichthyoplankton, and fish products.

  7. Facilitating functional annotation of chicken microarray data

    Directory of Open Access Journals (Sweden)

    Gresham Cathy R

    2009-10-01

    Full Text Available Abstract Background Modeling results from chicken microarray studies is challenging for researchers due to little functional annotation associated with these arrays. The Affymetrix GenChip chicken genome array, one of the biggest arrays that serve as a key research tool for the study of chicken functional genomics, is among the few arrays that link gene products to Gene Ontology (GO. However the GO annotation data presented by Affymetrix is incomplete, for example, they do not show references linked to manually annotated functions. In addition, there is no tool that facilitates microarray researchers to directly retrieve functional annotations for their datasets from the annotated arrays. This costs researchers amount of time in searching multiple GO databases for functional information. Results We have improved the breadth of functional annotations of the gene products associated with probesets on the Affymetrix chicken genome array by 45% and the quality of annotation by 14%. We have also identified the most significant diseases and disorders, different types of genes, and known drug targets represented on Affymetrix chicken genome array. To facilitate functional annotation of other arrays and microarray experimental datasets we developed an Array GO Mapper (AGOM tool to help researchers to quickly retrieve corresponding functional information for their dataset. Conclusion Results from this study will directly facilitate annotation of other chicken arrays and microarray experimental datasets. Researchers will be able to quickly model their microarray dataset into more reliable biological functional information by using AGOM tool. The disease, disorders, gene types and drug targets revealed in the study will allow researchers to learn more about how genes function in complex biological systems and may lead to new drug discovery and development of therapies. The GO annotation data generated will be available for public use via AgBase website and

  8. Investigation of cellular signalling responses to non-ionising radiation in melanocytes by microarray analysis

    International Nuclear Information System (INIS)

    Boyle, G.M.; Pedley, J.; Martyn, A.C.; Fraser, L.M.; Banducci, K.J.; Parsons, P.G.; Breit, S.N.

    2003-01-01

    Melanoma is a highly aggressive cancer resulting from the abnormal proliferation and spread of specialised pigment cells in the skin, known as melanocytes. Extensive epidemiological and molecular evidence suggests that a major risk factor for melanoma formation is exposure to non-ionising radiation in the form of solar ultra-violet (UV) light. However, the exact role of solar UV in the development of melanoma is unclear. To elucidate the molecular events that occur in melanocytes following solar UV exposure and determine how they lead to melanoma development, cDNA microarray analysis was used to analyse the gene expression profile of normal melanocytes, melanocytes exposed to simulated solar UV and melanoma cells. The development of cDNA microarray technology has allowed gene expression profiling at the mRNA level to be conducted for many thousands of genes simultaneously by hybridising an array of known sequences with labelled cDNA reverse transcribed form the sample RNA. Gene expression analysis was performed for over 13,000 genes. More than 500 genes were identified as differentially expressed in melanocytes following a single UV exposure, although overall there was a general suppression of transcription. Genes that were up-regulated included oncogenes and cytoskeletal genes; in contrast, genes encoding protein tyrosine kinases and apoptosis effectors were down-regulated. Many of the genes identified as being differentially expressed represent novel UV-regulated targets. Repeated exposure to solar UV resulted in the elevation in expression of a novel member of the transforming growth factor-b (TGF-b) superfamily, the Macrophage Inhibitory Cytokine-1 (MIC-1). Our results have shown that MIC-1 is up-regulated by solar UV in melanocytes, and is highly expressed (>3 fold) in a number of metastatic melanoma cell lines (31/61) in comparison to primary melanocytes. Furthermore functional, dimerised MIC-1 was found to be secreted by melanocytes, and secreted levels were

  9. Factorial microarray analysis of zebra mussel (Dreissena polymorpha: Dreissenidae, Bivalvia adhesion

    Directory of Open Access Journals (Sweden)

    Faisal Mohamed

    2010-05-01

    Full Text Available Abstract Background The zebra mussel (Dreissena polymorpha has been well known for its expertise in attaching to substances under the water. Studies in past decades on this underwater adhesion focused on the adhesive protein isolated from the byssogenesis apparatus of the zebra mussel. However, the mechanism of the initiation, maintenance, and determination of the attachment process remains largely unknown. Results In this study, we used a zebra mussel cDNA microarray previously developed in our lab and a factorial analysis to identify the genes that were involved in response to the changes of four factors: temperature (Factor A, current velocity (Factor B, dissolved oxygen (Factor C, and byssogenesis status (Factor D. Twenty probes in the microarray were found to be modified by one of the factors. The transcription products of four selected genes, DPFP-BG20_A01, EGP-BG97/192_B06, EGP-BG13_G05, and NH-BG17_C09 were unique to the zebra mussel foot based on the results of quantitative reverse transcription PCR (qRT-PCR. The expression profiles of these four genes under the attachment and non-attachment were also confirmed by qRT-PCR and the result is accordant to that from microarray assay. The in situ hybridization with the RNA probes of two identified genes DPFP-BG20_A01 and EGP-BG97/192_B06 indicated that both of them were expressed by a type of exocrine gland cell located in the middle part of the zebra mussel foot. Conclusions The results of this study suggested that the changes of D. polymorpha byssogenesis status and the environmental factors can dramatically affect the expression profiles of the genes unique to the foot. It turns out that the factorial design and analysis of the microarray experiment is a reliable method to identify the influence of multiple factors on the expression profiles of the probesets in the microarray; therein it provides a powerful tool to reveal the mechanism of zebra mussel underwater attachment.

  10. Factorial microarray analysis of zebra mussel (Dreissena polymorpha: Dreissenidae, Bivalvia) adhesion.

    Science.gov (United States)

    Xu, Wei; Faisal, Mohamed

    2010-05-28

    The zebra mussel (Dreissena polymorpha) has been well known for its expertise in attaching to substances under the water. Studies in past decades on this underwater adhesion focused on the adhesive protein isolated from the byssogenesis apparatus of the zebra mussel. However, the mechanism of the initiation, maintenance, and determination of the attachment process remains largely unknown. In this study, we used a zebra mussel cDNA microarray previously developed in our lab and a factorial analysis to identify the genes that were involved in response to the changes of four factors: temperature (Factor A), current velocity (Factor B), dissolved oxygen (Factor C), and byssogenesis status (Factor D). Twenty probes in the microarray were found to be modified by one of the factors. The transcription products of four selected genes, DPFP-BG20_A01, EGP-BG97/192_B06, EGP-BG13_G05, and NH-BG17_C09 were unique to the zebra mussel foot based on the results of quantitative reverse transcription PCR (qRT-PCR). The expression profiles of these four genes under the attachment and non-attachment were also confirmed by qRT-PCR and the result is accordant to that from microarray assay. The in situ hybridization with the RNA probes of two identified genes DPFP-BG20_A01 and EGP-BG97/192_B06 indicated that both of them were expressed by a type of exocrine gland cell located in the middle part of the zebra mussel foot. The results of this study suggested that the changes of D. polymorpha byssogenesis status and the environmental factors can dramatically affect the expression profiles of the genes unique to the foot. It turns out that the factorial design and analysis of the microarray experiment is a reliable method to identify the influence of multiple factors on the expression profiles of the probesets in the microarray; therein it provides a powerful tool to reveal the mechanism of zebra mussel underwater attachment.

  11. Image Processing of Porous Silicon Microarray in Refractive Index Change Detection.

    Science.gov (United States)

    Guo, Zhiqing; Jia, Zhenhong; Yang, Jie; Kasabov, Nikola; Li, Chuanxi

    2017-06-08

    A new method for extracting the dots is proposed by the reflected light image of porous silicon (PSi) microarray utilization in this paper. The method consists of three parts: pretreatment, tilt correction and spot segmentation. First, based on the characteristics of different components in HSV (Hue, Saturation, Value) space, a special pretreatment is proposed for the reflected light image to obtain the contour edges of the array cells in the image. Second, through the geometric relationship of the target object between the initial external rectangle and the minimum bounding rectangle (MBR), a new tilt correction algorithm based on the MBR is proposed to adjust the image. Third, based on the specific requirements of the reflected light image segmentation, the array cells are segmented into dots as large as possible and the distance between the dots is equal in the corrected image. Experimental results show that the pretreatment part of this method can effectively avoid the influence of complex background and complete the binarization processing of the image. The tilt correction algorithm has a shorter computation time, which makes it highly suitable for tilt correction of reflected light images. The segmentation algorithm makes the dots in a regular arrangement, excludes the edges and the bright spots. This method could be utilized in the fast, accurate and automatic dots extraction of the PSi microarray reflected light image.

  12. SiPM as miniaturised optical biosensor for DNA-microarray applications

    Directory of Open Access Journals (Sweden)

    M.F. Santangelo

    2015-12-01

    Full Text Available A miniaturized optical biosensor for low-level fluorescence emitted by DNA strands labelled with CY5 is showed. Aim of this work is to demonstrate that a Si-based photodetector, having a low noise and a high sensitivity, can replace traditional detection systems in DNA-microarray applications. The photodetector used is a photomultiplier (SiPM, with 25 pixels. It exhibits a higher sensitivity than commercial optical readers and we experimentally found a detection limit for spotted dried samples of ∼1 nM. We measured the fluorescence signal in different operating conditions (angle of analysis, fluorophores concentrations, solution volumes and support. Once fixed the angle of analysis, for samples spotted on Al-TEOS slide dried, the system is proportional to the concentration of the analyte in the sample and is linear in the range 1 nM–1 μM. For solutions, the range of linearity ranges from 100 fM to 10 nM. The system potentialities and the device low costs suggest it as basic component for the design and fabrication of a cheap, easy and portable optical system. Keywords: Optical Biosensor, SiPM, DNA microarray, Fluorophore detection

  13. Cellular neural networks, the Navier-Stokes equation, and microarray image reconstruction.

    Science.gov (United States)

    Zineddin, Bachar; Wang, Zidong; Liu, Xiaohui

    2011-11-01

    Although the last decade has witnessed a great deal of improvements achieved for the microarray technology, many major developments in all the main stages of this technology, including image processing, are still needed. Some hardware implementations of microarray image processing have been proposed in the literature and proved to be promising alternatives to the currently available software systems. However, the main drawback of those proposed approaches is the unsuitable addressing of the quantification of the gene spot in a realistic way without any assumption about the image surface. Our aim in this paper is to present a new image-reconstruction algorithm using the cellular neural network that solves the Navier-Stokes equation. This algorithm offers a robust method for estimating the background signal within the gene-spot region. The MATCNN toolbox for Matlab is used to test the proposed method. Quantitative comparisons are carried out, i.e., in terms of objective criteria, between our approach and some other available methods. It is shown that the proposed algorithm gives highly accurate and realistic measurements in a fully automated manner within a remarkably efficient time.

  14. Image Processing of Porous Silicon Microarray in Refractive Index Change Detection

    Directory of Open Access Journals (Sweden)

    Zhiqing Guo

    2017-06-01

    Full Text Available A new method for extracting the dots is proposed by the reflected light image of porous silicon (PSi microarray utilization in this paper. The method consists of three parts: pretreatment, tilt correction and spot segmentation. First, based on the characteristics of different components in HSV (Hue, Saturation, Value space, a special pretreatment is proposed for the reflected light image to obtain the contour edges of the array cells in the image. Second, through the geometric relationship of the target object between the initial external rectangle and the minimum bounding rectangle (MBR, a new tilt correction algorithm based on the MBR is proposed to adjust the image. Third, based on the specific requirements of the reflected light image segmentation, the array cells are segmented into dots as large as possible and the distance between the dots is equal in the corrected image. Experimental results show that the pretreatment part of this method can effectively avoid the influence of complex background and complete the binarization processing of the image. The tilt correction algorithm has a shorter computation time, which makes it highly suitable for tilt correction of reflected light images. The segmentation algorithm makes the dots in a regular arrangement, excludes the edges and the bright spots. This method could be utilized in the fast, accurate and automatic dots extraction of the PSi microarray reflected light image.

  15. Cloning and analysis of the mouse Fanconi anemia group a cDNA and an overlapping penta zinc finger cDNA

    NARCIS (Netherlands)

    Wong, JCY; Alon, N; Norga, K; Kruyt, FAE; Youssoufian, H; Buchwald, M

    2000-01-01

    Despite the cloning of four disease-associated genes for Fanconi anemia (FA), the molecular pathogenesis of FA remains largely unknown. To study FA complementation group A using the mouse as a mode I system, we cloned and characterized the mouse homolog of the human FANCA cDNA, The mouse cDNA

  16. Systematic validation and atomic force microscopy of non-covalent short oligonucleotide barcode microarrays.

    Directory of Open Access Journals (Sweden)

    Michael A Cook

    Full Text Available BACKGROUND: Molecular barcode arrays provide a powerful means to analyze cellular phenotypes in parallel through detection of short (20-60 base unique sequence tags, or "barcodes", associated with each strain or clone in a collection. However, costs of current methods for microarray construction, whether by in situ oligonucleotide synthesis or ex situ coupling of modified oligonucleotides to the slide surface are often prohibitive to large-scale analyses. METHODOLOGY/PRINCIPAL FINDINGS: Here we demonstrate that unmodified 20mer oligonucleotide probes printed on conventional surfaces show comparable hybridization signals to covalently linked 5'-amino-modified probes. As a test case, we undertook systematic cell size analysis of the budding yeast Saccharomyces cerevisiae genome-wide deletion collection by size separation of the deletion pool followed by determination of strain abundance in size fractions by barcode arrays. We demonstrate that the properties of a 13K unique feature spotted 20 mer oligonucleotide barcode microarray compare favorably with an analogous covalently-linked oligonucleotide array. Further, cell size profiles obtained with the size selection/barcode array approach recapitulate previous cell size measurements of individual deletion strains. Finally, through atomic force microscopy (AFM, we characterize the mechanism of hybridization to unmodified barcode probes on the slide surface. CONCLUSIONS/SIGNIFICANCE: These studies push the lower limit of probe size in genome-scale unmodified oligonucleotide microarray construction and demonstrate a versatile, cost-effective and reliable method for molecular barcode analysis.

  17. Preparation of a differentially expressed, full-length cDNA expression library by RecA-mediated triple-strand formation with subtractively enriched cDNA fragments

    NARCIS (Netherlands)

    Hakvoort, T. B.; Spijkers, J. A.; Vermeulen, J. L.; Lamers, W. H.

    1996-01-01

    We have developed a fast and general method to obtain an enriched, full-length cDNA expression library with subtractively enriched cDNA fragments. The procedure relies on RecA-mediated triple-helix formation of single-stranded cDNA fragments with a double-stranded cDNA plasmid library. The complexes

  18. Generation and analysis of large-scale expressed sequence tags (ESTs from a full-length enriched cDNA library of porcine backfat tissue

    Directory of Open Access Journals (Sweden)

    Lee Hae-Young

    2006-02-01

    Full Text Available Abstract Background Genome research in farm animals will expand our basic knowledge of the genetic control of complex traits, and the results will be applied in the livestock industry to improve meat quality and productivity, as well as to reduce the incidence of disease. A combination of quantitative trait locus mapping and microarray analysis is a useful approach to reduce the overall effort needed to identify genes associated with quantitative traits of interest. Results We constructed a full-length enriched cDNA library from porcine backfat tissue. The estimated average size of the cDNA inserts was 1.7 kb, and the cDNA fullness ratio was 70%. In total, we deposited 16,110 high-quality sequences in the dbEST division of GenBank (accession numbers: DT319652-DT335761. For all the expressed sequence tags (ESTs, approximately 10.9 Mb of porcine sequence were generated with an average length of 674 bp per EST (range: 200–952 bp. Clustering and assembly of these ESTs resulted in a total of 5,008 unique sequences with 1,776 contigs (35.46% and 3,232 singleton (65.54% ESTs. From a total of 5,008 unique sequences, 3,154 (62.98% were similar to other sequences, and 1,854 (37.02% were identified as having no hit or low identity (Sus scrofa. Gene ontology (GO annotation of unique sequences showed that approximately 31.7, 32.3, and 30.8% were assigned molecular function, biological process, and cellular component GO terms, respectively. A total of 1,854 putative novel transcripts resulted after comparison and filtering with the TIGR SsGI; these included a large percentage of singletons (80.64% and a small proportion of contigs (13.36%. Conclusion The sequence data generated in this study will provide valuable information for studying expression profiles using EST-based microarrays and assist in the condensation of current pig TCs into clusters representing longer stretches of cDNA sequences. The isolation of genes expressed in backfat tissue is the

  19. The Immunome of Colon Cancer: Functional In Silico Analysis of Antigenic Proteins Deduced from IgG Microarray Profiling

    Directory of Open Access Journals (Sweden)

    Johana A. Luna Coronell

    2018-02-01

    Full Text Available Characterization of the colon cancer immunome and its autoantibody signature from differentially-reactive antigens (DIRAGs could provide insights into aberrant cellular mechanisms or enriched networks associated with diseases. The purpose of this study was to characterize the antibody profile of plasma samples from 32 colorectal cancer (CRC patients and 32 controls using proteins isolated from 15,417 human cDNA expression clones on microarrays. 671 unique DIRAGs were identified and 632 were more highly reactive in CRC samples. Bioinformatics analyses reveal that compared to control samples, the immunoproteomic IgG profiling of CRC samples is mainly associated with cell death, survival, and proliferation pathways, especially proteins involved in EIF2 and mTOR signaling. Ribosomal proteins (e.g., RPL7, RPL22, and RPL27A and CRC-related genes such as APC, AXIN1, E2F4, MSH2, PMS2, and TP53 were highly enriched. In addition, differential pathways were observed between the CRC and control samples. Furthermore, 103 DIRAGs were reported in the SEREX antigen database, demonstrating our ability to identify known and new reactive antigens. We also found an overlap of 7 antigens with 48 “CRC genes.” These data indicate that immunomics profiling on protein microarrays is able to reveal the complexity of immune responses in cancerous diseases and faithfully reflects the underlying pathology. Keywords: Autoantibody tumor biomarker, Cancer immunology, Colorectal cancer, Immunomics, Protein microarray

  20. Models of spots and flares

    International Nuclear Information System (INIS)

    Mullan, D.J.

    1983-01-01

    Laboratory experiments in recent years have shown that there are many more ways to drive a plasma out of equilibrium than to preserve equilibrium. In that sense, it is perhaps easier to understand why flares should occur in a stellar atmosphere than why a long-lived feature such as a dark spot should persist. The author summarizes work on the equilibrium structure of cool spots in the sun and stars. Since spots involve complex interactions between convective flows and magnetic fields, he needs to refer to observations for help in identifying the dominant processes which should enter into the modelling. His summary therefore begins by discussing certain relevant properties of spots in the solar atmosphere. The next sections deal with the magnetic fields in spots, the stability of spots, spot cooling and missing flux. The author concludes that spots should be viewed not simply as cool areas, but rather as engines which do the work of converting the energy of convective flows into flare-compatible form. (Auth.)

  1. Black-spot poison ivy.

    Science.gov (United States)

    Schram, Sarah E; Willey, Andrea; Lee, Peter K; Bohjanen, Kimberly A; Warshaw, Erin M

    2008-01-01

    In black-spot poison ivy dermatitis, a black lacquerlike substance forms on the skin when poison ivy resin is exposed to air. Although the Toxicodendron group of plants is estimated to be the most common cause of allergic contact dermatitis in the United States, black-spot poison ivy dermatitis is relatively rare.

  2. Rocky Mountain spotted fever, Colombia.

    Science.gov (United States)

    Hidalgo, Marylin; Orejuela, Leonora; Fuya, Patricia; Carrillo, Pilar; Hernandez, Jorge; Parra, Edgar; Keng, Colette; Small, Melissa; Olano, Juan P; Bouyer, Donald; Castaneda, Elizabeth; Walker, David; Valbuena, Gustavo

    2007-07-01

    We investigated 2 fatal cases of Rocky Mountain spotted fever that occurred in 2003 and 2004 near the same locality in Colombia where the disease was first reported in the 1930s. A retrospective serosurvey of febrile patients showed that > 21% of the serum samples had antibodies aaainst spotted fever group rickettsiae.

  3. Advances in spot curing technology

    International Nuclear Information System (INIS)

    Burga, R.

    1999-01-01

    A brief review of spot curing technology was presented. The process which a spot of energy of a specific wavelength bandwidth and irradiance is used to cause a coating, encapsulant or adhesive to change from a liquid to a solid state

  4. DNA Microarrays in Comparative Genomics and Transcriptomics

    DEFF Research Database (Denmark)

    Willenbrock, Hanni

    2007-01-01

    at identifying the exact breakpoints where DNA has been gained or lost. In this thesis, three popular methods are compared and a realistic simulation model is presented for generating artificial data with known breakpoints and known DNA copy number. By using simulated data, we obtain a realistic evaluation......During the past few years, innovations in the DNA sequencing technology has led to an explosion in available DNA sequence information. This has revolutionized biological research and promoted the development of high throughput analysis methods that can take advantage of the vast amount of sequence...... data. For this, the DNA microarray technology has gained enormous popularity due to its ability to measure the presence or the activity of thousands of genes simultaneously. Microarrays for high throughput data analyses are not limited to a few organisms but may be applied to everything from bacteria...

  5. Immobilization Techniques for Microarray: Challenges and Applications

    Directory of Open Access Journals (Sweden)

    Satish Balasaheb Nimse

    2014-11-01

    Full Text Available The highly programmable positioning of molecules (biomolecules, nanoparticles, nanobeads, nanocomposites materials on surfaces has potential applications in the fields of biosensors, biomolecular electronics, and nanodevices. However, the conventional techniques including self-assembled monolayers fail to position the molecules on the nanometer scale to produce highly organized monolayers on the surface. The present article elaborates different techniques for the immobilization of the biomolecules on the surface to produce microarrays and their diagnostic applications. The advantages and the drawbacks of various methods are compared. This article also sheds light on the applications of the different technologies for the detection and discrimination of viral/bacterial genotypes and the detection of the biomarkers. A brief survey with 115 references covering the last 10 years on the biological applications of microarrays in various fields is also provided.

  6. Mining meiosis and gametogenesis with DNA microarrays.

    Science.gov (United States)

    Schlecht, Ulrich; Primig, Michael

    2003-04-01

    Gametogenesis is a key developmental process that involves complex transcriptional regulation of numerous genes including many that are conserved between unicellular eukaryotes and mammals. Recent expression-profiling experiments using microarrays have provided insight into the co-ordinated transcription of several hundred genes during mitotic growth and meiotic development in budding and fission yeast. Furthermore, microarray-based studies have identified numerous loci that are regulated during the cell cycle or expressed in a germ-cell specific manner in eukaryotic model systems like Caenorhabditis elegans, Mus musculus as well as Homo sapiens. The unprecedented amount of information produced by post-genome biology has spawned novel approaches to organizing biological knowledge using currently available information technology. This review outlines experiments that contribute to an emerging comprehensive picture of the molecular machinery governing sexual reproduction in eukaryotes.

  7. Facilitating RNA structure prediction with microarrays.

    Science.gov (United States)

    Kierzek, Elzbieta; Kierzek, Ryszard; Turner, Douglas H; Catrina, Irina E

    2006-01-17

    Determining RNA secondary structure is important for understanding structure-function relationships and identifying potential drug targets. This paper reports the use of microarrays with heptamer 2'-O-methyl oligoribonucleotides to probe the secondary structure of an RNA and thereby improve the prediction of that secondary structure. When experimental constraints from hybridization results are added to a free-energy minimization algorithm, the prediction of the secondary structure of Escherichia coli 5S rRNA improves from 27 to 92% of the known canonical base pairs. Optimization of buffer conditions for hybridization and application of 2'-O-methyl-2-thiouridine to enhance binding and improve discrimination between AU and GU pairs are also described. The results suggest that probing RNA with oligonucleotide microarrays can facilitate determination of secondary structure.

  8. cDNA sequences of two apolipoproteins from lamprey

    International Nuclear Information System (INIS)

    Pontes, M.; Xu, X.; Graham, D.; Riley, M.; Doolittle, R.F.

    1987-01-01

    The messages for two small but abundant apolipoproteins found in lamprey blood plasma were cloned with the aid of oligonucleotide probes based on amino-terminal sequences. In both cases, numerous clones were identified in a lamprey liver cDNA library, consistent with the great abundance of these proteins in lamprey blood. One of the cDNAs (LAL1) has a coding region of 105 amino acids that corresponds to a 21-residue signal peptide, a putative 8-residue propeptide, and the 76-residue mature protein found in blood. The other cDNA (LAL2) codes for a total of 191 residues, the first 23 of which constitute a signal peptide. The two proteins, which occur in the high-density lipoprotein fraction of ultracentrifuged plasma, have amino acid compositions similar to those of apolipoproteins found in mammalian blood; computer analysis indicates that the sequences are largely helix-permissive. When the sequences were searched against an amino acid sequence data base, rat apolipoprotein IV was the best matching candidate in both cases. Although a reasonable alignment can be made with that sequence and LAL1, definitive assignment of the two lamprey proteins to typical mammalian classes cannot be made at this point

  9. Infectious Maize rayado fino virus from Cloned cDNA.

    Science.gov (United States)

    Edwards, Michael C; Weiland, John J; Todd, Jane; Stewart, Lucy R

    2015-06-01

    A full-length cDNA clone was produced from a U.S. isolate of Maize rayado fino virus (MRFV), the type member of the genus Marafivirus within the family Tymoviridae. Infectivity of transcripts derived from cDNA clones was demonstrated by infection of maize plants and protoplasts, as well as by transmission via the known leafhopper vectors Dalbulus maidis and Graminella nigrifrons that transmit the virus in a persistent-propagative manner. Infection of maize plants through vascular puncture inoculation of seed with transcript RNA resulted in the induction of fine stipple stripe symptoms typical of those produced by wild-type MRFV and a frequency of infection comparable with that of the wild type. Northern and Western blotting confirmed the production of MRFV-specific RNAs and proteins in infected plants and protoplasts. An unanticipated increase in subgenomic RNA synthesis over levels in infected plants was observed in protoplasts infected with either wild-type or cloned virus. A conserved cleavage site motif previously demonstrated to function in both Oat blue dwarf virus capsid protein and tymoviral nonstructural protein processing was identified near the amino terminus of the MRFV replicase polyprotein, suggesting that cleavage at this site also may occur.

  10. Tissue Microarray Analysis Applied to Bone Diagenesis

    OpenAIRE

    Barrios Mello, Rafael; Regis Silva, Maria Regina; Seixas Alves, Maria Teresa; Evison, Martin; Guimarães, Marco Aurélio; Francisco, Rafaella Arrabaça; Dias Astolphi, Rafael; Miazato Iwamura, Edna Sadayo

    2017-01-01

    Taphonomic processes affecting bone post mortem are important in forensic, archaeological and palaeontological investigations. In this study, the application of tissue microarray (TMA) analysis to a sample of femoral bone specimens from 20 exhumed individuals of known period of burial and age at death is described. TMA allows multiplexing of subsamples, permitting standardized comparative analysis of adjacent sections in 3-D and of representative cross-sections of a large number of specimens....

  11. Deep learning for tissue microarray image-based outcome prediction in patients with colorectal cancer

    Science.gov (United States)

    Bychkov, Dmitrii; Turkki, Riku; Haglund, Caj; Linder, Nina; Lundin, Johan

    2016-03-01

    Recent advances in computer vision enable increasingly accurate automated pattern classification. In the current study we evaluate whether a convolutional neural network (CNN) can be trained to predict disease outcome in patients with colorectal cancer based on images of tumor tissue microarray samples. We compare the prognostic accuracy of CNN features extracted from the whole, unsegmented tissue microarray spot image, with that of CNN features extracted from the epithelial and non-epithelial compartments, respectively. The prognostic accuracy of visually assessed histologic grade is used as a reference. The image data set consists of digitized hematoxylin-eosin (H and E) stained tissue microarray samples obtained from 180 patients with colorectal cancer. The patient samples represent a variety of histological grades, have data available on a series of clinicopathological variables including long-term outcome and ground truth annotations performed by experts. The CNN features extracted from images of the epithelial tissue compartment significantly predicted outcome (hazard ratio (HR) 2.08; CI95% 1.04-4.16; area under the curve (AUC) 0.66) in a test set of 60 patients, as compared to the CNN features extracted from unsegmented images (HR 1.67; CI95% 0.84-3.31, AUC 0.57) and visually assessed histologic grade (HR 1.96; CI95% 0.99-3.88, AUC 0.61). As a conclusion, a deep-learning classifier can be trained to predict outcome of colorectal cancer based on images of H and E stained tissue microarray samples and the CNN features extracted from the epithelial compartment only resulted in a prognostic discrimination comparable to that of visually determined histologic grade.

  12. Production of DNA microarray and expression analysis of genes from Xylella fastidiosa in different culture media

    Directory of Open Access Journals (Sweden)

    Regiane de Fátima Travensolo

    2009-06-01

    Full Text Available DNA Microarray was developed to monitor the expression of many genes from Xylella fastidiosa, allowing the side by-side comparison of two situations in a single experiment. The experiments were performed using X. fastidiosa cells grown in two culture media: BCYE and XDM2. The primers were synthesized, spotted onto glass slides and the array was hybridized against fluorescently labeled cDNAs. The emitted signals were quantified, normalized and the data were statistically analyzed to verify the differentially expressed genes. According to the data, 104 genes were differentially expressed in XDM2 and 30 genes in BCYE media. The present study showed that DNA microarray technique efficiently differentiate the expressed genes under different conditions.DNA Microarray foi desenvolvida para monitorar a expressão de muitos genes de Xylella fastidiosa, permitindo a comparação de duas situações distintas em um único experimento. Os experimentos foram feitos utilizando células de X. fastidiosa cultivada em dois meios de cultura: BCYE e XDM2. Pares de oligonucleotídeos iniciadores foram sintetizados, depositados em lâminas de vidro e o arranjo foi hibridizado contra cDNAs marcados fluorescentemente. Os sinais emitidos foram quantificados, normalizados e os dados foram estatisticamente analisados para verificar os genes diferencialmente expressos. De acordo com nossos dados, 104 genes foram diferencialmente expressos para o meio de cultura XDM2 e 30 genes para o BCYE. No presente estudo, nós demonstramos que a técnica de DNA microarrays eficientemente diferencia genes expressos sob diferentes condições de cultivo.

  13. Can subtle changes in gene expression be consistently detected with different microarray platforms?

    Directory of Open Access Journals (Sweden)

    Kuiper Rowan

    2008-03-01

    Full Text Available Abstract Background The comparability of gene expression data generated with different microarray platforms is still a matter of concern. Here we address the performance and the overlap in the detection of differentially expressed genes for five different microarray platforms in a challenging biological context where differences in gene expression are few and subtle. Results Gene expression profiles in the hippocampus of five wild-type and five transgenic δC-doublecortin-like kinase mice were evaluated with five microarray platforms: Applied Biosystems, Affymetrix, Agilent, Illumina, LGTC home-spotted arrays. Using a fixed false discovery rate of 10% we detected surprising differences between the number of differentially expressed genes per platform. Four genes were selected by ABI, 130 by Affymetrix, 3,051 by Agilent, 54 by Illumina, and 13 by LGTC. Two genes were found significantly differentially expressed by all platforms and the four genes identified by the ABI platform were found by at least three other platforms. Quantitative RT-PCR analysis confirmed 20 out of 28 of the genes detected by two or more platforms and 8 out of 15 of the genes detected by Agilent only. We observed improved correlations between platforms when ranking the genes based on the significance level than with a fixed statistical cut-off. We demonstrate significant overlap in the affected gene sets identified by the different platforms, although biological processes were represented by only partially overlapping sets of genes. Aberrances in GABA-ergic signalling in the transgenic mice were consistently found by all platforms. Conclusion The different microarray platforms give partially complementary views on biological processes affected. Our data indicate that when analyzing samples with only subtle differences in gene expression the use of two different platforms might be more attractive than increasing the number of replicates. Commercial two-color platforms seem to

  14. Geiger mode avalanche photodiodes for microarray systems

    Science.gov (United States)

    Phelan, Don; Jackson, Carl; Redfern, R. Michael; Morrison, Alan P.; Mathewson, Alan

    2002-06-01

    New Geiger Mode Avalanche Photodiodes (GM-APD) have been designed and characterized specifically for use in microarray systems. Critical parameters such as excess reverse bias voltage, hold-off time and optimum operating temperature have been experimentally determined for these photon-counting devices. The photon detection probability, dark count rate and afterpulsing probability have been measured under different operating conditions. An active- quench circuit (AQC) is presented for operating these GM- APDs. This circuit is relatively simple, robust and has such benefits as reducing average power dissipation and afterpulsing. Arrays of these GM-APDs have already been designed and together with AQCs open up the possibility of having a solid-state microarray detector that enables parallel analysis on a single chip. Another advantage of these GM-APDs over current technology is their low voltage CMOS compatibility which could allow for the fabrication of an AQC on the same device. Small are detectors have already been employed in the time-resolved detection of fluorescence from labeled proteins. It is envisaged that operating these new GM-APDs with this active-quench circuit will have numerous applications for the detection of fluorescence in microarray systems.

