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Sample records for genes determine stability

  1. Determining postural stability

    Science.gov (United States)

    Lieberman, Erez (Inventor); Forth, Katharine E. (Inventor); Paloski, William H. (Inventor)

    2011-01-01

    A method for determining postural stability of a person can include acquiring a plurality of pressure data points over a period of time from at least one pressure sensor. The method can also include the step of identifying a postural state for each pressure data point to generate a plurality of postural states. The method can include the step of determining a postural state of the person at a point in time based on at least the plurality of postural states.

  2. Emulsion stability: determination from turbidity

    Energy Technology Data Exchange (ETDEWEB)

    Reddy, S.R.; Fogler, H.S.

    1981-01-01

    The relationship between particle size and concentration and turbidity has been developed for a polydispersed system. The stability of acoustically prepared emulsions of C36H74 in water were determined from turbidimetry and found to be in agreement with the stability determined by the freezing method. The turbidimetry method can be used for determining the stability of various emulsions easily and inexpensively. 11 references.

  3. Determinants of Price Stabilization in IPOs

    Directory of Open Access Journals (Sweden)

    Antonio Gledson de Carvalho

    2010-12-01

    Full Text Available In the most common mechanism for price stabilization in IPOs, the underwriter distributes stocks in excess of what was contracted (overallotment and eventually covers this short naked position by purchasing stocks in the secondary market (Aftermarket short covering , ASC. This mechanism can be used to avoid price drop or price volatility. This article provides a description of such activity in Brazil. We investigate the determinants of price stabilization in three aspects: amount overallotted, occurrence of ASC and its intensity. Our results indicate that price stabilization is an important activity in Brazilian IPOs and quite similar to that occurring in the US. The three different aspects of price stabilization have different determinants. The amount overallotted depends only on the ex-ante demand conditions. ASC occurs mostly on IPOs characterized by high risk, low ex-ante demand and carried by reputable underwriters. The intensity of the ASC increases with the riskiness and decreases with the ex-ante demand. None of the existing models fully explain these results.

  4. Transcriptional delay stabilizes bistable gene networks

    Science.gov (United States)

    Gupta, Chinmaya; López, José Manuel; Ott, William; Josić, Krešimir; Bennett, Matthew R.

    2014-01-01

    Transcriptional delay can significantly impact the dynamics of gene networks. Here we examine how such delay affects bistable systems. We investigate several stochastic models of bistable gene networks and find that increasing delay dramatically increases the mean residence times near stable states. To explain this, we introduce a non-Markovian, analytically tractable reduced model. The model shows that stabilization is the consequence of an increased number of failed transitions between stable states. Each of the bistable systems that we simulate behaves in this manner. PMID:23952450

  5. Engineering stability in gene networks by autoregulation

    Science.gov (United States)

    Becskei, Attila; Serrano, Luis

    2000-06-01

    The genetic and biochemical networks which underlie such things as homeostasis in metabolism and the developmental programs of living cells, must withstand considerable variations and random perturbations of biochemical parameters. These occur as transient changes in, for example, transcription, translation, and RNA and protein degradation. The intensity and duration of these perturbations differ between cells in a population. The unique state of cells, and thus the diversity in a population, is owing to the different environmental stimuli the individual cells experience and the inherent stochastic nature of biochemical processes (for example, refs 5 and 6). It has been proposed, but not demonstrated, that autoregulatory, negative feedback loops in gene circuits provide stability, thereby limiting the range over which the concentrations of network components fluctuate. Here we have designed and constructed simple gene circuits consisting of a regulator and transcriptional repressor modules in Escherichia coli and we show the gain of stability produced by negative feedback.

  6. DETERMINATION OF STABILITY CONSTANTS OF MANGANESE (II ...

    African Journals Online (AJOL)

    DR. AMINU

    Keywords: Amino acids, dissociation constant, potentiometry, stability constant. INTRODUCTION ... constants of manganese (II) amino acid complexes using potentiometer. .... Principles of Biochemistry Third Edition,. Worth publishers, 41 ...

  7. Determination of Stability from Multicomponent Pesticide Mixes.

    Science.gov (United States)

    Dorweiler, Kelly J; Gurav, Jagdish N; Walbridge, James S; Ghatge, Vishwas S; Savant, Rahul H

    2016-08-10

    A study was conducted to evaluate the stability of 528 pesticides, metabolites, and contaminants prepared in large multicomponent mixes to enhance laboratory efficiency by allowing maximum use of the useful shelf life of the mixtures. Accelerated aging at 50 °C simulated 6 month, 1 year, and 2 year storage periods at -20 °C. Initial mixture composition was based on the instrument of analysis. After preliminary stability data had been obtained, mixtures were reformulated and re-evaluated. In all, 344 compounds showed satisfactory stability across all treatment groups, 100 compounds showed statistically significant changes between the control and the 6 month simulated storage period (27 with losses >20%), and the remainder showed borderline stability or were tested in one protocol. Stability behavior for organophosphates agreed with the proposed reaction mechanism responsible for acetylcholinesterase inhibition. A small number of compounds increased in response over time, suggesting the occurrence of degradation of precursor pesticides into these respective compounds.

  8. Stability and Hopf Bifurcation Analysis of a Gene Expression Model with Diffusion and Time Delay

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    Yahong Peng

    2014-01-01

    Full Text Available We consider a model for gene expression with one or two time delays and diffusion. The local stability and delay-induced Hopf bifurcation are investigated. We also derive the formulas determining the direction and the stability of Hopf bifurcations by calculating the normal form on the center manifold.

  9. Determining Semantically Related Significant Genes.

    Science.gov (United States)

    Taha, Kamal

    2014-01-01

    GO relation embodies some aspects of existence dependency. If GO term xis existence-dependent on GO term y, the presence of y implies the presence of x. Therefore, the genes annotated with the function of the GO term y are usually functionally and semantically related to the genes annotated with the function of the GO term x. A large number of gene set enrichment analysis methods have been developed in recent years for analyzing gene sets enrichment. However, most of these methods overlook the structural dependencies between GO terms in GO graph by not considering the concept of existence dependency. We propose in this paper a biological search engine called RSGSearch that identifies enriched sets of genes annotated with different functions using the concept of existence dependency. We observe that GO term xcannot be existence-dependent on GO term y, if x- and y- have the same specificity (biological characteristics). After encoding into a numeric format the contributions of GO terms annotating target genes to the semantics of their lowest common ancestors (LCAs), RSGSearch uses microarray experiment to identify the most significant LCA that annotates the result genes. We evaluated RSGSearch experimentally and compared it with five gene set enrichment systems. Results showed marked improvement.

  10. Stability of waves using the Fredholm determinant

    CERN Document Server

    Karambal, Issa

    2011-01-01

    We present a new method for reducing the Fredholm determinant associated with an underlying Birman-Schwinger operator to a finite dimensional determinant. Moreover, we compute explicitly the connection between the Fredholm determinant and the Evans function for travelling wave problems of all orders, in one dimension

  11. Determination of the frequency of polymorphisms in genes related to the genome stability maintenance of the population residing at Monte Alegre, PA (Brazil) municipality; Determinacao da frequencia de polimorfismos em genes relacionados a manutencao da estabilidade do genoma na populacao residente no municipio de Monte Alegre, PA

    Energy Technology Data Exchange (ETDEWEB)

    Hozumi, Cristiny Gomes

    2010-07-01

    The human exposure to ionizing radiation coming from natural sources is an inherent feature of human life on earth, for man and all living things have always been exposed to these sources. Ionizing radiation is a known genotoxic agent which can affect the genomic stability and genes related to DNA repair may play a role when they have committed certain polymorphism. This study aimed to analyze the frequency of polymorphisms (SNPs) in genes of DNA repair and cell cycle control: hOGG1 (Ser326Cys), XRCC3 (Thr241 Met) and p53 (Arg72Pro) in saliva samples from a population located Monte Alegre, state of Para were collected in August 2008 and 40 samples of men and 46 samples of women, adding a total of 86 samples. By RFLP was determined the frequency of homozygous genotypes and / or heterozygous for polymorphic genes. The I)OGG1 gene was 5% of the allele 326Cys, XRCC3 gene found about 21 % of the allele 241 Met and p53 gene showed 40.8% of the 72Pro allele. And the genotype frequencies of individuals for the three genes were 91.04%, 88.06% and 59.7% for homozygous wild genotype, 5.97%, 11.94% and 22.39% for heterozygote genotype and 2,99%, zero and 17:91% for homozygous polymorphic hOGG1 genes respectively, XRCC3, p53. These values are similar to those found in previous studies. The influence of these polymorphisms, which are involved in DNA repair and consequent genotoxicity induced by radiation depends on dose and exposure factors such as smoking, which is statistically a factor in public health surveillance in the region. This study gathered information and molecular epidemiology in Monte Alegre, that help to characterization of local population. (author)

  12. Stability Indicating HPTLC Determination of Meloxicam.

    Science.gov (United States)

    Desai, Namita; Amin, Purnima

    2008-09-01

    A simple, selective, precise and stability-indicating high-performance thin layer chromatographic method of analysis of meloxicam both as a bulk drug and in formulation has been developed. The mobile phase selected was ethyl acetate:cyclohexane:glacial acetic acid (6.5:3.5:0.02% v/v/v). The calibration curve of the drug was linear in the range of 100-500 ng. The spectrodensitometric analysis was carried out in the absorbance mode at 353 nm. The mean (±RSD) values of slope, correlation coefficient and intercept were 3183.8±0.358, 0.9996±0.0321 and 13012±7.1 respectively. The system precision and the method precision studies were carried out with RSD of 0.83 and 1.89 respectively. The limit of detection and quantitation were 30 ng and 99 ng respectively. The mean percent recovery was found to be 100.3%. The method was used to analyze meloxicam from marketed tablet formulation in the presence of commonly used excipients.

  13. Superposition of transcriptional behaviors determines gene state.

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    Sol Efroni

    Full Text Available We introduce a novel technique to determine the expression state of a gene from quantitative information measuring its expression. Adopting a productive abstraction from current thinking in molecular biology, we consider two expression states for a gene--Up or Down. We determine this state by using a statistical model that assumes the data behaves as a combination of two biological distributions. Given a cohort of hybridizations, our algorithm predicts, for the single reading, the probability of each gene's being in an Up or a Down state in each hybridization. Using a series of publicly available gene expression data sets, we demonstrate that our algorithm outperforms the prevalent algorithm. We also show that our algorithm can be used in conjunction with expression adjustment techniques to produce a more biologically sound gene-state call. The technique we present here enables a routine update, where the continuously evolving expression level adjustments feed into gene-state calculations. The technique can be applied in almost any multi-sample gene expression experiment, and holds equal promise for protein abundance experiments.

  14. Major role for mRNA stability in shaping the kinetics of gene induction

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    Zeller Karen I

    2010-04-01

    Full Text Available Abstract Background mRNA levels in cells are determined by the relative rates of RNA production and degradation. Yet, to date, most analyses of gene expression profiles were focused on mechanisms which regulate transcription, while the role of mRNA stability in modulating transcriptional networks was to a large extent overlooked. In particular, kinetic waves in transcriptional responses are usually interpreted as resulting from sequential activation of transcription factors. Results In this study, we examined on a global scale the role of mRNA stability in shaping the kinetics of gene response. Analyzing numerous expression datasets we revealed a striking global anti-correlation between rapidity of induction and mRNA stability, fitting the prediction of a kinetic mathematical model. In contrast, the relationship between kinetics and stability was less significant when gene suppression was analyzed. Frequently, mRNAs that are stable under standard conditions were very rapidly down-regulated following stimulation. Such effect cannot be explained even by a complete shut-off of transcription, and therefore indicates intense modulation of RNA stability. Conclusion Taken together, our results demonstrate the key role of mRNA stability in determining induction kinetics in mammalian transcriptional networks.

  15. κMicroarray analysis of relative gene expression stability for selection of internal reference genes in the rhesus macaque brain

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    Urbanski Henryk F

    2010-06-01

    Full Text Available Abstract Background Normalization of gene expression data refers to the comparison of expression values using reference standards that are consistent across all conditions of an experiment. In PCR studies, genes designated as "housekeeping genes" have been used as internal reference genes under the assumption that their expression is stable and independent of experimental conditions. However, verification of this assumption is rarely performed. Here we assess the use of gene microarray analysis to facilitate selection of internal reference sequences with higher expression stability across experimental conditions than can be expected using traditional selection methods. We recently demonstrated that relative gene expression from qRT-PCR data normalized using GAPDH, ALG9 and RPL13A expression values mirrored relative expression using quantile normalization in Robust Multichip Analysis (RMA on the Affymetrix® GeneChip® rhesus Macaque Genome Array. Having shown that qRT-PCR and Affymetrix® GeneChip® data from the same hormone replacement therapy (HRT study yielded concordant results, we used quantile-normalized gene microarray data to identify the most stably expressed among probe sets for prospective internal reference genes across three brain regions from the HRT study and an additional study of normally menstruating rhesus macaques (cycle study. Gene selection was limited to 575 previously published human "housekeeping" genes. Twelve animals were used per study, and three brain regions were analyzed from each animal. Gene expression stabilities were determined using geNorm, NormFinder and BestKeeper software packages. Results Sequences co-annotated for ribosomal protein S27a (RPS27A, and ubiquitin were among the most stably expressed under all conditions and selection criteria used for both studies. Higher annotation quality on the human GeneChip® facilitated more targeted analysis than could be accomplished using the rhesus GeneChip®. In

  16. On the difficulty of determining tearing mode stability

    Energy Technology Data Exchange (ETDEWEB)

    Bishop, C.M.; Connor, J.W.; Hastie, R.J.; Cowley, S.C. (AEA Technology, Culham (United Kingdom))

    1991-04-01

    The effect of local pressure gradients and of a local flattening of the pressure profile (p' {yields} 0) around the resonant surface of a tearing mode is investigated in toroidal geometry. It is shown that the stability index {Delta}', calculated from the ideal outer region, is modified by local profile changes in a way reminiscent of the favourable curvature stabilization of linear and non-linear tearing mode layer theory. If the width of the region of pressure flattening is of the order of the linear resistive layer width, the stabilization from the ideal outer region compensates for the loss of pressure gradient stabilization from the layer, and the overall stability of the mode is largely unaffected. For pressure flattening over a larger region, however, the mode can be strongly destabilized. Since the flattening region may then still be too small to resolve experimentally, this result implies the essential difficulty of determining the tearing mode stability of experimental profiles. (Author).

  17. Determining the Effective Density and Stabilizer Layer Thickness of Sterically Stabilized Nanoparticles

    Science.gov (United States)

    2016-01-01

    A series of model sterically stabilized diblock copolymer nanoparticles has been designed to aid the development of analytical protocols in order to determine two key parameters: the effective particle density and the steric stabilizer layer thickness. The former parameter is essential for high resolution particle size analysis based on analytical (ultra)centrifugation techniques (e.g., disk centrifuge photosedimentometry, DCP), whereas the latter parameter is of fundamental importance in determining the effectiveness of steric stabilization as a colloid stability mechanism. The diblock copolymer nanoparticles were prepared via polymerization-induced self-assembly (PISA) using RAFT aqueous emulsion polymerization: this approach affords relatively narrow particle size distributions and enables the mean particle diameter and the stabilizer layer thickness to be adjusted independently via systematic variation of the mean degree of polymerization of the hydrophobic and hydrophilic blocks, respectively. The hydrophobic core-forming block was poly(2,2,2-trifluoroethyl methacrylate) [PTFEMA], which was selected for its relatively high density. The hydrophilic stabilizer block was poly(glycerol monomethacrylate) [PGMA], which is a well-known non-ionic polymer that remains water-soluble over a wide range of temperatures. Four series of PGMAx–PTFEMAy nanoparticles were prepared (x = 28, 43, 63, and 98, y = 100–1400) and characterized via transmission electron microscopy (TEM), dynamic light scattering (DLS), and small-angle X-ray scattering (SAXS). It was found that the degree of polymerization of both the PGMA stabilizer and core-forming PTFEMA had a strong influence on the mean particle diameter, which ranged from 20 to 250 nm. Furthermore, SAXS was used to determine radii of gyration of 1.46 to 2.69 nm for the solvated PGMA stabilizer blocks. Thus, the mean effective density of these sterically stabilized particles was calculated and determined to lie between 1.19 g

  18. Top mass determination, Higgs inflation, and vacuum stability

    CERN Document Server

    Branchina, Vincenzo; Platania, Alessia

    2014-01-01

    The possibility that new physics beyond the Standard Model (SM) appears only at the Planck scale $M_P$ is often considered. However, it is usually argued that new physics interactions at $M_P$ do not affect the SM stability phase diagram, so the latter is obtained neglecting these terms. According to this diagram, for the current experimental values of the top and Higgs masses, our universe lives in a metastable state (with very long lifetime), near the edge of stability. Contrary to these expectations, however, we show that the stability phase diagram strongly depends on new physics and that, despite claims to the contrary, a more precise determination of the top (as well as of the Higgs) mass will not allow to discriminate between stability, metastability or criticality of the electroweak vacuum. At the same time, we show that the conditions needed for the realization of Higgs inflation scenarios (all obtained neglecting new physics) are too sensitive to the presence of new interactions at $M_P$. Therefore,...

  19. Online attitude determination of a passively magnetically stabilized spacecraft

    Science.gov (United States)

    Burton, R.; Rock, S.; Springmann, J.; Cutler, J.

    2017-04-01

    An online attitude determination filter is developed for a nano satellite that has no onboard attitude sensors or gyros. Specifically, the attitude of NASA Ames Research Center's O/OREOS, a passively magnetically stabilized 3U CubeSat, is determined using only an estimate of the solar vector obtained from solar panel currents. The filter is based upon the existing multiplicative extended Kalman filter (MEKF) but instead of relying on gyros to drive the motion model, the filter instead incorporates a model of the spacecraft's attitude dynamics in the motion model. An attitude determination accuracy of five degrees is demonstrated, a performance verified using flight data from the University of Michigan's RAX-1. Although the filter was designed for the specific problem of a satellite without gyros or attitude determination it could also be used to provide smoothing of noisy gyro signals or to provide a backup in the event of gyro failures.

  20. Stability analysis of a model gene network links aging, stress resistance, and negligible senescence.

    Science.gov (United States)

    Kogan, Valeria; Molodtsov, Ivan; Menshikov, Leonid I; Shmookler Reis, Robert J; Fedichev, Peter

    2015-08-28

    Several animal species are considered to exhibit what is called negligible senescence, i.e. they do not show signs of functional decline or any increase of mortality with age. Recent studies in naked mole rat and long-lived sea urchins showed that these species do not alter their gene-expression profiles with age as much as other organisms do. This is consistent with exceptional endurance of naked mole rat tissues to various genotoxic stresses. We conjectured, therefore, that the lifelong transcriptional stability of an organism may be a key determinant of longevity. We analyzed the stability of a simple genetic-network model and found that under most common circumstances, such a gene network is inherently unstable. Over a time it undergoes an exponential accumulation of gene-regulation deviations leading to death. However, should the repair systems be sufficiently effective, the gene network can stabilize so that gene damage remains constrained along with mortality of the organism. We investigate the relationship between stress-resistance and aging and suggest that the unstable regime may provide a mathematical basis for the Gompertz "law" of aging in many species. At the same time, this model accounts for the apparently age-independent mortality observed in some exceptionally long-lived animals.

  1. Age-related changes in relative expression stability of commonly used housekeeping genes in selected porcine tissues

    Directory of Open Access Journals (Sweden)

    Looft Christian

    2011-10-01

    Full Text Available Abstract Background Gene expression analysis using real-time RT-PCR (qRT-PCR is increasingly important in biological research due to the high-throughput and accuracy of qRT-PCR. For accurate and reliable gene expression analysis, normalization of gene expression data against housekeeping genes or internal control genes is required. The stability of reference genes has a tremendous effect on the results of relative quantification of gene expression by qRT-PCR. The expression stability of reference genes could vary according to tissues, age of individuals and experimental conditions. In the pig however, very little information is available on the expression stability of reference genes. The aim of this research was therefore to develop a new set of reference genes which can be used for normalization of mRNA expression data of genes expressed in varieties of porcine tissues at different ages. Results The mRNA expression stability of nine commonly used reference genes (B2M, BLM, GAPDH, HPRT1, PPIA, RPL4, SDHA, TBP and YWHAZ was determined in varieties of tissues collected from newborn, young and adult pigs. geNorm, NormFinder and BestKeeper software were used to rank the genes according to their stability. geNorm software revealed that RPL4, PPIA and YWHAZ showed high stability in newborn and adult pigs, while B2M, YWHAZ and SDHA showed high stability in young pigs. In all cases, GAPDH showed the least stability in geNorm. NormFinder revealed that TBP was the most stable gene in newborn and young pigs, while PPIA was most stable in adult pigs. Moreover, geNorm software suggested that the geometric mean of three most stable gene would be the suitable combination for accurate normalization of gene expression study. Conclusions Although, there was discrepancy in the ranking order of reference genes obtained by different analysing software methods, the geometric mean of the RPL4, PPIA and YWHAZ seems to be the most appropriate combination of

  2. Stability Depends on Positive Autoregulation in Boolean Gene Regulatory Networks

    Science.gov (United States)

    Pinho, Ricardo; Garcia, Victor; Irimia, Manuel; Feldman, Marcus W.

    2014-01-01

    Network motifs have been identified as building blocks of regulatory networks, including gene regulatory networks (GRNs). The most basic motif, autoregulation, has been associated with bistability (when positive) and with homeostasis and robustness to noise (when negative), but its general importance in network behavior is poorly understood. Moreover, how specific autoregulatory motifs are selected during evolution and how this relates to robustness is largely unknown. Here, we used a class of GRN models, Boolean networks, to investigate the relationship between autoregulation and network stability and robustness under various conditions. We ran evolutionary simulation experiments for different models of selection, including mutation and recombination. Each generation simulated the development of a population of organisms modeled by GRNs. We found that stability and robustness positively correlate with autoregulation; in all investigated scenarios, stable networks had mostly positive autoregulation. Assuming biological networks correspond to stable networks, these results suggest that biological networks should often be dominated by positive autoregulatory loops. This seems to be the case for most studied eukaryotic transcription factor networks, including those in yeast, flies and mammals. PMID:25375153

  3. Stability depends on positive autoregulation in Boolean gene regulatory networks.

    Directory of Open Access Journals (Sweden)

    Ricardo Pinho

    2014-11-01

    Full Text Available Network motifs have been identified as building blocks of regulatory networks, including gene regulatory networks (GRNs. The most basic motif, autoregulation, has been associated with bistability (when positive and with homeostasis and robustness to noise (when negative, but its general importance in network behavior is poorly understood. Moreover, how specific autoregulatory motifs are selected during evolution and how this relates to robustness is largely unknown. Here, we used a class of GRN models, Boolean networks, to investigate the relationship between autoregulation and network stability and robustness under various conditions. We ran evolutionary simulation experiments for different models of selection, including mutation and recombination. Each generation simulated the development of a population of organisms modeled by GRNs. We found that stability and robustness positively correlate with autoregulation; in all investigated scenarios, stable networks had mostly positive autoregulation. Assuming biological networks correspond to stable networks, these results suggest that biological networks should often be dominated by positive autoregulatory loops. This seems to be the case for most studied eukaryotic transcription factor networks, including those in yeast, flies and mammals.

  4. Recommended Reference Genes for Quantitative PCR Analysis in Soybean Have Variable Stabilities during Diverse Biotic Stresses.

    Science.gov (United States)

    Bansal, Raman; Mittapelly, Priyanka; Cassone, Bryan J; Mamidala, Praveen; Redinbaugh, Margaret G; Michel, Andy

    2015-01-01

    For real-time reverse transcription-PCR (qRT-PCR) in soybean, reference genes in different tissues, developmental stages, various cultivars, and under stress conditions have been suggested but their usefulness for research on soybean under various biotic stresses occurring in North-Central U.S. is not known. Here, we investigated the expression stabilities of ten previously recommended reference genes (ABCT, CYP, EF1A, FBOX, GPDH, RPL30, TUA4, TUB4, TUA5, and UNK2) in soybean under biotic stress from Bean pod mottle virus (BPMV), powdery mildew (PMD), soybean aphid (SBA), and two-spotted spider mite (TSSM). BPMV, PMD, SBA, and TSSM are amongst the most common pest problems on soybean in North-Central U.S. and other regions. Reference gene stability was determined using three software algorithms (geNorm, NormFinder, BestKeeper) and a web-based tool (RefFinder). Reference genes showed variability in their expression as well as stability across various stressors and the best reference genes were stress-dependent. ABCT and FBOX were found to be the most stable in soybean under both BPMV and SBA stress but these genes had only minimal to moderate stability during PMD and TSSM stress. Expression of TUA4 and CYP was found to be most stable during PMD stress; TUB4 and TUA4 were stable under TSSM stress. Under various biotic stresses on soybean analyzed, GPDH expression was found to be consistently unstable. For all biotic stressors on soybean, we obtained pairwise variation (V2/3) values less than 0.15 which suggested that combined use of the two most stable reference genes would be sufficient for normalization. Further, we demonstrated the utility of normalizing the qRT-PCR data for target genes using the most stable reference genes validated in current study. Following of the recommendations from our current study will enable an accurate and reliable normalization of qRT-PCR data in soybean under biotic stress.

  5. Integration and inheritance stability of foreign Bt toxin gene in the bivalent insectresistant transgenic cotton plants

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Genetic and expressional stability of Bt toxin gene is crucial for the breeding of insect-resistant transgenic cotton varieties and their commercialization. Genomic Southern blot analysis of R3, R4 and R5 generations of bivalent transgenic insect-resistant cotton plants was done in order to determine the integration, the copy number and the inheritance stability of Bt toxin gene in the transgenic cotton plants. The results indicated that there was a 4.7 kb positive band in the Southern blot when the genomic DNA of the bivalent transgenic insect-resistant cotton plants and the positive control (the plasmid) were digested with HindⅢ respectively. This result proved that the Bt toxin gene had been integrated into the genome of the cotton in full length.There is only one Xho I restriction site in the Bt toxin gene.Southern blot analysis indicated that many copies of Bt toxin gene had been integrated into the genome of the cotton when the genomic DNA of transgenic plants was digested with Xho I. Among them, there were four copies (about 17.7, 8,5.5 and 4.7 kb in size) existing in all the tested plants of R3,R4 and R5 generations. The preliminary conclusion was that there were more than four copies of Bt toxin gene integrated into the genome of the cotton, among them, more than one copy can express and inherit steadily. This result provides a scientific basis for the breeding of the bivalent insect-resistant transgenic cotton plants and its commercialization.``

  6. Prediction of Factors Determining Changes in Stability in Protein Mutants

    OpenAIRE

    Parthiban, Vijayarangakannan

    2006-01-01

    Analysing the factors behind protein stability is a key research topic in molecular biology and has direct implications on protein structure prediction and protein-protein docking solutions. Protein stability upon point mutations were analysed using a distance dependant pair potential representing mainly through-space interactions and torsion angle potential representing neighbouring effects as a basic statistical mechanical setup for the analysis. The synergetic effect of accessible surface ...

  7. [Elucidation of key genes in sex determination in genetics teaching].

    Science.gov (United States)

    Li, Meng; He, Zhumei

    2014-06-01

    Sex is an important and complex feature of organisms, which is controlled by the genetic and environmental factors. The genetic factors, i.e., genes, are vital in sex determination. However, not all the related genes play the same roles, and some key genes play a vital role in the sex determination and differentiation. With the development of the modern genetics, a great progress on the key genes has been made in sex determination. In this review, we summarize the mechanism of sex determination and the strategy of how to study the key genes in sex determination. It will help us to understand the mechanism of sex determination better in the teaching of genetics.

  8. Single-Gene Determinants of Epilepsy Comorbidity.

    Science.gov (United States)

    Noebels, Jeffrey L

    2015-11-02

    Common somatic conditions are bound to occur by chance in individuals with neurological disorders as prevalent as epilepsy, but when biological links underlying the comorbidity can be uncovered, the relationship may provide clues into the origin and mechanisms of both. The expanding list of monogenic epilepsies and their associated clinical features offer a remarkable opportunity to mine the epilepsy genome for coordinate neurodevelopmental phenotypes and examine their pathogenic mechanisms. Defined single-gene-linked epilepsy syndromes identified to date include all of the most frequently cited comorbidities, such as cognitive disorders, autism, migraine, mood disorders, late-onset dementia, and even premature lethality. Gene-linked comorbidities may be aggravated by, or independent of, seizure history. Mutations in these genes establish clear biological links between abnormal neuronal synchronization and a variety of neurobehavioral disorders, and critically substantiate the definition of epilepsy as a complex spectrum disorder. Mapping the neural circuitry of epilepsy comorbidities and understanding their single-gene risk should substantially clarify this challenging aspect of clinical epilepsy management.

  9. Reliable Selection and Holistic Stability Evaluation of Reference Genes for Rice Under 22 Different Experimental Conditions.

    Science.gov (United States)

    Wang, Zhaohai; Wang, Ya; Yang, Jing; Hu, Keke; An, Baoguang; Deng, Xiaolong; Li, Yangsheng

    2016-07-01

    Stable and uniform expression of reference genes across samples plays a key role in accurate normalization of gene expression by reverse-transcription quantitative polymerase chain reaction (RT-qPCR). For rice study, there is still a lack of validation and recommendation of appropriate reference genes with high stability depending on experimental conditions. Eleven candidate reference genes potentially owning high stability were evaluated by geNorm and NormFinder for their expression stability in 22 various experimental conditions. Best combinations of multiple reference genes were recommended depending on experimental conditions, and the holistic stability of reference genes was also evaluated. Reference genes would become more variable and thus needed to be critically selected in experimental groups of tissues, heat, 6-benzylamino purine, and drought, but they were comparatively stable under cold, wound, and ultraviolet-B stresses. Triosephosphate isomerase (TI), profilin-2 (Profilin-2), ubiquitin-conjugating enzyme E2 (UBC), endothelial differentiation factor (Edf), and ADP-ribosylation factor (ARF) were stable in most of our experimental conditions. No universal reference gene showed good stability in all experimental conditions. To get accurate expression result, suitable combination of multiple reference genes for a specific experimental condition would be a better choice. This study provided an application guideline to select stable reference genes for rice gene expression study.