  15. Classification across gene expression microarray studies

    Directory of Open Access Journals (Sweden)

    Kuner Ruprecht

    2009-12-01

    Full Text Available Abstract Background The increasing number of gene expression microarray studies represents an important resource in biomedical research. As a result, gene expression based diagnosis has entered clinical practice for patient stratification in breast cancer. However, the integration and combined analysis of microarray studies remains still a challenge. We assessed the potential benefit of data integration on the classification accuracy and systematically evaluated the generalization performance of selected methods on four breast cancer studies comprising almost 1000 independent samples. To this end, we introduced an evaluation framework which aims to establish good statistical practice and a graphical way to monitor differences. The classification goal was to correctly predict estrogen receptor status (negative/positive and histological grade (low/high of each tumor sample in an independent study which was not used for the training. For the classification we chose support vector machines (SVM, predictive analysis of microarrays (PAM, random forest (RF and k-top scoring pairs (kTSP. Guided by considerations relevant for classification across studies we developed a generalization of kTSP which we evaluated in addition. Our derived version (DV aims to improve the robustness of the intrinsic invariance of kTSP with respect to technologies and preprocessing. Results For each individual study the generalization error was benchmarked via complete cross-validation and was found to be similar for all classification methods. The misclassification rates were substantially higher in classification across studies, when each single study was used as an independent test set while all remaining studies were combined for the training of the classifier. However, with increasing number of independent microarray studies used in the training, the overall classification performance improved. DV performed better than the average and showed slightly less variance. In

  16. Spotting psychopaths using technology.

    Science.gov (United States)

    Hulbert, Sarah; Adeli, Hojjat

    2015-01-01

    For the past three and a half decades, the Psychopathy Checklist-Revised (PCL-R) and the self-report Psychopathic Personality Inventory-Revised (PPI-R) have been the standard measures for the diagnosis of psychopathy. Technological approaches can enhance these diagnostic methodologies. The purpose of this paper is to present a state-of-the-art review of various technological approaches for spotting psychopathy, such as electroencephalogram (EEG), magnetic resonance imaging (MRI), functional MRI (fMRI), transcranial magnetic stimulation (TMS), and other measures. Results of EEG event-related potential (ERP) experiments support the theory that impaired amygdala function may be responsible for abnormal fear processing in psychopathy, which can ultimately manifest as psychopathic traits, as outlined by the PCL-R or PPI-R. Imaging studies, in general, point to reduced fear processing capabilities in psychopathic individuals. While the human element, introduced through researcher/participant interactions, can be argued as unequivocally necessary for diagnosis, these purely objective technological approaches have proven to be useful in conjunction with the subjective interviewing and questionnaire methods for differentiating psychopaths from non-psychopaths. Furthermore, these technologies are more robust than behavioral measures, which have been shown to fail.

  17. A rapid method for screening arrayed plasmid cDNA library by PCR

    International Nuclear Information System (INIS)

    Hu Yingchun; Zhang Kaitai; Wu Dechang; Li Gang; Xiang Xiaoqiong

    1999-01-01

    Objective: To develop a PCR-based method for rapid and effective screening of arrayed plasmid cDNA library. Methods: The plasmid cDNA library was arrayed and screened by PCR with a particular set of primers. Results: Four positive clones were obtained through about one week. Conclusion: This method can be applied to screening not only normal cDNA clones, but also cDNA clones-containing small size fragments. This method offers significant advantages over traditional screening method in terms of sensitivity, specificity and efficiency

  18. Functional cloning using pFB retroviral cDNA expression libraries.

    Science.gov (United States)

    Felts, Katherine A; Chen, Keith; Zaharee, Kim; Sundar, Latha; Limjoco, Jamie; Miller, Anna; Vaillancourt, Peter

    2002-09-01

    Retroviral cDNA expression libraries allow the efficient introduction of complex cDNA libraries into virtually any mitotic cell type for screening based on gene function. The cDNA copy number per cell can be easily controlled by adjusting the multiplicity of infection, thus cell populations may be generated in which >90% of infected cells contain one to three cDNAs. We describe the isolation of two known oncogenes and one cell-surface receptor from a human Burkitt's lymphoma (Daudi) cDNA library inserted into the high-titer retroviral vector pFB.

  19. Characterisation and correction of signal fluctuations in successive acquisitions of microarray images

    Directory of Open Access Journals (Sweden)

    François Nicolas

    2009-03-01

    Full Text Available Abstract Background There are many sources of variation in dual labelled microarray experiments, including data acquisition and image processing. The final interpretation of experiments strongly relies on the accuracy of the measurement of the signal intensity. For low intensity spots in particular, accurately estimating gene expression variations remains a challenge as signal measurement is, in this case, highly subject to fluctuations. Results To evaluate the fluctuations in the fluorescence intensities of spots, we used series of successive scans, at the same settings, of whole genome arrays. We measured the decrease in fluorescence and we evaluated the influence of different parameters (PMT gain, resolution and chemistry of the slide on the signal variability, at the level of the array as a whole and by intensity interval. Moreover, we assessed the effect of averaging scans on the fluctuations. We found that the extent of photo-bleaching was low and we established that 1 the fluorescence fluctuation is linked to the resolution e.g. it depends on the number of pixels in the spot 2 the fluorescence fluctuation increases as the scanner voltage increases and, moreover, is higher for the red as opposed to the green fluorescence which can introduce bias in the analysis 3 the signal variability is linked to the intensity level, it is higher for low intensities 4 the heterogeneity of the spots and the variability of the signal and the intensity ratios decrease when two or three scans are averaged. Conclusion Protocols consisting of two scans, one at low and one at high PMT gains, or multiple scans (ten scans can introduce bias or be difficult to implement. We found that averaging two, or at most three, acquisitions of microarrays scanned at moderate photomultiplier settings (PMT gain is sufficient to significantly improve the accuracy (quality of the data and particularly those for spots having low intensities and we propose this as a general

  20. Spot Welding of Honeycomb Structures

    Science.gov (United States)

    Cohal, V.

    2017-08-01

    Honeycomb structures are used to prepare meals water jet cutting machines for textile. These honeycomb structures are made of stainless steel sheet thickness of 0.1-0.2 mm. Corrugated sheet metal strips are between two gears with special tooth profile. Hexagonal cells for obtaining these strips are welded points between them. Spot welding device is three electrodes in the upper part, which carries three welding points across the width of the strip of corrugated sheet metal. Spot welding device filled with press and advance mechanisms. The paper presents the values of the regime for spot welding.

  1. Evaluation of toxicity of the mycotoxin citrinin using yeast ORF DNA microarray and Oligo DNA microarray

    Directory of Open Access Journals (Sweden)

    Nobumasa Hitoshi

    2007-04-01

    Full Text Available Abstract Background Mycotoxins are fungal secondary metabolites commonly present in feed and food, and are widely regarded as hazardous contaminants. Citrinin, one of the very well known mycotoxins that was first isolated from Penicillium citrinum, is produced by more than 10 kinds of fungi, and is possibly spread all over the world. However, the information on the action mechanism of the toxin is limited. Thus, we investigated the citrinin-induced genomic response for evaluating its toxicity. Results Citrinin inhibited growth of yeast cells at a concentration higher than 100 ppm. We monitored the citrinin-induced mRNA expression profiles in yeast using the ORF DNA microarray and Oligo DNA microarray, and the expression profiles were compared with those of the other stress-inducing agents. Results obtained from both microarray experiments clustered together, but were different from those of the mycotoxin patulin. The oxidative stress response genes – AADs, FLR1, OYE3, GRE2, and MET17 – were significantly induced. In the functional category, expression of genes involved in "metabolism", "cell rescue, defense and virulence", and "energy" were significantly activated. In the category of "metabolism", genes involved in the glutathione synthesis pathway were activated, and in the category of "cell rescue, defense and virulence", the ABC transporter genes were induced. To alleviate the induced stress, these cells might pump out the citrinin after modification with glutathione. While, the citrinin treatment did not induce the genes involved in the DNA repair. Conclusion Results from both microarray studies suggest that citrinin treatment induced oxidative stress in yeast cells. The genotoxicity was less severe than the patulin, suggesting that citrinin is less toxic than patulin. The reproducibility of the expression profiles was much better with the Oligo DNA microarray. However, the Oligo DNA microarray did not completely overcome cross

  2. Multiplex Detection and Genotyping of Point Mutations Involved in Charcot-Marie-Tooth Disease Using a Hairpin Microarray-Based Assay

    Directory of Open Access Journals (Sweden)

    Yasser Baaj

    2009-01-01

    Full Text Available We previously developed a highly specific method for detecting SNPs with a microarray-based system using stem-loop probes. In this paper we demonstrate that coupling a multiplexing procedure with our microarray method is possible for the simultaneous detection and genotyping of four point mutations, in three different genes, involved in Charcot-Marie-Tooth disease. DNA from healthy individuals and patients was amplified, labeled with Cy3 by multiplex PCR; and hybridized to microarrays. Spot signal intensities were 18 to 74 times greater for perfect matches than for mismatched target sequences differing by a single nucleotide (discrimination ratio for “homozygous” DNA from healthy individuals. “Heterozygous” mutant DNA samples gave signal intensity ratios close to 1 at the positions of the mutations as expected. Genotyping by this method was therefore reliable. This system now combines the principle of highly specific genotyping based on stem-loop structure probes with the advantages of multiplex analysis.

  3. A label-free, fluorescence based assay for microarray

    Science.gov (United States)

    Niu, Sanjun

    DNA chip technology has drawn tremendous attention since it emerged in the mid 90's as a method that expedites gene sequencing by over 100-fold. DNA chip, also called DNA microarray, is a combinatorial technology in which different single-stranded DNA (ssDNA) molecules of known sequences are immobilized at specific spots. The immobilized ssDNA strands are called probes. In application, the chip is exposed to a solution containing ssDNA of unknown sequence, called targets, which are labeled with fluorescent dyes. Due to specific molecular recognition among the base pairs in the DNA, the binding or hybridization occurs only when the probe and target sequences are complementary. The nucleotide sequence of the target is determined by imaging the fluorescence from the spots. The uncertainty of background in signal detection and statistical error in data analysis, primarily due to the error in the DNA amplification process and statistical distribution of the tags in the target DNA, have become the fundamental barriers in bringing the technology into application for clinical diagnostics. Furthermore, the dye and tagging process are expensive, making the cost of DNA chips inhibitive for clinical testing. These limitations and challenges make it difficult to implement DNA chip methods as a diagnostic tool in a pathology laboratory. The objective of this dissertation research is to provide an alternative approach that will address the above challenges. In this research, a label-free assay is designed and studied. Polystyrene (PS), a commonly used polymeric material, serves as the fluorescence agent. Probe ssDNA is covalently immobilized on polystyrene thin film that is supported by a reflecting substrate. When this chip is exposed to excitation light, fluorescence light intensity from PS is detected as the signal. Since the optical constants and conformations of ssDNA and dsDNA (double stranded DNA) are different, the measured fluorescence from PS changes for the same

  4. High-Throughput Quantification of SH2 Domain-Phosphopeptide Interactions with Cellulose-Peptide Conjugate Microarrays.

    Science.gov (United States)

    Engelmann, Brett W

    2017-01-01

    The Src Homology 2 (SH2) domain family primarily recognizes phosphorylated tyrosine (pY) containing peptide motifs. The relative affinity preferences among competing SH2 domains for phosphopeptide ligands define "specificity space," and underpins many functional pY mediated interactions within signaling networks. The degree of promiscuity exhibited and the dynamic range of affinities supported by individual domains or phosphopeptides is best resolved by a carefully executed and controlled quantitative high-throughput experiment. Here, I describe the fabrication and application of a cellulose-peptide conjugate microarray (CPCMA) platform to the quantitative analysis of SH2 domain specificity space. Included herein are instructions for optimal experimental design with special attention paid to common sources of systematic error, phosphopeptide SPOT synthesis, microarray fabrication, analyte titrations, data capture, and analysis.

  5. Isolation of Microarray-Grade Total RNA, MicroRNA, and DNA from a Single PAXgene Blood RNA Tube

    DEFF Research Database (Denmark)

    Kruhøffer, Mogens; Andersen, Lars Dyrskjøt; Voss, Thorsten

    2007-01-01

    We have developed a procedure for isolation of microRNA and genomic DNA in addition to total RNA from whole blood stabilized in PAXgene Blood RNA tubes. The procedure is based on automatic extraction on a BioRobot MDx and includes isolation of DNA from a fraction of the stabilized blood...... and recovery of small RNA species that are otherwise lost. The procedure presented here is suitable for large-scale experiments and is amenable to further automation. Procured total RNA and DNA was tested using Affymetrix Expression and single-nucleotide polymorphism GeneChips, respectively, and isolated micro......RNA was tested using spotted locked nucleic acid-based microarrays. We conclude that the yield and quality of total RNA, microRNA, and DNA from a single PAXgene blood RNA tube is sufficient for downstream microarray analysis....

  6. Expression analysis of a ''Cucurbita'' cDNA encoding endonuclease

    International Nuclear Information System (INIS)

    Szopa, J.

    1995-01-01

    The nuclear matrices of plant cell nuclei display intrinsic nuclease activity which consists in nicking supercoiled DNA. A cDNA encoding a 32 kDa endonuclease has been cloned and sequenced. The nucleotide and deduced amino-acid sequences show high homology to known 14-3-3-protein sequences from other sources. The amino-acid sequence shows agreement with consensus sequences for potential phosphorylation by protein kinase A and C and for calcium, lipid and membrane-binding sites. The nucleotide-binding site is also present within the conserved part of the sequence. By Northern blot analysis, the differential expression of the corresponding mRNA was detected; it was the strongest in sink tissues. The endonuclease activity found on DNA-polyacrylamide gel electrophoresis coincided with mRNA content and was the highest in tuber. (author). 22 refs, 6 figs

  7. Isolation of full-length putative rat lysophospholipase cDNA using improved methods for mRNA isolation and cDNA cloning

    International Nuclear Information System (INIS)

    Han, J.H.; Stratowa, C.; Rutter, W.J.

    1987-01-01

    The authors have cloned a full-length putative rat pancreatic lysophospholipase cDNA by an improved mRNA isolation method and cDNA cloning strategy using [ 32 P]-labelled nucleotides. These new methods allow the construction of a cDNA library from the adult rat pancreas in which the majority of recombinant clones contained complete sequences for the corresponding mRNAs. A previously recognized but unidentified long and relatively rare cDNA clone containing the entire sequence from the cap site at the 5' end to the poly(A) tail at the 3' end of the mRNA was isolated by single-step screening of the library. The size, amino acid composition, and the activity of the protein expressed in heterologous cells strongly suggest this mRNA codes for lysophospholipase

  8. Normalization for triple-target microarray experiments

    Directory of Open Access Journals (Sweden)

    Magniette Frederic

    2008-04-01

    Full Text Available Abstract Background Most microarray studies are made using labelling with one or two dyes which allows the hybridization of one or two samples on the same slide. In such experiments, the most frequently used dyes are Cy3 and Cy5. Recent improvements in the technology (dye-labelling, scanner and, image analysis allow hybridization up to four samples simultaneously. The two additional dyes are Alexa488 and Alexa494. The triple-target or four-target technology is very promising, since it allows more flexibility in the design of experiments, an increase in the statistical power when comparing gene expressions induced by different conditions and a scaled down number of slides. However, there have been few methods proposed for statistical analysis of such data. Moreover the lowess correction of the global dye effect is available for only two-color experiments, and even if its application can be derived, it does not allow simultaneous correction of the raw data. Results We propose a two-step normalization procedure for triple-target experiments. First the dye bleeding is evaluated and corrected if necessary. Then the signal in each channel is normalized using a generalized lowess procedure to correct a global dye bias. The normalization procedure is validated using triple-self experiments and by comparing the results of triple-target and two-color experiments. Although the focus is on triple-target microarrays, the proposed method can be used to normalize p differently labelled targets co-hybridized on a same array, for any value of p greater than 2. Conclusion The proposed normalization procedure is effective: the technical biases are reduced, the number of false positives is under control in the analysis of differentially expressed genes, and the triple-target experiments are more powerful than the corresponding two-color experiments. There is room for improving the microarray experiments by simultaneously hybridizing more than two samples.

  9. Design of a covalently bonded glycosphingolipid microarray

    DEFF Research Database (Denmark)

    Arigi, Emma; Blixt, Klas Ola; Buschard, Karsten

    2012-01-01

    , the major classes of plant and fungal GSLs. In this work, a prototype "universal" GSL-based covalent microarray has been designed, and preliminary evaluation of its potential utility in assaying protein-GSL binding interactions investigated. An essential step in development involved the enzymatic release...... of the fatty acyl moiety of the ceramide aglycone of selected mammalian GSLs with sphingolipid N-deacylase (SCDase). Derivatization of the free amino group of a typical lyso-GSL, lyso-G(M1), with a prototype linker assembled from succinimidyl-[(N-maleimidopropionamido)-diethyleneglycol] ester and 2...

  10. Linking probe thermodynamics to microarray quantification

    International Nuclear Information System (INIS)

    Li, Shuzhao; Pozhitkov, Alexander; Brouwer, Marius

    2010-01-01

    Understanding the difference in probe properties holds the key to absolute quantification of DNA microarrays. So far, Langmuir-like models have failed to link sequence-specific properties to hybridization signals in the presence of a complex hybridization background. Data from washing experiments indicate that the post-hybridization washing has no major effect on the specifically bound targets, which give the final signals. Thus, the amount of specific targets bound to probes is likely determined before washing, by the competition against nonspecific binding. Our competitive hybridization model is a viable alternative to Langmuir-like models. (comment)

  11. Molecular cloning, characterization and functional analysis of QRFP in orange-spotted grouper (Epinephelus coioides).

    Science.gov (United States)

    Shu, Hu; Chen, Huapu; Liu, Yun; Yang, Lidong; Yang, Yuqing; Zhang, Haifa

    2014-10-01

    The peptide QRFP plays an important role in the regulation of vertebrate feeding behavior. In this study, we cloned the full length cDNA of a QRFP precursor in a teleost fish, the orange-spotted grouper (Epinephelus coioides). Sequence analysis has shown that the functional regions of QRFP in other vertebrates (QRFP-25 and QRFP-7) are conserved in orange-spotted grouper. RT-PCR demonstrated that the pre-processed mRNA of QRFP is widely expressed in orange-spotted grouper. Three days of food deprivation did not change the hypothalamic pre-processed QRFP expression. However, QRFP expression significantly increased when the fish were reefed after three days of fasting. Intraperitoneal injection of QRFP-25 peptide to orange-spotted grouper suppressed expression of orexin, but elevated expression of pro-opiomelanocortin (POMC) in the hypothalamus. We also investigated the effects of QRFP-25 on the expression of reproductive genes. The peptide suppressed the expression of seabream-type gonadotropin-releasing hormones (sbGnRH), luteinizing hormone beta subunit (LHβ) and follicle-stimulating hormone beta subunit (FSHβ) in vivo, as well as inhibited the expression of LHβ and FSHβ in pituitary cells in primary culture. Our results indicate that QRFP may play an inhibitory role in the regulation of feeding behavior and reproduction in orange-spotted grouper. Copyright © 2014 Elsevier Inc. All rights reserved.

  12. Generation of an infectious clone of a new Korean isolate of apple chlorotic leaf spot virus (ACLSV) driven by dual 35S and T7 promoters in a versatile binary vector

    Science.gov (United States)

    The full-length sequence of a new isolate of Apple chlorotic leaf spot virus (ACLSV) from Korea was divergent, but most closely related to the Japanese isolate A4, at 84% nucleotide identity. The full-length cDNA of the Korean isolate of ACLSV was cloned into a binary vector downstream of the bacter...

  13. Cloning and functional expression of a human pancreatic islet glucose-transporter cDNA

    International Nuclear Information System (INIS)

    Permutt, M.A.; Koranyi, L.; Keller, K.; Lacy, P.E.; Scharp, D.W.; Mueckler, M.

    1989-01-01

    Previous studies have suggested that pancreatic islet glucose transport is mediated by a high-K m , low-affinity facilitated transporter similar to that expressed in liver. To determine the relationship between islet and liver glucose transporters, liver-type glucose-transporter cDNA clones were isolated from a human liver cDNA library. The liver-type glucose-transporter cDNA clone hybridized to mRNA transcripts of the same size in human liver and pancreatic islet RNA. A cDNA library was prepared from purified human pancreatic islet tissue and screened with human liver-type glucose-transporter cDNA. The authors isolated two overlapping cDNA clones encompassing 2600 base pairs, which encode a pancreatic islet protein identical in sequence to that of the putative liver-type glucose-transporter protein. Xenopus oocytes injected with synthetic mRNA transcribed from a full-length cDNA construct exhibited increased uptake of 2-deoxyglucose, confirming the functional identity of the clone. These cDNA clones can now be used to study regulation of expression of the gene and to assess the role of inherited defects in this gene as a candidate for inherited susceptibility to non-insulin-dependent diabetes mellitus

  14. Peptidomics combined with cDNA library unravel the diversity of centipede venom

    DEFF Research Database (Denmark)

    Rong, Mingqiang; Yang, Shilong; Wen, Bo

    2015-01-01

    of centipede venom. In the present study, we use peptidomics combined with cDNA library to uncover the diversity of centipede Scolopendra subspinipes mutilans L. Koch. 192 peptides were identified by LC-MS/MS and 79 precursors were deduced by cDNA library. Surprisingly, the signal peptides of centipede toxins...

  15. Construction of full-length cDNA library of white flower Salvia ...

    African Journals Online (AJOL)

    In order to screen and isolate secondary metabolite biosynthesis related gene, we construct a cDNA library of white flower Salvia miltiorrhiza bge. f.alba. High quality of total RNA was successfully isolated from roots of white flower S. miltiorrhiza using modified CTAB method. Double strand cDNA was cloned into pDNR-LIB ...

  16. Design of an Enterobacteriaceae Pan-genome Microarray Chip

    DEFF Research Database (Denmark)

    Lukjancenko, Oksana; Ussery, David

    2010-01-01

    -density microarray chip has been designed, using 116 Enterobacteriaceae genome sequences, taking into account the enteric pan-genome. Probes for the microarray were checked in silico and performance of the chip, based on experimental strains from four different genera, demonstrate a relatively high ability...... to distinguish those strains on genus, species, and pathotype/serovar levels. Additionally, the microarray performed well when investigating which genes were found in a given strain of interest. The Enterobacteriaceae pan-genome microarray, based on 116 genomes, provides a valuable tool for determination...

  17. Numerical optimisation in spot detector design

    NARCIS (Netherlands)

    van der Heijden, Ferdinand; Apperloo, W.; Spreeuwers, Lieuwe Jan

    1997-01-01

    Spots are image details resulting from objects, the projections of which are so small that the inner structure of these objects cannot be resolved from their image. Spot detectors are image operators aiming at the detection and localisation of spots in the image. Most spot detectors can be tuned

  18. Managing emerging threats to spotted owls

    Science.gov (United States)

    Ho Yi Wan; Joseph L. Ganey; Christina D. Vojta; Samuel A. Cushman

    2018-01-01

    The 3 spotted owl (Strix occidentalis) subspecies in North America (i.e., northern spotted owl [S. o. caurina], California spotted owl [S. o. occidentalis], Mexican spotted owl [S. o. lucida]) have all experienced population declines over the past century due to habitat loss and fragmentation from logging. Now, the emerging influences of climate change, high-severity...

  19. 9 CFR 149.4 - Spot audit.

    Science.gov (United States)

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Spot audit. 149.4 Section 149.4... LIVESTOCK IMPROVEMENT VOLUNTARY TRICHINAE CERTIFICATION PROGRAM § 149.4 Spot audit. (a) In addition to regularly scheduled site audits, certified production sites will be subject to spot audits. (1) Random spot...

  20. Comparing transformation methods for DNA microarray data

    Directory of Open Access Journals (Sweden)

    Zwinderman Aeilko H

    2004-06-01

    Full Text Available Abstract Background When DNA microarray data are used for gene clustering, genotype/phenotype correlation studies, or tissue classification the signal intensities are usually transformed and normalized in several steps in order to improve comparability and signal/noise ratio. These steps may include subtraction of an estimated background signal, subtracting the reference signal, smoothing (to account for nonlinear measurement effects, and more. Different authors use different approaches, and it is generally not clear to users which method they should prefer. Results We used the ratio between biological variance and measurement variance (which is an F-like statistic as a quality measure for transformation methods, and we demonstrate a method for maximizing that variance ratio on real data. We explore a number of transformations issues, including Box-Cox transformation, baseline shift, partial subtraction of the log-reference signal and smoothing. It appears that the optimal choice of parameters for the transformation methods depends on the data. Further, the behavior of the variance ratio, under the null hypothesis of zero biological variance, appears to depend on the choice of parameters. Conclusions The use of replicates in microarray experiments is important. Adjustment for the null-hypothesis behavior of the variance ratio is critical to the selection of transformation method.

  1. Molecular cloning and mammalian expression of human beta 2-glycoprotein I cDNA

    DEFF Research Database (Denmark)

    Kristensen, Torsten; Schousboe, Inger; Boel, Espen

    1991-01-01

    Human β2-glycoprotein (β2gpI) cDNA was isolated from a liver cDNA library and sequenced. The cDNA encoded a 19-residue hydrophobic signal peptide followed by the mature β2gpI of 326 amino acid residues. In liver and in the hepatoma cell line HepG2 there are two mRNA species of about 1.4 and 4.3 kb......, respectively, hybridizing specifically with the β2gpI cDNA. Upon isoelectric focusing, recombinant β2gpI obtained from expression of β2gpI cDNA in baby hamster kidney cells showed the same pattern of bands as β2gpI isolated from plasma, and at least 5 polypeptides were visible...

  2. Calibration and assessment of channel-specific biases in microarray data with extended dynamical range.

    Science.gov (United States)

    Bengtsson, Henrik; Jönsson, Göran; Vallon-Christersson, Johan

    2004-11-12

    Non-linearities in observed log-ratios of gene expressions, also known as intensity dependent log-ratios, can often be accounted for by global biases in the two channels being compared. Any step in a microarray process may introduce such offsets and in this article we study the biases introduced by the microarray scanner and the image analysis software. By scanning the same spotted oligonucleotide microarray at different photomultiplier tube (PMT) gains, we have identified a channel-specific bias present in two-channel microarray data. For the scanners analyzed it was in the range of 15-25 (out of 65,535). The observed bias was very stable between subsequent scans of the same array although the PMT gain was greatly adjusted. This indicates that the bias does not originate from a step preceding the scanner detector parts. The bias varies slightly between arrays. When comparing estimates based on data from the same array, but from different scanners, we have found that different scanners introduce different amounts of bias. So do various image analysis methods. We propose a scanning protocol and a constrained affine model that allows us to identify and estimate the bias in each channel. Backward transformation removes the bias and brings the channels to the same scale. The result is that systematic effects such as intensity dependent log-ratios are removed, but also that signal densities become much more similar. The average scan, which has a larger dynamical range and greater signal-to-noise ratio than individual scans, can then be obtained. The study shows that microarray scanners may introduce a significant bias in each channel. Such biases have to be calibrated for, otherwise systematic effects such as intensity dependent log-ratios will be observed. The proposed scanning protocol and calibration method is simple to use and is useful for evaluating scanner biases or for obtaining calibrated measurements with extended dynamical range and better precision. The

  3. Microarray Analysis of the Developing Rat Mandible

    Institute of Scientific and Technical Information of China (English)

    Hideo KABURAGI; Naoyuki SUGANO; Maiko OSHIKAWA; Ryosuke KOSHI; Naoki SENDA; Kazuhiro KAWAMOTO; Koichi ITO

    2007-01-01

    To analyze the molecular events that occur in the developing mandible, we examined the expression of 8803 genes from samples taken at different time points during rat postnatal mandible development.Total RNA was extracted from the mandibles of 1-day-old, 1-week-old, and 2-week-old rats. Complementary RNA (cRNA) was synthesized from cDNA and biotinylated. Fragmented cRNA was hybridized to RGU34A GeneChip arrays. Among the 8803 genes tested, 4344 were detectable. We identified 148 genes with significantly increased expression, and 19 genes with significantly decreased expression. A comprehensive analysis appears to be an effective method of studying the complex process of development.

  4. On the origin of delta spots

    International Nuclear Information System (INIS)

    Tang, F.

    1983-01-01

    Mount Wilson sunspot drawings from 1966 through 1980 were used in conjunction with Hα filtergrams from Big Bear Solar Observatory to examine the origin of delta spots, spots with bipolar umbrae within one penumbra. Of the six cases we studied, five were formed by the union of non-paired spots. They are either shoved into one another by two neighboring growing bipoles or by a new spot born piggy-back style on an existing spot of opposite polarity. Proper motions of the growing spots take on curvilinear paths around one another to avoid a collision. This is the shear motion observed in delta spots (Tanaka, 1979). In the remaining case, the delta spot was formed by spots that emerged as a pair. Our findings indicate no intrinsic differences in the formation or the behavior between delta spots of normal magnetic configuration. (orig.)

  5. Prognostic meta-signature of breast cancer developed by two-stage mixture modeling of microarray data

    Directory of Open Access Journals (Sweden)

    Ghosh Debashis

    2004-12-01

    Full Text Available Abstract Background An increasing number of studies have profiled tumor specimens using distinct microarray platforms and analysis techniques. With the accumulating amount of microarray data, one of the most intriguing yet challenging tasks is to develop robust statistical models to integrate the findings. Results By applying a two-stage Bayesian mixture modeling strategy, we were able to assimilate and analyze four independent microarray studies to derive an inter-study validated "meta-signature" associated with breast cancer prognosis. Combining multiple studies (n = 305 samples on a common probability scale, we developed a 90-gene meta-signature, which strongly associated with survival in breast cancer patients. Given the set of independent studies using different microarray platforms which included spotted cDNAs, Affymetrix GeneChip, and inkjet oligonucleotides, the individually identified classifiers yielded gene sets predictive of survival in each study cohort. The study-specific gene signatures, however, had minimal overlap with each other, and performed poorly in pairwise cross-validation. The meta-signature, on the other hand, accommodated such heterogeneity and achieved comparable or better prognostic performance when compared with the individual signatures. Further by comparing to a global standardization method, the mixture model based data transformation demonstrated superior properties for data integration and provided solid basis for building classifiers at the second stage. Functional annotation revealed that genes involved in cell cycle and signal transduction activities were over-represented in the meta-signature. Conclusion The mixture modeling approach unifies disparate gene expression data on a common probability scale allowing for robust, inter-study validated prognostic signatures to be obtained. With the emerging utility of microarrays for cancer prognosis, it will be important to establish paradigms to meta

  6. Accounting for one-channel depletion improves missing value imputation in 2-dye microarray data.

    Science.gov (United States)

    Ritz, Cecilia; Edén, Patrik

    2008-01-19

    For 2-dye microarray platforms, some missing values may arise from an un-measurably low RNA expression in one channel only. Information of such "one-channel depletion" is so far not included in algorithms for imputation of missing values. Calculating the mean deviation between imputed values and duplicate controls in five datasets, we show that KNN-based imputation gives a systematic bias of the imputed expression values of one-channel depleted spots. Evaluating the correction of this bias by cross-validation showed that the mean square deviation between imputed values and duplicates were reduced up to 51%, depending on dataset. By including more information in the imputation step, we more accurately estimate missing expression values.

  7. CDNA library from the Latex of Hevea brasiliensis

    Directory of Open Access Journals (Sweden)

    Wilaiwan Chotigeat

    2010-12-01

    Full Text Available Latex from Hevea brasiliensis contains 30-50% (w/w of natural rubber (cis-1,4-polyisoprene, the important rawmaterial for many rubber industries. We have constructed a cDNA library from the latex of H. brasiliensis to investigate theexpressed genes and molecular events in the latex. We analyzed 412 expressed sequence tags (ESTs. More than 90% of theEST clones showed homology to previously described sequences in public databases. Functional classification of the ESTsshowed that the largest category were proteins of unknown function (30.1%, 11.4% of ESTs encoded for rubber synthesisrelatedproteins (RS and 8.5% for defense or stress related proteins (DS. Those with no significant homology to knownsequences (NSH accounted for 8.7%, primary metabolism (PM and gene expression and RNA metabolism were 7.8% and6.6%, respectively. Other categories included, protein synthesis-related proteins (6.6%, chromatin and DNA metabolism(CDM 3.9%, energy metabolism (EM 3.4%, cellular transport (CT 3.2%, cell structure (CS 3.2%, signal transduction (ST2.2%, secondary metabolism (SM 1.7%, protein fate (PF 2.2%, and reproductive proteins (RP 0.7%.

  8. Identification of genes related to high royal jelly production in the honey bee (Apis mellifera) using microarray analysis.

    Science.gov (United States)

    Nie, Hongyi; Liu, Xiaoyan; Pan, Jiao; Li, Wenfeng; Li, Zhiguo; Zhang, Shaowu; Chen, Shenglu; Miao, Xiaoqing; Zheng, Nenggan; Su, Songkun

    2017-01-01

    China is the largest royal jelly producer and exporter in the world, and high royal jelly-yielding strains have been bred in the country for approximately three decades. However, information on the molecular mechanism underlying high royal jelly production is scarce. Here, a cDNA microarray was used to screen and identify differentially expressed genes (DEGs) to obtain an overview on the changes in gene expression levels between high and low royal jelly producing bees. We developed a honey bee gene chip that covered 11,689 genes, and this chip was hybridised with cDNA generated from RNA isolated from heads of nursing bees. A total of 369 DEGs were identified between high and low royal jelly producing bees. Amongst these DEGs, 201 (54.47%) genes were up-regulated, whereas 168 (45.53%) were down-regulated in high royal jelly-yielding bees. Gene ontology (GO) analyses showed that they are mainly involved in four key biological processes, and pathway analyses revealed that they belong to a total of 46 biological pathways. These results provide a genetic basis for further studies on the molecular mechanisms involved in high royal jelly production.

  9. Identification of genes related to high royal jelly production in the honey bee (Apis mellifera using microarray analysis

    Directory of Open Access Journals (Sweden)

    Hongyi Nie

    2017-10-01

    Full Text Available Abstract China is the largest royal jelly producer and exporter in the world, and high royal jelly-yielding strains have been bred in the country for approximately three decades. However, information on the molecular mechanism underlying high royal jelly production is scarce. Here, a cDNA microarray was used to screen and identify differentially expressed genes (DEGs to obtain an overview on the changes in gene expression levels between high and low royal jelly producing bees. We developed a honey bee gene chip that covered 11,689 genes, and this chip was hybridised with cDNA generated from RNA isolated from heads of nursing bees. A total of 369 DEGs were identified between high and low royal jelly producing bees. Amongst these DEGs, 201 (54.47% genes were up-regulated, whereas 168 (45.53% were down-regulated in high royal jelly-yielding bees. Gene ontology (GO analyses showed that they are mainly involved in four key biological processes, and pathway analyses revealed that they belong to a total of 46 biological pathways. These results provide a genetic basis for further studies on the molecular mechanisms involved in high royal jelly production.

  10. [Study of generational risk in deafness inflicted couples using deafness gene microarray technique].

    Science.gov (United States)

    Wang, Ping; Zhao, Jia; Yu, Shu-yuan; Jin, Peng; Zhu, Wei; DU, Bo

    2011-06-01

    To explored the significance of screening the gene mutations of deafness related in deaf-mute (deaf & dumb) family using DNA microarray. Total of 52 couples of deaf-mute were recruited from Changchun deaf-mute community. With an average age of (58.3 ± 6.7) years old (x(-) ± s). Blood samples were obtained with informed consent. Their genomic DNA was extracted from peripheral blood and PCR was performed. Nine of hot spot mutations in four most common deafness pathologic gene were examined with the DNA microarray, including GJB2, GJB3, PDS and mtDNA 12S rRNA genes. At the same time, the results were verified with the traditional methods of sequencing. Fifty of normal people served as a control group. All patients were diagnosed non-syndromic sensorineural hearing loss by subjective pure tone audiometry. Thirty-two of 104 cases appeared GJB2 gene mutation (30.7%), the mutation sites included 35delG, 176del16, 235delC and 299delAT. Eighteen of 32 cases of GJB2 mutations were 235delC (59.1%). Seven of 104 cases appeared SLC26A4 gene IVS7-2 A > G mutation. Questionnaire survey and gene diagnosis revealed that four of 52 families have deaf offspring (7.6%). When a couple carries the same gene mutation, the risk of their children deafness was 100%. The results were confirmed with the traditional methods of sequencing. There is a high risk of deafness if a deaf-mute family is planning to have a new baby. It is very important and helpful to avoid deaf newborns again in deaf-mute family by DNA microarray.

  11. Poisson's spot and Gouy phase

    Science.gov (United States)

    da Paz, I. G.; Soldati, Rodolfo; Cabral, L. A.; de Oliveira, J. G. G.; Sampaio, Marcos

    2016-12-01

    Recently there have been experimental results on Poisson spot matter-wave interferometry followed by theoretical models describing the relative importance of the wave and particle behaviors for the phenomenon. We propose an analytical theoretical model for Poisson's spot with matter waves based on the Babinet principle, in which we use the results for free propagation and single-slit diffraction. We take into account effects of loss of coherence and finite detection area using the propagator for a quantum particle interacting with an environment. We observe that the matter-wave Gouy phase plays a role in the existence of the central peak and thus corroborates the predominantly wavelike character of the Poisson's spot. Our model shows remarkable agreement with the experimental data for deuterium (D2) molecules.

  12. Shared probe design and existing microarray reanalysis using PICKY

    Directory of Open Access Journals (Sweden)

    Chou Hui-Hsien

    2010-04-01

    Full Text Available Abstract Background Large genomes contain families of highly similar genes that cannot be individually identified by microarray probes. This limitation is due to thermodynamic restrictions and cannot be resolved by any computational method. Since gene annotations are updated more frequently than microarrays, another common issue facing microarray users is that existing microarrays must be routinely reanalyzed to determine probes that are still useful with respect to the updated annotations. Results PICKY 2.0 can design shared probes for sets of genes that cannot be individually identified using unique probes. PICKY 2.0 uses novel algorithms to track sharable regions among genes and to strictly distinguish them from other highly similar but nontarget regions during thermodynamic comparisons. Therefore, PICKY does not sacrifice the quality of shared probes when choosing them. The latest PICKY 2.1 includes the new capability to reanalyze existing microarray probes against updated gene sets to determine probes that are still valid to use. In addition, more precise nonlinear salt effect estimates and other improvements are added, making PICKY 2.1 more versatile to microarray users. Conclusions Shared probes allow expressed gene family members to be detected; this capability is generally more desirable than not knowing anything about these genes. Shared probes also enable the design of cross-genome microarrays, which facilitate multiple species identification in environmental samples. The new nonlinear salt effect calculation significantly increases the precision of probes at a lower buffer salt concentration, and the probe reanalysis function improves existing microarray result interpretations.

  13. A Critical Perspective On Microarray Breast Cancer Gene Expression Profiling

    NARCIS (Netherlands)

    Sontrop, H.M.J.

    2015-01-01

    Microarrays offer biologists an exciting tool that allows the simultaneous assessment of gene expression levels for thousands of genes at once. At the time of their inception, microarrays were hailed as the new dawn in cancer biology and oncology practice with the hope that within a decade diseases

  14. The Importance of Normalization on Large and Heterogeneous Microarray Datasets

    Science.gov (United States)

    DNA microarray technology is a powerful functional genomics tool increasingly used for investigating global gene expression in environmental studies. Microarrays can also be used in identifying biological networks, as they give insight on the complex gene-to-gene interactions, ne...

  15. The application of DNA microarrays in gene expression analysis

    NARCIS (Netherlands)

    Hal, van N.L.W.; Vorst, O.; Houwelingen, van A.M.M.L.; Kok, E.J.; Peijnenburg, A.A.C.M.; Aharoni, A.; Tunen, van A.J.; Keijer, J.

    2000-01-01

    DNA microarray technology is a new and powerful technology that will substantially increase the speed of molecular biological research. This paper gives a survey of DNA microarray technology and its use in gene expression studies. The technical aspects and their potential improvements are discussed.

  16. Laser based spot weld characterization

    Science.gov (United States)

    Jonietz, Florian; Myrach, Philipp; Rethmeier, Michael; Suwala, Hubert; Ziegler, Mathias

    2016-02-01

    Spot welding is one of the most important joining technologies, especially in the automotive industry. Hitherto, the quality of spot welded joints is tested mainly by random destructive tests. A nondestructive testing technique offers the benefit of cost reduction of the testing procedure and optimization of the fabrication process, because every joint could be examined. This would lead to a reduced number of spot welded joints, as redundancies could be avoided. In the procedure described here, the spot welded joint between two zinc-coated steel sheets (HX340LAD+Z100MB or HC340LA+ZE 50/50) is heated optically on one side. Laser radiation and flash light are used as heat sources. The melted zone, the so called "weld nugget" provides the mechanical stability of the connection, but also constitutes a thermal bridge between the sheets. Due to the better thermal contact, the spot welded joint reveals a thermal behavior different from the surrounding material, where the heat transfer between the two sheets is much lower. The difference in the transient thermal behavior is measured with time resolved thermography. Hence, the size of the thermal contact between the two sheets is determined, which is directly correlated to the size of the weld nugget, indicating the quality of the spot weld. The method performs well in transmission with laser radiation and flash light. With laser radiation, it works even in reflection geometry, thus offering the possibility of testing with just one-sided accessibility. By using heating with collimated laser radiation, not only contact-free, but also remote testing is feasible. A further convenience compared to similar thermographic approaches is the applicability on bare steel sheets without any optical coating for emissivity correction. For this purpose, a proper way of emissivity correction was established.

  17. Uses of Dendrimers for DNA Microarrays

    Directory of Open Access Journals (Sweden)

    Jean-Pierre Majoral

    2006-08-01

    Full Text Available Biosensors such as DNA microarrays and microchips are gaining an increasingimportance in medicinal, forensic, and environmental analyses. Such devices are based onthe detection of supramolecular interactions called hybridizations that occur betweencomplementary oligonucleotides, one linked to a solid surface (the probe, and the other oneto be analyzed (the target. This paper focuses on the improvements that hyperbranched andperfectly defined nanomolecules called dendrimers can provide to this methodology. Twomain uses of dendrimers for such purpose have been described up to now; either thedendrimer is used as linker between the solid surface and the probe oligonucleotide, or thedendrimer is used as a multilabeled entity linked to the target oligonucleotide. In the firstcase the dendrimer generally induces a higher loading of probes and an easier hybridization,due to moving away the solid phase. In the second case the high number of localized labels(generally fluorescent induces an increased sensitivity, allowing the detection of smallquantities of biological entities.

  18. Fiscal 2000 report on result of the full-length cDNA structure analysis; 2000 nendo kanzen cho cDNA kozo kaiseki seika hokokusho

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    2001-03-01

    This paper explains the results of research on full-length cDNA structure analysis for the period from April, 2000 to March, 2001. The outline of human genome sequence was published in June, 2000. In Japan, human gene analysis was such that, as the basic technology of the bio industry, a millennium project was decided in the budget of fiscal 2000. The full-length cDNA structure analysis is the core of the project. The libraries of cDNA were prepared using full-length and more than 4-5kbp-long cDNAs by oligo-capping method. It began from determining partial sequence data at end cDNA, and then, with new clones selected therefrom, full-length human cDNA sequence data were determined. The partial sequence data determined by fiscal 2000 were 1,035,000 clones while the full-length sequence data were 12,144 clones. The sequence data obtained were analyzed by homology search and translated into amino acid coding sequences, with predictions conducted on protein functions. A clustering method was examined that selects new clones from partial sequences. Database was constructed on gene expression profiles and disease-related gene sequence data. (NEDO)

  19. RICD: A rice indica cDNA database resource for rice functional genomics

    Directory of Open Access Journals (Sweden)

    Zhang Qifa

    2008-11-01

    Full Text Available Abstract Background The Oryza sativa L. indica subspecies is the most widely cultivated rice. During the last few years, we have collected over 20,000 putative full-length cDNAs and over 40,000 ESTs isolated from various cDNA libraries of two indica varieties Guangluai 4 and Minghui 63. A database of the rice indica cDNAs was therefore built to provide a comprehensive web data source for searching and retrieving the indica cDNA clones. Results Rice Indica cDNA Database (RICD is an online MySQL-PHP driven database with a user-friendly web interface. It allows investigators to query the cDNA clones by keyword, genome position, nucleotide or protein sequence, and putative function. It also provides a series of information, including sequences, protein domain annotations, similarity search results, SNPs and InDels information, and hyperlinks to gene annotation in both The Rice Annotation Project Database (RAP-DB and The TIGR Rice Genome Annotation Resource, expression atlas in RiceGE and variation report in Gramene of each cDNA. Conclusion The online rice indica cDNA database provides cDNA resource with comprehensive information to researchers for functional analysis of indica subspecies and for comparative genomics. The RICD database is available through our website http://www.ncgr.ac.cn/ricd.

  20. Bystander effect: Biological endpoints and microarray analysis

    Energy Technology Data Exchange (ETDEWEB)

    Chaudhry, M. Ahmad [Department of Medical Laboratory and Radiation Sciences, College of Nursing and Health Sciences, University of Vermont, 302 Rowell Building, Burlington, VT 05405 (United States) and DNA Microarray Facility, University of Vermont, Burlington, VT 05405 (United States)]. E-mail: mchaudhr@uvm.edu

    2006-05-11

    In cell populations exposed to ionizing radiation, the biological effects occur in a much larger proportion of cells than are estimated to be traversed by radiation. It has been suggested that irradiated cells are capable of providing signals to the neighboring unirradiated cells resulting in damage to these cells. This phenomenon is termed the bystander effect. The bystander effect induces persistent, long-term, transmissible changes that result in delayed death and neoplastic transformation. Because the bystander effect is relevant to carcinogenesis, it could have significant implications for risk estimation for radiation exposure. The nature of the bystander effect signal and how it impacts the unirradiated cells remains to be elucidated. Examination of the changes in gene expression could provide clues to understanding the bystander effect and could define the signaling pathways involved in sustaining damage to these cells. The microarray technology serves as a tool to gain insight into the molecular pathways leading to bystander effect. Using medium from irradiated normal human diploid lung fibroblasts as a model system we examined gene expression alterations in bystander cells. The microarray data revealed that the radiation-induced gene expression profile in irradiated cells is different from unirradiated bystander cells suggesting that the pathways leading to biological effects in the bystander cells are different from the directly irradiated cells. The genes known to be responsive to ionizing radiation were observed in irradiated cells. Several genes were upregulated in cells receiving media from irradiated cells. Surprisingly no genes were found to be downregulated in these cells. A number of genes belonging to extracellular signaling, growth factors and several receptors were identified in bystander cells. Interestingly 15 genes involved in the cell communication processes were found to be upregulated. The induction of receptors and the cell

  1. Bystander effect: Biological endpoints and microarray analysis

    International Nuclear Information System (INIS)

    Chaudhry, M. Ahmad

    2006-01-01

    In cell populations exposed to ionizing radiation, the biological effects occur in a much larger proportion of cells than are estimated to be traversed by radiation. It has been suggested that irradiated cells are capable of providing signals to the neighboring unirradiated cells resulting in damage to these cells. This phenomenon is termed the bystander effect. The bystander effect induces persistent, long-term, transmissible changes that result in delayed death and neoplastic transformation. Because the bystander effect is relevant to carcinogenesis, it could have significant implications for risk estimation for radiation exposure. The nature of the bystander effect signal and how it impacts the unirradiated cells remains to be elucidated. Examination of the changes in gene expression could provide clues to understanding the bystander effect and could define the signaling pathways involved in sustaining damage to these cells. The microarray technology serves as a tool to gain insight into the molecular pathways leading to bystander effect. Using medium from irradiated normal human diploid lung fibroblasts as a model system we examined gene expression alterations in bystander cells. The microarray data revealed that the radiation-induced gene expression profile in irradiated cells is different from unirradiated bystander cells suggesting that the pathways leading to biological effects in the bystander cells are different from the directly irradiated cells. The genes known to be responsive to ionizing radiation were observed in irradiated cells. Several genes were upregulated in cells receiving media from irradiated cells. Surprisingly no genes were found to be downregulated in these cells. A number of genes belonging to extracellular signaling, growth factors and several receptors were identified in bystander cells. Interestingly 15 genes involved in the cell communication processes were found to be upregulated. The induction of receptors and the cell

  2. Lipid Microarray Biosensor for Biotoxin Detection.

    Energy Technology Data Exchange (ETDEWEB)

    Singh, Anup K.; Throckmorton, Daniel J.; Moran-Mirabal, Jose C.; Edel, Joshua B.; Meyer, Grant D.; Craighead, Harold G.

    2006-05-01

    We present the use of micron-sized lipid domains, patterned onto planar substrates and within microfluidic channels, to assay the binding of bacterial toxins via total internal reflection fluorescence microscopy (TIRFM). The lipid domains were patterned using a polymer lift-off technique and consisted of ganglioside-populated DSPC:cholesterol supported lipid bilayers (SLBs). Lipid patterns were formed on the substrates by vesicle fusion followed by polymer lift-off, which revealed micron-sized SLBs containing either ganglioside GT1b or GM1. The ganglioside-populated SLB arrays were then exposed to either Cholera toxin subunit B (CTB) or Tetanus toxin fragment C (TTC). Binding was assayed on planar substrates by TIRFM down to 1 nM concentration for CTB and 100 nM for TTC. Apparent binding constants extracted from three different models applied to the binding curves suggest that binding of a protein to a lipid-based receptor is strongly affected by the lipid composition of the SLB and by the substrate on which the bilayer is formed. Patterning of SLBs inside microfluidic channels also allowed the preparation of lipid domains with different compositions on a single device. Arrays within microfluidic channels were used to achieve segregation and selective binding from a binary mixture of the toxin fragments in one device. The binding and segregation within the microfluidic channels was assayed with epifluorescence as proof of concept. We propose that the method used for patterning the lipid microarrays on planar substrates and within microfluidic channels can be easily adapted to proteins or nucleic acids and can be used for biosensor applications and cell stimulation assays under different flow conditions. KEYWORDS. Microarray, ganglioside, polymer lift-off, cholera toxin, tetanus toxin, TIRFM, binding constant.4

  3. Is this Red Spot the Blue Spot (locus ceruleum)?

    Energy Technology Data Exchange (ETDEWEB)

    Choe, Won Sick; Lee, Yu Kyung; Lee, Min Kyung; Hwang, Kyung Hoon [Gachon University Gil Hospital, Incheon (Korea, Republic of)

    2010-06-15

    The authors report brain images of 18F-FDG-PET in a case of schizophrenia. The images showed strikingly increased bilateral uptake in the locus ceruleum. The locus ceruleum is called the blue spot and known to be a center of the norepinephrinergic system.

  4. Is this Red Spot the Blue Spot (locus ceruleum)?

    International Nuclear Information System (INIS)

    Choe, Won Sick; Lee, Yu Kyung; Lee, Min Kyung; Hwang, Kyung Hoon

    2010-01-01

    The authors report brain images of 18F-FDG-PET in a case of schizophrenia. The images showed strikingly increased bilateral uptake in the locus ceruleum. The locus ceruleum is called the blue spot and known to be a center of the norepinephrinergic system.

  5. Statistical hot spot analysis of reactor cores

    International Nuclear Information System (INIS)

    Schaefer, H.

    1974-05-01

    This report is an introduction into statistical hot spot analysis. After the definition of the term 'hot spot' a statistical analysis is outlined. The mathematical method is presented, especially the formula concerning the probability of no hot spots in a reactor core is evaluated. A discussion with the boundary conditions of a statistical hot spot analysis is given (technological limits, nominal situation, uncertainties). The application of the hot spot analysis to the linear power of pellets and the temperature rise in cooling channels is demonstrated with respect to the test zone of KNK II. Basic values, such as probability of no hot spots, hot spot potential, expected hot spot diagram and cumulative distribution function of hot spots, are discussed. It is shown, that the risk of hot channels can be dispersed equally over all subassemblies by an adequate choice of the nominal temperature distribution in the core

  6. Construction and characterization of cDNA library for IRM-2 mice

    International Nuclear Information System (INIS)

    Wang Qin; Li Jin; Song Li; Liu Qiang; Yue Jingyin; Mu Chuanjie; Tang Weisheng; Fan Feiyue

    2010-01-01

    Objective: To screen and isolate the radioresistance related genes of IRM-2 mice. Methods: cDNA library of IRM-2 mice was constructed by SMART technique. Total RNA was isolated from spleens of IRM-2 male mice. The first-strand cDNA was synthesized by using PowerScript reverse transcriptase, and double-strand cDNA was synthesized and amplified by long PCR. The PCR products were purified, digested with restriction enzyme Sfi I. The ds-cDNA fragment less than 500 bp was fractionated and ligated to the Sfi I-digested pDNR-LIB vector. The ligation mixture was transformed into E. coil DH5 α by electroporation transformation to generate the unamplified cDNA library. The quality of cDNA library was identified by PCR technique. 130 clones from cDNA library were sequenced and compared with GenBank database. Results: The cDNA library contained 2.25 x 10 6 independent clones with an average insert size of 1.2 kb. The ratio of recombination and full-length was 95% and 55%, respectively. 21 pieces of EST sequences from cDNA library were not the same as the known mice genes and registered into GenBank EST database, with registered number DW474856-DW474876. Conclusions: cDNA library of IRM-2 mice has been constructed successfully. 21 pieces of EST implies that radioresistance correlative genes may be in IRM-2 mice, which will lay a foundation for isolating and identifying radioresistance related genes in further study. (authors)

  7. Versatile High Resolution Oligosaccharide Microarrays for Plant Glycobiology and Cell Wall Research

    DEFF Research Database (Denmark)

    Pedersen, Henriette Lodberg; Fangel, Jonatan Ulrik; McCleary, Barry

    2012-01-01

    Microarrays are powerful tools for high throughput analysis, and hundreds or thousands of molecular interactions can be assessed simultaneously using very small amounts of analytes. Nucleotide microarrays are well established in plant research, but carbohydrate microarrays are much less establish...

  8. Isolation of cDNA clones coding for human tissue factor: primary structure of the protein and cDNA

    International Nuclear Information System (INIS)

    Spicer, E.K.; Horton, R.; Bloem, L.

    1987-01-01

    Tissue factor is a membrane-bound procoagulant protein that activates the extrinsic pathway of blood coagulation in the presence of factor VII and calcium. λ Phage containing the tissue factor gene were isolated from a human placental cDNA library. The amino acid sequence deduced from the nucleotide sequence of the cDNAs indicates that tissue factor is synthesized as a higher molecular weight precursor with a leader sequence of 32 amino acids, while the mature protein is a single polypeptide chain composed of 263 residues. The derived primary structure of tissue factor has been confirmed by comparison to protein and peptide sequence data. The sequence of the mature protein suggests that there are three distinct domains: extracellular, residues 1-219; hydrophobic, residues 220-242; and cytoplasmic, residues 243-263. Three potential N-linked carbohydrate attachment sites occur in the extracellular domain. The amino acid sequence of tissue factor shows no significant homology with the vitamin K-dependent serine proteases, coagulation cofactors, or any other protein in the National Biomedical Research Foundation sequence data bank (Washington, DC)

  9. Microarray-based analysis of differential gene expression between infective and noninfective larvae of Strongyloides stercoralis.

    Directory of Open Access Journals (Sweden)

    Roshan Ramanathan

    2011-05-01

    Full Text Available Differences between noninfective first-stage (L1 and infective third-stage (L3i larvae of parasitic nematode Strongyloides stercoralis at the molecular level are relatively uncharacterized. DNA microarrays were developed and utilized for this purpose.Oligonucleotide hybridization probes for the array were designed to bind 3,571 putative mRNA transcripts predicted by analysis of 11,335 expressed sequence tags (ESTs obtained as part of the Nematode EST project. RNA obtained from S. stercoralis L3i and L1 was co-hybridized to each array after labeling the individual samples with different fluorescent tags. Bioinformatic predictions of gene function were developed using a novel cDNA Annotation System software. We identified 935 differentially expressed genes (469 L3i-biased; 466 L1-biased having two-fold expression differences or greater and microarray signals with a p value<0.01. Based on a functional analysis, L1 larvae have a larger number of genes putatively involved in transcription (p = 0.004, and L3i larvae have biased expression of putative heat shock proteins (such as hsp-90. Genes with products known to be immunoreactive in S. stercoralis-infected humans (such as SsIR and NIE had L3i biased expression. Abundantly expressed L3i contigs of interest included S. stercoralis orthologs of cytochrome oxidase ucr 2.1 and hsp-90, which may be potential chemotherapeutic targets. The S. stercoralis ortholog of fatty acid and retinol binding protein-1, successfully used in a vaccine against Ancylostoma ceylanicum, was identified among the 25 most highly expressed L3i genes. The sperm-containing glycoprotein domain, utilized in a vaccine against the nematode Cooperia punctata, was exclusively found in L3i biased genes and may be a valuable S. stercoralis target of interest.A new DNA microarray tool for the examination of S. stercoralis biology has been developed and provides new and valuable insights regarding differences between infective and

  10. Three-dimensional (3D) plasma micro-nanotextured slides for high performance biomolecule microarrays: Comparison with epoxy-silane coated glass slides.

    Science.gov (United States)

    Tsougeni, Katerina; Ellinas, Kosmas; Koukouvinos, George; Petrou, Panagiota S; Tserepi, Angeliki; Kakabakos, Sotirios E; Gogolides, Evangelos

    2018-05-01

    Glass slides coated with a poly(methyl methacrylate) layer and plasma micro-nanotextured to acquire 3D topography (referred as 3D micro-nanotextured slides) were evaluated as substrates for biomolecule microarrays. Their performance is compared with that of epoxy-coated glass slides. We found that the proposed three-dimensional (3D) slides offered significant improvements in terms of spot intensity, homogeneity, and reproducibility. In particular, they provided higher spot intensity, by a factor of at least 1.5, and significantly improved spot homogeneity when compared to the epoxy-silane coated ones (intra-spot and between spot coefficients of variation ranging between 5 and 15% for the 3D micro-nanotextured slides and between 25 and 85% for the epoxy-silane coated ones). The latter was to a great extent the result of a strong "coffee-ring" effect observed for the spots created on the epoxy-coated slides; a phenomenon that was severely reduced in the 3D micro-nanotextured slides. The 3D micro-nanotextured slides offered in addition higher signal to noise ratio values over a wide range of protein probe concentrations and shelf-life over one year without requirement for specific storage conditions. Finally, the protocols employed for protein probe immobilization were extremely simple. Copyright © 2018 Elsevier B.V. All rights reserved.

  11. Rocky Mountain spotted fever, Panama.

    Science.gov (United States)

    Estripeaut, Dora; Aramburú, María Gabriela; Sáez-Llorens, Xavier; Thompson, Herbert A; Dasch, Gregory A; Paddock, Christopher D; Zaki, Sherif; Eremeeva, Marina E

    2007-11-01

    We describe a fatal pediatric case of Rocky Mountain spotted fever in Panama, the first, to our knowledge, since the 1950s. Diagnosis was established by immunohistochemistry, PCR, and isolation of Rickettsia rickettsii from postmortem tissues. Molecular typing demonstrated strong relatedness of the isolate to strains of R. rickettsii from Central and South America.

  12. Gene expression patterns during the larval development of European sea bass (dicentrarchus labrax) by microarray analysis.

    Science.gov (United States)

    Darias, M J; Zambonino-Infante, J L; Hugot, K; Cahu, C L; Mazurais, D

    2008-01-01

    During the larval period, marine teleosts undergo very fast growth and dramatic changes in morphology, metabolism, and behavior to accomplish their metamorphosis into juvenile fish. Regulation of gene expression is widely thought to be a key mechanism underlying the management of the biological processes required for harmonious development over this phase of life. To provide an overall analysis of gene expression in the whole body during sea bass larval development, we monitored the expression of 6,626 distinct genes at 10 different points in time between 7 and 43 days post-hatching (dph) by using heterologous hybridization of a rainbow trout cDNA microarray. The differentially expressed genes (n = 485) could be grouped into two categories: genes that were generally up-expressed early, between 7 and 23 dph, and genes up-expressed between 25 and 43 dph. Interestingly, among the genes regulated during the larval period, those related to organogenesis, energy pathways, biosynthesis, and digestion were over-represented compared with total set of analyzed genes. We discuss the quantitative regulation of whole-body contents of these specific transcripts with regard to the ontogenesis and maturation of essential functions that take place over larval development. Our study is the first utilization of a transcriptomic approach in sea bass and reveals dynamic changes in gene expression patterns in relation to marine finfish larval development.

  13. Protective Effect of Gwakhyangjeonggisan Herbal Acupuncture Solution in Glioblastoma Cells: Microarray Analysis of Gene Expression

    Directory of Open Access Journals (Sweden)

    Hong-Seok Lee

    2005-12-01

    Full Text Available Objectives : Neurological disorders have been one of main therapeutic targets of acupuncture. The present study investigated the protective effects of Gwakhyangjeonggisan herbal acupuncture solution (GHAS. Methods : We performed 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide (MTT assay in glioblastoma cells, and did microarray analysis with cells exposed to reactive oxigen species (ROS of hydrogen peroxide by 8.0 k Human cDNA, with cut-off level of 2-fold changes in gene expression. Results : MTT assay showed protective effect of GHAS on the glioblastoma cells exposed to hydrogen peroxide. When glioblastoma cells were exposed to hydrogen peroxide, 24 genes were downregulated. When the cells were pretreated with GHAS before exposure to hydrogen peroxide, 46 genes were downregulated. Many of the genes downregulated by hydrogen peroxide stimulation were decreased in the amount of downregulation or reversed to upregulation. Conclusions : The gene expression changes observed in the present study are supposed to be related to the protective molecular mechanism of GHAS in the glioblastoma cells exposed to ROS stress.

  14. Microarray analysis reveals genetic pathways modulated by tipifarnib in acute myeloid leukemia

    International Nuclear Information System (INIS)

    Raponi, Mitch; Belly, Robert T; Karp, Judith E; Lancet, Jeffrey E; Atkins, David; Wang, Yixin

    2004-01-01

    Farnesyl protein transferase inhibitors (FTIs) were originally developed to inhibit oncogenic ras, however it is now clear that there are several other potential targets for this drug class. The FTI tipifarnib (ZARNESTRA™, R115777) has recently demonstrated clinical responses in adults with refractory and relapsed acute leukemias. This study was conducted to identify genetic markers and pathways that are regulated by tipifarnib in acute myeloid leukemia (AML). Tipifarnib-mediated gene expression changes in 3 AML cell lines and bone marrow samples from two patients with AML were analyzed on a cDNA microarray containing approximately 7000 human genes. Pathways associated with these expression changes were identified using the Ingenuity Pathway Analysis tool. The expression analysis identified a common set of genes that were regulated by tipifarnib in three leukemic cell lines and in leukemic blast cells isolated from two patients who had been treated with tipifarnib. Association of modulated genes with biological functional groups identified several pathways affected by tipifarnib including cell signaling, cytoskeletal organization, immunity, and apoptosis. Gene expression changes were verified in a subset of genes using real time RT-PCR. Additionally, regulation of apoptotic genes was found to correlate with increased Annexin V staining in the THP-1 cell line but not in the HL-60 cell line. The genetic networks derived from these studies illuminate some of the biological pathways affected by FTI treatment while providing a proof of principle for identifying candidate genes that might be used as surrogate biomarkers of drug activity

  15. A general framework for optimization of probes for gene expression microarray and its application to the fungus Podospora anserina.

    Science.gov (United States)

    Bidard, Frédérique; Imbeaud, Sandrine; Reymond, Nancie; Lespinet, Olivier; Silar, Philippe; Clavé, Corinne; Delacroix, Hervé; Berteaux-Lecellier, Véronique; Debuchy, Robert

    2010-06-18

    The development of new microarray technologies makes custom long oligonucleotide arrays affordable for many experimental applications, notably gene expression analyses. Reliable results depend on probe design quality and selection. Probe design strategy should cope with the limited accuracy of de novo gene prediction programs, and annotation up-dating. We present a novel in silico procedure which addresses these issues and includes experimental screening, as an empirical approach is the best strategy to identify optimal probes in the in silico outcome. We used four criteria for in silico probe selection: cross-hybridization, hairpin stability, probe location relative to coding sequence end and intron position. This latter criterion is critical when exon-intron gene structure predictions for intron-rich genes are inaccurate. For each coding sequence (CDS), we selected a sub-set of four probes. These probes were included in a test microarray, which was used to evaluate the hybridization behavior of each probe. The best probe for each CDS was selected according to three experimental criteria: signal-to-noise ratio, signal reproducibility, and representative signal intensities. This procedure was applied for the development of a gene expression Agilent platform for the filamentous fungus Podospora anserina and the selection of a single 60-mer probe for each of the 10,556 P. anserina CDS. A reliable gene expression microarray version based on the Agilent 44K platform was developed with four spot replicates of each probe to increase statistical significance of analysis.