  10. Stability and immunogenicity properties of the gene-silencing polypurine reverse Hoogsteen hairpins.

    Science.gov (United States)

    Villalobos, Xenia; Rodríguez, Laura; Prévot, Jeanne; Oleaga, Carlota; Ciudad, Carlos J; Noé, Véronique

    2014-01-06

    Gene silencing by either small-interference RNAs (siRNA) or antisense oligodeoxynucleotides (aODN) is widely used in biomedical research. However, their use as therapeutic agents is hindered by two important limitations: their low stability and the activation of the innate immune response. Recently, we developed a new type of molecule to decrease gene expression named polypurine reverse Hoogsteen hairpins (PPRHs) that bind to polypyrimidine targets in the DNA. Herein, stability experiments performed in mouse, human, and fetal calf serum and in PC3 cells revealed that the half-life of PPRHs is much longer than that of siRNAs in all cases. Usage of PPRHs with a nicked-circular structure increased the binding affinity to their target sequence and their half-life in FCS when bound to the target. Regarding the innate immune response, we determined that the levels of the transcription factors IRF3 and its phosphorylated form, as well as NF-κB were increased by siRNAs and not by PPRHs; that the expression levels of several proinflammatory cytokines including IL-6, TNF-α, IFN-α, IFN-ß, IL-1ß, and IL-18 were not significantly increased by PPRHs; and that the cleavage and activation of the proteolytic enzyme caspase-1 was not triggered by PPRHs. These determinations indicated that PPRHs, unlike siRNAs, do not activate the innate inflammatory response.

  11. Experimental bifurcation analysis of an impact oscillator – Determining stability

    DEFF Research Database (Denmark)

    Bureau, Emil; Schilder, Frank; Elmegård, Michael

    2014-01-01

    We propose and investigate three different methods for assessing stability of dynamical equilibrium states during experimental bifurcation analysis, using a control-based continuation method. The idea is to modify or turn off the control at an equilibrium state and study the resulting behavior. A...

  12. Electron Delocalization Determined Anomalous Stability in Small Water Rings

    CERN Document Server

    Wang, Bo; Dai, Xing; Song, Ruixia; Meng, Yan; Wang, Zhigang; Zhang, Ruiqin

    2013-01-01

    Water clusters are known to form through hydrogen bonding. However, this study shows that the formation of very small water clusters significantly deviates from this mechanism and instead involves both hydrogen bonding and electron delocalization. Our density functional theory calculations show that small water rings (H2O)n of n=3 or 4 show strong electron delocalization originating from both the hydrogen and oxygen atoms and extending to the ring center. This is very different from larger rings. Further energy decomposition of rings with n=3-6 demonstrates an upward trend in the polarization component but an decrease in the electrostatic and exchange repulsion components, presenting a minimum and accounting for 33% of interaction energy at n=3. This significantly promotes stability of the small water rings. Our findings provide a comprehensive analysis and improve our understanding of the stability characteristics of water clusters.

  13. The first determination of DNA sequence of a specific gene.

    Science.gov (United States)

    Inouye, Masayori

    2016-05-10

    How and when the first DNA sequence of a gene was determined? In 1977, F. Sanger came up with an innovative technology to sequence DNA by using chain terminators, and determined the entire DNA sequence of the 5375-base genome of bacteriophage φX 174 (Sanger et al., 1977). While this Sanger's achievement has been recognized as the first DNA sequencing of genes, we had determined DNA sequence of a gene, albeit a partial sequence, 11 years before the Sanger's DNA sequence (Okada et al., 1966).

  14. Genotypic sex determination in teleosts: insights from the testis-determining amhy gene.

    Science.gov (United States)

    Hattori, Ricardo Shohei; Strüssmann, Carlos Augusto; Fernandino, Juan Ignacio; Somoza, Gustavo Manuel

    2013-10-01

    The master sex-determining genes identified so far in fishes are clearly not conserved, as evidenced by several unrelated genes reported to play critical roles in sex determination. In this study, we reviewed the molecular process of sex determination in the Patagonian pejerrey Odontesthes hatcheri, an emerging model due to the recent discovery that a Y-chromosome linked, duplicated copy of the anti-Müllerian hormone gene, amhy plays a pivotal role in sex determination. A comparative analysis with other newly found sex-determining genes of teleost fish, DMY/dmrt1bY, sdY, amhr2, and gsdf(Y) is performed and alternative ideas are proposed to explain the mechanism involved in the rise of various types of non-homologous sex-determining genes.

  15. Local/bulk determinants of conformational stability of exchangeable apolipoproteins.

    Science.gov (United States)

    Dergunov, Alexander D

    2011-09-01

    GuHCl-induced denaturation of human plasma apoA-I, apoA-II, apoA-IV, apoE3 and three recombinant apoE isoforms in solution and discoidal complexes with phosphatidylcholine (only plasma proteins) was studied. The protein conformational stability (ΔG(H(2)O)) and a slope of linear dependence of free energy of unfolding on GuHCl concentration (m-value) were estimated with the three equilibrium schemes. The data for all proteins, except apoA-II, fit with the three-state model, thus evidencing two-domain structure. The predicted folding rate of the four apoE in solution correlated with conformational stability. The dependence disappeared at the inclusion of apoA-I and apoA-IV into analysis and the m-values, adjusted for residue number in helices (m(rh)), differed between those for apoE and apoA-I/apoA-IV. However, the m(rh)-values for six proteins correlated positively with the fractional change in accessible surface area at unfolding for Phe, Lys and Asn, while negatively for Arg, Ala and Gly residues. The difference between the adjusted ΔG(rh)(H(2)O) values for apolipoproteins in complexes and in solution decreased at the increase of reduced temperature (T(obs)-T(t))/T(t). The induction of intrinsic disorder by arginine residues may be of primary importance in metabolism and function of exchangeable apolipoproteins, while their stability in nascent discoidal HDL is controlled by the physical state of phosphatidylcholine. Copyright © 2011 Elsevier B.V. All rights reserved.

  16. Evolutionary stabilization of the gene-3-protein of phage fd reveals the principles that govern the thermodynamic stability of two-domain proteins.

    Science.gov (United States)

    Martin, Andreas; Schmid, Franz X

    2003-05-09

    The gene-3-protein (G3P) of filamentous phage is essential for their propagation. It consists of three domains. The CT domain anchors G3P in the phage coat, the N2 domain binds to the F pilus of Escherichia coli and thus initiates infection, and the N1 domain continues by interacting with the TolA receptor. Phage are thus only infective when the three domains of G3P are tightly linked, and this requirement is exploited by Proside, an in vitro selection method for proteins with increased stability. In Proside, a repertoire of variants of the protein to be stabilized is inserted between the N2 and the CT domains of G3P. Stabilized variants can be selected because they resist cleavage by a protease and thus maintain the essential linkage between the domains. The method is limited by the proteolytic stability of G3P itself. We improved the stability of G3P by subjecting the phage without a guest protein to rounds of random in vivo mutagenesis and proteolytic Proside selections. Variants of G3P with one to four mutations were selected, and the temperature at which the corresponding phage became accessible for a protease increased in a stepwise manner from 40 degrees C to almost 60 degrees C. The N1-N2 fragments of wild-type gene-3-protein and of the four selected variants were purified and their stabilities towards thermal and denaturant-induced unfolding were determined. In the biphasic transitions of these proteins domain dissociation and unfolding of N2 occur in a concerted reaction in the first step, followed by the independent unfolding of domain N1 in the second step. N2 is thus less stable than N1, and it unfolds when the interactions with N1 are broken. The strongest stabilizations were caused by mutations in domain N2, in particular in its hinge subdomain, which provides many stabilizing interactions between the N1 and N2 domains. These results reveal how the individual domains and their assembly contribute to the overall stability of two-domain proteins and

  17. Evaluation of pancreatin stability through enzyme activity determination

    Directory of Open Access Journals (Sweden)

    Terra Gleysson De Paula

    2016-09-01

    Full Text Available Pancreatin is a biotechnological product containing an enzyme complex, obtained from porcine pancreas, that is employed in treating pancreatic diseases. Experiments regarding the stability of the pharmaceutical formulation containing pancreatin were performed using standard binary mixtures with 6 excipients in a 1:1 ratio (m/m and a commercial formulation. To accomplish these goals, samples were stored for 1, 3 and 6 months at 40 ± 1 °C and 75 ± 5 % relative humidity (RH and 40 ± 1 °C and 0 % RH. Stress testing was also performed. All samples were analyzed to evaluate the α-amylase, lipase and protease activities through UV/Vis spectrophotometry. The results revealed that the excipient proprieties and the storage conditions affected enzyme stability. Humidity was a strong influencing factor in the reduction of α-amylase and protease activities. Stress testing indicated that pH 9.0 and UV light did not induce substantial alterations in enzyme activity.

  18. Codon Usage and 3' UTR Length Determine Maternal mRNA Stability in Zebrafish.

    Science.gov (United States)

    Mishima, Yuichiro; Tomari, Yukihide

    2016-03-17

    The control of mRNA stability plays a central role in regulating gene expression. In metazoans, the earliest stages of development are driven by maternally supplied mRNAs. The degradation of these maternal mRNAs is critical for promoting the maternal-to-zygotic transition of developmental programs, although the underlying mechanisms are poorly understood in vertebrates. Here, we characterized maternal mRNA degradation pathways in zebrafish using a transcriptome analysis and systematic reporter assays. Our data demonstrate that ORFs enriched with uncommon codons promote deadenylation by the CCR4-NOT complex in a translation-dependent manner. This codon-mediated mRNA decay is conditional on the context of the 3' UTR, with long 3' UTRs conferring resistance to deadenylation. These results indicate that the combined effect of codon usage and 3' UTR length determines the stability of maternal mRNAs in zebrafish embryos. Our study thus highlights the codon-mediated mRNA decay as a conserved regulatory mechanism in eukaryotes.

  19. Analytic theory for the determination of velocity and stability of bubbles in a Hele-Shaw cell. Part 2: Stability

    Science.gov (United States)

    Tanveer, Saleh

    1989-01-01

    The analysis is extended to determine the linear stability of a bubble in a Hele-Shaw cell analytically. Only the solution branch corresponding to largest possible bubble velocity U for given surface tension is found to be stable, while all the others are unstable, in accordance with earlier numerical results.

  20. CORPORATE GOVERNANCE – DETERMINANT OF MOLDOVAN BANKING SYSTEM STABILITY

    Directory of Open Access Journals (Sweden)

    Dorina CLICHICI

    2015-04-01

    Full Text Available Recent events recorded in the banking sector of the Republic of Moldova – establishing special administration in three commercial banks by the National Bank of Moldova (NBM – have highlighted the significant role which the quality of corporate governance presents for banking system. The aim of this paper is to identify the challenges of corporate governance for the stability of the Moldovan banking system and destabilizing effects of weak corporate governance structures within banking institutions and analyze the consequences of corporate governance deficiencies. For achieving the goal the author conducted a detailed analysis of normative acts regulating the banking system in order to identify the existing gaps regarding this subject and analyzed recent performances of commercial banks in Moldova. Despite some progress in addressing the recommendations of the International Monetary Fund (IMF and the satisfactory reported performance of banks, there are serious governance problems in several banks including the largest ones.

  1. Identification of the two rotavirus genes determining neutralization specificities

    Energy Technology Data Exchange (ETDEWEB)

    Offit, P.A.; Blavat, G.

    1986-01-01

    Bovine rotavirus NCDV and simian rotavirus SA-11 represent two distinct rotavirus serotypes. A genetic approach was used to determine which viral gene segments segregated with serotype-specific viral neutralization. There were 16 reassortant rotarviruses derived by coinfection of MA-104 cells in vitro with the SA-11 and NCDV strains. The parental origin of reassortant rotavirus double-stranded RNA segments was determined by gene segment mobility in polyacrylamide gels and by hybridization with radioactively labeled parental viral transcripts. The authors found that two rotavirus gene segments found previously to code for outer capsid proteins vp3 and vp7 cosegreated with virus neutralization specificities.

  2. Stability of Reference Genes for Messenger RNA Quantification by Real-Time PCR in Mouse Dextran Sodium Sulfate Experimental Colitis.

    Directory of Open Access Journals (Sweden)

    Nour Eissa

    Full Text Available Many animal models have been developed to characterize the complexity of colonic inflammation. In dextran sodium sulfate (DSS experimental colitis in mice the choice of reference genes is critical for accurate quantification of target genes using quantitative real time PCR (RT-qPCR. No studies have addressed the performance of reference genes in mice DSS-experimental colitis. This study aimed to determine the stability of reference genes expression (RGE in DSS-experimental murine colitis.Colitis was induced in male C57BL/6 mice using DSS5% for 5 days, control group received water. RNA was extracted from inflamed and non-inflamed colon. Using RT-qPCR, comparative analysis of 13 RGE was performed according to predefined criteria and relative colonic TNF-α and IL-1β gene expression was determined by calculating the difference in the threshold cycle.Colitis significantly altered the stability of mucosal RGE. Commonly used glyceraldehyde-3-phosphate dehydrogenase (Gapdh, β-actin (Actb, or β2-microglobulin (β2m showed the highest variability within the inflamed and control groups. Conversely, TATA-box-binding protein (Tbp and eukaryotic translation elongation factor 2 (Eef2 were not affected by inflammation and were the most stable genes. Normalization of colonic TNF-α and IL-1β mRNA levels was dependent on the reference gene used. Depending on the genes used to normalize the data, statistical significance varied from significant when TBP / Eef2 were used to non-significant when Gapdh, Actb or β2m were used.This study highlights the appropriate choice of RGE to ensure adequate normalization of RT-qPCR data when using this model. Suboptimal RGE may explain controversial results from published studies. We recommend using Tbp and Eef2 instead of Gapdh, Actb or β2m as reference genes.

  3. Mitochondria as determinant of nucleotide pools and chromosomal stability

    Energy Technology Data Exchange (ETDEWEB)

    Desler, Claus; Munch-Petersen, Birgitte [Department of Science, Systems and Models, Roskilde University, 4000 Roskilde (Denmark); Stevnsner, Tinna [Department of Molecular Biology, University of Aarhus, 8000 Aarhus (Denmark); Matsui, Sei-Ichi; Kulawiec, Mariola; Singh, Keshav K. [Department og Cancer Genetics, Roswell Park Cancer Institute, Buffalo, NY 14263 (United States); Rasmussen, Lene Juel [Department of Science, Systems and Models, Roskilde University, 4000 Roskilde (Denmark)], E-mail: ljr@ruc.dk

    2007-12-01

    Mitochondrial function plays an important role in multiple human diseases and mutations in the mitochondrial genome have been detected in nearly every type of cancer investigated to date. However, the mechanism underlying the interrelation is unknown. We used human cell lines depleted of mitochondrial DNA as models and analyzed the outcome of mitochondrial dysfunction on major cellular repair activities. We show that the deoxyribonucleoside triphosphate (dNTP) pools are affected, most prominently we detect a 3-fold reduction of the dTTP pool when normalized to the number of cells in S-phase. It is known that imbalanced dNTP pools are mutagenic and in accordance, we show that mitochondrial dysfunction results in chromosomal instability, which can explain its role in tumor development. We did not find any straightforward correlation between ATP levels and dNTP pools in cells with defective mitochondrial activity. Our results suggest that mitochondria are central players in maintaining genomic stability and in controlling essential nuclear processes such as upholding a balanced supply of nucleotides.