  16. A general framework for optimization of probes for gene expression microarray and its application to the fungus Podospora anserina

    Directory of Open Access Journals (Sweden)

    Bidard Frédérique

    2010-06-01

    Full Text Available Abstract Background The development of new microarray technologies makes custom long oligonucleotide arrays affordable for many experimental applications, notably gene expression analyses. Reliable results depend on probe design quality and selection. Probe design strategy should cope with the limited accuracy of de novo gene prediction programs, and annotation up-dating. We present a novel in silico procedure which addresses these issues and includes experimental screening, as an empirical approach is the best strategy to identify optimal probes in the in silico outcome. Findings We used four criteria for in silico probe selection: cross-hybridization, hairpin stability, probe location relative to coding sequence end and intron position. This latter criterion is critical when exon-intron gene structure predictions for intron-rich genes are inaccurate. For each coding sequence (CDS, we selected a sub-set of four probes. These probes were included in a test microarray, which was used to evaluate the hybridization behavior of each probe. The best probe for each CDS was selected according to three experimental criteria: signal-to-noise ratio, signal reproducibility, and representative signal intensities. This procedure was applied for the development of a gene expression Agilent platform for the filamentous fungus Podospora anserina and the selection of a single 60-mer probe for each of the 10,556 P. anserina CDS. Conclusions A reliable gene expression microarray version based on the Agilent 44K platform was developed with four spot replicates of each probe to increase statistical significance of analysis.

  17. Development and validation of a microarray for the investigation of the CAZymes encoded by the human gut microbiome.

    Directory of Open Access Journals (Sweden)

    Abdessamad El Kaoutari

    Full Text Available Distal gut bacteria play a pivotal role in the digestion of dietary polysaccharides by producing a large number of carbohydrate-active enzymes (CAZymes that the host otherwise does not produce. We report here the design of a custom microarray that we used to spot non-redundant DNA probes for more than 6,500 genes encoding glycoside hydrolases and lyases selected from 174 reference genomes from distal gut bacteria. The custom microarray was tested and validated by the hybridization of bacterial DNA extracted from the stool samples of lean, obese and anorexic individuals. Our results suggest that a microarray-based study can detect genes from low-abundance bacteria better than metagenomic-based studies. A striking example was the finding that a gene encoding a GH6-family cellulase was present in all subjects examined, whereas metagenomic studies have consistently failed to detect this gene in both human and animal gut microbiomes. In addition, an examination of eight stool samples allowed the identification of a corresponding CAZome core containing 46 families of glycoside hydrolases and polysaccharide lyases, which suggests the functional stability of the gut microbiota despite large taxonomical variations between individuals.

  18. Using microarrays to identify positional candidate genes for QTL: the case study of ACTH response in pigs.

    Science.gov (United States)

    Jouffe, Vincent; Rowe, Suzanne; Liaubet, Laurence; Buitenhuis, Bart; Hornshøj, Henrik; SanCristobal, Magali; Mormède, Pierre; de Koning, D J

    2009-07-16

    Microarray studies can supplement QTL studies by suggesting potential candidate genes in the QTL regions, which by themselves are too large to provide a limited selection of candidate genes. Here we provide a case study where we explore ways to integrate QTL data and microarray data for the pig, which has only a partial genome sequence. We outline various procedures to localize differentially expressed genes on the pig genome and link this with information on published QTL. The starting point is a set of 237 differentially expressed cDNA clones in adrenal tissue from two pig breeds, before and after treatment with adrenocorticotropic hormone (ACTH). Different approaches to localize the differentially expressed (DE) genes to the pig genome showed different levels of success and a clear lack of concordance for some genes between the various approaches. For a focused analysis on 12 genes, overlapping QTL from the public domain were presented. Also, differentially expressed genes underlying QTL for ACTH response were described. Using the latest version of the draft sequence, the differentially expressed genes were mapped to the pig genome. This enabled co-location of DE genes and previously studied QTL regions, but the draft genome sequence is still incomplete and will contain many errors. A further step to explore links between DE genes and QTL at the pathway level was largely unsuccessful due to the lack of annotation of the pig genome. This could be improved by further comparative mapping analyses but this would be time consuming. This paper provides a case study for the integration of QTL data and microarray data for a species with limited genome sequence information and annotation. The results illustrate the challenges that must be addressed but also provide a roadmap for future work that is applicable to other non-model species.

  19. Microintaglio Printing for Soft Lithography-Based in Situ Microarrays

    Directory of Open Access Journals (Sweden)

    Manish Biyani

    2015-07-01

    Full Text Available Advances in lithographic approaches to fabricating bio-microarrays have been extensively explored over the last two decades. However, the need for pattern flexibility, a high density, a high resolution, affordability and on-demand fabrication is promoting the development of unconventional routes for microarray fabrication. This review highlights the development and uses of a new molecular lithography approach, called “microintaglio printing technology”, for large-scale bio-microarray fabrication using a microreactor array (µRA-based chip consisting of uniformly-arranged, femtoliter-size µRA molds. In this method, a single-molecule-amplified DNA microarray pattern is self-assembled onto a µRA mold and subsequently converted into a messenger RNA or protein microarray pattern by simultaneously producing and transferring (immobilizing a messenger RNA or a protein from a µRA mold to a glass surface. Microintaglio printing allows the self-assembly and patterning of in situ-synthesized biomolecules into high-density (kilo-giga-density, ordered arrays on a chip surface with µm-order precision. This holistic aim, which is difficult to achieve using conventional printing and microarray approaches, is expected to revolutionize and reshape proteomics. This review is not written comprehensively, but rather substantively, highlighting the versatility of microintaglio printing for developing a prerequisite platform for microarray technology for the postgenomic era.

  20. Sequence of a cloned cDNA encoding human ribosomal protein S11

    Energy Technology Data Exchange (ETDEWEB)

    Lott, J B; Mackie, G A

    1988-02-11

    The authors have isolated a cloned cDNA that encodes human ribosomal protein (rp) S11 by screening a human fibroblast cDNA library with a labelled 204 bp DNA fragment encompassing residues 212-416 of pRS11, a rat rp Sll cDNA clone. The human rp S11 cloned cDNA consists of 15 residues of the 5' leader, the entire coding sequence and all 51 residues of the 3' untranslated region. The predicted amino acid sequence of 158 residues is identical to rat rpS11. The nucleotide sequence in the coding region differs, however, from that in rat in the first position in two codons and in the third position in 44 codons.

  1. Mouse tetranectin: cDNA sequence, tissue-specific expression, and chromosomal mapping

    DEFF Research Database (Denmark)

    Ibaraki, K; Kozak, C A; Wewer, U M

    1995-01-01

    regulation, mouse tetranectin cDNA was cloned from a 16-day-old mouse embryo library. Sequence analysis revealed a 992-bp cDNA with an open reading frame of 606 bp, which is identical in length to the human tetranectin cDNA. The deduced amino acid sequence showed high homology to the human cDNA with 76......(s) of tetranectin. The sequence analysis revealed a difference in both sequence and size of the noncoding regions between mouse and human cDNAs. Northern analysis of the various tissues from mouse, rat, and cow showed the major transcript(s) to be approximately 1 kb, which is similar in size to that observed...

  2. Reverse transcription using random pentadecamer primers increases yield and quality of resulting cDNA

    DEFF Research Database (Denmark)

    Stangegaard, Michael; Dufva, I.H.; Dufva, Hans Martin

    2006-01-01

    oligonucleotides (pentadecamers) consistently, yielded at least 2 fold as much cDNA as did random hexamers using either-poly(A) RNA or an amplified version of messenger RNA (aRNA) as a template. The cDNA generated using pentadecamers did not differ in size distribution or the amount of incorporated label compared...... with cDNA generated with random hexamers. The increased efficiency of priming using random pentadecamers resulted in reverse transcription of > 80% of the template aRNA, while random hexamers induced reverse transcription of only 40% of the template aRNA. This suggests a better coverage...... that random pentadecamers can replace random hexamers in reverse transcription reactions on both poly(A) RNA and amplified RNA, resulting in higher cDNA yields and quality....

  3. Development of Gene Expression Fingerprints for Identification of Environmental Contaminants Using cDNA Arrays

    National Research Council Canada - National Science Library

    Inouye, L

    2004-01-01

    ...) to develop cDNA array-based assays that map gene expression from contaminant exposures. Results substantiate that distinct gene expression profiles exist for major contaminant classes such as PARs, PCBs, and PCDD/Fs...

  4. 3G vector-primer plasmid for constructing full-length-enriched cDNA libraries.

    Science.gov (United States)

    Zheng, Dong; Zhou, Yanna; Zhang, Zidong; Li, Zaiyu; Liu, Xuedong

    2008-09-01

    We designed a 3G vector-primer plasmid for the generation of full-length-enriched complementary DNA (cDNA) libraries. By employing the terminal transferase activity of reverse transcriptase and the modified strand replacement method, this plasmid (assembled with a polydT end and a deoxyguanosine [dG] end) combines priming full-length cDNA strand synthesis and directional cDNA cloning. As a result, the number of steps involved in cDNA library preparation is decreased while simplifying downstream gene manipulation, sequencing, and subcloning. The 3G vector-primer plasmid method yields fully represented plasmid primed libraries that are equivalent to those made by the SMART (switching mechanism at 5' end of RNA transcript) approach.

  5. The spot market and the spot price: applicability and limitations

    International Nuclear Information System (INIS)

    White, G. Jr.

    1987-01-01

    The subject of spot prices and their relationship to long-term contracting is addressed. The author is associated with Nuexco, which originally was called the Nuclear Exchange Corporation. They use the term Exchange Value which originated in the idea that Nuexco operated an exchange 'bank' - those with too much uranium could 'bank it', those with short-term needs could borrow from the 'bank'. If the borrower repaid slightly more or less the difference was settled using the 'exchange value'. This became used for longer-term transactions and now settling the monthly value is an important part of Nuexco's activities. The exact nature of the Exchange Value is defined. Now more and more buyers are insisting on spot market related pricing even where this is not meaningfully related to uranium production costs. (U.K.)

  6. Cloning of cDNA encoding steroid 11β-hydroxylase (P450c11)

    International Nuclear Information System (INIS)

    Chua, S.C.; Szabo, P.; Vitek, A.; Grzeschik, K.H.; John, M.; White, P.C.

    1987-01-01

    The authors have isolated bovine and human adrenal cDNA clones encoding the adrenal cytochrome P-450 specific for 11β-hydroxylation (P450c11). A bovine adrenal cDNA library constructed in the bacteriophage λ vector gt10 was probed with a previously isolated cDNA clone corresponding to part of the 3' untranslated region of the 4.2-kilobase (kb) mRNA encoding P450c11. Several clones with 3.2-kb cDNA inserts were isolated. Sequence analysis showed that they overlapped the original probe by 300 base pairs (bp). Combined cDNA and RNA sequence data demonstrated a continuous open reading frame of 1509 bases. P450c11 is predicted to contain 479 amino acid residues in the mature protein in addition to a 24-residue amino-terminal mitochondrial signal sequence. A bovine clone was used to isolate a homologous clone with a 3.5-kb insert from a human adrenal cDNA library. A region of 1100 bp was 81% homologous to 769 bp of the coding sequence of the bovine cDNA except for a 400-bp segment presumed to be an unprocessed intron. Hybridization of the human cDNA to DNA from a panel of human-rodent somatic cell hybrid lines and in situ hybridization to metaphase spreads of human chromosomes localized the gene to the middle of the long arm of chromosome 8. These data should be useful in developing reagents for heterozygote detection and prenatal diagnosis of 11β-hydroxylase deficiency, the second most frequent cause of congenital adrenal hyperplasia

  7. CDNA cloning, characterization and expression of an endosperm-specific barley peroxidase

    DEFF Research Database (Denmark)

    Rasmussen, Søren Kjærsgård; Welinder, K.G.; Hejgaard, J.

    1991-01-01

    A barley peroxidase (BP 1) of pI ca. 8.5 and M(r) 37000 has been purified from mature barley grains. Using antibodies towards peroxidase BP 1, a cDNA clone (pcR7) was isolated from cDNA expression library. The nucleotide sequence of pcR7 gave a derived amino acid sequence identical to the 158 C...

  8. cDNA fingerprinting of osteoprogenitor cells to isolate differentiation stage-specific genes.

    OpenAIRE

    Candeliere, G A; Rao, Y; Floh, A; Sandler, S D; Aubin, J E

    1999-01-01

    A cDNA fingerprinting strategy was developed to identify genes based on their differential expression pattern during osteoblast development. Preliminary biological and molecular staging of cDNA pools prepared by global amplification PCR allowed discrim-inating choices to be made in selection of expressed sequence tags (ESTs) to be isolated. Sequencing of selected ESTs confirmed that both known and novel genes can be isolated from any developmental stage of interest, e.g. from primitive progen...

  9. cDNA cloning and mRNA expression of heat shock protein 70 gene ...

    African Journals Online (AJOL)

    In this study, the full-length heat shock protein 70 of Tegillarca granosa was cloned from cDNA library by rapid amplification of cDNA end (RACE). The open reading frame (ORF) of heat shock protein 70 was 1968 bp, and it encoded a protein of 655 amino acids with a predicted molecular weight of 71.48 kDa and an ...

  10. [Primary culture of cat intestinal epithelial cell and construction of its cDNA library].

    Science.gov (United States)

    Ye, L; Gui-Hua, Z; Kun, Y; Hong-Fa, W; Ting, X; Gong-Zhen, L; Wei-Xia, Z; Yong, C

    2017-04-12

    Objective To establish the primary cat intestinal epithelial cells (IECs) culture methods and construct the cDNA library for the following yeast two-hybrid experiment, so as to screen the virulence interaction factors among the final host. Methods The primary cat IECs were cultured by the tissue cultivation and combined digestion with collagenase XI and dispase I separately. Then the cat IECs cultured was identified with the morphological observation and cyto-keratin detection, by using goat anti-cyto-keratin monoclonal antibodies. The mRNA of cat IECs was isolated and used as the template to synthesize the first strand cDNA by SMART™ technology, and then the double-strand cDNAs were acquired by LD-PCR, which were subsequently cloned into the plasmid PGADT7-Rec to construct yeast two-hybrid cDNA library in the yeast strain Y187 by homologous recombination. Matchmaker™ Insert Check PCR was used to detect the size distribution of cDNA fragments after the capacity calculation of the cDNA library. Results The comparison of the two cultivation methods indicated that the combined digestion of collagenase XI and dispase I was more effective than the tissue cultivation. The cat IECs system of continuous culture was established and the cat IECs with high purity were harvested for constructing the yeast two-hybrid cDNA library. The library contained 1.1×10 6 independent clones. The titer was 2.8×10 9 cfu/ml. The size of inserted fragments was among 0.5-2.0 kb. Conclusion The yeast two-hybrid cDNA library of cat IECs meets the requirements of further screen research, and this study lays the foundation of screening the Toxoplasma gondii virulence interaction factors among the cDNA libraries of its final hosts.

  11. Laser Pyrometer For Spot Temperature Measurements

    Science.gov (United States)

    Elleman, D. D.; Allen, J. L.; Lee, M. C.

    1988-01-01

    Laser pyrometer makes temperature map by scanning measuring spot across target. Scanning laser pyrometer passively measures radiation emitted by scanned spot on target and calibrated by similar passive measurement on blackbody of known temperature. Laser beam turned on for active measurements of reflectances of target spot and reflectance standard. From measurements, temperature of target spot inferred. Pyrometer useful for non-contact measurement of temperature distributions in processing of materials.

  12. Laser Spot Detection Based on Reaction Diffusion

    OpenAIRE

    Alejandro Vázquez-Otero; Danila Khikhlukha; J. M. Solano-Altamirano; Raquel Dormido; Natividad Duro

    2016-01-01

    Center-location of a laser spot is a problem of interest when the laser is used for processing and performing measurements. Measurement quality depends on correctly determining the location of the laser spot. Hence, improving and proposing algorithms for the correct location of the spots are fundamental issues in laser-based measurements. In this paper we introduce a Reaction Diffusion (RD) system as the main computational framework for robustly finding laser spot centers. The method presente...

  13. The application of DNA microarrays in gene expression analysis.

    Science.gov (United States)

    van Hal, N L; Vorst, O; van Houwelingen, A M; Kok, E J; Peijnenburg, A; Aharoni, A; van Tunen, A J; Keijer, J

    2000-03-31

    DNA microarray technology is a new and powerful technology that will substantially increase the speed of molecular biological research. This paper gives a survey of DNA microarray technology and its use in gene expression studies. The technical aspects and their potential improvements are discussed. These comprise array manufacturing and design, array hybridisation, scanning, and data handling. Furthermore, it is discussed how DNA microarrays can be applied in the working fields of: safety, functionality and health of food and gene discovery and pathway engineering in plants.

  14. LEDGF/p75 Deficiency Increases Deletions at the HIV-1 cDNA Ends.

    Science.gov (United States)

    Bueno, Murilo T D; Reyes, Daniel; Llano, Manuel

    2017-09-15

    Processing of unintegrated linear HIV-1 cDNA by the host DNA repair system results in its degradation and/or circularization. As a consequence, deficient viral cDNA integration generally leads to an increase in the levels of HIV-1 cDNA circles containing one or two long terminal repeats (LTRs). Intriguingly, impaired HIV-1 integration in LEDGF/p75-deficient cells does not result in a correspondent increase in viral cDNA circles. We postulate that increased degradation of unintegrated linear viral cDNA in cells lacking the lens epithelium-derived growth factor (LEDGF/p75) account for this inconsistency. To evaluate this hypothesis, we characterized the nucleotide sequence spanning 2-LTR junctions isolated from LEDGF/p75-deficient and control cells. LEDGF/p75 deficiency resulted in a significant increase in the frequency of 2-LTRs harboring large deletions. Of note, these deletions were dependent on the 3' processing activity of integrase and were not originated by aberrant reverse transcription. Our findings suggest a novel role of LEDGF/p75 in protecting the unintegrated 3' processed linear HIV-1 cDNA from exonucleolytic degradation.

  15. Purification of Single-Stranded cDNA Based on RNA Degradation Treatment and Adsorption Chromatography.

    Science.gov (United States)

    Trujillo-Esquivel, Elías; Franco, Bernardo; Flores-Martínez, Alberto; Ponce-Noyola, Patricia; Mora-Montes, Héctor M

    2016-08-02

    Analysis of gene expression is a common research tool to study networks controlling gene expression, the role of genes with unknown function, and environmentally induced responses of organisms. Most of the analytical tools used to analyze gene expression rely on accurate cDNA synthesis and quantification to obtain reproducible and quantifiable results. Thus far, most commercial kits for isolation and purification of cDNA target double-stranded molecules, which do not accurately represent the abundance of transcripts. In the present report, we provide a simple and fast method to purify single-stranded cDNA, exhibiting high purity and yield. This method is based on the treatment with RNase H and RNase A after cDNA synthesis, followed by separation in silica spin-columns and ethanol precipitation. In addition, our method avoids the use of DNase I to eliminate genomic DNA from RNA preparations, which improves cDNA yield. As a case report, our method proved to be useful in the purification of single-stranded cDNA from the pathogenic fungus Sporothrix schenckii.

  16. Construction of a T7 Human Lung Cancer cDNA Library

    Directory of Open Access Journals (Sweden)

    Wentao YUE

    2008-10-01

    Full Text Available Background and objective Currently, only a limited numbers of tumor markers for non small lung cancer (NSCLC diagnosis, new biomarker, such as serum autoantibody may improve the early detection of lung cancer. Our objective is construction human lung squamous carcinoma and adenocarcinoma T7 phage display cDNA library from the tissues of NSCLC patients. Methods mRNA was isolated from a pool of total RNA extract from NSCLC tissues obtained from 5 adenocarcinomas and 5 squamous carcinomas, and then mRNA was reverse transcribed into double stranded cDNA. After digestion, the cDNA was inserted into T7Select 10-3 vector. The phage display cDNA library was constructed by package reaction in vitro and plate proliferation. Plaque assay and PCR were used to evaluate the library.Results Two T7 phage display cDNA library were established. Plaque assay show the titer of lung squamas carcinoma library was 1.8×106 pfu, and the adenocarcinoma library was 5×106 pfu. The phage titer of the amplified library were 3.2×1010 pfu/mL and 2.5×1010 pfu/mL. PCR amplification of random plaque show insert ratio were 100% (24/24 in adenocarcinoma library and 95.8% in human lung squamas carcinoma library (23/24. Insert range from 300 bp to 1 500 bp. Conclusion Two phage display cDNA library from NSCLC were constructed.

  17. Construction of Infectious cDNA Clone of a Chrysanthemum stunt viroid Korean Isolate

    Directory of Open Access Journals (Sweden)

    Ju-Yeon Yoon

    2014-03-01

    Full Text Available Chrysanthemum stunt viroid (CSVd, a noncoding infectious RNA molecule, causes seriously economic losses of chrysanthemum for 3 or 4 years after its first infection. Monomeric cDNA clones of CSVd isolate SK1 (CSVd-SK1 were constructed in the plasmids pGEM-T easy vector and pUC19 vector. Linear positive-sense transcripts synthesized in vitro from the full-length monomeric cDNA clones of CSVd-SK1 could infect systemically tomato seedlings and chrysanthemum plants, suggesting that the linear CSVd RNA transcribed from the cDNA clones could be replicated as efficiently as circular CSVd in host species. However, direct inoculation of plasmid cDNA clones containing full-length monomeric cDNA of CSVd-SK1 failed to infect tomato and chrysanthemum and linear negative-sense transcripts from the plasmid DNAs were not infectious in the two plant species. The cDNA sequences of progeny viroid in systemically infected tomato and chrysanthemum showed a few substitutions at a specific nucleotide position, but there were no deletions and insertions in the sequences of the CSVd progeny from tomato and chrysanthemum plants.

  18. 7 CFR 1421.11 - Spot checks.

    Science.gov (United States)

    2010-01-01

    ... 7 Agriculture 10 2010-01-01 2010-01-01 false Spot checks. 1421.11 Section 1421.11 Agriculture... ASSISTANCE LOANS AND LOAN DEFICIENCY PAYMENTS FOR 2008 THROUGH 2012 General § 1421.11 Spot checks. (a) CCC... CCC access to the farm and storage facility as necessary to conduct collateral inspections, or “spot...

  19. 21 CFR 886.1435 - Maxwell spot.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Maxwell spot. 886.1435 Section 886.1435 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES OPHTHALMIC DEVICES Diagnostic Devices § 886.1435 Maxwell spot. (a) Identification. A Maxwell spot is an AC...

  20. Tissue Microarray TechnologyA Brief Review

    Directory of Open Access Journals (Sweden)

    Ramya S Vokuda

    2018-01-01

    Full Text Available In this era of modern revolutionisation in the field of medical laboratory technology, everyone is aiming at taking the innovations from laboratory to bed side. One such technique that is most relevant to the pathologic community is Tissue Microarray (TMA technology. This is becoming quite popular amongst all the members of this family, right from laboratory scientists to clinicians and residents to technologists. The reason for this technique to gain popularity is attributed to its cost effectiveness and time saving protocols. Though, every technique is accompanied by disadvantages, the benefits out number them. This technique is very versatile as many downstream molecular assays such as immunohistochemistry, cytogenetic studies, Fluorescent In situ-Hybridisation (FISH etc., can be carried out on a single slide with multiple numbers of samples. It is a very practical approach that aids effectively to identify novel biomarkers in cancer diagnostics and therapeutics. It helps in assessing the molecular markers on a large scale very quickly. Also, the quality assurance protocols in pathological laboratory has exploited TMA to a great extent. However, the application of TMA technology is beyond oncology. This review shall focus on the different aspects of this technology such as construction of TMA, instrumentation, types, advantages and disadvantages and utilisation of the technique in various disease conditions.

  1. Tissue Microarray Analysis Applied to Bone Diagenesis.

    Science.gov (United States)

    Mello, Rafael Barrios; Silva, Maria Regina Regis; Alves, Maria Teresa Seixas; Evison, Martin Paul; Guimarães, Marco Aurelio; Francisco, Rafaella Arrabaca; Astolphi, Rafael Dias; Iwamura, Edna Sadayo Miazato

    2017-01-04

    Taphonomic processes affecting bone post mortem are important in forensic, archaeological and palaeontological investigations. In this study, the application of tissue microarray (TMA) analysis to a sample of femoral bone specimens from 20 exhumed individuals of known period of burial and age at death is described. TMA allows multiplexing of subsamples, permitting standardized comparative analysis of adjacent sections in 3-D and of representative cross-sections of a large number of specimens. Standard hematoxylin and eosin, periodic acid-Schiff and silver methenamine, and picrosirius red staining, and CD31 and CD34 immunohistochemistry were applied to TMA sections. Osteocyte and osteocyte lacuna counts, percent bone matrix loss, and fungal spheroid element counts could be measured and collagen fibre bundles observed in all specimens. Decalcification with 7% nitric acid proceeded more rapidly than with 0.5 M EDTA and may offer better preservation of histological and cellular structure. No endothelial cells could be detected using CD31 and CD34 immunohistochemistry. Correlation between osteocytes per lacuna and age at death may reflect reported age-related responses to microdamage. Methodological limitations and caveats, and results of the TMA analysis of post mortem diagenesis in bone are discussed, and implications for DNA survival and recovery considered.

  2. Transcriptome analysis of zebrafish embryogenesis using microarrays.

    Directory of Open Access Journals (Sweden)

    Sinnakaruppan Mathavan

    2005-08-01

    Full Text Available Zebrafish (Danio rerio is a well-recognized model for the study of vertebrate developmental genetics, yet at the same time little is known about the transcriptional events that underlie zebrafish embryogenesis. Here we have employed microarray analysis to study the temporal activity of developmentally regulated genes during zebrafish embryogenesis. Transcriptome analysis at 12 different embryonic time points covering five different developmental stages (maternal, blastula, gastrula, segmentation, and pharyngula revealed a highly dynamic transcriptional profile. Hierarchical clustering, stage-specific clustering, and algorithms to detect onset and peak of gene expression revealed clearly demarcated transcript clusters with maximum gene activity at distinct developmental stages as well as co-regulated expression of gene groups involved in dedicated functions such as organogenesis. Our study also revealed a previously unidentified cohort of genes that are transcribed prior to the mid-blastula transition, a time point earlier than when the zygotic genome was traditionally thought to become active. Here we provide, for the first time to our knowledge, a comprehensive list of developmentally regulated zebrafish genes and their expression profiles during embryogenesis, including novel information on the temporal expression of several thousand previously uncharacterized genes. The expression data generated from this study are accessible to all interested scientists from our institute resource database (http://giscompute.gis.a-star.edu.sg/~govind/zebrafish/data_download.html.

  3. Cloning and analysis of the mouse Fanconi anemia group A cDNA and an overlapping penta zinc finger cDNA.

    Science.gov (United States)

    Wong, J C; Alon, N; Norga, K; Kruyt, F A; Youssoufian, H; Buchwald, M

    2000-08-01

    Despite the cloning of four disease-associated genes for Fanconi anemia (FA), the molecular pathogenesis of FA remains largely unknown. To study FA complementation group A using the mouse as a model system, we cloned and characterized the mouse homolog of the human FANCA cDNA. The mouse cDNA (Fanca) encodes a 161-kDa protein that shares 65% amino acid sequence identity with human FANCA. Fanca is located at the distal region of mouse chromosome 8 and has a ubiquitous pattern of expression in embryonic and adult tissues. Expression of the mouse cDNA in human FA-A cells restores the cellular drug sensitivity to normal levels. Thus, the expression pattern, protein structure, chromosomal location, and function of FANCA are conserved in the mouse. We also isolated a novel zinc finger protein, Zfp276, which has five C(2)H(2) domains. Interestingly, Zfp276 is situated in the Fanca locus, and the 3'UTR of its cDNA overlaps with the last four exons of Fanca in a tail-to-tail manner. Zfp276 is expressed in the same tissues as Fanca, but does not complement the mitomycin C (MMC)-sensitive phenotype of FA-A cells. The overlapping genomic organization between Zfp276 and Fanca may have relevance to the disease phenotype of FA. Copyright 2000 Academic Press.

  4. A NASBA on microgel-tethered molecular-beacon microarray for real-time microbial molecular diagnostics.

    Science.gov (United States)

    Ma, Y; Dai, X; Hong, T; Munk, G B; Libera, M

    2016-12-19

    Despite their many advantages and successes, molecular beacon (MB) hybridization probes have not been extensively used in microarray formats because of the complicating probe-substrate interactions that increase the background intensity. We have previously shown that tethering to surface-patterned microgels is an effective means for localizing MB probes to specific surface locations in a microarray format while simultaneously maintaining them in as water-like an environment as possible and minimizing probe-surface interactions. Here we extend this approach to include both real-time detection together with integrated NASBA amplification. We fabricate small (∼250 μm × 250 μm) simplex, duplex, and five-plex assays with microarray spots of controllable size (∼20 μm diameter), position, and shape to detect bacteria and fungi in a bloodstream-infection model. The targets, primers, and microgel-tethered probes can be combined in a single isothermal reaction chamber with no post-amplification labelling. We extract total RNA from clinical blood samples and differentiate between Gram-positive and Gram-negative bloodstream infection in a duplex assay to detect RNA- amplicons. The sensitivity based on our current protocols in a simplex assay to detect specific ribosomal RNA sequences within total RNA extracted from S. aureus and E. coli cultures corresponds to tens of bacteria per ml. We furthermore show that the platform can detect RNA- amplicons from synthetic target DNA with 1 fM sensitivity in sample volumes that contain about 12 000 DNA molecules. These experiments demonstrate an alternative approach that can enable rapid and real-time microarray-based molecular diagnostics.

  5. Spot på samtalen:

    DEFF Research Database (Denmark)

    Danneris, Sophie; Jensen, Tanja Dall; Caswell, Dorte

    Spot på samtalen sætter fokus på det, der konkret foregår i samtaler mellem borgere og de beskæftigelsesfaglige medarbejdere i jobcentrene. Da de udsatte grupper i mange tilfælde er langt fra arbejdsmarkedet, er interessen rettet mod, hvilke forhold i kontakten med beskæftigelsessystemet, der...... har betydning hvilke indsatser ledige modtager, men også hvordan de modtager dem. Her rettes blikket mod den centrale del af den beskæftigelsespolitiske indsats som samtalerne udgør. I Spot på samtalen er blikket rettet mod de dynamikker, mønstre og mekanismer, der kommer i spil i samtalerne i...

  6. A Drosophila wing spot test

    International Nuclear Information System (INIS)

    Ayaki, Toshikazu; Yoshikawa, Isao; Niikawa, Norio; Hoshi, Masaharu.

    1986-01-01

    A Drosophila wing spot test system was used to investigate the effects of low doses of X-rays, gamma rays, and both 2.3 and 14.1 MeV neutrons on somatic chromosome mutation (SCM) induction. The incidence of SCM was significantly increased with any type of radiation, with evident linear dose-response relationship within the range of 3 to 20 cGy. It was estimated that relative biological effectiveness value for SCM induction of 2.3 MeV neutrons to X-rays and gamma rays is much higher than that of 14.1 MeV neutrons to those photons (2.4 vs 8.0). The Drosophila wing spot test system seems to become a promising in vivo experimental method for higher animals in terms of the lack of necessity for a marvelously large number of materials required in conventional test system. (Namekawa, K.)

  7. Managing Rocky Mountain spotted fever.

    Science.gov (United States)

    Minniear, Timothy D; Buckingham, Steven C

    2009-11-01

    Rocky Mountain spotted fever is caused by the tick-borne bacterium Rickettsia rickettsii. Symptoms range from moderate illness to severe illness, including cardiovascular compromise, coma and death. The disease is prevalent in most of the USA, especially during warmer months. The trademark presentation is fever and rash with a history of tick bite, although tick exposure is unappreciated in over a third of cases. Other signature symptoms include headache and abdominal pain. The antibiotic therapy of choice for R. rickettsii infection is doxycycline. Preventive measures for Rocky Mountain spotted fever and other tick-borne diseases include: wearing long-sleeved, light colored clothing; checking for tick attachment and removing attached ticks promptly; applying topical insect repellent; and treating clothing with permethrin.

  8. Measurement of laser spot quality

    Science.gov (United States)

    Milster, T. D.; Treptau, J. P.

    1991-01-01

    Several ways of measuring spot quality are compared. We examine in detail various figures of merit such as full width at half maximum (FWHM), full width at 1/(e exp 2) maximum, Strehl ratio, and encircled energy. Our application is optical data storage, but results can be applied to other areas like space communications and high energy lasers. We found that the optimum figure of merit in many cases is Strehl ratio.

  9. Sweet Spots and Door Stops

    Science.gov (United States)

    Thompson, Michael; Tsui, Stella; Leung, Chi Fan

    2011-01-01

    A sweet spot is referred to in sport as the perfect place to strike a ball with a racquet or bat. It is the point of contact between bat and ball where maximum results can be produced with minimal effort from the hand of the player. Similar physics can be applied to the less inspiring examples of door stops; the perfect position of a door stop is…

  10. Glare Spot Phase Doppler Anemometry

    OpenAIRE

    Hespel, Camille; Ren, Kuan Fang; Gréhan, Gérard; Onofri, Fabrice

    2006-01-01

    International audience; The Phase Doppler anemometry has been developed to measure simultaneously the velocity and the size of droplets. The measurement of the refractive index is also necessary since it depends on the temperature and the composition of the particle and its measurement permits both to increase the quality of the diameter measurement and to obtain information on the temperature and/or the composition of the droplets. In this paper, we introduce a Glare Spot Phase Doppler Anemo...

  11. Cell-Based Microarrays for In Vitro Toxicology

    Science.gov (United States)

    Wegener, Joachim

    2015-07-01

    DNA/RNA and protein microarrays have proven their outstanding bioanalytical performance throughout the past decades, given the unprecedented level of parallelization by which molecular recognition assays can be performed and analyzed. Cell microarrays (CMAs) make use of similar construction principles. They are applied to profile a given cell population with respect to the expression of specific molecular markers and also to measure functional cell responses to drugs and chemicals. This review focuses on the use of cell-based microarrays for assessing the cytotoxicity of drugs, toxins, or chemicals in general. It also summarizes CMA construction principles with respect to the cell types that are used for such microarrays, the readout parameters to assess toxicity, and the various formats that have been established and applied. The review ends with a critical comparison of CMAs and well-established microtiter plate (MTP) approaches.

  12. High throughput screening of starch structures using carbohydrate microarrays

    DEFF Research Database (Denmark)

    Tanackovic, Vanja; Rydahl, Maja Gro; Pedersen, Henriette Lodberg

    2016-01-01

    In this study we introduce the starch-recognising carbohydrate binding module family 20 (CBM20) from Aspergillus niger for screening biological variations in starch molecular structure using high throughput carbohydrate microarray technology. Defined linear, branched and phosphorylated...

  13. Microarray of DNA probes on carboxylate functional beads surface

    Institute of Scientific and Technical Information of China (English)

    黄承志; 李原芳; 黄新华; 范美坤

    2000-01-01

    The microarray of DNA probes with 5’ -NH2 and 5’ -Tex/3’ -NH2 modified terminus on 10 um carboxylate functional beads surface in the presence of 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC) is characterized in the preseni paper. it was found that the microarray capacity of DNA probes on the beads surface depends on the pH of the aqueous solution, the concentra-tion of DNA probe and the total surface area of the beads. On optimal conditions, the minimum distance of 20 mer single-stranded DNA probe microarrayed on beads surface is about 14 nm, while that of 20 mer double-stranded DNA probes is about 27 nm. If the probe length increases from 20 mer to 35 mer, its microarray density decreases correspondingly. Mechanism study shows that the binding mode of DNA probes on the beads surface is nearly parallel to the beads surface.

  14. Microarray of DNA probes on carboxylate functional beads surface

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    The microarray of DNA probes with 5′-NH2 and 5′-Tex/3′-NH2 modified terminus on 10 m m carboxylate functional beads surface in the presence of 1-ethyl-3-(3-dimethylaminopropyl)- carbodiimide (EDC) is characterized in the present paper. It was found that the microarray capacity of DNA probes on the beads surface depends on the pH of the aqueous solution, the concentration of DNA probe and the total surface area of the beads. On optimal conditions, the minimum distance of 20 mer single-stranded DNA probe microarrayed on beads surface is about 14 nm, while that of 20 mer double-stranded DNA probes is about 27 nm. If the probe length increases from 20 mer to 35 mer, its microarray density decreases correspondingly. Mechanism study shows that the binding mode of DNA probes on the beads surface is nearly parallel to the beads surface.

  15. Rapid Diagnosis of Bacterial Meningitis Using a Microarray

    Directory of Open Access Journals (Sweden)

    Ren-Jy Ben

    2008-06-01

    Conclusion: The microarray method provides a more accurate and rapid diagnostic tool for bacterial meningitis compared to traditional culture methods. Clinical application of this new technique may reduce the potential risk of delay in treatment.

  16. Variance estimation in the analysis of microarray data

    KAUST Repository

    Wang, Yuedong; Ma, Yanyuan; Carroll, Raymond J.

    2009-01-01

    Microarrays are one of the most widely used high throughput technologies. One of the main problems in the area is that conventional estimates of the variances that are required in the t-statistic and other statistics are unreliable owing

  17. Novel Protein Microarray Technology to Examine Men with Prostate Cancer

    National Research Council Canada - National Science Library

    Lilja, Hans

    2005-01-01

    The authors developed a novel macro and nanoporous silicon surface for protein microarrays to facilitate high-throughput biomarker discovery, and high-density protein-chip array analyses of complex biological samples...

  18. Justifications shape ethical blind spots.

    Science.gov (United States)

    Pittarello, Andrea; Leib, Margarita; Gordon-Hecker, Tom; Shalvi, Shaul

    2015-06-01

    To some extent, unethical behavior results from people's limited attention to ethical considerations, which results in an ethical blind spot. Here, we focus on the role of ambiguity in shaping people's ethical blind spots, which in turn lead to their ethical failures. We suggest that in ambiguous settings, individuals' attention shifts toward tempting information, which determines the magnitude of their lies. Employing a novel ambiguous-dice paradigm, we asked participants to report the outcome of the die roll appearing closest to the location of a previously presented fixation cross on a computer screen; this outcome would determine their pay. We varied the value of the die second closest to the fixation cross to be either higher (i.e., tempting) or lower (i.e., not tempting) than the die closest to the fixation cross. Results of two experiments revealed that in ambiguous settings, people's incorrect responses were self-serving. Tracking participants' eye movements demonstrated that people's ethical blind spots are shaped by increased attention toward tempting information. © The Author(s) 2015.

  19. Efficacy Testing of H56 cDNA Tattoo Immunization against Tuberculosis in a Mouse Model.

    Science.gov (United States)

    Platteel, Anouk C M; Nieuwenhuizen, Natalie E; Domaszewska, Teresa; Schürer, Stefanie; Zedler, Ulrike; Brinkmann, Volker; Sijts, Alice J A M; Kaufmann, Stefan H E

    2017-01-01

    Tuberculosis (TB), caused by Mycobacterium tuberculosis ( Mtb ), remains a global threat. The only approved vaccine against TB, Mycobacterium bovis bacillus Calmette-Guérin (BCG), provides insufficient protection and, being a live vaccine, can cause disseminated disease in immunocompromised individuals. Previously, we found that intradermal cDNA tattoo immunization with cDNA of tetanus toxoid fragment C domain 1 fused to cDNA of the fusion protein H56, comprising the Mtb antigens Ag85B, ESAT-6, and Rv2660c, induced antigen-specific CD8 + T cell responses in vivo . As cDNA tattoo immunization would be safer than a live vaccine in immunocompromised patients, we tested the protective efficacy of intradermal tattoo immunization against TB with H56 cDNA, as well as with H56_E, a construct optimized for epitope processing in a mouse model. As Mtb antigens can be used in combination with BCG to boost immune responses, we also tested the protective efficacy of heterologous prime-boost, using dermal tattoo immunization with H56_E cDNA to boost BCG immunization in mice. Dermal H56 and H56_E cDNA immunization induced H56-specific CD4 + and CD8 + T cell responses and Ag85B-specific IgG antibodies, but did not reduce bacterial loads, although immunization with H56_E ameliorated lung pathology. Both subcutaneous and intradermal immunization with BCG resulted in broad cellular immune responses, with increased frequencies of CD4 + T effector memory cells, T follicular helper cells, and germinal center B cells, and resulted in reduced bacterial loads and lung pathology. Heterologous vaccination with BCG/H56_E cDNA induced increased H56-specific CD4 + and CD8 + T cell cytokine responses compared to vaccination with BCG alone, and lung pathology was significantly decreased in BCG/H56_E cDNA immunized mice compared to unvaccinated controls. However, bacterial loads were not decreased after heterologous vaccination compared to BCG alone. CD4 + T cells responding to Ag85B- and ESAT-6

  20. Resistance Spot Welding of dissimilar Steels

    Directory of Open Access Journals (Sweden)

    Ladislav Kolařík

    2012-01-01

    Full Text Available This paper presents an analysis of the properties of resistance spot welds between low carbon steel and austenitic CrNi stainless steel. The thickness of the welded dissimilar materials was 2 mm. A DeltaSpot welding gun with a process tape was used for welding the dissimilar steels. Resistance spot welds were produced with various welding parameters (welding currents ranging from 7 to 8 kA. Light microscopy, microhardness measurements across the welded joints, and EDX analysis were used to evaluate the quality of the resistance spot welds. The results confirm the applicability of DeltaSpot welding for this combination of materials.

  1. Addressable droplet microarrays for single cell protein analysis.

    Science.gov (United States)

    Salehi-Reyhani, Ali; Burgin, Edward; Ces, Oscar; Willison, Keith R; Klug, David R

    2014-11-07

    Addressable droplet microarrays are potentially attractive as a way to achieve miniaturised, reduced volume, high sensitivity analyses without the need to fabricate microfluidic devices or small volume chambers. We report a practical method for producing oil-encapsulated addressable droplet microarrays which can be used for such analyses. To demonstrate their utility, we undertake a series of single cell analyses, to determine the variation in copy number of p53 proteins in cells of a human cancer cell line.

  2. Microarrays for Universal Detection and Identification of Phytoplasmas

    DEFF Research Database (Denmark)

    Nicolaisen, Mogens; Nyskjold, Henriette; Bertaccini, Assunta

    2013-01-01

    Detection and identification of phytoplasmas is a laborious process often involving nested PCR followed by restriction enzyme analysis and fine-resolution gel electrophoresis. To improve throughput, other methods are needed. Microarray technology offers a generic assay that can potentially detect...... and differentiate all types of phytoplasmas in one assay. The present protocol describes a microarray-based method for identification of phytoplasmas to 16Sr group level....

  3. Spot formation of radiation particles by electrochemical etching

    International Nuclear Information System (INIS)

    Nozaki, Tetsuya

    1999-01-01

    An electrochemical etching (ECE) spot formation from the top of chemical etching (CE) spot was confirmed by a series of experiments. One of polycarbonate (Iupilon) could not make the spot, because ECE spot had grown up before the microscope confirming the CE spot. Clear CEC spots by α-ray and neutron were found on Harzlas and Baryotrak, both improvements of CR-39. Under the same etching conditions, the growth of ECE spot on Harzlas was more rapid than Baryotrak, but both spots were almost the same. All CE spot by α-ray produced the CEC spots, but a part of CE circle spot by neutron formed them. (S.Y.)

  4. A comparative analysis of DNA barcode microarray feature size

    Directory of Open Access Journals (Sweden)

    Smith Andrew M

    2009-10-01

    Full Text Available Abstract Background Microarrays are an invaluable tool in many modern genomic studies. It is generally perceived that decreasing the size of microarray features leads to arrays with higher resolution (due to greater feature density, but this increase in resolution can compromise sensitivity. Results We demonstrate that barcode microarrays with smaller features are equally capable of detecting variation in DNA barcode intensity when compared to larger feature sizes within a specific microarray platform. The barcodes used in this study are the well-characterized set derived from the Yeast KnockOut (YKO collection used for screens of pooled yeast (Saccharomyces cerevisiae deletion mutants. We treated these pools with the glycosylation inhibitor tunicamycin as a test compound. Three generations of barcode microarrays at 30, 8 and 5 μm features sizes independently identified the primary target of tunicamycin to be ALG7. Conclusion We show that the data obtained with 5 μm feature size is of comparable quality to the 30 μm size and propose that further shrinking of features could yield barcode microarrays with equal or greater resolving power and, more importantly, higher density.

  5. Assessing Bacterial Interactions Using Carbohydrate-Based Microarrays

    Directory of Open Access Journals (Sweden)

    Andrea Flannery

    2015-12-01

    Full Text Available Carbohydrates play a crucial role in host-microorganism interactions and many host glycoconjugates are receptors or co-receptors for microbial binding. Host glycosylation varies with species and location in the body, and this contributes to species specificity and tropism of commensal and pathogenic bacteria. Additionally, bacterial glycosylation is often the first bacterial molecular species encountered and responded to by the host system. Accordingly, characterising and identifying the exact structures involved in these critical interactions is an important priority in deciphering microbial pathogenesis. Carbohydrate-based microarray platforms have been an underused tool for screening bacterial interactions with specific carbohydrate structures, but they are growing in popularity in recent years. In this review, we discuss carbohydrate-based microarrays that have been profiled with whole bacteria, recombinantly expressed adhesins or serum antibodies. Three main types of carbohydrate-based microarray platform are considered; (i conventional carbohydrate or glycan microarrays; (ii whole mucin microarrays; and (iii microarrays constructed from bacterial polysaccharides or their components. Determining the nature of the interactions between bacteria and host can help clarify the molecular mechanisms of carbohydrate-mediated interactions in microbial pathogenesis, infectious disease and host immune response and may lead to new strategies to boost therapeutic treatments.

  6. Computational prediction of protein hot spot residues.

    Science.gov (United States)

    Morrow, John Kenneth; Zhang, Shuxing

    2012-01-01

    Most biological processes involve multiple proteins interacting with each other. It has been recently discovered that certain residues in these protein-protein interactions, which are called hot spots, contribute more significantly to binding affinity than others. Hot spot residues have unique and diverse energetic properties that make them challenging yet important targets in the modulation of protein-protein complexes. Design of therapeutic agents that interact with hot spot residues has proven to be a valid methodology in disrupting unwanted protein-protein interactions. Using biological methods to determine which residues are hot spots can be costly and time consuming. Recent advances in computational approaches to predict hot spots have incorporated a myriad of features, and have shown increasing predictive successes. Here we review the state of knowledge around protein-protein interactions, hot spots, and give an overview of multiple in silico prediction techniques of hot spot residues.

  7. Computational Prediction of Hot Spot Residues

    Science.gov (United States)

    Morrow, John Kenneth; Zhang, Shuxing

    2013-01-01

    Most biological processes involve multiple proteins interacting with each other. It has been recently discovered that certain residues in these protein-protein interactions, which are called hot spots, contribute more significantly to binding affinity than others. Hot spot residues have unique and diverse energetic properties that make them challenging yet important targets in the modulation of protein-protein complexes. Design of therapeutic agents that interact with hot spot residues has proven to be a valid methodology in disrupting unwanted protein-protein interactions. Using biological methods to determine which residues are hot spots can be costly and time consuming. Recent advances in computational approaches to predict hot spots have incorporated a myriad of features, and have shown increasing predictive successes. Here we review the state of knowledge around protein-protein interactions, hot spots, and give an overview of multiple in silico prediction techniques of hot spot residues. PMID:22316154

  8. cDNA sequencing improves the detection of P53 missense mutations in colorectal cancer

    International Nuclear Information System (INIS)

    Szybka, Malgorzata; Kordek, Radzislaw; Zakrzewska, Magdalena; Rieske, Piotr; Pasz-Walczak, Grazyna; Kulczycka-Wojdala, Dominika; Zawlik, Izabela; Stawski, Robert; Jesionek-Kupnicka, Dorota; Liberski, Pawel P

    2009-01-01

    Recently published data showed discrepancies beteween P53 cDNA and DNA sequencing in glioblastomas. We hypothesised that similar discrepancies may be observed in other human cancers. To this end, we analyzed 23 colorectal cancers for P53 mutations and gene expression using both DNA and cDNA sequencing, real-time PCR and immunohistochemistry. We found P53 gene mutations in 16 cases (15 missense and 1 nonsense). Two of the 15 cases with missense mutations showed alterations based only on cDNA, and not DNA sequencing. Moreover, in 6 of the 15 cases with a cDNA mutation those mutations were difficult to detect in the DNA sequencing, so the results of DNA analysis alone could be misinterpreted if the cDNA sequencing results had not also been available. In all those 15 cases, we observed a higher ratio of the mutated to the wild type template by cDNA analysis, but not by the DNA analysis. Interestingly, a similar overexpression of P53 mRNA was present in samples with and without P53 mutations. In terms of colorectal cancer, those discrepancies might be explained under three conditions: 1, overexpression of mutated P53 mRNA in cancer cells as compared with normal cells; 2, a higher content of cells without P53 mutation (normal cells and cells showing K-RAS and/or APC but not P53 mutation) in samples presenting P53 mutation; 3, heterozygous or hemizygous mutations of P53 gene. Additionally, for heterozygous mutations unknown mechanism(s) causing selective overproduction of mutated allele should also be considered. Our data offer new clues for studying discrepancy in P53 cDNA and DNA sequencing analysis

  9. ESA uncovers Geminga's `hot spot'

    Science.gov (United States)

    2004-07-01

    16 July 2004 Astronomers using ESA’s X-ray observatory XMM-Newton have detected a small, bright ‘hot spot’ on the surface of the neutron star called Geminga, 500 light-years away. The hot spot is the size of a football field and is caused by the same mechanism producing Geminga’s X-ray tails. This discovery identifies the missing link between the X-ray and gamma-ray emission from Geminga. hi-res Size hi-res: 1284 kb Credits: ESA, P. Caraveo (IASF, Milan) Geminga's hot spot This figure shows the effects of charged particles accelerated in the magnetosphere of Geminga. Panel (a) shows an image taken with the EPIC instrument on board the XMM-Newton observatory. The bright tails, made of particles kicked out by Geminga’s strong magnetic field, trail the neutron star as it moves about in space. Panel (b) shows how electrically charged particles interact with Geminga’s magnetic field. For example, if electrons (blue) are kicked out by the star, positrons (in red) hit the star’s magnetic poles like in an ‘own goal’. Panel (c) illustrates the size of Geminga’s magnetic field (blue) compared to that of the star itself at the centre (purple). The magnetic field is tilted with respect to Geminga’s rotation axis (red). Panel (d) shows the magnetic poles of Geminga, where charged particles hit the surface of the star, creating a two-million degrees hot spot, a region much hotter than the surroundings. As the star spins on its rotation axis, the hot spot comes into view and then disappears, causing the periodic colour change seen by XMM-Newton. An animated version of the entire sequence can be found at: Click here for animated GIF [low resolution, animated GIF, 5536 KB] Click here for AVI [high resolution, AVI with DIVX compression, 19128 KB] hi-res Size hi-res: 371 kb Credits: ESA, P. Caraveo (IASF, Milan) Geminga's hot spot, panel (a) Panel (a) shows an image taken with the EPIC instrument on board the XMM-Newton observatory. The bright tails, made of

  10. Microarray profiling of mononuclear peripheral blood cells identifies novel candidate genes related to chemoradiation response in rectal cancer.

    Directory of Open Access Journals (Sweden)

    Pablo Palma

    Full Text Available Preoperative chemoradiation significantly improves oncological outcome in locally advanced rectal cancer. However there is no effective method of predicting tumor response to chemoradiation in these patients. Peripheral blood mononuclear cells have emerged recently as pathology markers of cancer and other diseases, making possible their use as therapy predictors. Furthermore, the importance of the immune response in radiosensivity of solid organs led us to hypothesized that microarray gene expression profiling of peripheral blood mononuclear cells could identify patients with response to chemoradiation in rectal cancer. Thirty five 35 patients with locally advanced rectal cancer were recruited initially to perform the study. Peripheral blood samples were obtained before neaodjuvant treatment. RNA was extracted and purified to obtain cDNA and cRNA for hybridization of microarrays included in Human WG CodeLink bioarrays. Quantitative real time PCR was used to validate microarray experiment data. Results were correlated with pathological response, according to Mandard´s criteria and final UICC Stage (patients with tumor regression grade 1-2 and downstaging being defined as responders and patients with grade 3-5 and no downstaging as non-responders. Twenty seven out of 35 patients were finally included in the study. We performed a multiple t-test using Significance Analysis of Microarrays, to find those genes differing significantly in expression, between responders (n = 11 and non-responders (n = 16 to CRT. The differently expressed genes were: BC 035656.1, CIR, PRDM2, CAPG, FALZ, HLA-DPB2, NUPL2, and ZFP36. The measurement of FALZ (p = 0.029 gene expression level determined by qRT-PCR, showed statistically significant differences between the two groups. Gene expression profiling reveals novel genes in peripheral blood samples of mononuclear cells that could predict responders and non-responders to chemoradiation in patients with

  11. cDNA cloning, sequence analysis, and chromosomal localization of the gene for human carnitine palmitoyltransferase

    International Nuclear Information System (INIS)

    Finocchiaro, G.; Taroni, F.; Martin, A.L.; Colombo, I.; Tarelli, G.T.; DiDonato, S.; Rocchi, M.

    1991-01-01

    The authors have cloned and sequenced a cDNA encoding human liver carnitine palmitoyltransferase an inner mitochondrial membrane enzyme that plays a major role in the fatty acid oxidation pathway. Mixed oligonucleotide primers whose sequences were deduced from one tryptic peptide obtained from purified CPTase were used in a polymerase chain reaction, allowing the amplification of a 0.12-kilobase fragment of human genomic DNA encoding such a peptide. A 60-base-pair (bp) oligonucleotide synthesized on the basis of the sequence from this fragment was used for the screening of a cDNA library from human liver and hybridized to a cDNA insert of 2255 bp. This cDNA contains an open reading frame of 1974 bp that encodes a protein of 658 amino acid residues including 25 residues of an NH 2 -terminal leader peptide. The assignment of this open reading frame to human liver CPTase is confirmed by matches to seven different amino acid sequences of tryptic peptides derived from pure human CPTase and by the 82.2% homology with the amino acid sequence of rat CPTase. The NH 2 -terminal region of CPTase contains a leucine-proline motif that is shared by carnitine acetyl- and octanoyltransferases and by choline acetyltransferase. The gene encoding CPTase was assigned to human chromosome 1, region 1q12-1pter, by hybridization of CPTase cDNA with a DNA panel of 19 human-hanster somatic cell hybrids

  12. Molecular cloning and nucleotide sequence of cDNA for human liver arginase

    International Nuclear Information System (INIS)

    Haraguchi, Y.; Takiguchi, M.; Amaya, Y.; Kawamoto, S.; Matsuda, I.; Mori, M.

    1987-01-01

    Arginase (EC3.5.3.1) catalyzes the last step of the urea cycle in the liver of ureotelic animals. Inherited deficiency of the enzyme results in argininemia, an autosomal recessive disorder characterized by hyperammonemia. To facilitate investigation of the enzyme and gene structures and to elucidate the nature of the mutation in argininemia, the authors isolated cDNA clones for human liver arginase. Oligo(dT)-primed and random primer human liver cDNA libraries in λ gt11 were screened using isolated rat arginase cDNA as a probe. Two of the positive clones, designated λ hARG6 and λ hARG109, contained an overlapping cDNA sequence with an open reading frame encoding a polypeptide of 322 amino acid residues (predicted M/sub r/, 34,732), a 5'-untranslated sequence of 56 base pairs, a 3'-untranslated sequence of 423 base pairs, and a poly(A) segment. Arginase activity was detected in Escherichia coli cells transformed with the plasmid carrying λ hARG6 cDNA insert. RNA gel blot analysis of human liver RNA showed a single mRNA of 1.6 kilobases. The predicted amino acid sequence of human liver arginase is 87% and 41% identical with those of the rat liver and yeast enzymes, respectively. There are several highly conserved segments among the human, rat, and yeast enzymes

  13. Sequence of a cDNA encoding turtle high mobility group 1 protein.

    Science.gov (United States)

    Zheng, Jifang; Hu, Bi; Wu, Duansheng

    2005-07-01

    In order to understand sequence information about turtle HMG1 gene, a cDNA encoding HMG1 protein of the Chinese soft-shell turtle (Pelodiscus sinensis) was amplified by RT-PCR from kidney total RNA, and was cloned, sequenced and analyzed. The results revealed that the open reading frame (ORF) of turtle HMG1 cDNA is 606 bp long. The ORF codifies 202 amino acid residues, from which two DNA-binding domains and one polyacidic region are derived. The DNA-binding domains share higher amino acid identity with homologues sequences of chicken (96.5%) and mammalian (74%) than homologues sequence of rainbow trout (67%). The polyacidic region shows 84.6% amino acid homology with the equivalent region of chicken HMG1 cDNA. Turtle HMG1 protein contains 3 Cys residues located at completely conserved positions. Conservation in sequence and structure suggests that the functions of turtle HMG1 cDNA may be highly conserved during evolution. To our knowledge, this is the first report of HMG1 cDNA sequence in any reptilian.

  14. Watermarking spot colors in packaging

    Science.gov (United States)

    Reed, Alastair; Filler, TomáÅ.¡; Falkenstern, Kristyn; Bai, Yang

    2015-03-01

    In January 2014, Digimarc announced Digimarc® Barcode for the packaging industry to improve the check-out efficiency and customer experience for retailers. Digimarc Barcode is a machine readable code that carries the same information as a traditional Universal Product Code (UPC) and is introduced by adding a robust digital watermark to the package design. It is imperceptible to the human eye but can be read by a modern barcode scanner at the Point of Sale (POS) station. Compared to a traditional linear barcode, Digimarc Barcode covers the whole package with minimal impact on the graphic design. This significantly improves the Items per Minute (IPM) metric, which retailers use to track the checkout efficiency since it closely relates to their profitability. Increasing IPM by a few percent could lead to potential savings of millions of dollars for retailers, giving them a strong incentive to add the Digimarc Barcode to their packages. Testing performed by Digimarc showed increases in IPM of at least 33% using the Digimarc Barcode, compared to using a traditional barcode. A method of watermarking print ready image data used in the commercial packaging industry is described. A significant proportion of packages are printed using spot colors, therefore spot colors needs to be supported by an embedder for Digimarc Barcode. Digimarc Barcode supports the PANTONE spot color system, which is commonly used in the packaging industry. The Digimarc Barcode embedder allows a user to insert the UPC code in an image while minimizing perceptibility to the Human Visual System (HVS). The Digimarc Barcode is inserted in the printing ink domain, using an Adobe Photoshop plug-in as the last step before printing. Since Photoshop is an industry standard widely used by pre-press shops in the packaging industry, a Digimarc Barcode can be easily inserted and proofed.

  15. Collaboration spotting for dental science.

    Science.gov (United States)

    Leonardi, E; Agocs, A; Fragkiskos, S; Kasfikis, N; Le Goff, J M; Cristalli, M P; Luzzi, V; Polimeni, A

    2014-10-06

    The goal of the Collaboration Spotting project is to create an automatic system to collect information about publications and patents related to a given technology, to identify the key players involved, and to highlight collaborations and related technologies. The collected information can be visualized in a web browser as interactive graphical maps showing in an intuitive way the players and their collaborations (Sociogram) and the relations among the technologies (Technogram). We propose to use the system to study technologies related to Dental Science. In order to create a Sociogram, we create a logical filter based on a set of keywords related to the technology under study. This filter is used to extract a list of publications from the Web of Science™ database. The list is validated by an expert in the technology and sent to CERN where it is inserted in the Collaboration Spotting database. Here, an automatic software system uses the data to generate the final maps. We studied a set of recent technologies related to bone regeneration procedures of oro--maxillo--facial critical size defects, namely the use of Porous HydroxyApatite (HA) as a bone substitute alone (bone graft) or as a tridimensional support (scaffold) for insemination and differentiation ex--vivo of Mesenchymal Stem Cells. We produced the Sociograms for these technologies and the resulting maps are now accessible on--line. The Collaboration Spotting system allows the automatic creation of interactive maps to show the current and historical state of research on a specific technology. These maps are an ideal tool both for researchers who want to assess the state--of--the--art in a given technology, and for research organizations who want to evaluate their contribution to the technological development in a given field. We demonstrated that the system can be used for Dental Science and produced the maps for an initial set of technologies in this field. We now plan to enlarge the set of mapped

  16. Collaboration Spotting for oral medicine.

    Science.gov (United States)

    Leonardi, E; Agocs, A; Fragkiskos, S; Kasfikis, N; Le Goff, J M; Cristalli, M P; Luzzi, V; Polimeni, A

    2014-09-01

    The goal of the Collaboration Spotting project is to create an automatic system to collect information about publications and patents related to a given technology, to identify the key players involved, and to highlight collaborations and related technologies. The collected information can be visualized in a web browser as interactive graphical maps showing in an intuitive way the players and their collaborations (Sociogram) and the relations among the technologies (Technogram). We propose to use the system to study technologies related to oral medicine. In order to create a sociogram, we create a logical filter based on a set of keywords related to the technology under study. This filter is used to extract a list of publications from the Web of Science™ database. The list is validated by an expert in the technology and sent to CERN where it is inserted in the Collaboration Spotting database. Here, an automatic software system uses the data to generate the final maps. We studied a set of recent technologies related to bone regeneration procedures of oro-maxillo-facial critical size defects, namely the use of porous hydroxyapatite (HA) as a bone substitute alone (bone graft) or as a tridimensional support (scaffold) for insemination and differentiation ex vivo of mesenchymal stem cells. We produced the sociograms for these technologies and the resulting maps are now accessible on-line. The Collaboration Spotting system allows the automatic creation of interactive maps to show the current and historical state of research on a specific technology. These maps are an ideal tool both for researchers who want to assess the state-of-the-art in a given technology, and for research organizations who want to evaluate their contribution to the technological development in a given field. We demonstrated that the system can be used in oral medicine as is produced the maps for an initial set of technologies in this field. We now plan to enlarge the set of mapped technologies in

  17. Energy is not Coffee. An assessment of blind spots on energy spot-markets

    International Nuclear Information System (INIS)

    Jepma, C.J.; Spijker, E.; Van der Gaast, W.; De Jong, F.; Overmars, P.

    2006-01-01

    This study was to be the first in a series of studies on the title subject. It specifically focuses on the differences and similarities with a number of other spot-markets and aims to frame the energy spot markets and their potential development into a broader perspective. Main conclusion is that energy spot-markets differ from several other physical and non-physical spot-markets in many ways. This implies that 'perfect' energy spot-markets may inherently be (much) less perfect than other spot-markets that have approximated the stage of theoretical perfection

  18. Analysis of human HPRT- deletion mutants by the microarray-CGH (comparative genomic hybridization)

    International Nuclear Information System (INIS)

    Kodaira, M.; Sasaki, K.; Tagawa, H.; Omine, H.; Kushiro, J.; Takahashi, N.; Katayama, H.

    2003-01-01

    We are trying to evaluate genetic effects of radiation on human using mutation frequency as an indicator. For the efficient detection of mutations, it is important to understand the mechanism and the characteristics of radiation-induced mutations. We have started the analysis of hypoxanthine-guanine phosphoribosyl transferase (HPRT) mutants induced by X-ray in order to clarify the deletion size and the mutation-distribution. We analyzed 39 human X-ray induced HPRT-deletion mutants by using the microarray-CGH. The array for this analysis contains 57 BAC clones covering as much as possible of the 4Mb of the 5' side and 10Mb of the 3' side of the HPRT gene based on the NCBI genome database. DNA from parent strain and each HPRT-mutant strain are labeled with Cy5 and Cy3 respectively, and were mixed and hybridized on the array. Fluorescent intensity ratio of the obtained spots was analyzed using software we developed to identify clones corresponding to the deletion region. The deletion in these strains ranged up to 3.5 Mb on the 5' side and 6 Mb on the 3' side of the HPRT gene. Deletions in 13 strains ended around BAC clones located at about 3 Mb on the 5' side. On the 3' side, deletions extended up to the specific clones located at 1.5 Mb in 11 strains. The mutations seem to be complex on the 3' end of deletion; some accompanied duplications with deletions and others could not be explained by one mutation event. We need to confirm these results, taking into account the experimental reproducibility and the accuracy of the published genetic map. The results of the research using the microarray-CGH help us to search the regions where deletions are easily induced and to identify the factors affecting the range of deletions

  19. Assessing probe-specific dye and slide biases in two-color microarray data

    Directory of Open Access Journals (Sweden)

    Goldberg Zelanna

    2008-07-01

    Full Text Available Abstract Background A primary reason for using two-color microarrays is that the use of two samples labeled with different dyes on the same slide, that bind to probes on the same spot, is supposed to adjust for many factors that introduce noise and errors into the analysis. Most users assume that any differences between the dyes can be adjusted out by standard methods of normalization, so that measures such as log ratios on the same slide are reliable measures of comparative expression. However, even after the normalization, there are still probe specific dye and slide variation among the data. We define a method to quantify the amount of the dye-by-probe and slide-by-probe interaction. This serves as a diagnostic, both visual and numeric, of the existence of probe-specific dye bias. We show how this improved the performance of two-color array analysis for arrays for genomic analysis of biological samples ranging from rice to human tissue. Results We develop a procedure for quantifying the extent of probe-specific dye and slide bias in two-color microarrays. The primary output is a graphical diagnostic of the extent of the bias which called ECDF (Empirical Cumulative Distribution Function, though numerical results are also obtained. Conclusion We show that the dye and slide biases were high for human and rice genomic arrays in two gene expression facilities, even after the standard intensity-based normalization, and describe how this diagnostic allowed the problems causing the probe-specific bias to be addressed, and resulted in important improvements in performance. The R package LMGene which contains the method described in this paper has been available to download from Bioconductor.