  4. Thermal stability of ladderane lipids as determined by hydrous pyrolysis

    Science.gov (United States)

    Jaeschke, A.; Lewan, M.D.; Hopmans, E.C.; Schouten, S.; Sinninghe, Damste J.S.

    2008-01-01

    Anaerobic ammonium oxidation (anammox) has been recognized as a major process resulting in loss of fixed inorganic nitrogen in the marine environment. Ladderane lipids, membrane lipids unique to anammox bacteria, have been used as markers for the detection of anammox in marine settings. However, the fate of ladderane lipids after sediment burial and maturation is unknown. In this study, anammox bacterial cell material was artificially matured by hydrous pyrolysis at constant temperatures ranging from 120 to 365 ??C for 72 h to study the stability of ladderane lipids during progressive dia- and catagenesis. HPLC-MS/MS analysis revealed that structural alterations of ladderane lipids already occurred at 120 ??C. At temperatures >140 ??C, ladderane lipids were absent and only more thermally stable products could be detected, i.e., ladderane derivatives in which some of the cyclobutane rings were opened. These diagenetic products of ladderane lipids were still detectable up to temperatures of 260 ??C using GC-MS. Thus, ladderane lipids are unlikely to occur in ancient sediments and sedimentary rocks, but specific diagenetic products of ladderane lipids will likely be present in sediments and sedimentary rocks of relatively low maturity (i.e., C31 hopane 22S/(22S + 22R) ratio 0.5). ?? 2008 Elsevier Ltd.

  5. Relative stability of network states in Boolean network models of gene regulation in development.

    Science.gov (United States)

    Zhou, Joseph Xu; Samal, Areejit; d'Hérouël, Aymeric Fouquier; Price, Nathan D; Huang, Sui

    2016-01-01

    Progress in cell type reprogramming has revived the interest in Waddington's concept of the epigenetic landscape. Recently researchers developed the quasi-potential theory to represent the Waddington's landscape. The Quasi-potential U(x), derived from interactions in the gene regulatory network (GRN) of a cell, quantifies the relative stability of network states, which determine the effort required for state transitions in a multi-stable dynamical system. However, quasi-potential landscapes, originally developed for continuous systems, are not suitable for discrete-valued networks which are important tools to study complex systems. In this paper, we provide a framework to quantify the landscape for discrete Boolean networks (BNs). We apply our framework to study pancreas cell differentiation where an ensemble of BN models is considered based on the structure of a minimal GRN for pancreas development. We impose biologically motivated structural constraints (corresponding to specific type of Boolean functions) and dynamical constraints (corresponding to stable attractor states) to limit the space of BN models for pancreas development. In addition, we enforce a novel functional constraint corresponding to the relative ordering of attractor states in BN models to restrict the space of BN models to the biological relevant class. We find that BNs with canalyzing/sign-compatible Boolean functions best capture the dynamics of pancreas cell differentiation. This framework can also determine the genes' influence on cell state transitions, and thus can facilitate the rational design of cell reprogramming protocols.

  6. Analysis of the CoIE1 stability determinant Rcd.

    Science.gov (United States)

    Sharpe, M E; Chatwin, H M; Macpherson, C; Withers, H L; Summers, D K

    1999-08-01

    Multimer formation is an important cause of instability for many multicopy plasmids. Plasmid CoIE1 is maintained stably because multimers are converted to monomers by Xer-mediated site-specific recombination at the cer site. However, multimer resolution is not the whole story; inactivation of a promoter (Pcer) within cer causes plasmid instability even though recombination is unaffected. The promoter directs the synthesis of a short transcript (Rcd) which is proposed to delay the division of multimer-containing cells. Mapping of the 5' terminus of Rcd confirms that transcription initiates from Pcer. The 3' terminus shows considerable heterogeneity, consistent with a primary transcript of 95 nt being degraded via intermediates of 79 and 70 nt. Secondary structure predictions for Rcd are presented. Of four mutations which abolish Rcd-mediated growth inhibition, one reduces the activity of Pcer while the other three map to the rcd coding sequence and reduce the steady-state level of the transcript. RNA folding analysis suggests that these three mutant transcripts adopt a common secondary structure in which the major stem-loop differs from that of wild-type Rcd. A survey of 24 cer-like multimer resolution sites revealed six which contain Pcer-like sequences. The putative transcripts from these sites have similar predicted secondary structures to Rcd and contain a highly conserved 15 base sequence. To test the hypothesis that Rcd acts as an anti-sense RNA, interacting with its target gene(s) through the 15 nt sequence, we used DNA hybridization and sequence analysis to find matches to this sequence in the Escherichia coli chromosome. Our failure to find plausible anti-sense targets has led to the suggestion that Rcd may interact directly with a protein target.

  7. Determination of the stability of diluted allergen extracts using a concentration step prior to EAST inhibition

    NARCIS (Netherlands)

    Niemeijer, NR; Kauffman, HF; DeMonchy, JGR; Meijer, G.

    1996-01-01

    Background Generally the stability of diluted allergen extracts, as used for skin testing, provocation testing and immunotherapy can not be measured using a normal enzyme allergosorbent test (EAST) inhibition method. Objective The aim of this study was to determine the stability of diluted allergen

  8. Stability and toxicity of empty or gene-loaded lipopolysaccharide-amine nanopolymersomes

    Directory of Open Access Journals (Sweden)

    Wang QM

    2015-01-01

    Full Text Available Qinmei Wang,1,* Ying Chen,1,* Lichun Wang,1 Xinchun Zhang,2 Hongzhang Huang,2 Wei Teng2 1Key Laboratory on Assisted Circulation, Ministry of Health, Cardiovascular Division, First Affiliated Hospital, Sun Yat-sen University, Guangzhou, People’s Republic of China; 2Hospital of Stomatology, Institute of Stomatological Research, Guanghua School of Stomatology, Sun Yat-sen University, Guangzhou, People’s Republic of China *These authors contributed equally to this work Abstract: Successful in vivo gene delivery mediated by nonviral vectors requires efficient extracellular and intracellular gene delivery, but few studies have given attention to the former. That is why numerous gene delivery systems have succeeded in vitro, while the expected clinical success has not come about. To realize efficient extracellular gene delivery, the stability of vectors and/or their complexes with genes in body fluids is first required, which prevents loaded genes from premature unloading and degradation. Furthermore, the storage stability of vectors under common conditions is important for their widespread applications. Lipopolysaccharide-amine nanopolymersomes (NPs, a gene vector developed by our group recently, have higher than 95% in vitro transfection efficiency in mesenchymal stem cells when delivering pEGFP, and induce significant angiogenesis in zebrafish when delivering plasmid encoding vascular endothelial growth factor deoxyribonucleic acid (pVEGF. To reveal their extracellular delivery ability and storage stability, in this study their stability in various simulant physiological environments and storage conditions was systematically studied by monitoring their changes in disassembly, size, zeta potential, and transfection efficiency. Additionally, damage to the mitochondria of mesenchymal stem cells was evaluated. Results show that NPs and plasmid deoxyribonucleic acid (pDNA-loaded NPs (pNPs have acceptable stability against dilution, anions, salts, p

  9. Characterization of sex determination and sex differentiation genes in Latimeria.

    Directory of Open Access Journals (Sweden)

    Mariko Forconi

    Full Text Available Genes involved in sex determination and differentiation have been identified in mice, humans, chickens, reptiles, amphibians and teleost fishes. However, little is known of their functional conservation, and it is unclear whether there is a common set of genes shared by all vertebrates. Coelacanths, basal Sarcopterygians and unique "living fossils", could help establish an inventory of the ancestral genes involved in these important developmental processes and provide insights into their components. In this study 33 genes from the genome of Latimeria chalumnae and from the liver and testis transcriptomes of Latimeria menadoensis, implicated in sex determination and differentiation, were identified and characterized and their expression levels measured. Interesting findings were obtained for GSDF, previously identified only in teleosts and now characterized for the first time in the sarcopterygian lineage; FGF9, which is not found in teleosts; and DMRT1, whose expression in adult gonads has recently been related to maintenance of sexual identity. The gene repertoire and testis-specific gene expression documented in coelacanths demonstrate a greater similarity to modern fishes and point to unexpected changes in the gene regulatory network governing sexual development.

  10. Characterization of sex determination and sex differentiation genes in Latimeria.

    Science.gov (United States)

    Forconi, Mariko; Canapa, Adriana; Barucca, Marco; Biscotti, Maria A; Capriglione, Teresa; Buonocore, Francesco; Fausto, Anna M; Makapedua, Daisy M; Pallavicini, Alberto; Gerdol, Marco; De Moro, Gianluca; Scapigliati, Giuseppe; Olmo, Ettore; Schartl, Manfred

    2013-01-01

    Genes involved in sex determination and differentiation have been identified in mice, humans, chickens, reptiles, amphibians and teleost fishes. However, little is known of their functional conservation, and it is unclear whether there is a common set of genes shared by all vertebrates. Coelacanths, basal Sarcopterygians and unique "living fossils", could help establish an inventory of the ancestral genes involved in these important developmental processes and provide insights into their components. In this study 33 genes from the genome of Latimeria chalumnae and from the liver and testis transcriptomes of Latimeria menadoensis, implicated in sex determination and differentiation, were identified and characterized and their expression levels measured. Interesting findings were obtained for GSDF, previously identified only in teleosts and now characterized for the first time in the sarcopterygian lineage; FGF9, which is not found in teleosts; and DMRT1, whose expression in adult gonads has recently been related to maintenance of sexual identity. The gene repertoire and testis-specific gene expression documented in coelacanths demonstrate a greater similarity to modern fishes and point to unexpected changes in the gene regulatory network governing sexual development.

  11. Determining optimal parameters in magnetic spacecraft stabilization via attitude feedback

    Science.gov (United States)

    Bruni, Renato; Celani, Fabio

    2016-10-01

    The attitude control of a spacecraft using magnetorquers can be achieved by a feedback control law which has four design parameters. However, the practical determination of appropriate values for these parameters is a critical open issue. We propose here an innovative systematic approach for finding these values: they should be those that minimize the convergence time to the desired attitude. This a particularly diffcult optimization problem, for several reasons: 1) such time cannot be expressed in analytical form as a function of parameters and initial conditions; 2) design parameters may range over very wide intervals; 3) convergence time depends also on the initial conditions of the spacecraft, which are not known in advance. To overcome these diffculties, we present a solution approach based on derivative-free optimization. These algorithms do not need to write analytically the objective function: they only need to compute it in a number of points. We also propose a fast probing technique to identify which regions of the search space have to be explored densely. Finally, we formulate a min-max model to find robust parameters, namely design parameters that minimize convergence time under the worst initial conditions. Results are very promising.

  12. NOVEL HYBRID GENE VECTOR STABILIZED BY CROSS-LINKING WITH GOLD NANOPARTICLES

    Institute of Scientific and Technical Information of China (English)

    You-xiang Wang; Ying Zhu; Jia-cong Shen

    2008-01-01

    Enhanced stability of polyplexes in physiological condition was an important prerequisite for successful systemic gene delivery. Herein novel method was reported to develop stable gene vector by nanotechnology. Thiolated polyplexes were constructed and then cross-linked with gold nanoparticles (AuNPs) by gold-thiol interactions. TEM pictures showed that AuNPs were attached to the shell of spherical polyplexes. The hybrid gene vector was stable enough in physiological condition and maintained efficient transfection, which showed great potential in gene delivery research and application.

  13. Determination of internal controls for quantitative gene expression of Isochrysis zhangjiangensis at nitrogen stress condition

    Science.gov (United States)

    Wu, Shuang; Zhou, Jiannan; Cao, Xupeng; Xue, Song

    2016-02-01

    Isochrysis zhangjiangensis is a potential marine microalga for biodiesel production, which accumulates lipid under nitrogen limitation conditions, but the mechanism on molecular level is veiled. Quantitative real-time polymerase chain reaction (qPCR) provides the possibility to investigate the gene expression levels, and a valid reference for data normalization is an essential prerequisite for firing up the analysis. In this study, five housekeeping genes, actin (ACT), α-tubulin (TUA), ß-tubulin (TUB), ubiquitin (UBI), 18S rRNA (18S) and one target gene, diacylglycerol acyltransferase (DGAT), were used for determining the reference. By analyzing the stabilities based on calculation of the stability index and on operating the two types of software, geNorm and bestkeeper, it showed that the reference genes widely used in higher plant and microalgae, such as UBI, TUA and 18S, were not the most stable ones in nitrogen-stressed I. zhangjiangensis, and thus are not suitable for exploring the mRNA expression levels under these experimental conditions. Our results show that ACT together with TUB is the most feasible internal control for investigating gene expression under nitrogen-stressed conditions. Our findings will contribute not only to future qPCR studies of I. zhangjiangensis, but also to verification of comparative transcriptomics studies of the microalgae under similar conditions.

  14. Stability studies of chitosan-DNA-FAP-B nanoparticles for gene delivery to lung epithelial cells.

    Science.gov (United States)

    Mohammadi, Zohreh; Dorkoosh, Farid Abedin; Hosseinkhani, Saman; Amini, Tina; Rahimi, Amir Abbas; Najafabadi, Abdolhossein Rouholamini; Tehrani, Morteza Rafiee

    2012-03-01

    A successful gene delivery system requires efficiency and stability during storage. Stability studies are imperative for nanomedicines containing biotechnological products such as plasmids and targeting peptides. Chitosan-DNA-FAP-B nanoparticles are novel non-viral vectors for specific gene delivery to the lung epithelial cells. In this study, the storage stability of chitosan-DNA-FAP-B nanoparticles at -20, 5 and 24 °C was examined. Size, zeta potential and transfection efficiency of these nano-particles in storage were also evaluated. Stability studies showed that chitosan-DNA-FAP-B nanoparticles were stable after 1 month when stored at -20 °C and retained their initial size, zeta potential and transfection efficiency. However, their stability was not desirable at 5 and 24 °C. Based on these results, it can be concluded that chitosan-DNA-FAP-B nanoparticles can be a promising candidate for gene delivery to lung epithelial cells with good storage stability at -20 °C during 1 month.