  20. Styl RFLP recognized by a human IRBP cDNA localized to chromosome 10

    Energy Technology Data Exchange (ETDEWEB)

    Chin, K S; Mathew, C G.P.; Fong, S L; Bridges, C D; Ponder, B A.J.

    1988-02-25

    A 2184 bp cDNA (H.4 IRBP) encoding human interstitial retinol-biding protein isolated from a human retina cDNA library in lambdagt10 by screening with a bovine IRBP cDNA probe. Styl identifies a 2-allele polymorphism with bands at 2.3 kb (Cl) and 1.95 kb (C2) and invariant bands at 1.1, 1.0 and 0.8kb. Codominant segregation was observed in two informative families. The RFLP was mapped to chromosome 10 using somatic cell hybrids. In situ hybridization suggests regional assignments near p11.2 -q11.2 with a secondary site of hybridization at q24-25.

  1. Solar 'hot spots' are still hot

    Science.gov (United States)

    Bai, Taeil

    1990-01-01

    Longitude distributions of solar flares are not random but show evidence for active zones (or hot spots) where flares are concentrated. According to a previous study, two hot spots in the northern hemisphere, which rotate with a synodic period of about 26.72 days, produced the majority of major flares, during solar cycles 20 and 21. The more prominent of these two hot spots is found to be still active during the rising part of cycle 22, producing the majority of northern hemisphere major flares. The synodic rotation period of this hot spot is 26.727 + or - 0.007 days. There is also evidence for hot spots in the southern hemisphere. Two hot spots separated by 180 deg are found to rotate with a period of 29.407 days, with one of them having persisted in the same locations during cycles 19-22 and the other, during cycles 20-22.

  2. Variable-spot ion beam figuring

    International Nuclear Information System (INIS)

    Wu, Lixiang; Qiu, Keqiang; Fu, Shaojun

    2016-01-01

    This paper introduces a new scheme of ion beam figuring (IBF), or rather variable-spot IBF, which is conducted at a constant scanning velocity with variable-spot ion beam collimated by a variable diaphragm. It aims at improving the reachability and adaptation of the figuring process within the limits of machine dynamics by varying the ion beam spot size instead of the scanning velocity. In contrast to the dwell time algorithm in the conventional IBF, the variable-spot IBF adopts a new algorithm, which consists of the scan path programming and the trajectory optimization using pattern search. In this algorithm, instead of the dwell time, a new concept, integral etching time, is proposed to interpret the process of variable-spot IBF. We conducted simulations to verify its feasibility and practicality. The simulation results indicate the variable-spot IBF is a promising alternative to the conventional approach.

  3. Solar hot spots are still hot

    International Nuclear Information System (INIS)

    Bai, T.

    1990-01-01

    Longitude distributions of solar flares are not random but show evidence for active zones (or hot spots) where flares are concentrated. According to a previous study, two hot spots in the northern hemisphere, which rotate with a synodic period of about 26.72 days, produced the majority of major flares, during solar cycles 20 and 21. The more prominent of these two hot spots is found to be still active during the rising part of cycle 22, producing the majority of northern hemisphere major flares. The synodic rotation period of this hot spot is 26.727 + or - 0.007 days. There is also evidence for hot spots in the southern hemisphere. Two hot spots separated by 180 deg are found to rotate with a period of 29.407 days, with one of them having persisted in the same locations during cycles 19-22 and the other, during cycles 20-22. 14 refs

  4. Spot and Runway Departure Advisor

    Science.gov (United States)

    Jung, Yoon Chul

    2013-01-01

    The Spot and Runway Departure Advisor (SARDA) is a research prototype of a decision support tool for ATC tower controllers to assist in manging and controlling traffic on the surface of an airport. SARDA employs a scheduler to generate an optimal runway schedule and gate push-back - spot release sequence and schedule that improves efficiency of surface operations. The advisories for ATC tower controllers are displayed on an Electronic Flight Strip (EFS) system. The human-in-the-loop simulation of the SARDA tool was conducted for east operations of Dallas-Ft. Worth International Airport (DFW) to evaluate performance of the SARDA tool and human factors, such as situational awareness and workload. The results indicates noticeable taxi delay reduction and fuel savings by using the SARDA tool. Reduction in controller workload were also observed throughout the scenario runs. The future plan includes modeling and simulation of the ramp operations of the Charlotte International Airport, and develop a decision support tool for the ramp controllers.

  5. X-ray spot filmer

    International Nuclear Information System (INIS)

    1975-01-01

    An X-ray apparatus is described which includes a spot filmer for feeding sheets of unexposed film one at a time into a vacuum evacuable cassette for exposure, and for returning exposed film sheets to an exposed film magazine. The spot filmer has a housing defining a light-tight enclosure. The film magazines are insertable through a door into the housing and into a film feed mechanism. The film feed mechanism unlatches, opens and positions the magazines; it then feeds a sheet of unexposed film into the vacuum evacuable cassette, releases the film sheet so the cassette can position the film sheet for exposure, and closes the film magazines. An orthogonal drive system positions the vacuum evacuable cassette to expose selected film sheet portions and returns the cassette to a retracted position. The film feed mechanism opens the magazines, feeds the exposed film sheet into the exposed film magazine, and closes the magazines. A film identification system is provided for forming an identifying image on a marginal portion of each film sheet

  6. Glare Spot Phase Doppler Anemometry

    Science.gov (United States)

    Hespel, Camille; Ren, Kuanfang; Gréhan, Gérard; Onofri, Fabrice

    2007-06-01

    The Phase Doppler anemometry has been developed to measure simultaneously the velocity and the size of droplets. The measurement of the refractive index would be also interesting since it depends on the temperature and the composition of the particle and its measurement permits both to increase the quality of the diameter measurement and to obtain information on the temperature and/or the composition of the droplets. In this paper, we introduce a Glare Spot Phase Doppler Anemometry which uses two large beams. In this case, the images of the particle formed by the reflected and refracted light, known as glare spots, are separated in space. When a particle passes in the probe volume, the two parts in a signal obtained by a detector in forward direction are then separated in time. If two detectors are used the phase differences between two signals, the distance and the intensity ratio of reflected and refracted parts can be obtained and they provide rich information about the particle diameter and its refractive index, as well as its velocity. This paper is devoted to the numerical study of such a configuration with two theoretical models: geometrical optics and rigorous electromagnetism solution.

  7. Oil futures and spot markets

    International Nuclear Information System (INIS)

    Samii, M.V.

    1992-01-01

    In the last decade, the oil futures market has risen to prominence and has become a major factor in influencing oil market psychology and the crude oil market. On a normal day, over 92 thousand contracts, the equivalent of 92 million barrels per day, change hands on the New York Mercantile Exchange, NYMEX. This market has provided a vehicle for hedging against risk. At the same time, it has also created opportunities for speculation. Those who previously were unable to participate in oil market transactions can now become involved through the futures market. The large number of participants in the future market and the availability of information has made this market more efficient and transparent, relative to the crude oil market. While there has been considerable in-depth analysis of other future markets, relatively little theoretical attention has focused on that of oil. This paper looks at the following issues. First, what is the relationship between futures and spot oil prices? And secondly, are futures prices a good predictor of spot crude prices in the future? (author)

  8. Significance analysis of lexical bias in microarray data

    Directory of Open Access Journals (Sweden)

    Falkow Stanley

    2003-04-01

    Full Text Available Abstract Background Genes that are determined to be significantly differentially regulated in microarray analyses often appear to have functional commonalities, such as being components of the same biochemical pathway. This results in certain words being under- or overrepresented in the list of genes. Distinguishing between biologically meaningful trends and artifacts of annotation and analysis procedures is of the utmost importance, as only true biological trends are of interest for further experimentation. A number of sophisticated methods for identification of significant lexical trends are currently available, but these methods are generally too cumbersome for practical use by most microarray users. Results We have developed a tool, LACK, for calculating the statistical significance of apparent lexical bias in microarray datasets. The frequency of a user-specified list of search terms in a list of genes which are differentially regulated is assessed for statistical significance by comparison to randomly generated datasets. The simplicity of the input files and user interface targets the average microarray user who wishes to have a statistical measure of apparent lexical trends in analyzed datasets without the need for bioinformatics skills. The software is available as Perl source or a Windows executable. Conclusion We have used LACK in our laboratory to generate biological hypotheses based on our microarray data. We demonstrate the program's utility using an example in which we confirm significant upregulation of SPI-2 pathogenicity island of Salmonella enterica serovar Typhimurium by the cation chelator dipyridyl.

  9. A Fisheye Viewer for microarray-based gene expression data.

    Science.gov (United States)

    Wu, Min; Thao, Cheng; Mu, Xiangming; Munson, Ethan V

    2006-10-13

    Microarray has been widely used to measure the relative amounts of every mRNA transcript from the genome in a single scan. Biologists have been accustomed to reading their experimental data directly from tables. However, microarray data are quite large and are stored in a series of files in a machine-readable format, so direct reading of the full data set is not feasible. The challenge is to design a user interface that allows biologists to usefully view large tables of raw microarray-based gene expression data. This paper presents one such interface--an electronic table (E-table) that uses fisheye distortion technology. The Fisheye Viewer for microarray-based gene expression data has been successfully developed to view MIAME data stored in the MAGE-ML format. The viewer can be downloaded from the project web site http://polaris.imt.uwm.edu:7777/fisheye/. The fisheye viewer was implemented in Java so that it could run on multiple platforms. We implemented the E-table by adapting JTable, a default table implementation in the Java Swing user interface library. Fisheye views use variable magnification to balance magnification for easy viewing and compression for maximizing the amount of data on the screen. This Fisheye Viewer is a lightweight but useful tool for biologists to quickly overview the raw microarray-based gene expression data in an E-table.

  10. A fisheye viewer for microarray-based gene expression data

    Directory of Open Access Journals (Sweden)

    Munson Ethan V

    2006-10-01

    Full Text Available Abstract Background Microarray has been widely used to measure the relative amounts of every mRNA transcript from the genome in a single scan. Biologists have been accustomed to reading their experimental data directly from tables. However, microarray data are quite large and are stored in a series of files in a machine-readable format, so direct reading of the full data set is not feasible. The challenge is to design a user interface that allows biologists to usefully view large tables of raw microarray-based gene expression data. This paper presents one such interface – an electronic table (E-table that uses fisheye distortion technology. Results The Fisheye Viewer for microarray-based gene expression data has been successfully developed to view MIAME data stored in the MAGE-ML format. The viewer can be downloaded from the project web site http://polaris.imt.uwm.edu:7777/fisheye/. The fisheye viewer was implemented in Java so that it could run on multiple platforms. We implemented the E-table by adapting JTable, a default table implementation in the Java Swing user interface library. Fisheye views use variable magnification to balance magnification for easy viewing and compression for maximizing the amount of data on the screen. Conclusion This Fisheye Viewer is a lightweight but useful tool for biologists to quickly overview the raw microarray-based gene expression data in an E-table.

  11. Advanced Data Mining of Leukemia Cells Micro-Arrays

    Directory of Open Access Journals (Sweden)

    Richard S. Segall

    2009-12-01

    Full Text Available This paper provides continuation and extensions of previous research by Segall and Pierce (2009a that discussed data mining for micro-array databases of Leukemia cells for primarily self-organized maps (SOM. As Segall and Pierce (2009a and Segall and Pierce (2009b the results of applying data mining are shown and discussed for the data categories of microarray databases of HL60, Jurkat, NB4 and U937 Leukemia cells that are also described in this article. First, a background section is provided on the work of others pertaining to the applications of data mining to micro-array databases of Leukemia cells and micro-array databases in general. As noted in predecessor article by Segall and Pierce (2009a, micro-array databases are one of the most popular functional genomics tools in use today. This research in this paper is intended to use advanced data mining technologies for better interpretations and knowledge discovery as generated by the patterns of gene expressions of HL60, Jurkat, NB4 and U937 Leukemia cells. The advanced data mining performed entailed using other data mining tools such as cubic clustering criterion, variable importance rankings, decision trees, and more detailed examinations of data mining statistics and study of other self-organized maps (SOM clustering regions of workspace as generated by SAS Enterprise Miner version 4. Conclusions and future directions of the research are also presented.

  12. Probe Selection for DNA Microarrays using OligoWiz

    DEFF Research Database (Denmark)

    Wernersson, Rasmus; Juncker, Agnieszka; Nielsen, Henrik Bjørn

    2007-01-01

    Nucleotide abundance measurements using DNA microarray technology are possible only if appropriate probes complementary to the target nucleotides can be identified. Here we present a protocol for selecting DNA probes for microarrays using the OligoWiz application. OligoWiz is a client-server appl......Nucleotide abundance measurements using DNA microarray technology are possible only if appropriate probes complementary to the target nucleotides can be identified. Here we present a protocol for selecting DNA probes for microarrays using the OligoWiz application. OligoWiz is a client......-server application that offers a detailed graphical interface and real-time user interaction on the client side, and massive computer power and a large collection of species databases (400, summer 2007) on the server side. Probes are selected according to five weighted scores: cross-hybridization, deltaT(m), folding...... computer skills and can be executed from any Internet-connected computer. The probe selection procedure for a standard microarray design targeting all yeast transcripts can be completed in 1 h....

  13. Microarray-based screening of heat shock protein inhibitors.

    Science.gov (United States)

    Schax, Emilia; Walter, Johanna-Gabriela; Märzhäuser, Helene; Stahl, Frank; Scheper, Thomas; Agard, David A; Eichner, Simone; Kirschning, Andreas; Zeilinger, Carsten

    2014-06-20

    Based on the importance of heat shock proteins (HSPs) in diseases such as cancer, Alzheimer's disease or malaria, inhibitors of these chaperons are needed. Today's state-of-the-art techniques to identify HSP inhibitors are performed in microplate format, requiring large amounts of proteins and potential inhibitors. In contrast, we have developed a miniaturized protein microarray-based assay to identify novel inhibitors, allowing analysis with 300 pmol of protein. The assay is based on competitive binding of fluorescence-labeled ATP and potential inhibitors to the ATP-binding site of HSP. Therefore, the developed microarray enables the parallel analysis of different ATP-binding proteins on a single microarray. We have demonstrated the possibility of multiplexing by immobilizing full-length human HSP90α and HtpG of Helicobacter pylori on microarrays. Fluorescence-labeled ATP was competed by novel geldanamycin/reblastatin derivatives with IC50 values in the range of 0.5 nM to 4 μM and Z(*)-factors between 0.60 and 0.96. Our results demonstrate the potential of a target-oriented multiplexed protein microarray to identify novel inhibitors for different members of the HSP90 family. Copyright © 2014 Elsevier B.V. All rights reserved.

  14. cDNA cloning of rat and human medium chain acyl-CoA dehydrogenase (MCAD)

    International Nuclear Information System (INIS)

    Matsubara, Y.; Kraus, J.P.; Rosenberg, L.E.; Tanaka, K.

    1986-01-01

    MCAD is one of three mitochondrial flavoenzymes which catalyze the first step in the β-oxidation of straight chain fatty acids. It is a tetramer with a subunit Mr of 45 kDa. MCAD is synthesized in the cytosol as a 49 kDa precursor polypeptide (pMCAD), imported into mitochondria, and cleaved to the mature form. Genetic deficiency of MCAD causes recurrent episodes of hypoglycemic coma accompanied by medium chain dicarboxylic aciduria. Employing a novel approach, the authors now report isolation of partial rat and human cDNA clones encoding pMCAD. mRNA encoding pMCAD was purified to near homogeneity by polysome immunoadsorption using polyclonal monospecific antibody. Single-stranded [ 32 P]labeled cDNA probe was synthesized using the enriched mRNA as template, and was used to screen directly 16,000 colonies from a total rat liver cDNA library constructed in pBR322. One clone (600 bp) was detected by in situ hybridization. Hybrid-selected translation with this cDNA yielded a 49 kDa polypeptide indistinguishable in size from rat pMCAD and immunoprecipitable with anti-MCAD antibody. Using the rat cDNA as probe, 43,000 colonies from a human liver cDNA library were screened. Four identical positive clones (400 bp) were isolated and positively identified by hybrid-selected translation and immunoprecipitation. The sizes of rat and human mRNAs encoding pMCAD were 2.2 kb and 2.4 kb, respectively, as determined by Northern blotting

  15. Cloning, sequencing, and expression of cDNA for human β-glucuronidase

    International Nuclear Information System (INIS)

    Oshima, A.; Kyle, J.W.; Miller, R.D.

    1987-01-01

    The authors report here the cDNA sequence for human placental β-glucuronidase (β-D-glucuronoside glucuronosohydrolase, EC 3.2.1.31) and demonstrate expression of the human enzyme in transfected COS cells. They also sequenced a partial cDNA clone from human fibroblasts that contained a 153-base-pair deletion within the coding sequence and found a second type of cDNA clone from placenta that contained the same deletion. Nuclease S1 mapping studies demonstrated two types of mRNAs in human placenta that corresponded to the two types of cDNA clones isolated. The NH 2 -terminal amino acid sequence determined for human spleen β-glucuronidase agreed with that inferred from the DNA sequence of the two placental clones, beginning at amino acid 23, suggesting a cleaved signal sequence of 22 amino acids. When transfected into COS cells, plasmids containing either placental clone expressed an immunoprecipitable protein that contained N-linked oligosaccharides as evidenced by sensitivity to endoglycosidase F. However, only transfection with the clone containing the 153-base-pair segment led to expression of human β-glucuronidase activity. These studies provide the sequence for the full-length cDNA for human β-glucuronidase, demonstrate the existence of two populations of mRNA for β-glucuronidase in human placenta, only one of which specifies a catalytically active enzyme, and illustrate the importance of expression studies in verifying that a cDNA is functionally full-length

  16. Display of a Maize cDNA library on baculovirus infected insect cells

    Directory of Open Access Journals (Sweden)

    Jones Ian M

    2008-08-01

    Full Text Available Abstract Background Maize is a good model system for cereal crop genetics and development because of its rich genetic heritage and well-characterized morphology. The sequencing of its genome is well advanced, and new technologies for efficient proteomic analysis are needed. Baculovirus expression systems have been used for the last twenty years to express in insect cells a wide variety of eukaryotic proteins that require complex folding or extensive posttranslational modification. More recently, baculovirus display technologies based on the expression of foreign sequences on the surface of Autographa californica (AcMNPV have been developed. We investigated the potential of a display methodology for a cDNA library of maize young seedlings. Results We constructed a full-length cDNA library of young maize etiolated seedlings in the transfer vector pAcTMVSVG. The library contained a total of 2.5 × 105 independent clones. Expression of two known maize proteins, calreticulin and auxin binding protein (ABP1, was shown by western blot analysis of protein extracts from insect cells infected with the cDNA library. Display of the two proteins in infected insect cells was shown by selective biopanning using magnetic cell sorting and demonstrated proof of concept that the baculovirus maize cDNA display library could be used to identify and isolate proteins. Conclusion The maize cDNA library constructed in this study relies on the novel technology of baculovirus display and is unique in currently published cDNA libraries. Produced to demonstrate proof of principle, it opens the way for the development of a eukaryotic in vivo display tool which would be ideally suited for rapid screening of the maize proteome for binding partners, such as proteins involved in hormone regulation or defence.

  17. Integrating sustainable hunting in biodiversity protection in Central Africa: hot spots, weak spots, and strong spots.

    Directory of Open Access Journals (Sweden)

    John E Fa

    Full Text Available Wild animals are a primary source of protein (bushmeat for people living in or near tropical forests. Ideally, the effect of bushmeat harvests should be monitored closely by making regular estimates of offtake rate and size of stock available for exploitation. However, in practice, this is possible in very few situations because it requires both of these aspects to be readily measurable, and even in the best case, entails very considerable time and effort. As alternative, in this study, we use high-resolution, environmental favorability models for terrestrial mammals (N = 165 in Central Africa to map areas of high species richness (hot spots and hunting susceptibility. Favorability models distinguish localities with environmental conditions that favor the species' existence from those with detrimental characteristics for its presence. We develop an index for assessing Potential Hunting Sustainability (PHS of each species based on their ecological characteristics (population density, habitat breadth, rarity and vulnerability, weighted according to restrictive and permissive assumptions of how species' characteristics are combined. Species are classified into five main hunting sustainability classes using fuzzy logic. Using the accumulated favorability values of all species, and their PHS values, we finally identify weak spots, defined as high diversity regions of especial hunting vulnerability for wildlife, as well as strong spots, defined as high diversity areas of high hunting sustainability potential. Our study uses relatively simple models that employ easily obtainable data of a species' ecological characteristics to assess the impacts of hunting in tropical regions. It provides information for management by charting the geography of where species are more or less likely to be at risk of extinction from hunting.

  18. PreP+07: improvements of a user friendly tool to preprocess and analyse microarray data

    Directory of Open Access Journals (Sweden)

    Claros M Gonzalo

    2009-01-01

    Full Text Available Abstract Background Nowadays, microarray gene expression analysis is a widely used technology that scientists handle but whose final interpretation usually requires the participation of a specialist. The need for this participation is due to the requirement of some background in statistics that most users lack or have a very vague notion of. Moreover, programming skills could also be essential to analyse these data. An interactive, easy to use application seems therefore necessary to help researchers to extract full information from data and analyse them in a simple, powerful and confident way. Results PreP+07 is a standalone Windows XP application that presents a friendly interface for spot filtration, inter- and intra-slide normalization, duplicate resolution, dye-swapping, error removal and statistical analyses. Additionally, it contains two unique implementation of the procedures – double scan and Supervised Lowess-, a complete set of graphical representations – MA plot, RG plot, QQ plot, PP plot, PN plot – and can deal with many data formats, such as tabulated text, GenePix GPR and ArrayPRO. PreP+07 performance has been compared with the equivalent functions in Bioconductor using a tomato chip with 13056 spots. The number of differentially expressed genes considering p-values coming from the PreP+07 and Bioconductor Limma packages were statistically identical when the data set was only normalized; however, a slight variability was appreciated when the data was both normalized and scaled. Conclusion PreP+07 implementation provides a high degree of freedom in selecting and organizing a small set of widely used data processing protocols, and can handle many data formats. Its reliability has been proven so that a laboratory researcher can afford a statistical pre-processing of his/her microarray results and obtain a list of differentially expressed genes using PreP+07 without any programming skills. All of this gives support to scientists

  19. Detection of reverse transcriptase termination sites using cDNA ligation and massive parallel sequencing

    DEFF Research Database (Denmark)

    Kielpinski, Lukasz J; Boyd, Mette; Sandelin, Albin

    2013-01-01

    Detection of reverse transcriptase termination sites is important in many different applications, such as structural probing of RNAs, rapid amplification of cDNA 5' ends (5' RACE), cap analysis of gene expression, and detection of RNA modifications and protein-RNA cross-links. The throughput...... of these methods can be increased by applying massive parallel sequencing technologies.Here, we describe a versatile method for detection of reverse transcriptase termination sites based on ligation of an adapter to the 3' end of cDNA with bacteriophage TS2126 RNA ligase (CircLigase™). In the following PCR...

  20. The nucleotide sequence of human transition protein 1 cDNA

    Energy Technology Data Exchange (ETDEWEB)

    Luerssen, H; Hoyer-Fender, S; Engel, W [Universitaet Goettingen (West Germany)

    1988-08-11

    The authors have screened a human testis cDNA library with an oligonucleotide of 81 mer prepared according to a part of the published nucleotide sequence of the rat transition protein TP 1. They have isolated a cDNA clone with the length of 441 bp containing the coding region of 162 bp for human transition protein 1. There is about 84% homology in the coding region of the sequence compared to rat. The human cDNA-clone encodes a polypeptide of 54 amino acids of which 7 are different to that of rat.

  1. Avoiding cross hybridization by choosing nonredundant targets on cDNA arrays

    DEFF Research Database (Denmark)

    Nielsen, Henrik Bjørn; Knudsen, Steen

    2002-01-01

    PROBEWIZ designs PCR primers for amplifying probes for cDNA arrays. The probes are designed to have minimal homology to other expressed sequences from a given organism. The primer selection is based on user-defined penalties for homology, primer quality, and proximity to the 3' end.......PROBEWIZ designs PCR primers for amplifying probes for cDNA arrays. The probes are designed to have minimal homology to other expressed sequences from a given organism. The primer selection is based on user-defined penalties for homology, primer quality, and proximity to the 3' end....

  2. Isolation and expression of a pea vicilin cDNA in the yeast Saccharomyces cerevisiae.

    OpenAIRE

    Watson, M D; Lambert, N; Delauney, A; Yarwood, J N; Croy, R R; Gatehouse, J A; Wright, D J; Boulter, D

    1988-01-01

    A cDNA clone containing the complete coding sequence for vicilin from pea (Pisum sativum L.) was isolated. It specifies a 50,000-Mr protein that in pea is neither post-translationally processed nor glycosylated. The cDNA clone was expressed in yeast from a 2 micron plasmid by using the yeast phosphoglycerate kinase promoter and initiator codon. The resultant fusion protein, which contains the first 16 amino acid residues of phosphoglycerate kinase in addition to the vicilin sequence, was puri...

  3. Ionospheric hot spot at high latitudes

    International Nuclear Information System (INIS)

    Schunk, R.W.; Sojka, J.J.

    1982-01-01

    A hot spot (or spots) can occur in the high-latitude ionosphere depending on the plasma convection pattern. The hot spot corresponds to a small magnetic local time-magnetic latitude region of elevated ion temperatures located near the dusk and/or dawn meridians. For asymmetric convection electric field patterns, with enhanced flow in either the dusk or dawn sector of the polar cap, a single hot spot should occur in association with the strong convection cell. However, on geomagnetically disturbed days, two strong convection cells can occur, and hence, two hot spots should exist. The hot spot should be detectable when the electric field in the strong convection cell exceeds about 40 mV m -1 . For electric fields of the order of 100 mV m -1 in the convection cell, the ion temperature in the hot spot is greatest at low altitudes, reaching 4000 0 K at 160 km, and decreases with altitude in the F-region. An ionospheric hot spot (or spots) can be expected at all seasons and for a wide range of solar cycle conditions

  4. Identification of novel candidate target genes in amplicons of Glioblastoma multiforme tumors detected by expression and CGH microarray profiling

    Directory of Open Access Journals (Sweden)

    Hernández-Moneo Jose-Luis

    2006-09-01

    Full Text Available Abstract Background Conventional cytogenetic and comparative genomic hybridization (CGH studies in brain malignancies have shown that glioblastoma multiforme (GBM is characterized by complex structural and numerical alterations. However, the limited resolution of these techniques has precluded the precise identification of detailed specific gene copy number alterations. Results We performed a genome-wide survey of gene copy number changes in 20 primary GBMs by CGH on cDNA microarrays. A novel amplicon at 4p15, and previously uncharacterized amplicons at 13q32-34 and 1q32 were detected and are analyzed here. These amplicons contained amplified genes not previously reported. Other amplified regions containg well-known oncogenes in GBMs were also detected at 7p12 (EGFR, 7q21 (CDK6, 4q12 (PDGFRA, and 12q13-15 (MDM2 and CDK4. In order to identify the putative target genes of the amplifications, and to determine the changes in gene expression levels associated with copy number change events, we carried out parallel gene expression profiling analyses using the same cDNA microarrays. We detected overexpression of the novel amplified genes SLA/LP and STIM2 (4p15, and TNFSF13B and COL4A2 (13q32-34. Some of the candidate target genes of amplification (EGFR, CDK6, MDM2, CDK4, and TNFSF13B were tested in an independent set of 111 primary GBMs by using FISH and immunohistological assays. The novel candidate 13q-amplification target TNFSF13B was amplified in 8% of the tumors, and showed protein expression in 20% of the GBMs. Conclusion This high-resolution analysis allowed us to propose novel candidate target genes such as STIM2 at 4p15, and TNFSF13B or COL4A2 at 13q32-34 that could potentially contribute to the pathogenesis of these tumors and which would require futher investigations. We showed that overexpression of the amplified genes could be attributable to gene dosage and speculate that deregulation of those genes could be important in the development

  5. DNA microarray data and contextual analysis of correlation graphs

    Directory of Open Access Journals (Sweden)

    Hingamp Pascal

    2003-04-01

    Full Text Available Abstract Background DNA microarrays are used to produce large sets of expression measurements from which specific biological information is sought. Their analysis requires efficient and reliable algorithms for dimensional reduction, classification and annotation. Results We study networks of co-expressed genes obtained from DNA microarray experiments. The mathematical concept of curvature on graphs is used to group genes or samples into clusters to which relevant gene or sample annotations are automatically assigned. Application to publicly available yeast and human lymphoma data demonstrates the reliability of the method in spite of its simplicity, especially with respect to the small number of parameters involved. Conclusions We provide a method for automatically determining relevant gene clusters among the many genes monitored with microarrays. The automatic annotations and the graphical interface improve the readability of the data. A C++ implementation, called Trixy, is available from http://tagc.univ-mrs.fr/bioinformatics/trixy.html.

  6. A Versatile Microarray Platform for Capturing Rare Cells

    Science.gov (United States)

    Brinkmann, Falko; Hirtz, Michael; Haller, Anna; Gorges, Tobias M.; Vellekoop, Michael J.; Riethdorf, Sabine; Müller, Volkmar; Pantel, Klaus; Fuchs, Harald

    2015-10-01

    Analyses of rare events occurring at extremely low frequencies in body fluids are still challenging. We established a versatile microarray-based platform able to capture single target cells from large background populations. As use case we chose the challenging application of detecting circulating tumor cells (CTCs) - about one cell in a billion normal blood cells. After incubation with an antibody cocktail, targeted cells are extracted on a microarray in a microfluidic chip. The accessibility of our platform allows for subsequent recovery of targets for further analysis. The microarray facilitates exclusion of false positive capture events by co-localization allowing for detection without fluorescent labelling. Analyzing blood samples from cancer patients with our platform reached and partly outreached gold standard performance, demonstrating feasibility for clinical application. Clinical researchers free choice of antibody cocktail without need for altered chip manufacturing or incubation protocol, allows virtual arbitrary targeting of capture species and therefore wide spread applications in biomedical sciences.

  7. SIGMA: A System for Integrative Genomic Microarray Analysis of Cancer Genomes

    Directory of Open Access Journals (Sweden)

    Davies Jonathan J

    2006-12-01

    Full Text Available Abstract Background The prevalence of high resolution profiling of genomes has created a need for the integrative analysis of information generated from multiple methodologies and platforms. Although the majority of data in the public domain are gene expression profiles, and expression analysis software are available, the increase of array CGH studies has enabled integration of high throughput genomic and gene expression datasets. However, tools for direct mining and analysis of array CGH data are limited. Hence, there is a great need for analytical and display software tailored to cross platform integrative analysis of cancer genomes. Results We have created a user-friendly java application to facilitate sophisticated visualization and analysis such as cross-tumor and cross-platform comparisons. To demonstrate the utility of this software, we assembled array CGH data representing Affymetrix SNP chip, Stanford cDNA arrays and whole genome tiling path array platforms for cross comparison. This cancer genome database contains 267 profiles from commonly used cancer cell lines representing 14 different tissue types. Conclusion In this study we have developed an application for the visualization and analysis of data from high resolution array CGH platforms that can be adapted for analysis of multiple types of high throughput genomic datasets. Furthermore, we invite researchers using array CGH technology to deposit both their raw and processed data, as this will be a continually expanding database of cancer genomes. This publicly available resource, the System for Integrative Genomic Microarray Analysis (SIGMA of cancer genomes, can be accessed at http://sigma.bccrc.ca.

  8. Comparative analysis of human conjunctival and corneal epithelial gene expression with oligonucleotide microarrays.

    Science.gov (United States)

    Turner, Helen C; Budak, Murat T; Akinci, M A Murat; Wolosin, J Mario

    2007-05-01

    To determine global mRNA expression levels in corneal and conjunctival epithelia and identify transcripts that exhibit preferential tissue expression. cDNA samples derived from human conjunctival and corneal epithelia were hybridized in three independent experiments to a commercial oligonucleotide array representing more than 22,000 transcripts. The resultant signal intensities and microarray software transcript present/absent calls were used in conjunction with the local pooled error (LPE) statistical method to identify transcripts that are preferentially or exclusively expressed in one of the two tissues at significant levels (expression >1% of the beta-actin level). EASE (Expression Analysis Systematic Explorer software) was used to identify biological systems comparatively overrepresented in either epithelium. Immuno-, and cytohistochemistry was performed to validate or expand on selected results of interest. The analysis identified 332 preferential and 93 exclusive significant corneal epithelial transcripts. The corresponding numbers of conjunctival epithelium transcripts were 592 and 211, respectively. The overrepresented biological processes in the cornea were related to cell adhesion and oxiredox equilibria and cytoprotection activities. In the conjunctiva, the biological processes that were most prominent were related to innate immunity and melanogenesis. Immunohistochemistry for antigen-presenting cells and melanocytes was consistent with these gene signatures. The transcript comparison identified a substantial number of genes that have either not been identified previously or are not known to be highly expressed in these two epithelia, including testican-1, ECM1, formin, CRTAC1, and NQO1 in the cornea and, in the conjunctiva, sPLA(2)-IIA, lipocalin 2, IGFBP3, multiple MCH class II proteins, and the Na-Pi cotransporter type IIb. Comparative gene expression profiling leads to the identification of many biological processes and previously unknown genes that

  9. AMDA: an R package for the automated microarray data analysis

    Directory of Open Access Journals (Sweden)

    Foti Maria

    2006-07-01

    Full Text Available Abstract Background Microarrays are routinely used to assess mRNA transcript levels on a genome-wide scale. Large amount of microarray datasets are now available in several databases, and new experiments are constantly being performed. In spite of this fact, few and limited tools exist for quickly and easily analyzing the results. Microarray analysis can be challenging for researchers without the necessary training and it can be time-consuming for service providers with many users. Results To address these problems we have developed an automated microarray data analysis (AMDA software, which provides scientists with an easy and integrated system for the analysis of Affymetrix microarray experiments. AMDA is free and it is available as an R package. It is based on the Bioconductor project that provides a number of powerful bioinformatics and microarray analysis tools. This automated pipeline integrates different functions available in the R and Bioconductor projects with newly developed functions. AMDA covers all of the steps, performing a full data analysis, including image analysis, quality controls, normalization, selection of differentially expressed genes, clustering, correspondence analysis and functional evaluation. Finally a LaTEX document is dynamically generated depending on the performed analysis steps. The generated report contains comments and analysis results as well as the references to several files for a deeper investigation. Conclusion AMDA is freely available as an R package under the GPL license. The package as well as an example analysis report can be downloaded in the Services/Bioinformatics section of the Genopolis http://www.genopolis.it/

  10. SU-E-T-510: Interplay Between Spots Sizes, Spot / Line Spacing and Motion in Spot Scanning Proton Therapy

    International Nuclear Information System (INIS)

    Lee, TK

    2015-01-01

    Purpose In proton beam configuration for spot scanning proton therapy (SSPT), one can define the spacing between spots and lines of scanning as a ratio of given spot size. If the spacing increases, the number of spots decreases which can potentially decrease scan time, and so can whole treatment time, and vice versa. However, if the spacing is too large, the uniformity of scanned field decreases. Also, the field uniformity can be affected by motion during SSPT beam delivery. In the present study, the interplay between spot/ line spacing and motion is investigated. Methods We used four Gaussian-shape spot sizes with 0.5cm, 1.0cm, 1.5cm, and 2.0cm FWHM, three spot/line spacing that creates uniform field profile which are 1/3*FWHM, σ/3*FWHM and 2/3*FWHM, and three random motion amplitudes within, +/−0.3mm, +/−0.5mm, and +/−1.0mm. We planned with 2Gy uniform single layer of 10×10cm2 and 20×20cm2 fields. Then, mean dose within 80% area of given field size, contrubuting MU per each spot assuming 1cGy/MU calibration for all spot sizes, number of spots and uniformity were calculated. Results The plans with spot/line spacing equal to or smaller than 2/3*FWHM without motion create ∼100% uniformity. However, it was found that the uniformity decreases with increased spacing, and it is more pronounced with smaller spot sizes, but is not affected by scanned field sizes. Conclusion It was found that the motion during proton beam delivery can alter the dose uniformity and the amount of alteration changes with spot size which changes with energy and spot/line spacing. Currently, robust evaluation in TPS (e.g. Eclipse system) performs range uncertainty evaluation using isocenter shift and CT calibration error. Based on presented study, it is recommended to add interplay effect evaluation to robust evaluation process. For future study, the additional interplay between the energy layers and motion is expected to present volumetric effect

  11. Label and Label-Free Detection Techniques for Protein Microarrays

    Directory of Open Access Journals (Sweden)

    Amir Syahir

    2015-04-01

    Full Text Available Protein microarray technology has gone through numerous innovative developments in recent decades. In this review, we focus on the development of protein detection methods embedded in the technology. Early microarrays utilized useful chromophores and versatile biochemical techniques dominated by high-throughput illumination. Recently, the realization of label-free techniques has been greatly advanced by the combination of knowledge in material sciences, computational design and nanofabrication. These rapidly advancing techniques aim to provide data without the intervention of label molecules. Here, we present a brief overview of this remarkable innovation from the perspectives of label and label-free techniques in transducing nano‑biological events.

  12. Advanced Data Mining of Leukemia Cells Micro-Arrays

    OpenAIRE

    Richard S. Segall; Ryan M. Pierce

    2009-01-01

    This paper provides continuation and extensions of previous research by Segall and Pierce (2009a) that discussed data mining for micro-array databases of Leukemia cells for primarily self-organized maps (SOM). As Segall and Pierce (2009a) and Segall and Pierce (2009b) the results of applying data mining are shown and discussed for the data categories of microarray databases of HL60, Jurkat, NB4 and U937 Leukemia cells that are also described in this article. First, a background section is pro...

  13. 5'-end sequences of budding yeast full-length cDNA clones and quality scores - Budding yeast cDNA sequencing project | LSDB Archive [Life Science Database Archive metadata

    Lifescience Database Archive (English)

    Full Text Available east_seq_qual.zip File URL: ftp://ftp.biosciencedbc.jp/archive/yeast_cdna/LATEST/...yeast_seq_qual.zip File size: 59.9MB Simple search URL http://togodb.biosciencedbc.jp/togodb/view/budding_yeast_cdna

  14. Cosmicflows-3: Cold Spot Repeller?

    Energy Technology Data Exchange (ETDEWEB)

    Courtois, Hélène M.; Graziani, Romain; Dupuy, Alexandra [University of Lyon, UCB Lyon 1, CNRS/IN2P3, IPN, Lyon (France); Tully, R. Brent [Institute for Astronomy, University of Hawaii, 2680 Woodlawn Drive, Honolulu, HI 96822 (United States); Hoffman, Yehuda [Racah Institute of Physics, Hebrew University, Jerusalem, 91904 (Israel); Pomarède, Daniel [Institut de Recherche sur les Lois Fondamentales de l’Univers, CEA, Université Paris-Saclay, F-91191 Gif-sur-Yvette (France)

    2017-09-20

    The three-dimensional gravitational velocity field within z ∼ 0.1 has been modeled with the Wiener filter methodology applied to the Cosmicflows-3 compilation of galaxy distances. The dominant features are a basin of attraction and two basins of repulsion. The major basin of attraction is an extension of the Shapley concentration of galaxies. One basin of repulsion, the Dipole Repeller, is located near the anti-apex of the cosmic microwave background dipole. The other basin of repulsion is in the proximate direction toward the “Cold Spot” irregularity in the cosmic microwave background. It has been speculated that a vast void might contribute to the amplitude of the Cold Spot from the integrated Sachs–Wolfe effect.

  15. Fabrication of Biomolecule Microarrays for Cell Immobilization Using Automated Microcontact Printing.

    Science.gov (United States)

    Foncy, Julie; Estève, Aurore; Degache, Amélie; Colin, Camille; Cau, Jean Christophe; Malaquin, Laurent; Vieu, Christophe; Trévisiol, Emmanuelle

    2018-01-01

    Biomolecule microarrays are generally produced by conventional microarrayer, i.e., by contact or inkjet printing. Microcontact printing represents an alternative way of deposition of biomolecules on solid supports but even if various biomolecules have been successfully microcontact printed, the production of biomolecule microarrays in routine by microcontact printing remains a challenging task and needs an effective, fast, robust, and low-cost automation process. Here, we describe the production of biomolecule microarrays composed of extracellular matrix protein for the fabrication of cell microarrays by using an automated microcontact printing device. Large scale cell microarrays can be reproducibly obtained by this method.

  16. 7 CFR 28.412 - Strict Middling Light Spotted Color.

    Science.gov (United States)

    2010-01-01

    ... 7 Agriculture 2 2010-01-01 2010-01-01 false Strict Middling Light Spotted Color. 28.412 Section 28... Light Spotted Color. Strict Middling Light Spotted Color is color which in spot or color, or both, is between Strict Middling Color and Strict Middling Spotted Color. ...

  17. 7 CFR 28.415 - Low Middling Light Spotted Color.

    Science.gov (United States)

    2010-01-01

    ... 7 Agriculture 2 2010-01-01 2010-01-01 false Low Middling Light Spotted Color. 28.415 Section 28... Spotted Color. Low Middling Light Spotted Color is color which in spot or color, or both, is between Low Middling Color and Low Middling Spotted Color. ...

  18. 7 CFR 28.411 - Good Middling Light Spotted Color.

    Science.gov (United States)

    2010-01-01

    ... 7 Agriculture 2 2010-01-01 2010-01-01 false Good Middling Light Spotted Color. 28.411 Section 28... Light Spotted Color. Good Middling Light Spotted Color is color which in spot or color, or both, is between Good Middling Color and Good Middling Spotted Color. ...

  19. 7 CFR 28.413 - Middling Light Spotted Color.

    Science.gov (United States)

    2010-01-01

    ... 7 Agriculture 2 2010-01-01 2010-01-01 false Middling Light Spotted Color. 28.413 Section 28.413... Spotted Color. Middling Light Spotted Color is color which in spot or color, or both, is between Middling Color and Middling Spotted Color. ...

  20. Characterization of ORF89 - A latency-related gene of white spot syndrome virus

    International Nuclear Information System (INIS)

    Hossain, M.S.; Khadijah, Siti; Kwang, Jimmy

    2004-01-01

    Open reading frame 89 (ORF89) is one of the three genes that are believed to be involved in the latent infection of white spot syndrome virus (WSSV). Here, we report the structure and functional characterization of ORF89. cDNA sequencing, 5' RLM-RACE, and 3' RLM-RACE showed that ORF89 gene is transcribed into an unspliced mRNA of 4436 nucleotides, which is predicted to encode a protein of 1437 amino acids. ORF89 expressed an approximately 165-kDa protein in Sf9 cells that localized in the nucleus. Amino acids 678-683 were found to be essential for nuclear localization. Cotransfection assays demonstrated that ORF89 protein repressed its own promoter as well as those of a protein kinase and the thymidine-thymidylate kinase genes of WSSV. SYBR Green real-time PCR indicated that the repression occurred at the transcriptional level

  1. Hot Spot Removal System: System description

    Energy Technology Data Exchange (ETDEWEB)

    NONE

    1997-09-01

    Hazardous wastes contaminated with radionuclides, chemicals, and explosives exist across the Department of Energy complex and need to be remediated due to environmental concerns. Currently, an opportunity is being developed to dramatically reduce remediation costs and to assist in the acceleration of schedules associated with these wastes by deploying a Hot Spot Removal System. Removing the hot spot from the waste site will remove risk driver(s) and enable another, more cost effective process/option/remedial alternative (i.e., capping) to be applied to the remainder of the site. The Hot Spot Removal System consists of a suite of technologies that will be utilized to locate and remove source terms. Components of the system can also be used in a variety of other cleanup activities. This Hot Spot Removal System Description document presents technologies that were considered for possible inclusion in the Hot Spot Removal System, technologies made available to the Hot Spot Removal System, industrial interest in the Hot Spot Removal System`s subsystems, the schedule required for the Hot Spot Removal System, the evaluation of the relevant technologies, and the recommendations for equipment and technologies as stated in the Plan section.

  2. Rocky Mountain spotted fever in children.

    Science.gov (United States)

    Woods, Charles R

    2013-04-01

    Rocky Mountain spotted fever is typically undifferentiated from many other infections in the first few days of illness. Treatment should not be delayed pending confirmation of infection when Rocky Mountain spotted fever is suspected. Doxycycline is the drug of choice even for infants and children less than 8 years old. Copyright © 2013 Elsevier Inc. All rights reserved.

  3. Rocky Mountain Spotted Fever: Statistics and Epidemiology

    Science.gov (United States)

    ... Search Form Controls Cancel Submit Search the CDC Rocky Mountain Spotted Fever (RMSF) Note: Javascript is disabled or is not ... message, please visit this page: About CDC.gov . Rocky Mountain Spotted Fever (RMSF) Transmission Signs and Symptoms Diagnosis and Testing ...

  4. Hot Spot Removal System: System description

    International Nuclear Information System (INIS)

    1997-09-01

    Hazardous wastes contaminated with radionuclides, chemicals, and explosives exist across the Department of Energy complex and need to be remediated due to environmental concerns. Currently, an opportunity is being developed to dramatically reduce remediation costs and to assist in the acceleration of schedules associated with these wastes by deploying a Hot Spot Removal System. Removing the hot spot from the waste site will remove risk driver(s) and enable another, more cost effective process/option/remedial alternative (i.e., capping) to be applied to the remainder of the site. The Hot Spot Removal System consists of a suite of technologies that will be utilized to locate and remove source terms. Components of the system can also be used in a variety of other cleanup activities. This Hot Spot Removal System Description document presents technologies that were considered for possible inclusion in the Hot Spot Removal System, technologies made available to the Hot Spot Removal System, industrial interest in the Hot Spot Removal System''s subsystems, the schedule required for the Hot Spot Removal System, the evaluation of the relevant technologies, and the recommendations for equipment and technologies as stated in the Plan section

  5. Cloning and sequencing of Indian Water buffalo (Bubalus bubalis) interleukin-3 cDNA

    KAUST Repository

    Sugumar, Thennarasu; Harishankar, M.; Dhinakar Raj, G.

    2011-01-01

    Full-length cDNA (435 bp) of the interleukin-3(IL-3) gene of the Indian water buffalo was amplified by reverse transcriptase-polymerase chain reaction and sequenced. This sequence had 96% nucleotide identity and 92% amino acid identity with bovine

  6. FIELD VALIDATION OF A SHEEPSHEAD MINNOW ESTROGEN-RESPONSIVE CDNA MACROARRAY

    Science.gov (United States)

    Hemmer, Michael J., Iris Knoebl, Becky L. Hemmer, Patrick Larkin, Peggy S. Harris and Nancy D. Denslow. In press. Field Validation of a Sheepshead Minnow Estrogen-Responsive cDNA Macroarray (Abstract). To be presented at the SETAC Fourth World Congress, 14-18 November 2004, Portl...

  7. Characterization and immunological identification of cDNA clones encoding two human DNA topoisomerase II isozymes

    International Nuclear Information System (INIS)

    Chung, T.D.Y.; Drake, F.H.; Tan, K.B.; Per, S.R.; Crooke, S.T.; Mirabelli, C.K.

    1989-01-01

    Several DNA topoisomerase II partial cDNA clones obtained from a human Raji-HN2 cDNA library were sequenced and two classes of nucleotide sequences were found. One member of the first class, SP1, was identical to an internal fragment of human HeLa cell Topo II cDNA described earlier. A member of the second class, SP11, shared extensive nucleotide (75%) and predicted peptide (92%) sequence similarities with the first two-thirds of HeLa Topo II. Each class of cDNAs hybridized to unique, nonoverlapping restriction enzyme fragments of genomic DNA from several human cell lines. Synthetic 24-mer oligonucleotide probes specific for each cDNA class hybridized to 6.5-kilobase mRNAs; furthermore, hybridization of probe specific for one class was not blocked by probe specific for the other. Antibodies raised against a synthetic SP1-encoded dodecapeptide specifically recognized the 170-kDa form of Topo II, while antibodies raised against the corresponding SP11-encoded dodecapeptide, or a second unique SP11-encoded tridecapeptide, selectively recognized the 180-kDa form of Topo II. These data provide genetic and immunochemical evidence for two Topo II isozymes

  8. Cloning a cDNA for the lysosomal alpha-glucosidase

    NARCIS (Netherlands)

    KONINGS, A.; HUPKES, P.; Versteeg, R.; Grosveld, G.; Reuser, A.; Galjaard, H.

    1984-01-01

    Messenger RNA was isolated from monkey testes and size-fractionated on sucrose gradients. In vitro translation of these mRNA fractions resulted in nascent, labeled alpha-glucosidase that could be precipitated with anti human alpha-glucosidase antiserum. A cDNA library was constructed from the most

  9. EXPRESSION PROFILING OF ESTROGENIC COMPOUNDS USING A SHEEPSHEAD MINNOW CDNA MACROARRAY

    Science.gov (United States)

    Larkin, Patrick, Leroy C. Folmar, Michael J. Hemmer, Arianna J. Poston and Nancy D. Denslow. 2003. Expression Profiling of Estrogenic Compounds Using a Sheepshead Minnow cDNA Macroarray. Environ. Health Perspect. 111(6):839-846. (ERL,GB 1171). A variety of anthropogenic c...

  10. Heterogeneity of rat tropoelastin mRNA revealed by cDNA cloning

    International Nuclear Information System (INIS)

    Pierce, R.A.; Deak, S.B.; Stolle, C.A.; Boyd, C.D.

    1990-01-01

    A λgt11 library constructed from poly(A+) RNA isolated from aortic tissue of neonatal rats was screened for rat tropoelastin cDNAs. The first, screen, utilizing a human tropoelastin cDNA clone, provided rat tropoelastin cDNAs spanning 2.3 kb of carboxy-terminal coding sequence and extended into the 3'-untranslated region. A subsequent screen using a 5' rat tropoelastin cDNA clone yielded clones extending into the amino-terminal signal sequence coding region. Sequence analysis of these clones has provided the complete derived amino acid sequence of rat tropoelastin and allowed alignment and comparison with published bovine cDNA sequence. While the overall structure of rat tropoelastin is similar to bovine sequence, numerous substitutions, deletions, and insertions demonstrated considerable heterogeneity between species. In particular, the pentapeptide repeat VPGVG, characteristic of all tropoelastins analyzed to date, is replaced in rat tropoelastin by a repeating pentapeptide, IPGVG. The hexapeptide repeat VGVAPG, the bovine elastin receptor binding peptide, is not encoded by rat tropoelastin cDNAs. Variations in coding sequence between rat tropoelastin CDNA clones were also found which may represent mRNA heterogeneity produced by alternative splicing of the rat tropoelastin pre-mRNA

  11. Complete amino acid sequence of human intestinal aminopeptidase N as deduced from cloned cDNA

    DEFF Research Database (Denmark)

    Cowell, G M; Kønigshøfer, E; Danielsen, E M

    1988-01-01

    The complete primary structure (967 amino acids) of an intestinal human aminopeptidase N (EC 3.4.11.2) was deduced from the sequence of a cDNA clone. Aminopeptidase N is anchored to the microvillar membrane via an uncleaved signal for membrane insertion. A domain constituting amino acid 250...

  12. Strategies to enhance immunogenicity of cDNA vaccine encoded antigens by modulation of antigen processing

    NARCIS (Netherlands)

    Platteel, Anouk C M; Marit de Groot, A; Andersen, Peter; Ovaa, Huib; Kloetzel, Peter M; Mishto, Michele; Sijts, Alice J A M

    2016-01-01

    Most vaccines are based on protective humoral responses while for intracellular pathogens CD8(+) T cells are regularly needed to provide protection. However, poor processing efficiency of antigens is often a limiting factor in CD8(+) T cell priming, hampering vaccine efficacy. The multistage cDNA

  13. cDNA, genomic cloning and sequence analysis of ribosomal protein ...

    African Journals Online (AJOL)

    enoh

    2012-03-13

    Mar 13, 2012 ... cDNA and the genomic sequence of RPS4X were cloned successfully from ... S4 genes plays a role in Turner syndrome; however, this ..... Project of Educational Committee of Sichuan Province ... Molecular biology of the cell.

  14. A putative peroxidase cDNA from turnip and analysis of the encoded protein sequence.

    Science.gov (United States)

    Romero-Gómez, S; Duarte-Vázquez, M A; García-Almendárez, B E; Mayorga-Martínez, L; Cervantes-Avilés, O; Regalado, C

    2008-12-01

    A putative peroxidase cDNA was isolated from turnip roots (Brassica napus L. var. purple top white globe) by reverse transcriptase-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE). Total RNA extracted from mature turnip roots was used as a template for RT-PCR, using a degenerated primer designed to amplify the highly conserved distal motif of plant peroxidases. The resulting partial sequence was used to design the rest of the specific primers for 5' and 3' RACE. Two cDNA fragments were purified, sequenced, and aligned with the partial sequence from RT-PCR, and a complete overlapping sequence was obtained and labeled as BbPA (Genbank Accession No. AY423440, named as podC). The full length cDNA is 1167bp long and contains a 1077bp open reading frame (ORF) encoding a 358 deduced amino acid peroxidase polypeptide. The putative peroxidase (BnPA) showed a calculated Mr of 34kDa, and isoelectric point (pI) of 4.5, with no significant identity with other reported turnip peroxidases. Sequence alignment showed that only three peroxidases have a significant identity with BnPA namely AtP29a (84%), and AtPA2 (81%) from Arabidopsis thaliana, and HRPA2 (82%) from horseradish (Armoracia rusticana). Work is in progress to clone this gene into an adequate host to study the specific role and possible biotechnological applications of this alternative peroxidase source.

  15. Construction and characterization of a cDNA library from human ...

    African Journals Online (AJOL)

    The tumor-suppressor gene p53 and its downstream genes consist of a complicated gene network, and the challenge to understand the network is to identify p53 downstream genes. In order to isolate and identify new p53 regulated genes, we constructed and characterized a normalized cDNA library from human brain ...

  16. cDNA, genomic sequence cloning and analysis of the ribosomal ...

    African Journals Online (AJOL)

    Ribosomal protein L37A (RPL37A) is a component of 60S large ribosomal subunit encoded by the RPL37A gene, which belongs to the family of ribosomal L37AE proteins, located in the cytoplasm. The complementary deoxyribonucleic acid (cDNA) and the genomic sequence of RPL37A were cloned successfully from giant ...

  17. Cloning and sequence analysis of cDNA coding for rat nucleolar protein C23

    International Nuclear Information System (INIS)

    Ghaffari, S.H.; Olson, M.O.J.

    1986-01-01

    Using synthetic oligonucleotides as primers and probes, the authors have isolated and sequenced cDNA clones encoding protein C23, a putative nucleolus organizer protein. Poly(A + ) RNA was isolated from rat Novikoff hepatoma cells and enriched in C23 mRNA by sucrose density gradient ultracentrifugation. Two deoxyoligonuleotides, a 48- and a 27-mer, were synthesized on the basis of amino acid sequence from the C-terminal half of protein C23 and cDNA sequence data from CHO cell protein. The 48-mer was used a primer for synthesis of cDNA which was then inserted into plasmid pUC9. Transformed bacterial colonies were screened by hybridization with 32 P labeled 27-mer. Two clones among 5000 gave a strong positive signal. Plasmid DNAs from these clones were purified and characterized by blotting and nucleotide sequence analysis. The length of C23 mRNA was estimated to be 3200 bases in a northern blot analysis. The sequence of a 267 b.p. insert shows high homology with the CHO cDNA with only 9 nucleotide differences and an identical amino acid sequence. These studies indicate that this region of the protein is highly conserved

  18. Isolation and characterization of two cDNA clones encoding for glutamate dehydrogenase in Nicotiana plumbaginifolia.

    Science.gov (United States)

    Ficarelli, A; Tassi, F; Restivo, F M

    1999-03-01

    We have isolated two full length cDNA clones encoding Nicotiana plumbaginifolia NADH-glutamate dehydrogenase. Both clones share amino acid boxes of homology corresponding to conserved GDH catalytic domains and putative mitochondrial targeting sequence. One clone shows a putative EF-hand loop. The level of the two transcripts is affected differently by carbon source.

  19. cDNA sequence and tissue expression analysis of glucokinase from ...

    African Journals Online (AJOL)

    Yomi

    2012-01-10

    Jan 10, 2012 ... distribution of GK mRNA in brain, mesenteric adipose tissue, spleen, white muscle and liver of grass ... expression profile of GK mRNA in liver normalized with β-actin level was 31, 454 and 649-fold compared .... Primers and expected products used for GK gene cDNA RT-PCR, RACE and real-time PCR.

  20. Cloning of oleosin, a putative new hazelnut allergen, using a hazelnut cDNA library

    NARCIS (Netherlands)

    Akkerdaas, Jaap H.; Schocker, Frauke; Vieths, Stefan; Versteeg, Serge; Zuidmeer, Laurian; Hefle, Sue L.; Aalberse, Rob C.; Richter, Klaus; Ferreira, Fatima; van Ree, Ronald

    2006-01-01

    The clinical presentation of non-pollen related allergy to hazelnut can be severe and systemic. So far, only a limited number of non-pollen related hazelnut allergens have been identified and characterized. The aim of this study was to identify and clone new hazelnut allergens. A lambda ZAP cDNA

  1. cDNA cloning and primary structure analysis of invariant chain in ...

    African Journals Online (AJOL)

    cDNA cloning and primary structure analysis of invariant chain in Chinese Pengze crucian carp. X Liu, W Yu, J Li, F Chen, S Liu, C Wu, J Xu. Abstract. Invariant chain (Ii) plays an important role in MHC class II molecules assembly and exogenous peptide presentation in vertebrates. Although mammalian Ii has been ...

  2. A Sol-gel Integrated Dual-readout Microarray Platform for Quantification and Identification of Prostate-specific Antigen.

    Science.gov (United States)

    Lee, SangWook; Lee, Jong Hyun; Kwon, Hyuck Gi; Laurell, Thomas; Jeong, Ok Chan; Kim, Soyoun

    2018-01-01

    Here, we report a sol-gel integrated affinity microarray for on-chip matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) that enables capture and identification of prostate?specific antigen (PSA) in samples. An anti-PSA antibody (H117) was mixed with a sol?gel, and the mixture was spotted onto a porous silicon (pSi) surface without additional surface modifications. The antibody easily penetrates the sol-gel macropore fluidic network structure, making possible high affinities. To assess the capture affinity of the platform, we performed a direct assay using fluorescein isothiocyanate-labeled PSA. Pure PSA was subjected to on-chip MALDI-TOF-MS analysis, yielding three clear mass peptide peaks (m/z = 1272, 1407, and 1872). The sol-gel microarray platform enables dual readout of PSA both fluorometric and MALDI-TOF MS analysis in biological samples. Here we report a useful method for a means for discovery of biomarkers in complex body fluids.

  3. Crack imaging by pulsed laser spot thermography

    International Nuclear Information System (INIS)

    Li, T; Almond, D P; Rees, D A S; Weekes, B

    2010-01-01

    A surface crack close to a spot heated by a laser beam impedes lateral heat flow and produces alterations to the shape of the thermal image of the spot that can be monitored by thermography. A full 3D simulation has been developed to simulate heat flow from a laser heated spot in the proximity of a crack. The modelling provided an understanding of the ways that different parameters affect the thermal images of laser heated spots. It also assisted in the development of an efficient image processing strategy for extracting the scanned cracks. Experimental results show that scanning pulsed laser spot thermography has considerable potential as a remote, non-contact crack imaging technique.

  4. Laser Spot Detection Based on Reaction Diffusion.

    Science.gov (United States)

    Vázquez-Otero, Alejandro; Khikhlukha, Danila; Solano-Altamirano, J M; Dormido, Raquel; Duro, Natividad

    2016-03-01

    Center-location of a laser spot is a problem of interest when the laser is used for processing and performing measurements. Measurement quality depends on correctly determining the location of the laser spot. Hence, improving and proposing algorithms for the correct location of the spots are fundamental issues in laser-based measurements. In this paper we introduce a Reaction Diffusion (RD) system as the main computational framework for robustly finding laser spot centers. The method presented is compared with a conventional approach for locating laser spots, and the experimental results indicate that RD-based computation generates reliable and precise solutions. These results confirm the flexibility of the new computational paradigm based on RD systems for addressing problems that can be reduced to a set of geometric operations.

  5. Laser Spot Detection Based on Reaction Diffusion

    Directory of Open Access Journals (Sweden)

    Alejandro Vázquez-Otero

    2016-03-01

    Full Text Available Center-location of a laser spot is a problem of interest when the laser is used for processing and performing measurements. Measurement quality depends on correctly determining the location of the laser spot. Hence, improving and proposing algorithms for the correct location of the spots are fundamental issues in laser-based measurements. In this paper we introduce a Reaction Diffusion (RD system as the main computational framework for robustly finding laser spot centers. The method presented is compared with a conventional approach for locating laser spots, and the experimental results indicate that RD-based computation generates reliable and precise solutions. These results confirm the flexibility of the new computational paradigm based on RD systems for addressing problems that can be reduced to a set of geometric operations.

  6. Rocky Mountain spotted fever in Argentina.

    Science.gov (United States)

    Paddock, Christopher D; Fernandez, Susana; Echenique, Gustavo A; Sumner, John W; Reeves, Will K; Zaki, Sherif R; Remondegui, Carlos E

    2008-04-01

    We describe the first molecular confirmation of Rickettsia rickettsii, the cause of Rocky Mountain spotted fever (RMSF), from a tick vector, Amblyomma cajennense, and from a cluster of fatal spotted fever cases in Argentina. Questing A. cajennense ticks were collected at or near sites of presumed or confirmed cases of spotted fever rickettsiosis in Jujuy Province and evaluated by polymerase chain reaction assays for spotted fever group rickettsiae. DNA of R. rickettsii was amplified from a pool of A. cajennense ticks and from tissues of one of four patients who died during 2003-2004 after illnesses characterized by high fever, severe headache, myalgias, and petechial rash. The diagnosis of spotted fever rickettsiosis was confirmed in the other patients by indirect immunofluorescence antibody and immunohistochemical staining techniques. These findings show the existence of RMSF in Argentina and emphasize the need for clinicians throughout the Americas to consider RMSF in patients with febrile rash illnesses.

  7. Sensitivity and fidelity of DNA microarray improved with integration of Amplified Differential Gene Expression (ADGE

    Directory of Open Access Journals (Sweden)

    Ile Kristina E

    2003-07-01

    Full Text Available Abstract Background The ADGE technique is a method designed to magnify the ratios of gene expression before detection. It improves the detection sensitivity to small change of gene expression and requires small amount of starting material. However, the throughput of ADGE is low. We integrated ADGE with DNA microarray (ADGE microarray and compared it with regular microarray. Results When ADGE was integrated with DNA microarray, a quantitative relationship of a power function between detected and input ratios was found. Because of ratio magnification, ADGE microarray was better able to detect small changes in gene expression in a drug resistant model cell line system. The PCR amplification of templates and efficient labeling reduced the requirement of starting material to as little as 125 ng of total RNA for one slide hybridization and enhanced the signal intensity. Integration of ratio magnification, template amplification and efficient labeling in ADGE microarray reduced artifacts in microarray data and improved detection fidelity. The results of ADGE microarray were less variable and more reproducible than those of regular microarray. A gene expression profile generated with ADGE microarray characterized the drug resistant phenotype, particularly with reference to glutathione, proliferation and kinase pathways. Conclusion ADGE microarray magnified the ratios of differential gene expression in a power function, improved the detection sensitivity and fidelity and reduced the requirement for starting material while maintaining high throughput. ADGE microarray generated a more informative expression pattern than regular microarray.

  8. Carbon Nanotube Microarrays Grown on Nanoflake Substrates

    Science.gov (United States)

    Schmidt, Howard K.; Hauge, Robert H.; Pint, Cary; Pheasant, Sean

    2013-01-01

    coated on U.S. currency. After deposition, the growth is carried out in a hot-filament chemical vapor deposition apparatus. A tungsten hot filament placed in the flow of H2 at a temperature greater than 1,600 C creates atomic hydrogen, which serves to reduce the Fe catalyst into a metallic state. The catalyst can now precipitate SWNTs in the presence of growth gases. The gases used for the experiments reported are C2H2, H2O, and H2, at rates of 2, 2, and 400 standard cubic centimeters per minute (sccm), respectively. In order to retain the flakes, a cage is constructed by spot welding stainless steel or copper mesh to form an enclosed area, in which the flakes are placed prior to growth. This allows growth gases and atomic hydrogen to reach the flakes, but does not allow the flakes, which rapidly nucleate SWNTs, to escape from the cage.

  9. Cloning and expression of a cDNA encoding human sterol carrier protein 2

    International Nuclear Information System (INIS)

    Yamamoto, Ritsu; Kallen, C.B.; Babalola, G.O.; Rennert, H.; Strauss, J.F. III; Billheimer, J.T.