  15. Secondary structures involving the poly(A tail and other 3’ sequences are major determinants of mRNA isoform stability in yeast

    Directory of Open Access Journals (Sweden)

    Zarmik Moqtaderi

    2014-04-01

    Full Text Available In Saccharomyces cerevisiae, previous measurements of mRNA stabilities have been determined on a per-gene basis. We and others have recently shown that yeast genes give rise to a highly heterogeneous population of mRNAs due to extensive alternative 3’ end formation. Typical genes can have fifty or more distinct mRNA isoforms with 3’ endpoints differing by as little as one and as many as hundreds of nucleotides. In our recent paper [Geisberg et al. Cell (2014 156: 812-824] we measured half-lives of individual mRNA isoforms in Saccharomyces cerevisiae by using the anchor away method for the rapid removal of Rpb1, the largest subunit of RNA Polymerase II, from the nucleus, followed by direct RNA sequencing of the cellular mRNA population over time. Combining these two methods allowed us to determine half-lives for more than 20,000 individual mRNA isoforms originating from nearly 5000 yeast genes. We discovered that different 3’ mRNA isoforms arising from the same gene can have widely different stabilities, and that such half-life variability across mRNA isoforms from a single gene is highly prevalent in yeast cells. Determining half-lives for many different mRNA isoforms from the same genes allowed us to identify hundreds of RNA sequence elements involved in the stabilization and destabilization of individual isoforms. In many cases, the poly(A tail is likely to participate in the formation of stability - enhancing secondary structures at mRNA 3’ ends. Our results point to an important role for mRNA structure at 3’ termini in governing transcript stability, likely by reducing the interaction of the mRNA with the degradation apparatus.

  16. Investigation of yeast genes possibly involved in mtDNA stability ...

    African Journals Online (AJOL)

    Phelim Isichei

    function and structure on mtDNA stability in yeast, our results did not support those ... most studied model organism for acquisition of basic ... RNA interference of genes involved in mtDNA replication ... polymerase, results in reduced mtDNA copy number but .... found that RNAi of 4 genes (M01E5.2, T27F6.5, T26A5.6.

  17. CFD Based Determination of Dynamic Stability Derivatives in Yaw for a Bird

    Institute of Scientific and Technical Information of China (English)

    M. A. Moelyadi; G. Sachs

    2007-01-01

    Dynamic yaw stability derivatives of a gull bird are determined using Computational Fluid Dynamics(CFD) method. Two kinds of motions are applied for calculating the dynamic yaw stability derivatives CNr and CNβ. The first one relates to a lateral translation and, separately, to a yaw rotation. The second one consists of a combined translational and rotational motion. To determine dynamic yaw stability derivatives, the simulation of an unsteady flow with a bird model showing a harmonic motion is performed. The flow solution for each time step is obtained by solving unsteady Euler equations based on a finite volume approach for a small reduced frequency. Then, an evaluation of unsteady forces and moments for one cycle is conducted using harmonic Fourier analysis. The results of the dynamic yaw stability derivatives for both simulations of the model show a good agreement.

  18. A new method to determine oxidative stability of vegetable fats and oils at simulated frying temperature

    Directory of Open Access Journals (Sweden)

    Gertz Christian

    2001-01-01

    Full Text Available A new procedure at simulated frying conditions in our laboratory was developed to monitor frying stability of fats and oils. Water-conditioned silica was prepared and added to the fresh vegetable oil, which was heated for two hours at 170°C. The oil stability at frying temperature was then evaluated by determining the amount of formed dimeric triglycerides The results obtained showed that the stability of the vegetable oils at frying temperature could not be explained by the fatty acid composition alone. Corn oil was observed to be more stable than soybean oil, and rapeseed oil was better than olive oil. It was also observed that crude, non-refined oils were found to have a better heat stability than refin-ed oils. To estimate the effectiveness of synthetic and naturally occurring antioxidants, namely various tocopherols, tocopherol acetate and phytosterol fractions, phenolic compounds like quercetin, oryzanol, ferulic acid, gallates, BHT, BHA and other compounds like ascorbic acid 6-palmitate and squalene were added to refined sunflower and rape seed oil, and their oxidative stability at elevated temperature (OSET values determined. Both linoleic and oleic rich oils gave comparable results for the activity of the various compounds. alpha-tocopherol, tocopherol esters and BHA had low effects on oil stability at frying temperature, while ascorbyl palmitate and some phytosterol fractions were found to have the most stabilizing activity under frying conditions.

  19. METHODS OF DETERMINING THE ENTITY’S RATING IN THE FINANCIAL STABILITY ANALYSIS

    Directory of Open Access Journals (Sweden)

    Neli MUNTEAN

    2016-09-01

    Full Text Available Financial stability analysis cannot provide a complete exercise without identifying the possibilities for quantitative measurement of this phenomenon. In the present article, we intended to present the methods of determining the rating of the entities that can be used to measure the financial stability, emphasizing at the same time the limits of each of these methods. This approach has helped us to demonstrate that there is no „best practice” for evaluation of the stability, but rather a complementarity of these techniques.

  20. Vertebrate sex-determining genes play musical chairs.

    Science.gov (United States)

    Pan, Qiaowei; Anderson, Jennifer; Bertho, Sylvain; Herpin, Amaury; Wilson, Catherine; Postlethwait, John H; Schartl, Manfred; Guiguen, Yann

    2016-01-01

    Sexual reproduction is one of the most highly conserved processes in evolution. However, the genetic and cellular mechanisms making the decision of whether the undifferentiated gonad of animal embryos develops either towards male or female are manifold and quite diverse. In vertebrates, sex-determining mechanisms range from environmental to simple or complex genetic mechanisms and different mechanisms have evolved repeatedly and independently. In species with simple genetic sex-determination, master sex-determining genes lying on sex chromosomes drive the gonadal differentiation process by switching on a developmental program, which ultimately leads to testicular or ovarian differentiation. So far, very few sex-determining genes have been identified in vertebrates and apart from mammals and birds, these genes are apparently not conserved over a larger number of related orders, families, genera, or even species. To fill this knowledge gap and to better explore genetic sex-determination, we propose a strategy (RAD-Sex) that makes use of next-generation sequencing technology to identify genetic markers that define sex-specific segments of the male or female genome. Copyright © 2016 Académie des sciences. All rights reserved.

  1. Stability

    Directory of Open Access Journals (Sweden)

    Nada S. Abdelwahab

    2017-05-01

    Full Text Available The present work concerns with the development of stability indicating the RP-HPLC method for simultaneous determination of guaifenesin (GUF and pseudoephedrine hydrochloride (PSH in the presence of guaifenesin related substance (Guaiacol. GUC, and in the presence of syrup excepients with minimum sample pre-treatment. In the developed RP-HPLC method efficient chromatographic separation was achieved for GUF, PSH, GUC and syrup excepients using ODS column as a stationary phase and methanol: water (50:50, v/v, pH = 4 with orthophosphoric acid as a mobile phase with a flow rate of 1 mL min−1 and UV detection at 210 nm. The chromatographic run time was approximately 10 min. Calibration curves were drawn relating the integrated area under peak to the corresponding concentrations of PSH, GUF and GUC in the range of 1–8, 1–20, 0.4–8 μg mL−1, respectively. The developed method has been validated and met the requirements delineated by ICH guidelines with respect to linearity, accuracy, precision, specificity and robustness. The validated method was successfully applied for determination of the studied drugs in triaminic chest congestion® syrup; moreover its results were statistically compared with those obtained by the official method and no significant difference was found between them.

  2. Influence of putative exopolysaccharide genes on Pseudomonas putida KT2440 biofilm stability

    DEFF Research Database (Denmark)

    Nilsson, Martin; Chiang, Wen-Chi; Fazli, Mustafa;

    2011-01-01

    . putida KT2440 biofilm. The gene clusters alg and bcs, which code for proteins mediating alginate and cellulose biosynthesis, were found to play minor roles in P. putida KT2440 biofilm formation and stability under the conditions tested. A P. putida KT2440 derivative devoid of any identifiable...

  3. Retroviral integration profiles: their determinants and implications for gene therapy

    Directory of Open Access Journals (Sweden)

    Kwang-il Lim

    2012-04-01

    Full Text Available Retroviruses have often been used for gene therapy because oftheir capacity for the long-term expression of transgenes via stableintegration into the host genome. However, retroviral integrationcan also result in the transformation of normal cells into cancercells, as demonstrated by the incidence of leukemia in a recentretroviral gene therapy trial in Europe. This unfortunate outcomehas led to the rapid initiation of studies examining variousbiological and pathological aspects of retroviral integration. Thisreview summarizes recent findings from these studies, includingthe global integration patterns of various types of retroviruses,viral and cellular determinants of integration, implications ofintegration for gene therapy and retrovirus-mediated infectiousdiseases, and strategies to shift integration to safe host genomicloci. A more comprehensive and mechanistic understanding ofretroviral integration processes will eventually make it possible togenerate safer retroviral vector platforms in the near future. [BMBreports 2012; 45(4: 207-212

  4. Codon optimality is a major determinant of mRNA stability.

    Science.gov (United States)

    Presnyak, Vladimir; Alhusaini, Najwa; Chen, Ying-Hsin; Martin, Sophie; Morris, Nathan; Kline, Nicholas; Olson, Sara; Weinberg, David; Baker, Kristian E; Graveley, Brenton R; Coller, Jeff

    2015-03-12

    mRNA degradation represents a critical regulated step in gene expression. Although the major pathways in turnover have been identified, accounting for disparate half-lives has been elusive. We show that codon optimality is one feature that contributes greatly to mRNA stability. Genome-wide RNA decay analysis revealed that stable mRNAs are enriched in codons designated optimal, whereas unstable mRNAs contain predominately non-optimal codons. Substitution of optimal codons with synonymous, non-optimal codons results in dramatic mRNA destabilization, whereas the converse substitution significantly increases stability. Further, we demonstrate that codon optimality impacts ribosome translocation, connecting the processes of translation elongation and decay through codon optimality. Finally, we show that optimal codon content accounts for the similar stabilities observed in mRNAs encoding proteins with coordinated physiological function. This work demonstrates that codon optimization exists as a mechanism to finely tune levels of mRNAs and, ultimately, proteins.

  5. Field Supervisory Test of DREB-Transgenic Populus: Salt Tolerance, Long-Term Gene Stability and Horizontal Gene Transfer

    Directory of Open Access Journals (Sweden)

    Nan Lu

    2014-05-01

    Full Text Available Improving saline resistance may be useful for reducing environmental susceptibility and improving yields in poplar plantations. However, the instability of genetically engineered traits and gene transfer reduce their usefulness and commercial value. To investigate whether the foreign gene is still present in the genome of receptor plants after seven years (i.e., long-term foreign gene stability and gene transfer, we randomly analyzed ten field-grown transgenic hybrid Populus ((Populus tomentosa × Populus bolleana × P. tomentosa carrying the DREB1 gene from Atriplex hortensis. The results of PCR and tissue culture experiments showed that AhDREB1 was present in the transgenic trees and was still expressed. However, the transcriptional expression level had decreased compared with that four years earlier. The PCR results also indicated no foreign gene in the genomic DNA of microorganisms in the soil near the transgenic poplars, indicating that no significant gene transfer had occurred from the transgenic poplars to the microorganisms at seven years after planting.

  6. Survival of Clostridium perfringens During Simulated Transport and Stability of Some Plasmid-borne Toxin Genes under Aerobic Conditions

    Directory of Open Access Journals (Sweden)

    Johansson K-E

    2005-12-01

    Full Text Available Clostridium perfringens is a pathogen of great concern in veterinary medicine, because it causes enteric diseases and different types of toxaemias in domesticated animals. It is important that bacteria in tissue samples, which have been collected in the field, survive and for the classification of C. perfringens into the correct toxin group, it is crucial that plasmid-borne genes are not lost during transportation or in the diagnostic laboratory. The objectives of this study were to investigate the survival of C. perfringens in a simulated transport of field samples and to determine the stability of the plasmid-borne toxin genes cpb1 and etx after storage at room temperature and at 4°C. Stability of the plasmid-borne genes cpb1 and etx of C. perfringens CCUG 2035, and cpb2 from C. perfringens CIP 106526, JF 2255 and 6 field isolates in aerobic atmosphere was also studied. Survival of C. perfringens was similar in all experiments. The cpb1 and etx genes were detected in all isolates from samples stored either at room temperature or at 4°C for 24–44 h. Repeated aerobic treatment of C. perfringens CCUG 2035 and CIP 106526 did not result in the loss of the plasmid-borne genes cpb1, cpb2 or etx. Plasmid-borne genes in C. perfringens were found to be more stable than generally reported. Therefore, C. perfringens toxinotyping by PCR can be performed reliably, as the risk of plasmid loss seems to be a minor problem.

  7. Genomic imprinting and maternal effect genes in haplodiploid sex determination.

    Science.gov (United States)

    van de Zande, L; Verhulst, E C

    2014-01-01

    The research into the Drosophila melanogaster sex-determining system has been at the basis of all further research on insect sex determination. This further research has made it clear that, for most insect species, the presence of sufficient functional Transformer (TRA) protein in the early embryonic stage is essential for female sexual development. In Hymenoptera, functional analysis of sex determination by knockdown studies of sex-determining genes has only been performed for 2 species. The first is the social insect species Apis mellifera, the honeybee, which has single-locus complementary sex determination (CSD). The other species is the parasitoid Nasonia vitripennis, the jewel wasp. Nasonia has a non-CSD sex-determining system, described as the maternal effect genomic imprinting sex determination system (MEGISD). Here, we describe the arguments that eventually led to the formulation of MEGISD and the experimental data that supported and refined this model. We evaluate the possibility that DNA methylation lies at the basis of MEGISD and briefly address the role of genomic imprinting in non-CSD sex determination in other Hymenoptera.

  8. New insights into structural determinants of prion protein folding and stability.

    Science.gov (United States)

    Benetti, Federico; Legname, Giuseppe

    2015-01-01

    Prions are the etiological agent of fatal neurodegenerative diseases called prion diseases or transmissible spongiform encephalopathies. These maladies can be sporadic, genetic or infectious disorders. Prions are due to post-translational modifications of the cellular prion protein leading to the formation of a β-sheet enriched conformer with altered biochemical properties. The molecular events causing prion formation in sporadic prion diseases are still elusive. Recently, we published a research elucidating the contribution of major structural determinants and environmental factors in prion protein folding and stability. Our study highlighted the crucial role of octarepeats in stabilizing prion protein; the presence of a highly enthalpically stable intermediate state in prion-susceptible species; and the role of disulfide bridge in preserving native fold thus avoiding the misfolding to a β-sheet enriched isoform. Taking advantage from these findings, in this work we present new insights into structural determinants of prion protein folding and stability.

  9. A gene signature to determine metastatic behavior in thymomas.

    Directory of Open Access Journals (Sweden)

    Yesim Gökmen-Polar

    Full Text Available PURPOSE: Thymoma represents one of the rarest of all malignancies. Stage and completeness of resection have been used to ascertain postoperative therapeutic strategies albeit with limited prognostic accuracy. A molecular classifier would be useful to improve the assessment of metastatic behaviour and optimize patient management. METHODS: qRT-PCR assay for 23 genes (19 test and four reference genes was performed on multi-institutional archival primary thymomas (n = 36. Gene expression levels were used to compute a signature, classifying tumors into classes 1 and 2, corresponding to low or high likelihood for metastases. The signature was validated in an independent multi-institutional cohort of patients (n = 75. RESULTS: A nine-gene signature that can predict metastatic behavior of thymomas was developed and validated. Using radial basis machine modeling in the training set, 5-year and 10-year metastasis-free survival rates were 77% and 26% for predicted low (class 1 and high (class 2 risk of metastasis (P = 0.0047, log-rank, respectively. For the validation set, 5-year metastasis-free survival rates were 97% and 30% for predicted low- and high-risk patients (P = 0.0004, log-rank, respectively. The 5-year metastasis-free survival rates for the validation set were 49% and 41% for Masaoka stages I/II and III/IV (P = 0.0537, log-rank, respectively. In univariate and multivariate Cox models evaluating common prognostic factors for thymoma metastasis, the nine-gene signature was the only independent indicator of metastases (P = 0.036. CONCLUSION: A nine-gene signature was established and validated which predicts the likelihood of metastasis more accurately than traditional staging. This further underscores the biologic determinants of the clinical course of thymoma and may improve patient management.

  10. Antioxidant study of quercetin and their metal complex and determination of stability constant by spectrophotometry method.

    Science.gov (United States)

    Ravichandran, R; Rajendran, M; Devapiriam, D

    2014-03-01

    Quercetin found chelate cadmium ions, scavenge free radicals produced by cadmium. Hence new complex, quercetin with cadmium was synthesised, and the synthesised complex structures were determined by UV-vis spectrophotometry, infrared spectroscopy, thermogravimetry and differential thermal analysis techniques (UV-vis, IR, TGA and DTA). The equilibrium stability constants of quercetin-cadmium complex were determined by Job's method. The determined stability constant value of quercetin-cadminum complex at pH 4.4 is 2.27×10(6) and at pH 7.4 is 7.80×10(6). It was found that the quercetin and cadmium ion form 1:1 complex in both pH 4.4 and pH 7.4. The structure of the compounds was elucidated on the basis of obtained results. Furthermore, the antioxidant activity of the free quercetin and quercetin-cadmium complexes were determined by DPPH and ABTS assays.

  11. The polarographic determination of stability constants of urea/crown ether complexes in methanol

    NARCIS (Netherlands)

    Zollinger, D.Ph.; Bos, M.; Veen-Blaauw, van A.M.W.; Linden, van der W.E.

    1984-01-01

    A general method for determination of the stability constants of complexes of crown ethers and related compounds with small organic molecules in polar solvents is described, based on an indirect polarographic procedure. Computerized evaluation of the data forms an essential part of the procedure.

  12. Quantitative determination of target gene with electrical sensor

    Science.gov (United States)

    Zhang, Xuzhi; Li, Qiufen; Jin, Xianshi; Jiang, Cheng; Lu, Yong; Tavallaie, Roya; Gooding, J. Justin

    2015-07-01

    Integrating loop-mediated isothermal amplification (LAMP) with capacitively coupled contactless conductivity detection (C4D), we have developed an electrical sensor for the simultaneous amplification and detection of specific sequence DNA. Using the O26-wzy gene as a model, the amount of initial target gene could be determined via the threshold time obtained by monitoring the progression of the LAMP reaction in real time. Using the optimal conditions, a detection limit of 12.5 copy/μL can be obtained within 30 min. Monitoring the LAMP reaction by C4D has not only all the advantages that existing electrochemical methods have, but also additional attractive features including being completely free of carryover contamination risk, high simplicity and extremely low cost. These benefits all arise from the fact that the electrodes are separated from the reaction solution, that is C4D is a contactless method. Hence in proof of principle, the new strategy promises a robust, simple, cost-effective and sensitive method for quantitative determination of a target gene, that is applicable either to specialized labs or at point-of-care.

  13. True Stability of Lubricants Determined Using the Ball-on-Disk Test

    Directory of Open Access Journals (Sweden)

    Angela Maria Tortora

    2016-01-01

    Full Text Available True stability of lubricants can be determined when there is minimum change in the contact area and also the intervention of wear debris in the contact zone. Here, we have used the ball-on-disk instrument with the migrating point contact, that is, relative motion between the ball and disk condition to fix the contact area and minimize the wear debris at the contact zone. The jump in the friction coefficient indicates the film failure, which appeared earlier for the motor oil 5W30 compared to 5W40. Such profile was not recorded in absence of relative motion. Therefore, 5W40 was considered to have a better lubricant stability than 5W30. Applying the same test condition to the natural lubricants shows that glycerol has better lubricant stability than glycerol-water mixture. Superior true lubricant stability by glycerol and 5W40 can be related to its high viscosity. However, they were less wear resistant compared to low viscosity lubricants like 5W30 and glycerol-water. We suspect the role of microscopic wear debris at the contact zone for this behavior although it should have been avoided in the migrating point contact condition. Overall, ball-on-disk instrument with a migrating point contact condition is an effective technique to determine the stability of lubricants.

  14. Low-level infrared laser modulates muscle repair and chromosome stabilization genes in myoblasts.

    Science.gov (United States)

    da Silva Neto Trajano, Larissa Alexsandra; Stumbo, Ana Carolina; da Silva, Camila Luna; Mencalha, Andre Luiz; Fonseca, Adenilson S

    2016-08-01

    Infrared laser therapy is used for skeletal muscle repair based on its biostimulative effect on satellite cells. However, shortening of telomere length limits regenerative potential in satellite cells, which occurs after each cell division cycle. Also, laser therapy could be more effective on non-physiologic tissues. This study evaluated low-level infrared laser exposure effects on mRNA expression from muscle injury repair and telomere stabilization genes in myoblasts in normal and stressful conditions. Laser fluences were those used in clinical protocols. C2C12 myoblast cultures were exposed to low-level infrared laser (10, 35, and 70 J/cm(2)) in standard or normal (10 %) and reduced (2 %) fetal bovine serum concentrations; total RNA was extracted for mRNA expression evaluation from muscle injury repair (MyoD and Pax7) and chromosome stabilization (TRF1 and TRF2) genes by real time quantitative polymerization chain reaction. Data show that low-level infrared laser increases the expression of MyoD and Pax7 in 10 J/cm(2) fluence, TRF1 expression in all fluences, and TRF2 expression in 70 J/cm(2) fluence in both 10 and 2 % fetal bovine serum. Low-level infrared laser increases mRNA expression from genes related to muscle repair and telomere stabilization in myoblasts in standard or normal and stressful conditions.

  15. A New Numerical Method of Determining Parameter Stability Bounds : SISO Linear System with Time-Varying Uncertainties Case

    Energy Technology Data Exchange (ETDEWEB)

    Kang, H.J. [Mando Machinery Co., Pyungtaek (Korea); Ko, J.W. [Yuhan College, Buchon (Korea); Park, M.N. [Yonsei University, Seoul (Korea)

    1999-06-01

    This note presents an efficient numerical method of determining a quadratic stability bound for SISO linear system with time-varying uncertainties. Based on the quadratic stability condition in Linear Matrix Inequalities (LMI) form, the proposed method gives a quadratic stability bound for each system parameter. As an example, the state feedback regulation problem is presented. (author). 12 refs.

  16. Comparative analysis of reference gene stability in human mesenchymal stromal cells during osteogenic differentiation.

    Science.gov (United States)

    Jacobi, Angela; Rauh, Juliane; Bernstein, Peter; Liebers, Cornelia; Zou, Xuenong; Stiehler, Maik

    2013-01-01

    Mesenchymal stromal cells (MSCs) are one of the most frequently used cell sources for tissue engineering strategies. Cultivation of osteogenic MSCs is a prerequisite for cell-based concepts that aim at bone regeneration. Quantitative real time reverse transcription polymerase chain reaction (qRT-PCR) analysis is a commonly used method for the examination of mRNA expression levels. However, data on suitable reference genes for osteogenically cultivated MSCs is scarce. Hence, the aim of the study was to compare the regulation of different potential reference genes in osteogenically stimulated MSCs. Human MSCs were isolated from bone marrow aspirates of N = 6 hematologically healthy individuals, expanded by polystyrene-adherence, and maintained with and without osteogenic supplements for 14 days. Cellular proliferation and osteogenic differentiation were assessed by total DNA quantification, cell-specific alkaline phosphatase (ALP) activity and by qualitative staining for ALP and alizarin red, respectively. mRNA expression levels of N = 32 potential reference genes were quantified using the human Endogenous Control TaqMan® assays. mRNA expression stability was calculated using geNorm. The combined use of the most stable reference genes and DNA-damage-inducible alpha, Pumilio homolog 1, and large ribosomal protein P0 significantly improved gene expression accuracy as compared to the use of the commonly used reference genes beta actin and glyceraldehyde-3-phosphate dehydrogenase during qRT-PCR-based target gene expression analysis of osteogenically stimulated MSCs. © 2013 American Institute of Chemical Engineers.

  17. Stability-indicating LC method for the simultaneous determination of lisinopril and hydrochlorothiazide.

    Science.gov (United States)

    de Diego, Marta; Soto, Jorge; Mennickent, Sigrid

    2014-01-01

    A simple and rapid stability-indicating liquid chromatographic method was developed and validated for the simultaneous determination of lisinopril and hydrochlorotiazide (HCTZ) in drug substances and dosage forms in the presence of degradation products. Forced degradation studies were conducted on the pure drugs under hydrolytic, oxidative, thermal and photolytic conditions. A chromatographic separation of the two drugs and its degradation products was achieved with an RP-18 column, using methanol, acetonitrile and phosphate buffer (pH 7.1; 0.05 M) (15:15:70, v/v/v) as mobile phase at a flow rate of 0.8 mL min(-1) and UV detection at 210 nm. Lisinopril and HCTZ were well resolved from its degradation products showing the stability-indicating capability of the method. The described method was linear over a range of 40-200 µg mL(-1) for lisinopril and 25-175 µg mL(-1) for HCTZ. The assay was also selective, accurate and precise for lisinopril and HCTZ determination. This method represents an alternative to the United States Pharmacopeia (USP) method showing shorter retention time. The method was successfully applied for determination of lisinopril and HCTZ in combined commercial tablets. The results showed that the proposed method was found to be suitable for quantitative determination and the stability study of lisinopril and HCTZ in pharmaceutical samples.

  18. Novel microscale approaches for easy, rapid determination of protein stability in academic and commercial settings.

    Science.gov (United States)

    Alexander, Crispin G; Wanner, Randy; Johnson, Christopher M; Breitsprecher, Dennis; Winter, Gerhard; Duhr, Stefan; Baaske, Philipp; Ferguson, Neil

    2014-12-01

    Chemical denaturant titrations can be used to accurately determine protein stability. However, data acquisition is typically labour intensive, has low throughput and is difficult to automate. These factors, combined with high protein consumption, have limited the adoption of chemical denaturant titrations in commercial settings. Thermal denaturation assays can be automated, sometimes with very high throughput. However, thermal denaturation assays are incompatible with proteins that aggregate at high temperatures and large extrapolation of stability parameters to physiological temperatures can introduce significant uncertainties. We used capillary-based instruments to measure chemical denaturant titrations by intrinsic fluorescence and microscale thermophoresis. This allowed higher throughput, consumed several hundred-fold less protein than conventional, cuvette-based methods yet maintained the high quality of the conventional approaches. We also established efficient strategies for automated, direct determination of protein stability at a range of temperatures via chemical denaturation, which has utility for characterising stability for proteins that are difficult to purify in high yield. This approach may also have merit for proteins that irreversibly denature or aggregate in classical thermal denaturation assays. We also developed procedures for affinity ranking of protein-ligand interactions from ligand-induced changes in chemical denaturation data, and proved the principle for this by correctly ranking the affinity of previously unreported peptide-PDZ domain interactions. The increased throughput, automation and low protein consumption of protein stability determinations afforded by using capillary-based methods to measure denaturant titrations, can help to revolutionise protein research. We believe that the strategies reported are likely to find wide applications in academia, biotherapeutic formulation and drug discovery programmes.

  19. Method for determining asphaltene stability of a hydrocarbon-containing material

    Science.gov (United States)

    Schabron, John F; Rovani, Jr., Joseph F

    2013-02-05

    A method for determining asphaltene stability in a hydrocarbon-containing material having solvated asphaltenes therein is disclosed. In at least one embodiment, it involves the steps of: (a) precipitating an amount of the asphaltenes from a liquid sample of the hydrocarbon-containing material with an alkane mobile phase solvent in a column; (b) dissolving a first amount and a second amount of the precipitated asphaltenes by changing the alkane mobile phase solvent to a final mobile phase solvent having a solubility parameter that is higher than the alkane mobile phase solvent; (c) monitoring the concentration of eluted fractions from the column; (d) creating a solubility profile of the dissolved asphaltenes in the hydrocarbon-containing material; and (e) determining one or more asphaltene stability parameters of the hydrocarbon-containing material.

  20. Pilocarpine-induced seizures trigger differential regulation of microRNA-stability related genes in rat hippocampal neurons

    Science.gov (United States)

    Kinjo, Erika R.; Higa, Guilherme S. V.; Santos, Bianca A.; de Sousa, Erica; Damico, Marcio V.; Walter, Lais T.; Morya, Edgard; Valle, Angela C.; Britto, Luiz R. G.; Kihara, Alexandre H.

    2016-01-01

    Epileptogenesis in the temporal lobe elicits regulation of gene expression and protein translation, leading to reorganization of neuronal networks. In this process, miRNAs were described as being regulated in a cell-specific manner, although mechanistics of miRNAs activity are poorly understood. The specificity of miRNAs on their target genes depends on their intracellular concentration, reflecting the balance of biosynthesis and degradation. Herein, we confirmed that pilocarpine application promptly (<30 min) induces status epilepticus (SE) as revealed by changes in rat electrocorticogram particularly in fast-beta range (21–30 Hz). SE simultaneously upregulated XRN2 and downregulated PAPD4 gene expression in the hippocampus, two genes related to miRNA degradation and stability, respectively. Moreover, SE decreased the number of XRN2-positive cells in the hilus, while reduced the number of PAPD4-positive cells in CA1. XRN2 and PAPD4 levels did not change in calretinin- and CamKII-positive cells, although it was possible to determine that PAPD4, but not XRN2, was upregulated in parvalbumin-positive cells, revealing that SE induction unbalances the accumulation of these functional-opposed proteins in inhibitory interneurons that directly innervate distinct domains of pyramidal cells. Therefore, we were able to disclose a possible mechanism underlying the differential regulation of miRNAs in specific neurons during epileptogenesis. PMID:26869208

  1. Uniqueness and local stability for the inverse scattering problem of determining the cavity

    Institute of Scientific and Technical Information of China (English)

    FENG; Lixin; MA; Fuming

    2005-01-01

    Considering a time-harmonic electromagnetic plane wave incident on an arbitrarily shaped open cavity embedded in infinite ground plane, the physical process is modelled by Maxwell's equations. We investigate the inverse problem of determining the shape of the open cavity from the information of the measured scattered field. Results on the uniqueness and the local stability of the inverse problem in the 2-dimensional TM (transverse magnetic) polarization are proved in this paper.

  2. Stability of Mandibular Position during Pronunciation on Determination of Occlusal Vertical Dimension

    OpenAIRE

    齋藤, 彰久; サイトウ, アキヒサ; Akihisa, SAITO

    2003-01-01

    This study was carried out to find out whether the mandibular position during pronunciation of Japanese syllable is effective in the determination of the vertical dimension. The stability of the mandibular positions during pronunciation of the Japanese /ashi/, /achu/, /aji/, /esu/ and the relationship between each position and the rest position were examined in 64 subjects (male : 31, female : 33). Each subject had a complete natural dentition with no marked occlusal abnormalities. The subjec...