    1991-01-01

    The authors report the cloning and expression of a cDNA encoding human sterol carrier protein 2 (SCP 2 ). The 1.3-kilobase (kb) cDNA contains an open reading frame which encompasses a 143-amino acid sequence which is 89% identical to the rat SCP 2 amino acid sequence. The deduced amino acid sequence of the polypeptide reveals a 20-residue amino-terminal leader sequence in front of the mature polypeptide, which contains a carboxyl-terminal tripeptide (Ala-Lys-Leu) related to the peroxisome targeting sequence. The expressed cDNA in COS-7 cells yields a 15.3-kDa polypeptide and increased amounts of a 13.2-kDa polypeptide, both reacting with a specific rabbit antiserum to rat liver SCP 2 . The cDNA insert hybridizes with 3.2- and 1.8-kb mRNA species in human liver poly(A) + RNA. In human fibroblasts and placenta the 1.8-kb mRNA was most abundant. Southern blot analysis suggests either that there are multiple copies of the SCP 2 gene in the human genome or that the SCP 2 gene is very large. Coexpression of the SCP 2 cDNA with expression vectors for cholesterol side-chain cleavage enzyme and adrenodoxin resulted in a 2.5-fold enhancement of progestin synthesis over that obtained with expression of the steroidogenic enzyme system alone. These findings are concordant with the notion that SCP 2 plays a role in regulating steroidogenesis, among other possible functions

  10. Primary culture of cat intestinal epithelial cells in vitro and the cDNA library construction.

    Science.gov (United States)

    Zhao, Gui Hua; Liu, Ye; Cheng, Yun Tang; Zhao, Qing Song; Qiu, Xiao; Xu, Chao; Xiao, Ting; Zhu, Song; Liu, Gong Zhen; Yin, Kun

    2018-06-26

    Felids are the only definitive hosts of Toxoplasma gondii. To lay a foundation for screening the T. gondii-felids interaction factors, we have developed a reproducible primary culture method for cat intestinal epithelial cells (IECs). The primary IECs were isolated from a new born cat's small intestine jejunum region without food ingress, and respectively in vitro cultured by tissue cultivation and combined digestion method with collagenase XI and dispase I, then purified by trypsinization. After identification, the ds cDNA of cat IECs was synthesized for constructing pGADT7 homogenization three-frame plasmid, and transformed into the yeast Y187 for generating the cDNA library. Our results indicated that cultivation of primary cat IECs relays on combined digestion to form polarized and confluent monolayers within 3 days with typical features of normal epithelial cells. The purified cells cultured by digestion method were identified to be nature intestinal epithelial cells using immunohistochemical analysis and were able to maintain viability for at least 15 passages. The homogenizable ds cDNA, which is synthesized from the total RNA extracted from our cultured IECs, distributed among 0.5-2.0 kb, and generated satisfying three-frame cDNA library with the capacity of 1.2 × 106 and the titer of 5.2 × 107 pfu/mL. Our results established an optimal method for the culturing and passage of cat IECs model in vitro, and laid a cDNA library foundation for the subsequent interaction factors screening by yeast two-hybrid.

  11. The tissue microarray OWL schema: An open-source tool for sharing tissue microarray data

    Directory of Open Access Journals (Sweden)

    Hyunseok P Kang

    2010-01-01

    Full Text Available Background: Tissue microarrays (TMAs are enormously useful tools for translational research, but incompatibilities in database systems between various researchers and institutions prevent the efficient sharing of data that could help realize their full potential. Resource Description Framework (RDF provides a flexible method to represent knowledge in triples, which take the form Subject- Predicate-Object. All data resources are described using Uniform Resource Identifiers (URIs, which are global in scope. We present an OWL (Web Ontology Language schema that expands upon the TMA data exchange specification to address this issue and assist in data sharing and integration. Methods: A minimal OWL schema was designed containing only concepts specific to TMA experiments. More general data elements were incorporated from predefined ontologies such as the NCI thesaurus. URIs were assigned using the Linked Data format. Results: We present examples of files utilizing the schema and conversion of XML data (similar to the TMA DES to OWL. Conclusion: By utilizing predefined ontologies and global unique identifiers, this OWL schema provides a solution to the limitations of XML, which represents concepts defined in a localized setting. This will help increase the utilization of tissue resources, facilitating collaborative translational research efforts.

  12. Performance of a polymer coated silicon microarray for simultaneous detection of food allergen-specific IgE and IgG4.

    Science.gov (United States)

    Sievers, S; Cretich, M; Gagni, P; Ahrens, B; Grishina, G; Sampson, H A; Niggemann, B; Chiari, M; Beyer, K

    2017-08-01

    Microarray-based component-resolved diagnostics (CRD) has become an accepted tool to detect allergen-specific IgE sensitization towards hundreds of allergens in parallel from one drop of serum. Nevertheless, specificity and sensitivity as well as a simultaneous detection of allergen-specific IgG 4 , as a potential parameter for tolerance development, remain to be optimized. We applied the recently introduced silicon chip coated with a functional polymer named copoly(DMA-NAS-MAPS) to the simultaneous detection of food allergen-specific IgE and IgG 4 , and compared it with ImmunoCAP and ImmunoCAP ISAC. Inter- and intraslide variation, linearity of signal and working range, sensitivity and application of internal calibrations for IgE and IgG 4 were assessed. Native and recombinant allergenic proteins from hen's egg and cow's milk were spotted on silicon chips coated with copoly(DMA-NAS-MAPS) along with known concentrations for human IgE and IgG 4 . A serum pool and 105 patient samples were assessed quantitatively and semi-quantitatively with the ImmunoCAP and ImmunoCAP ISAC and correlated with IgE- and IgG 4 -specific fluorescence on silicon microarrays. Allergen-specific IgE and IgG 4 were detected in parallel using two fluorescent dyes with no crosstalk. Results from the ImmunoCAP correlated better with microarray fluorescence than with ImmunoCAP ISAC except for the allergen ovomucoid. The working range of the silicon microarray for total hen's egg-specific IgE was comparable to the range of 0.1 to >100 kU A /L of the ImmunoCAP system, whereas for total cow's milk, the silicon microarray was less sensitive. Detectable allergen-specific IgG 4 could be determined only for low concentrations, but still correlated positively with ImmunoCAP results. We confirmed the ability of the polymer coated silicon microarray to be comparably sensitive to the ImmunoCAP ISAC for various food allergens. This suggests that the copoly(DMA-NAS-MAPS) microarray is a low-cost, self

  13. Integrating Biological Perspectives:. a Quantum Leap for Microarray Expression Analysis

    Science.gov (United States)

    Wanke, Dierk; Kilian, Joachim; Bloss, Ulrich; Mangelsen, Elke; Supper, Jochen; Harter, Klaus; Berendzen, Kenneth W.

    2009-02-01

    Biologists and bioinformatic scientists cope with the analysis of transcript abundance and the extraction of meaningful information from microarray expression data. By exploiting biological information accessible in public databases, we try to extend our current knowledge over the plant model organism Arabidopsis thaliana. Here, we give two examples of increasing the quality of information gained from large scale expression experiments by the integration of microarray-unrelated biological information: First, we utilize Arabidopsis microarray data to demonstrate that expression profiles are usually conserved between orthologous genes of different organisms. In an initial step of the analysis, orthology has to be inferred unambiguously, which then allows comparison of expression profiles between orthologs. We make use of the publicly available microarray expression data of Arabidopsis and barley, Hordeum vulgare. We found a generally positive correlation in expression trajectories between true orthologs although both organisms are only distantly related in evolutionary time scale. Second, extracting clusters of co-regulated genes implies similarities in transcriptional regulation via similar cis-regulatory elements (CREs). Vice versa approaches, where co-regulated gene clusters are found by investigating on CREs were not successful in general. Nonetheless, in some cases the presence of CREs in a defined position, orientation or CRE-combinations is positively correlated with co-regulated gene clusters. Here, we make use of genes involved in the phenylpropanoid biosynthetic pathway, to give one positive example for this approach.

  14. Kernel Based Nonlinear Dimensionality Reduction and Classification for Genomic Microarray

    Directory of Open Access Journals (Sweden)

    Lan Shu

    2008-07-01

    Full Text Available Genomic microarrays are powerful research tools in bioinformatics and modern medicinal research because they enable massively-parallel assays and simultaneous monitoring of thousands of gene expression of biological samples. However, a simple microarray experiment often leads to very high-dimensional data and a huge amount of information, the vast amount of data challenges researchers into extracting the important features and reducing the high dimensionality. In this paper, a nonlinear dimensionality reduction kernel method based locally linear embedding(LLE is proposed, and fuzzy K-nearest neighbors algorithm which denoises datasets will be introduced as a replacement to the classical LLE’s KNN algorithm. In addition, kernel method based support vector machine (SVM will be used to classify genomic microarray data sets in this paper. We demonstrate the application of the techniques to two published DNA microarray data sets. The experimental results confirm the superiority and high success rates of the presented method.

  15. The microarray detecting six fruit-tree viruses

    Czech Academy of Sciences Publication Activity Database

    Lenz, Ondřej; Petrzik, Karel; Špak, Josef

    2009-01-01

    Roč. 148, July (2009), s. 27 ISSN 1866-590X. [International Conference on Virus and other Graft Transmissible Diseases of Fruit Crops /21./. 05.07.2009-10.07.2009, Neustadt] R&D Projects: GA MŠk OC 853.001 Institutional research plan: CEZ:AV0Z50510513 Keywords : microarray * detection * virus Subject RIV: EE - Microbiology, Virology

  16. Dimension reduction methods for microarray data: a review

    Directory of Open Access Journals (Sweden)

    Rabia Aziz

    2017-03-01

    Full Text Available Dimension reduction has become inevitable for pre-processing of high dimensional data. “Gene expression microarray data” is an instance of such high dimensional data. Gene expression microarray data displays the maximum number of genes (features simultaneously at a molecular level with a very small number of samples. The copious numbers of genes are usually provided to a learning algorithm for producing a complete characterization of the classification task. However, most of the times the majority of the genes are irrelevant or redundant to the learning task. It will deteriorate the learning accuracy and training speed as well as lead to the problem of overfitting. Thus, dimension reduction of microarray data is a crucial preprocessing step for prediction and classification of disease. Various feature selection and feature extraction techniques have been proposed in the literature to identify the genes, that have direct impact on the various machine learning algorithms for classification and eliminate the remaining ones. This paper describes the taxonomy of dimension reduction methods with their characteristics, evaluation criteria, advantages and disadvantages. It also presents a review of numerous dimension reduction approaches for microarray data, mainly those methods that have been proposed over the past few years.

  17. GenePublisher: automated analysis of DNA microarray data

    DEFF Research Database (Denmark)

    Knudsen, Steen; Workman, Christopher; Sicheritz-Ponten, T.

    2003-01-01

    GenePublisher, a system for automatic analysis of data from DNA microarray experiments, has been implemented with a web interface at http://www.cbs.dtu.dk/services/GenePublisher. Raw data are uploaded to the server together with aspecification of the data. The server performs normalization...

  18. Towards a programmable magnetic bead microarray in a microfluidic channel

    DEFF Research Database (Denmark)

    Smistrup, Kristian; Bruus, Henrik; Hansen, Mikkel Fougt

    2007-01-01

    to use larger currents and obtain forces of longer range than from thin current lines at a given power limit. Guiding of magnetic beads in the hybrid magnetic separator and the construction of a programmable microarray of magnetic beads in the microfluidic channel by hydrodynamic focusing is presented....

  19. Comparison of Comparative Genomic Hybridization Technologies across Microarray Platforms

    Science.gov (United States)

    In the 2007 Association of Biomolecular Resource Facilities (ABRF) Microarray Research Group (MARG) project, we analyzed HL-60 DNA with five platforms: Agilent, Affymetrix 500K, Affymetrix U133 Plus 2.0, Illumina, and RPCI 19K BAC arrays. Copy number variation (CNV) was analyzed ...

  20. CONFIRMING MICROARRAY DATA--IS IT REALLY NECESSARY?

    Science.gov (United States)

    The generation of corroborative data has become a commonly used approach for ensuring the veracity of microarray data. Indeed, the need to conduct corroborative studies has now become official editorial policy for at least two journals, and several more are considering introducin...

  1. Microarrays: Molecular allergology and nanotechnology for personalised medicine (II).

    Science.gov (United States)

    Lucas, J M

    2010-01-01

    Progress in nanotechnology and DNA recombination techniques have produced tools for the diagnosis and investigation of allergy at molecular level. The most advanced examples of such progress are the microarray techniques, which have been expanded not only in research in the field of proteomics but also in application to the clinical setting. Microarrays of allergic components offer results relating to hundreds of allergenic components in a single test, and using a small amount of serum which can be obtained from capillary blood. The availability of new molecules will allow the development of panels including new allergenic components and sources, which will require evaluation for clinical use. Their application opens the door to component-based diagnosis, to the holistic perception of sensitisation as represented by molecular allergy, and to patient-centred medical practice by allowing great diagnostic accuracy and the definition of individualised immunotherapy for each patient. The present article reviews the application of allergenic component microarrays to allergology for diagnosis, management in the form of specific immunotherapy, and epidemiological studies. A review is also made of the use of protein and gene microarray techniques in basic research and in allergological diseases. Lastly, an evaluation is made of the challenges we face in introducing such techniques to clinical practice, and of the future perspectives of this new technology. Copyright 2010 SEICAP. Published by Elsevier Espana. All rights reserved.

  2. Broad spectrum microarray for fingerprint-based bacterial species identification

    Directory of Open Access Journals (Sweden)

    Frey Jürg E

    2010-02-01

    Full Text Available Abstract Background Microarrays are powerful tools for DNA-based molecular diagnostics and identification of pathogens. Most target a limited range of organisms and are based on only one or a very few genes for specific identification. Such microarrays are limited to organisms for which specific probes are available, and often have difficulty discriminating closely related taxa. We have developed an alternative broad-spectrum microarray that employs hybridisation fingerprints generated by high-density anonymous markers distributed over the entire genome for identification based on comparison to a reference database. Results A high-density microarray carrying 95,000 unique 13-mer probes was designed. Optimized methods were developed to deliver reproducible hybridisation patterns that enabled confident discrimination of bacteria at the species, subspecies, and strain levels. High correlation coefficients were achieved between replicates. A sub-selection of 12,071 probes, determined by ANOVA and class prediction analysis, enabled the discrimination of all samples in our panel. Mismatch probe hybridisation was observed but was found to have no effect on the discriminatory capacity of our system. Conclusions These results indicate the potential of our genome chip for reliable identification of a wide range of bacterial taxa at the subspecies level without laborious prior sequencing and probe design. With its high resolution capacity, our proof-of-principle chip demonstrates great potential as a tool for molecular diagnostics of broad taxonomic groups.

  3. Exploiting fluorescence for multiplex immunoassays on protein microarrays

    International Nuclear Information System (INIS)

    Herbáth, Melinda; Balogh, Andrea; Matkó, János; Papp, Krisztián; Prechl, József

    2014-01-01

    Protein microarray technology is becoming the method of choice for identifying protein interaction partners, detecting specific proteins, carbohydrates and lipids, or for characterizing protein interactions and serum antibodies in a massively parallel manner. Availability of the well-established instrumentation of DNA arrays and development of new fluorescent detection instruments promoted the spread of this technique. Fluorescent detection has the advantage of high sensitivity, specificity, simplicity and wide dynamic range required by most measurements. Fluorescence through specifically designed probes and an increasing variety of detection modes offers an excellent tool for such microarray platforms. Measuring for example the level of antibodies, their isotypes and/or antigen specificity simultaneously can offer more complex and comprehensive information about the investigated biological phenomenon, especially if we take into consideration that hundreds of samples can be measured in a single assay. Not only body fluids, but also cell lysates, extracted cellular components, and intact living cells can be analyzed on protein arrays for monitoring functional responses to printed samples on the surface. As a rapidly evolving area, protein microarray technology offers a great bulk of information and new depth of knowledge. These are the features that endow protein arrays with wide applicability and robust sample analyzing capability. On the whole, protein arrays are emerging new tools not just in proteomics, but glycomics, lipidomics, and are also important for immunological research. In this review we attempt to summarize the technical aspects of planar fluorescent microarray technology along with the description of its main immunological applications. (topical review)

  4. Development of DNA Microarrays for Metabolic Pathway and Bioprocess Monitoring

    Energy Technology Data Exchange (ETDEWEB)

    Gregory Stephanopoulos

    2004-07-31

    Transcriptional profiling experiments utilizing DNA microarrays to study the intracellular accumulation of PHB in Synechocystis has proved difficult in large part because strains that show significant differences in PHB which would justify global analysis of gene expression have not been isolated.

  5. SNP typing on the NanoChip electronic microarray

    DEFF Research Database (Denmark)

    Børsting, Claus; Sanchez Sanchez, Juan Jose; Morling, Niels

    2005-01-01

    We describe a single nucleotide polymorphism (SNP) typing protocol developed for the NanoChip electronic microarray. The NanoChip array consists of 100 electrodes covered by a thin hydrogel layer containing streptavidin. An electric currency can be applied to one, several, or all electrodes...

  6. Application of Microarray technology in research and diagnostics

    DEFF Research Database (Denmark)

    Helweg-Larsen, Rehannah Borup

    The overall purpose of this thesis is to evaluate the use of microarray analysis to investigate the transcriptome of human cancers and human follicular cells and define the correlation between expression of human genes and specific cancer types as well as the developmental competence of the oocyte...

  7. Bacterial identification and subtyping using DNA microarray and DNA sequencing.

    Science.gov (United States)

    Al-Khaldi, Sufian F; Mossoba, Magdi M; Allard, Marc M; Lienau, E Kurt; Brown, Eric D

    2012-01-01

    The era of fast and accurate discovery of biological sequence motifs in prokaryotic and eukaryotic cells is here. The co-evolution of direct genome sequencing and DNA microarray strategies not only will identify, isotype, and serotype pathogenic bacteria, but also it will aid in the discovery of new gene functions by detecting gene expressions in different diseases and environmental conditions. Microarray bacterial identification has made great advances in working with pure and mixed bacterial samples. The technological advances have moved beyond bacterial gene expression to include bacterial identification and isotyping. Application of new tools such as mid-infrared chemical imaging improves detection of hybridization in DNA microarrays. The research in this field is promising and future work will reveal the potential of infrared technology in bacterial identification. On the other hand, DNA sequencing by using 454 pyrosequencing is so cost effective that the promise of $1,000 per bacterial genome sequence is becoming a reality. Pyrosequencing technology is a simple to use technique that can produce accurate and quantitative analysis of DNA sequences with a great speed. The deposition of massive amounts of bacterial genomic information in databanks is creating fingerprint phylogenetic analysis that will ultimately replace several technologies such as Pulsed Field Gel Electrophoresis. In this chapter, we will review (1) the use of DNA microarray using fluorescence and infrared imaging detection for identification of pathogenic bacteria, and (2) use of pyrosequencing in DNA cluster analysis to fingerprint bacterial phylogenetic trees.

  8. Exploring Lactobacillus plantarum genome diversity by using microarrays

    NARCIS (Netherlands)

    Molenaar, D.; Bringel, F.; Schuren, F.H.; Vos, de W.M.; Siezen, R.J.; Kleerebezem, M.

    2005-01-01

    Lactobacillus plantarum is a versatile and flexible species that is encountered in a variety of niches and can utilize a broad range of fermentable carbon sources. To assess if this versatility is linked to a variable gene pool, microarrays containing a subset of small genomic fragments of L.

  9. See what you eat--broad GMO screening with microarrays.

    Science.gov (United States)

    von Götz, Franz

    2010-03-01

    Despite the controversy of whether genetically modified organisms (GMOs) are beneficial or harmful for humans, animals, and/or ecosystems, the number of cultivated GMOs is increasing every year. Many countries and federations have implemented safety and surveillance systems for GMOs. Potent testing technologies need to be developed and implemented to monitor the increasing number of GMOs. First, these GMO tests need to be comprehensive, i.e., should detect all, or at least the most important, GMOs on the market. This type of GMO screening requires a high degree of parallel tests or multiplexing. To date, DNA microarrays have the highest number of multiplexing capabilities when nucleic acids are analyzed. This trend article focuses on the evolution of DNA microarrays for GMO testing. Over the last 7 years, combinations of multiplex PCR detection and microarray detection have been developed to qualitatively assess the presence of GMOs. One example is the commercially available DualChip GMO (Eppendorf, Germany; http://www.eppendorf-biochip.com), which is the only GMO screening system successfully validated in a multicenter study. With use of innovative amplification techniques, promising steps have recently been taken to make GMO detection with microarrays quantitative.

  10. Microarray-Based Identification of Transcription Factor Target Genes

    NARCIS (Netherlands)

    Gorte, M.; Horstman, A.; Page, R.B.; Heidstra, R.; Stromberg, A.; Boutilier, K.A.

    2011-01-01

    Microarray analysis is widely used to identify transcriptional changes associated with genetic perturbation or signaling events. Here we describe its application in the identification of plant transcription factor target genes with emphasis on the design of suitable DNA constructs for controlling TF

  11. Employing image processing techniques for cancer detection using microarray images.

    Science.gov (United States)

    Dehghan Khalilabad, Nastaran; Hassanpour, Hamid

    2017-02-01

    Microarray technology is a powerful genomic tool for simultaneously studying and analyzing the behavior of thousands of genes. The analysis of images obtained from this technology plays a critical role in the detection and treatment of diseases. The aim of the current study is to develop an automated system for analyzing data from microarray images in order to detect cancerous cases. The proposed system consists of three main phases, namely image processing, data mining, and the detection of the disease. The image processing phase performs operations such as refining image rotation, gridding (locating genes) and extracting raw data from images the data mining includes normalizing the extracted data and selecting the more effective genes. Finally, via the extracted data, cancerous cell is recognized. To evaluate the performance of the proposed system, microarray database is employed which includes Breast cancer, Myeloid Leukemia and Lymphomas from the Stanford Microarray Database. The results indicate that the proposed system is able to identify the type of cancer from the data set with an accuracy of 95.45%, 94.11%, and 100%, respectively. Copyright © 2017 Elsevier Ltd. All rights reserved.

  12. Microarray-based RNA profiling of breast cancer

    DEFF Research Database (Denmark)

    Larsen, Martin J; Thomassen, Mads; Tan, Qihua

    2014-01-01

    analyzed the same 234 breast cancers on two different microarray platforms. One dataset contained known batch-effects associated with the fabrication procedure used. The aim was to assess the significance of correcting for systematic batch-effects when integrating data from different platforms. We here...

  13. Microarray expression profiling of human dental pulp from single subject.

    Science.gov (United States)

    Tete, Stefano; Mastrangelo, Filiberto; Scioletti, Anna Paola; Tranasi, Michelangelo; Raicu, Florina; Paolantonio, Michele; Stuppia, Liborio; Vinci, Raffaele; Gherlone, Enrico; Ciampoli, Cristian; Sberna, Maria Teresa; Conti, Pio

    2008-01-01

    Microarray is a recently developed simultaneous analysis of expression patterns of thousand of genes. The aim of this research was to evaluate the expression profile of human healthy dental pulp in order to find the presence of genes activated and encoding for proteins involved in the physiological process of human dental pulp. We report data obtained by analyzing expression profiles of human tooth pulp from single subjects, using an approach based on the amplification of the total RNA. Experiments were performed on a high-density array able to analyse about 21,000 oligonucleotide sequences of about 70 bases in duplicate, using an approach based on the amplification of the total RNA from the pulp of a single tooth. Obtained data were analyzed using the S.A.M. system (Significance Analysis of Microarray) and genes were merged according to their molecular functions and biological process by the Onto-Express software. The microarray analysis revealed 362 genes with specific pulp expression. Genes showing significant high expression were classified in genes involved in tooth development, protoncogenes, genes of collagen, DNAse, Metallopeptidases and Growth factors. We report a microarray analysis, carried out by extraction of total RNA from specimens of healthy human dental pulp tissue. This approach represents a powerful tool in the study of human normal and pathological pulp, allowing minimization of the genetic variability due to the pooling of samples from different individuals.

  14. Microarray analysis of the gene expression profile in triethylene ...

    African Journals Online (AJOL)

    Microarray analysis of the gene expression profile in triethylene glycol dimethacrylate-treated human dental pulp cells. ... Conclusions: Our results suggest that TEGDMA can change the many functions of hDPCs through large changes in gene expression levels and complex interactions with different signaling pathways.

  15. Comparison of gene coverage of mouse oligonucleotide microarray platforms

    Directory of Open Access Journals (Sweden)

    Medrano Juan F

    2006-03-01

    Full Text Available Abstract Background The increasing use of DNA microarrays for genetical genomics studies generates a need for platforms with complete coverage of the genome. We have compared the effective gene coverage in the mouse genome of different commercial and noncommercial oligonucleotide microarray platforms by performing an in-house gene annotation of probes. We only used information about probes that is available from vendors and followed a process that any researcher may take to find the gene targeted by a given probe. In order to make consistent comparisons between platforms, probes in each microarray were annotated with an Entrez Gene id and the chromosomal position for each gene was obtained from the UCSC Genome Browser Database. Gene coverage was estimated as the percentage of Entrez Genes with a unique position in the UCSC Genome database that is tested by a given microarray platform. Results A MySQL relational database was created to store the mapping information for 25,416 mouse genes and for the probes in five microarray platforms (gene coverage level in parenthesis: Affymetrix430 2.0 (75.6%, ABI Genome Survey (81.24%, Agilent (79.33%, Codelink (78.09%, Sentrix (90.47%; and four array-ready oligosets: Sigma (47.95%, Operon v.3 (69.89%, Operon v.4 (84.03%, and MEEBO (84.03%. The differences in coverage between platforms were highly conserved across chromosomes. Differences in the number of redundant and unspecific probes were also found among arrays. The database can be queried to compare specific genomic regions using a web interface. The software used to create, update and query the database is freely available as a toolbox named ArrayGene. Conclusion The software developed here allows researchers to create updated custom databases by using public or proprietary information on genes for any organisms. ArrayGene allows easy comparisons of gene coverage between microarray platforms for any region of the genome. The comparison presented here

  16. Workflows for microarray data processing in the Kepler environment

    Directory of Open Access Journals (Sweden)

    Stropp Thomas

    2012-05-01

    Full Text Available Abstract Background Microarray data analysis has been the subject of extensive and ongoing pipeline development due to its complexity, the availability of several options at each analysis step, and the development of new analysis demands, including integration with new data sources. Bioinformatics pipelines are usually custom built for different applications, making them typically difficult to modify, extend and repurpose. Scientific workflow systems are intended to address these issues by providing general-purpose frameworks in which to develop and execute such pipelines. The Kepler workflow environment is a well-established system under continual development that is employed in several areas of scientific research. Kepler provides a flexible graphical interface, featuring clear display of parameter values, for design and modification of workflows. It has capabilities for developing novel computational components in the R, Python, and Java programming languages, all of which are widely used for bioinformatics algorithm development, along with capabilities for invoking external applications and using web services. Results We developed a series of fully functional bioinformatics pipelines addressing common tasks in microarray processing in the Kepler workflow environment. These pipelines consist of a set of tools for GFF file processing of NimbleGen chromatin immunoprecipitation on microarray (ChIP-chip datasets and more comprehensive workflows for Affymetrix gene expression microarray bioinformatics and basic primer design for PCR experiments, which are often used to validate microarray results. Although functional in themselves, these workflows can be easily customized, extended, or repurposed to match the needs of specific projects and are designed to be a toolkit and starting point for specific applications. These workflows illustrate a workflow programming paradigm focusing on local resources (programs and data and therefore are close to

  17. Workflows for microarray data processing in the Kepler environment.

    Science.gov (United States)

    Stropp, Thomas; McPhillips, Timothy; Ludäscher, Bertram; Bieda, Mark

    2012-05-17

    Microarray data analysis has been the subject of extensive and ongoing pipeline development due to its complexity, the availability of several options at each analysis step, and the development of new analysis demands, including integration with new data sources. Bioinformatics pipelines are usually custom built for different applications, making them typically difficult to modify, extend and repurpose. Scientific workflow systems are intended to address these issues by providing general-purpose frameworks in which to develop and execute such pipelines. The Kepler workflow environment is a well-established system under continual development that is employed in several areas of scientific research. Kepler provides a flexible graphical interface, featuring clear display of parameter values, for design and modification of workflows. It has capabilities for developing novel computational components in the R, Python, and Java programming languages, all of which are widely used for bioinformatics algorithm development, along with capabilities for invoking external applications and using web services. We developed a series of fully functional bioinformatics pipelines addressing common tasks in microarray processing in the Kepler workflow environment. These pipelines consist of a set of tools for GFF file processing of NimbleGen chromatin immunoprecipitation on microarray (ChIP-chip) datasets and more comprehensive workflows for Affymetrix gene expression microarray bioinformatics and basic primer design for PCR experiments, which are often used to validate microarray results. Although functional in themselves, these workflows can be easily customized, extended, or repurposed to match the needs of specific projects and are designed to be a toolkit and starting point for specific applications. These workflows illustrate a workflow programming paradigm focusing on local resources (programs and data) and therefore are close to traditional shell scripting or R

  18. High quality protein microarray using in situ protein purification

    Directory of Open Access Journals (Sweden)

    Fleischmann Robert D

    2009-08-01

    Full Text Available Abstract Background In the postgenomic era, high throughput protein expression and protein microarray technologies have progressed markedly permitting screening of therapeutic reagents and discovery of novel protein functions. Hexa-histidine is one of the most commonly used fusion tags for protein expression due to its small size and convenient purification via immobilized metal ion affinity chromatography (IMAC. This purification process has been adapted to the protein microarray format, but the quality of in situ His-tagged protein purification on slides has not been systematically evaluated. We established methods to determine the level of purification of such proteins on metal chelate-modified slide surfaces. Optimized in situ purification of His-tagged recombinant proteins has the potential to become the new gold standard for cost-effective generation of high-quality and high-density protein microarrays. Results Two slide surfaces were examined, chelated Cu2+ slides suspended on a polyethylene glycol (PEG coating and chelated Ni2+ slides immobilized on a support without PEG coating. Using PEG-coated chelated Cu2+ slides, consistently higher purities of recombinant proteins were measured. An optimized wash buffer (PBST composed of 10 mM phosphate buffer, 2.7 mM KCl, 140 mM NaCl and 0.05% Tween 20, pH 7.4, further improved protein purity levels. Using Escherichia coli cell lysates expressing 90 recombinant Streptococcus pneumoniae proteins, 73 proteins were successfully immobilized, and 66 proteins were in situ purified with greater than 90% purity. We identified several antigens among the in situ-purified proteins via assays with anti-S. pneumoniae rabbit antibodies and a human patient antiserum, as a demonstration project of large scale microarray-based immunoproteomics profiling. The methodology is compatible with higher throughput formats of in vivo protein expression, eliminates the need for resin-based purification and circumvents

  19. Workflows for microarray data processing in the Kepler environment

    Science.gov (United States)

    2012-01-01

    Background Microarray data analysis has been the subject of extensive and ongoing pipeline development due to its complexity, the availability of several options at each analysis step, and the development of new analysis demands, including integration with new data sources. Bioinformatics pipelines are usually custom built for different applications, making them typically difficult to modify, extend and repurpose. Scientific workflow systems are intended to address these issues by providing general-purpose frameworks in which to develop and execute such pipelines. The Kepler workflow environment is a well-established system under continual development that is employed in several areas of scientific research. Kepler provides a flexible graphical interface, featuring clear display of parameter values, for design and modification of workflows. It has capabilities for developing novel computational components in the R, Python, and Java programming languages, all of which are widely used for bioinformatics algorithm development, along with capabilities for invoking external applications and using web services. Results We developed a series of fully functional bioinformatics pipelines addressing common tasks in microarray processing in the Kepler workflow environment. These pipelines consist of a set of tools for GFF file processing of NimbleGen chromatin immunoprecipitation on microarray (ChIP-chip) datasets and more comprehensive workflows for Affymetrix gene expression microarray bioinformatics and basic primer design for PCR experiments, which are often used to validate microarray results. Although functional in themselves, these workflows can be easily customized, extended, or repurposed to match the needs of specific projects and are designed to be a toolkit and starting point for specific applications. These workflows illustrate a workflow programming paradigm focusing on local resources (programs and data) and therefore are close to traditional shell scripting or

  20. Computational biology of genome expression and regulation--a review of microarray bioinformatics.

    Science.gov (United States)

    Wang, Junbai

    2008-01-01

    Microarray technology is being used widely in various biomedical research areas; the corresponding microarray data analysis is an essential step toward the best utilizing of array technologies. Here we review two components of the microarray data analysis: a low level of microarray data analysis that emphasizes the designing, the quality control, and the preprocessing of microarray experiments, then a high level of microarray data analysis that focuses on the domain-specific microarray applications such as tumor classification, biomarker prediction, analyzing array CGH experiments, and reverse engineering of gene expression networks. Additionally, we will review the recent development of building a predictive model in genome expression and regulation studies. This review may help biologists grasp a basic knowledge of microarray bioinformatics as well as its potential impact on the future evolvement of biomedical research fields.