  3. On Stability and the Spectrum Determined Growth Condition for Spatially Periodic Systems

    Science.gov (United States)

    2005-01-01

    On Stability and the Spectrum Determined Growth Condition for Spatially Periodic Systems Makan Fardad and Bassam Bamieh Abstract— We consider...difficult. This work is partially supported by AFOSR Grant FA9550-04-1-0207. M. Fardad and B. Bamieh are with the Department of Me- chanical and...Environmental Engineering, University of California, Santa Barbara, CA 93105-5070. email: fardad @engineering.ucsb.edu, bamieh@engineering.ucsb.edu. In this

  4. Differential distribution improves gene selection stability and has competitive classification performance for patient survival.

    Science.gov (United States)

    Strbenac, Dario; Mann, Graham J; Yang, Jean Y H; Ormerod, John T

    2016-07-27

    A consistent difference in average expression level, often referred to as differential expression (DE), has long been used to identify genes useful for classification. However, recent cancer studies have shown that when transcription factors or epigenetic signals become deregulated, a change in expression variability (DV) of target genes is frequently observed. This suggests that assessing the importance of genes by either differential expression or variability alone potentially misses sets of important biomarkers that could lead to improved predictions and treatments. Here, we describe a new approach for assessing the importance of genes based on differential distribution (DD), which combines information from differential expression and differential variability into a unified metric. We show that feature ranking and selection stability based on DD can perform two to three times better than DE or DV alone, and that DD yields equivalent error rates to DE and DV. Finally, assessing genes via differential distribution produces a complementary set of selected genes to DE and DV, potentially opening up new categories of biomarkers.

  5. A Subset of Autism-associated Genes Regulate the Structural Stability of Neurons

    Directory of Open Access Journals (Sweden)

    Yu-Chih Lin

    2016-11-01

    Full Text Available Autism spectrum disorder (ASD comprises a range of neurological conditions that affect individuals’ ability to communicate and interact with others. People with ASD often exhibit marked qualitative difficulties in social interaction, communication, and behavior. Alterations in neurite arborization and dendritic spine morphology, including size, shape, and number, are hallmarks of almost all neurological conditions, including ASD. As experimental evidence emerges in recent years, it becomes clear that although there is broad heterogeneity of identified autism risk genes, many of them converge into similar cellular pathways, including those regulating neurite outgrowth, synapse formation and spine stability, and synaptic plasticity. These mechanisms together regulate the structural stability of neurons and are vulnerable targets in ASD. In this review, we discuss the current understanding of those autism risk genes that affect the structural connectivity of neurons. We sub-categorize them into 1 cytoskeletal regulators, e.g. motors and small RhoGTPase regulators; 2 adhesion molecules, e.g. cadherins, NCAM, and neurexin superfamily; 3 cell surface receptors, e.g. glutamatergic receptors and receptor tyrosine kinases; 4 signaling molecules, e.g. protein kinases and phosphatases; and 5 synaptic proteins, e.g. vesicle and scaffolding proteins. Although the roles of some of these genes in maintaining neuronal structural stability are well studied, how mutations contribute to the autism phenotype is still largely unknown. Investigating whether and how the neuronal structure and function are affected when these genes are mutated will provide insights toward developing effective interventions aimed at improving the lives of people with autism and their families.

  6. Cell Cycle Regulates Nuclear Stability of AID and Determines the Cellular Response to AID.

    Directory of Open Access Journals (Sweden)

    Quy Le

    2015-09-01

    Full Text Available AID (Activation Induced Deaminase deaminates cytosines in DNA to initiate immunoglobulin gene diversification and to reprogram CpG methylation in early development. AID is potentially highly mutagenic, and it causes genomic instability evident as translocations in B cell malignancies. Here we show that AID is cell cycle regulated. By high content screening microscopy, we demonstrate that AID undergoes nuclear degradation more slowly in G1 phase than in S or G2-M phase, and that mutations that affect regulatory phosphorylation or catalytic activity can alter AID stability and abundance. We directly test the role of cell cycle regulation by fusing AID to tags that destabilize nuclear protein outside of G1 or S-G2/M phases. We show that enforced nuclear localization of AID in G1 phase accelerates somatic hypermutation and class switch recombination, and is well-tolerated; while nuclear AID compromises viability in S-G2/M phase cells. We identify AID derivatives that accelerate somatic hypermutation with minimal impact on viability, which will be useful tools for engineering genes and proteins by iterative mutagenesis and selection. Our results further suggest that use of cell cycle tags to regulate nuclear stability may be generally applicable to studying DNA repair and to engineering the genome.

  7. Cell Cycle Regulates Nuclear Stability of AID and Determines the Cellular Response to AID.

    Directory of Open Access Journals (Sweden)

    Quy Le

    2015-09-01

    Full Text Available AID (Activation Induced Deaminase deaminates cytosines in DNA to initiate immunoglobulin gene diversification and to reprogram CpG methylation in early development. AID is potentially highly mutagenic, and it causes genomic instability evident as translocations in B cell malignancies. Here we show that AID is cell cycle regulated. By high content screening microscopy, we demonstrate that AID undergoes nuclear degradation more slowly in G1 phase than in S or G2-M phase, and that mutations that affect regulatory phosphorylation or catalytic activity can alter AID stability and abundance. We directly test the role of cell cycle regulation by fusing AID to tags that destabilize nuclear protein outside of G1 or S-G2/M phases. We show that enforced nuclear localization of AID in G1 phase accelerates somatic hypermutation and class switch recombination, and is well-tolerated; while nuclear AID compromises viability in S-G2/M phase cells. We identify AID derivatives that accelerate somatic hypermutation with minimal impact on viability, which will be useful tools for engineering genes and proteins by iterative mutagenesis and selection. Our results further suggest that use of cell cycle tags to regulate nuclear stability may be generally applicable to studying DNA repair and to engineering the genome.

  8. Attitude determination for three-axis stabilized geostationary meteorological satellite image navigation

    Science.gov (United States)

    Wu, Yaguang; Wang, Zhigang

    2005-11-01

    To achieve the high accuracy of attitude determination for three-axis stabilized geostationary meteorological satellite image navigation, a new approach combined gyro with star trackers is proposed, and a real-time algorithm for attitude estimation is designed. This algorithm begins with a prediction for angular rate model errors induced by gyro drifting error, and ends with the extended Kalman filtering (EKF) for attitude estimation of three-axis. A Matlab-based time domain simulation model is developed to evaluate the attitude determination performance. Simulation results demonstrate that the proposed algorithm has characteristics of high accuracy, rapid convergence and strong robustness.

  9. Stabilization

    Directory of Open Access Journals (Sweden)

    Muhammad H. Al-Malack

    2016-07-01

    Full Text Available Fuel oil flyash (FFA produced in power and water desalination plants firing crude oils in the Kingdom of Saudi Arabia is being disposed in landfills, which increases the burden on the environment, therefore, FFA utilization must be encouraged. In the current research, the effect of adding FFA on the engineering properties of two indigenous soils, namely sand and marl, was investigated. FFA was added at concentrations of 5%, 10% and 15% to both soils with and without the addition of Portland cement. Mixtures of the stabilized soils were thoroughly evaluated using compaction, California Bearing Ratio (CBR, unconfined compressive strength (USC and durability tests. Results of these tests indicated that stabilized sand mixtures could not attain the ACI strength requirements. However, marl was found to satisfy the ACI strength requirement when only 5% of FFA was added together with 5% of cement. When the FFA was increased to 10% and 15%, the mixture’s strength was found to decrease to values below the ACI requirements. Results of the Toxicity Characteristics Leaching Procedure (TCLP, which was performed on samples that passed the ACI requirements, indicated that FFA must be cautiously used in soil stabilization.

  10. Pre-Analytical Determination of the Effect of Extended Warm or Cold Ischaemia on RNA Stability in the Human Ileum Mucosa.

    Directory of Open Access Journals (Sweden)

    Serene M L Lee

    Full Text Available The use of banked human tissue, obtained with informed consent after elective surgical procedures, represents a powerful model for understanding underlying mechanisms of diseases or therapeutic interventions and for establishing prognostic markers. However, donated tissues typically have varying times of warm ischaemia in situ due to blood arrest or cold ischaemia due to procurement and transportation. Hence, before using these tissues, it is important to carry out pre-analytical studies to ensure that they are representative of the in vivo state. In particular, tissues of the gastrointestinal tract have been thought to have low RNA stability. Therefore, this study aimed to determine if extended warm or cold ischaemia times and snap-freezing or banking in RNA stabilization solution affects RNA integrity or gene expression in human ileum mucosa. In short, ileum mucosa was collected for up to 1.5 h and 6 h of simulated warm or cold ischaemia respectively. Subsequently, RNA integrity and gene expressions were determined. It was found that RNA integrity remained high over the course of warm and cold ischaemia examined and there were in general no significant differences between snap-freezing and banking in RNA stabilization solution. Following the same trend, there were in general no significant changes in gene expressions measured (MYC, HIF1α, CDX, HMOX1 and IL1β. In conclusion, RNA in the ileum mucosa is maintained at a high integrity and has stable gene expression over the examined time course of warm or cold ischaemia when banked in RNA stabilization solution or snap-frozen in liquid nitrogen. As the average warm and cold ischaemia times imposed by surgery and the process of tissue banking are shorter than the time period examined in this study, human ileum mucosa samples collected after surgeries could be used for gene expression studies.

  11. Expression Stabilities of Candidate Reference Genes for RT-qPCR in Chinese Jujube (Ziziphus jujuba Mill. under a Variety of Conditions.

    Directory of Open Access Journals (Sweden)

    Jiaodi Bu

    Full Text Available Reverse transcription-quantitative real-time polymerase chain reaction (RT-qPCR is a powerful method for evaluating patterns of gene expression. Jujube whole-genome sequencing has been completed, and analysis of gene function, an important part of any follow-up study, requires the appropriate selection of reference genes. Indeed, suitable reference gene selection for RT-qPCR is critical for accurate normalization of target gene expression. In this study, the software packages geNorm and NormFinder were employed to examine the expression stabilities of nine candidate reference genes under a variety of conditions. Actin-depolymerizing factor 1 (ACT1, Histone-H3 (His3, and Polyadenylate-binding protein-interacting protein (PAIP were determined to be the most stably expressed genes during five stages of fruit development and ACT1, SiR-Fd, BTF3, and Tubulin alpha chain (TUA across different tissues/organs. Whereas ACT1, Basic Transcription factor 3 (BTF3, Glyceraldehyde-3-phosphate dehydrogenase (GADPH, and PAIP were the most stable under dark conditions. ACT1, PAIP, BTF3, and Elongation factor 1- gamma (EF1γ were the most stably expressed genes under phytoplasma infection. Among these genes, SiR-Fd and PAIP are here first reported as stable reference genes. When normalized using these most stable reference genes, the expression patterns of four target genes were found to be in accordance with physiological data, indicating that the reference genes selected in our study are suitable for use in such analyses. This study provides appropriate reference genes and corresponding primers for further RT-qPCR studies in Chinese jujube and emphasizes the importance of validating reference genes for gene expression analysis under variable experimental conditions.

  12. Determination of alpha-2-MRAP gene polymorphisms in nephrolithiasis patients.

    Science.gov (United States)

    Mehde, Atheer Awad; Mehdi, Wesen Adel; Yusof, Faridah; Raus, Raha Ahmed; Abidin, Zaima Azira Zainal; Ghazali, Hamid; Rahman, Azlina Abd

    2017-07-29

    The intron 5 insertion/deletion polymorphism of Alpha-2-macroglobulin receptor-associated protein gene (Alpha-2-MRAP) has been implicated in numerous diseases. The current study was designed to analyze the association of intron 5 insertion/deletion polymorphism of Alpha-2-MRAP with nephrolithiasis patients. PCR was conducted on genomic DNA of patients and control to look for Alpha-2-MRAP insertion/deletion polymorphism. Besides that, serum level of Alpha-2-MRAP, oxidative stress marker myeloperoxidase, Malondialdehyde (MDA), Advanced oxidation protein products (AOPP), and uric acid were determined. The D and I allele frequencies were 57.50% and 42.50% in patients, 77.50% and 22.50% in control, individually. The result showed that II genotype was associated with nephrolithiasis patients group. A significant decrease was observed in serum Alpha-2-MRAP,myeloperoxidase and TAS,while TOS,OSI,MDA,AOPP and uric acid were substantially increased in II and ID when compared to DD genotype in patients with nephrolithiasis. Our results demonstrate for the first time that patients with II genotype had an increased risk of stones. Also, the results demonstrate that I allele of the 5 insertion/deletion polymorphism in the Alpha-2-MRAP gene is related with an increase of oxidative stress in nephrolithiasis patients and may possibly impose a risk for cardiovascular diseases in patients with II genotype of Alpha-2-MRAP. Copyright © 2017. Published by Elsevier B.V.

  13. [Effects of knockout ECM25/YJL201W gene in brewing yeast on beer flavor stability].

    Science.gov (United States)

    Zhang, Yixin; Li, Qi; Shen, Wei; Xie, Yan; Gu, Guoxian

    2008-08-01

    The ECM25 deletion mutant of industrial brewing yeast, G03/a, was constructed by replacing the ECM25 gene with the kanMX gene. The transformant was verified to be genetically stable. The PCR analysis showed that ECM25 gene in the G-03/a was deleted. Under aerobic conditions of ll degrees C and 28 degrees C, compared with the host strain G-03, the excretive glutathione concentration of G-03/a increased by 21.4% and 14.7%, respectively. Strains G-03 and G-03/a were inoculated in flasks and cultivated continuously for 4 generations. Compared with the host strain G-03, the glutathione concentration in the main fermentation broth and final beer of strain G-03/a increased by 32.1% and 13.8%, the stability index (SI) increased by 7.7% and 5.3%, respectively, and the flavor resistance staling value (RSV value) in final beer increased by 45.0%. During EBC fermentation, the glutathione concentration in the main fermentation broth of strain G-03/a increased by 34.0%, compared with the host strain G-03. Furthermore, no significant difference in routine fermentation parameters was found. The strain G-03/a is proved to be an excellent anti-staling brewing yeast to improve beer flavor stability.

  14. Milks pigmentation with astaxanthin and determination of colour stability during short period cold storage.

    Science.gov (United States)

    Mezquita, Pedro Cerezal; Huerta, Blanca E Barragán; Ramírez, Jenifer C Palma; Hinojosa, Claudia P Ortíz

    2015-03-01

    Astaxanthin has been used as a colorant and antioxidant with excellent results, its application and stability in food matrices to human consumption has been little studied. The aim of this work was the incorporation of astaxanthin oleoresin to milks with different fat content, simulating the red-orange color that can impart apricot fruit. For astaxanthin determination by HPLC, a methodology was implemented for its extraction from the food matrix, followed by saponification with KOH. Milk samples were stored (5 ± 2 °C) and stability of color and astaxanthin content were determined by colorimetry and high performance liquid chromatography each 24 h for a week. Pigment degradation followed first-order kinetic with a constant degradation of 0.259 day(-1) and 0.104 day(-1), in whole and semi-skimmed milk, respectively. Chromaticity coordinates L*, a*, b* for different types of milk showed a low dispersion of their values during the storage time, indicating high stability of astaxanthin within the matrix.

  15. Stability Indicating RP-HPLC Method for Determination of Valsartan in Pure and Pharmaceutical Formulation

    Directory of Open Access Journals (Sweden)

    S. K. Patro

    2010-01-01

    Full Text Available A simple, rapid and accurate and stability indicating RP-HPLC method was developed for the determination of valsartan in pure and tablet forms. The method showed a linear response for concentrations in the range of 50-175 µg/mL using 0.01 M NH4H2PO4 (pH 3.5 buffer: methanol [50:50] as the mobile phase with detection at 210 nm and a flow rate of 1 mL/min and retention time 11.041 min. The method was statistically validated for accuracy, precision, linearity, ruggedness, robustness, forced degradation, solution stability and selectivity. Quantitative and recovery studies of the dosage form were also carried out and analyzed; the % RSD from recovery studies was found to be less than 1. Due to simplicity, rapidity and accuracy of the method, we believe that the method will be useful for routine quality control analysis.

  16. Determination of the oxidative stability by DSC of vegetable oils from the Amazonian area.

    Science.gov (United States)

    Pardauil, Juliana J R; Souza, Luiz K C; Molfetta, Fábio A; Zamian, José R; Rocha Filho, Geraldo N; da Costa, C E F

    2011-05-01

    Differential scanning calorimetry (DSC) and a Rancimat method apparatus were applied to evaluate the oxidative stability of buriti pulp oil (Mauritia flexuosa Mart), rubber seed oil (Hevea brasiliensis), and passion fruit oil (Passiflora edulis). The Rancimat measurements taken for the oxidative induction times were performed under isothermal conditions at 100°C and in an air atmosphere. The DSC technique involved the oxidation of oil samples in an oxygen-flow DSC cell. The DSC cell temperature was set at five different isothermal temperatures: 100, 110, 120, 130 and 140°C. During the oxidation reaction, an increase in heat was observed as a sharp exothermic curve. The value T(0) represents the oxidative induction time, which is determined from the downward extrapolated DSC oxidative curve verses the time axis. These curves indicate a good correlation between the DSC T(0) and oxidative stability index (OSI) values. The DSC method is useful because it consumes less time and less sample.

  17. Identification of two key genes controlling chill haze stability of beer in barley (Hordeum vulgare L).

    Science.gov (United States)

    Ye, Lingzhen; Huang, Yuqing; Dai, Fei; Ning, Huajiang; Li, Chengdao; Zhou, Meixue; Zhang, Guoping

    2015-06-11

    In bright beer, haze formation is a serious quality problem, degrading beer quality and reducing its shelf life. The quality of barley (Hordeum vulgare L) malt, as the main raw material for beer brewing, largely affects the colloidal stability of beer. In this study, the genetic mechanism of the factors affecting beer haze stability in barley was studied. Quantitative trait loci (QTL) analysis of alcohol chill haze (ACH) in beer was carried out using a Franklin/Yerong double haploid (DH) population. One QTL, named as qACH, was detected for ACH, and it was located on the position of about 108 cM in chromosome 4H and can explain about 20 % of the phenotypic variation. Two key haze active proteins, BATI-CMb and BATI-CMd were identified by proteomics analysis. Bioinformatics analysis showed that BATI-CMb and BATI-CMd had the same position as qACH in the chromosome. It may be deduced that BATI-CMb and BATI-CMd are candidate genes for qACH, controlling colloidal stability of beer. Polymorphism comparison between Yerong and Franklin in the nucleotide and amino acid sequence of BATI-CMb and BATI-CMd detected the corresponding gene specific markers, which could be used in marker-assisted selection for malt barley breeding. We identified a novel QTL, qACH controlling chill haze of beer, and two key haze active proteins, BATI-CMb and BATI-CMd. And further analysis showed that BATI-CMb and BATI-CMd might be the candidate genes associated with beer chill haze.

  18. An ultrasonic-accelerated oxidation method for determining the oxidative stability of biodiesel.

    Science.gov (United States)

    Avila Orozco, Francisco D; Sousa, Antonio C; Domini, Claudia E; Ugulino Araujo, Mario Cesar; Fernández Band, Beatriz S

    2013-05-01

    Biodiesel is considered an alternative energy because it is produced from fats and vegetable oils by means of transesterification. Furthermore, it consists of fatty acid alkyl esters (FAAS) which have a great influence on biodiesel fuel properties and in the storage lifetime of biodiesel itself. The biodiesel storage stability is directly related to the oxidative stability parameter (Induction Time - IT) which is determined by means of the Rancimat® method. This method uses condutimetric monitoring and induces the degradation of FAAS by heating the sample at a constant temperature. The European Committee for Standardization established a standard (EN 14214) to determine the oxidative stability of biodiesel, which requires it to reach a minimum induction period of 6h as tested by Rancimat® method at 110°C. In this research, we aimed at developing a fast and simple alternative method to determine the induction time (IT) based on the FAAS ultrasonic-accelerated oxidation. The sonodegradation of biodiesel samples was induced by means of an ultrasonic homogenizer fitted with an immersible horn at 480Watts of power and 20 duty cycles. The UV-Vis spectrometry was used to monitor the FAAS sonodegradation by measuring the absorbance at 270nm every 2. Biodiesel samples from different feedstock were studied in this work. In all cases, IT was established as the inflection point of the absorbance versus time curve. The induction time values of all biodiesel samples determined using the proposed method was in accordance with those measured through the Rancimat® reference method by showing a R(2)=0.998.

  19. Determination of Close Loop System Stability in Automobile Adaptive Cruise Control Systems

    Directory of Open Access Journals (Sweden)

    Owunna Ikechukwu

    2016-07-01

    Full Text Available The beginning of the 21st century sees auto makers pursuing research in advanced features like collision warning and avoidance system into their product. Automotive cruise control system has been undergoing development in EU since the PROMETHEUS programme in the late 1980’s, and has currently metamorphous into Adaptive Cruise Control (ACC technology which is presently emerging in the automotive market as a convenience function intended to reduce driver workload. Adaptive cruise control is the first of the new generation of advanced driver’s assistance devices to reach the market, which partially automates the driver’s task and bringing the drivers comfort into perspective. It allows the host vehicle to maintain a set speed and distance from preceding vehicles by a forward object detection sensor. The forward object detection sensor is the focal point of the ACC system, which determines and regulates vehicle acceleration and deceleration through a powertrain torque control system and an automatic brake control system. This study presents overview of adaptive cruise control system, operation principles and the advantages of integrating ACC system in automobiles. Also, the system must be stable for optimum performance, and stability of a close loop system which the cruise system is an example, was determined by calculating the controller gain (K1, K2, K3 and substituting into the characteristic equations. The stability of a close loop system for the values of K1, K2 and K3 when substituted into the characteristic equation produced a negative real part. To achieve stability in close loop systems, all the poles must have negative real values and this is in line with the values obtain for p1, p2 and p3. From the pole zero plots of 1 = (-7 ± 7.14, 2 = (-7± 11.60 and 3 = (-0.08 and -13.91, stability of the system was achieved

  20. CRISPR/Cas9-based genome editing for simultaneous interference with gene expression and protein stability

    DEFF Research Database (Denmark)

    Martinez, Virginia; Lauritsen, Ida; Hobel, Tonja

    2017-01-01

    Interference with genes is the foundation of reverse genetics and is key to manipulation of living cells for biomedical and biotechnological applications. However, classical genetic knockout and transcriptional knockdown technologies have different drawbacks and offer no control over existing...... protein levels. Here, we describe an efficient genome editing approach that affects specific protein abundances by changing the rates of both RNA synthesis and protein degradation, based on the two cross-kingdom control mechanisms CRISPRi and the N-end rule for protein stability. In addition, our approach...

  1. Control of gene expression during T cell activation: alternate regulation of mRNA transcription and mRNA stability

    Directory of Open Access Journals (Sweden)

    Gorospe Myriam

    2005-05-01

    Full Text Available Abstract Background Microarray technology has become highly valuable for identifying complex global changes in gene expression patterns. The effective correlation of observed changes in gene expression with shared transcription regulatory elements remains difficult to demonstrate convincingly. One reason for this difficulty may result from the intricate convergence of both transcriptional and mRNA turnover events which, together, directly influence steady-state mRNA levels. Results In order to investigate the relative contribution of gene transcription and changes in mRNA stability regulation to standard analyses of gene expression, we used two distinct microarray methods which individually measure nuclear gene transcription and changes in polyA mRNA gene expression. Gene expression profiles were obtained from both polyA mRNA (whole-cell and nuclear run-on (newly transcribed RNA across a time course of one hour following the activation of human Jurkat T cells with PMA plus ionomycin. Comparative analysis revealed that regulation of mRNA stability may account for as much as 50% of all measurements of changes in polyA mRNA in this system, as inferred by the absence of any corresponding regulation of nuclear gene transcription activity for these groups of genes. Genes which displayed dramatic elevations in both mRNA and nuclear run-on RNA were shown to be inhibited by Actinomycin D (ActD pre-treatment of cells while large numbers of genes regulated only through altered mRNA turnover (both up and down were ActD-resistant. Consistent patterns across the time course were observed for both transcribed and stability-regulated genes. Conclusion We propose that regulation of mRNA stability contributes significantly to the observed changes in gene expression in response to external stimuli, as measured by high throughput systems.

  2. On the Decay Ratio Determination in BWR Stability Analysis by Auto-Correlation Function Techniques

    Energy Technology Data Exchange (ETDEWEB)

    Behringer, K.; Hennig, D

    2002-11-01

    A novel auto-correlation function (ACF) method has been investigated for determining the oscillation frequency and the decay ratio in BWR stability analyses. The neutron signals are band-pass filtered to separate the oscillation peak in the power spectral density (PSD) from background. Two linear second-order oscillation models are considered. These models, corrected for signal filtering and including a background term under the peak in the PSD, are then least-squares fitted to the ACF of the previously filtered neutron signal, in order to determine the oscillation frequency and the decay ratio. Our method uses fast Fourier transform techniques with signal segmentation for filtering and ACF estimation. Gliding 'short-term' ACF estimates on a record allow the evaluation of uncertainties. Numerical results are given which have been obtained from neutron data of the recent Forsmark I and Forsmark II NEA benchmark project. Our results are compared with those obtained by other participants in the benchmark project. The present PSI report is an extended version of the publication K. Behringer, D. Hennig 'A novel auto-correlation function method for the determination of the decay ratio in BWR stability studies' (Behringer, Hennig, 2002)

  3. Stabilizing in vitro ultrasound-mediated gene transfection by regulating cavitation.

    Science.gov (United States)

    Lo, Chia-Wen; Desjouy, Cyril; Chen, Shing-Ru; Lee, Jyun-Lin; Inserra, Claude; Béra, Jean-Christophe; Chen, Wen-Shiang

    2014-03-01

    It is well known that acoustic cavitation can facilitate the inward transport of genetic materials across cell membranes (sonoporation). However, partially due to the unstationary behavior of the initiation and leveling of cavitation, the sonoporation effect is usually unstable, especially in low intensity conditions. A system which is able to regulate the cavitation level during sonication by modulating the applied acoustic intensity with a feedback loop is implemented and its effect on in vitro gene transfection is tested. The regulated system provided better time stability and reproducibility of the cavitation levels than the unregulated conditions. Cultured hepatoma cells (BNL) mixed with 10 μg luciferase plasmids are exposed to 1-MHz pulsed ultrasound with or without cavitation regulation, and the gene transfection efficiency and cell viability are subsequently assessed. Experimental results show that for all exposure intensities (low, medium, and high), stable and intensity dependent, although not higher, gene expression could be achieved in the regulated cavitation system than the unregulated conditions. The cavitation regulation system provides a better control of cavitation and its bioeffect which are crucial important for clinical applications of ultrasound-mediated gene transfection.

  4. Upgrading fuzzy logic by GA-PS to determine asphaltene stability in crude oil

    Directory of Open Access Journals (Sweden)

    Saeid Ahmadi

    2017-06-01

    Full Text Available Precipitation and deposition of asphaltene are undesirable phenomena that arise during petroleum production which give rise to a pronounced rate of increase in operational cost and adversely affect production rates as well. Hence, it is imperative to develop a mathematical model for the assessment of asphaltene stability in crude oil. In the present study, delta RI which constitutes the difference between refractive index of crude oil (RI and refractive index of crude oil at the onset of asphaltene precipitation (PRI is employed as the principal factor for determining the asphaltene stability of the region. Fuzzy logic is a potent tool capable of extracting the underlying dependency between SARA fractions (saturate, aromatic, resin, and asphaltene data and delta RI for the inexpensive and rapid diagnosis of asphaltene stability. In this study a novel strategy known as hybrid genetic algorithm-pattern search (GA-PS is suggested for the development of an optimal fuzzy logic model as a reliable alternative for the widely-applied subtractive clustering (SC method. While SC solely optimizes mean of input Gaussian membership functions (GMFs, GA-PS tool optimizes both mean and variance of input GMFs. Comparison between GA-PS and SC methods confirmed the capability of GA-PS for developing an optimal fuzzy logic model.

  5. ECOLOGICAL STABILITY AS A DETERMINANT OF NITRA REGION DEVELOPMENT IN SLOVAKIA

    Directory of Open Access Journals (Sweden)

    Marián KOTRLA

    2014-04-01

    Full Text Available Nitra region is evaluated based on the analysis of land use ecological stability in the present article, which is significant for its development, particularly in terms of increasing potential for tourism. The coefficient of ecological stability (CES was used to evaluate the ecological stability of the region as a relatively simple ecological indicator for quality of life determination in the region. Three basic methods were used of calculating CES according to the methodology Míchal (1982, Löw et al. (1984 and Miklós (1986. The districts of Nitra region differs by natural conditions from each other. There are represented three types of natural areas: foothill (Zlaté Moravce, upland (Nitra, Levice, Topoľčany and lowland area (Šaľa, Nové Zámky, Komárom. The highest value of CES was in the district of Zlaté Moravce. A higher value of CES was in the district of Topoľčany. The lowest value of CES was evaluated in the district of Šaľa, where do absent ecologically stable elements. Nitra region is classifies as an area of low to median ecological stability, which is critical to the improvement and development of the region need to make eco-stabilizing elements and eco-stabilizing management measures. On the basis of country originality can be expressed the impact of human activity on the landscape and its features. Originality coefficient of the cultural landscape for individual districts of Nitra region is following: Komárno and Levice - 0.16; Nitra - 0.17; Nové Zámky - 0.15; Šaľa - 0.07; Topoľčany - 0.56; Zlaté Moravce - 1.49. Slovakia is in terms of the authenticity of cultural landscapes evaluated coefficient of 2.01. Based on the analysis it can be concluded heterogeneity in the origin of cultural landscapes in the Nitra region.

  6. Stabilization of gene expression and cell morphology after explant recycling during fin explant culture in goldfish.

    Science.gov (United States)

    Chenais, Nathalie; Lareyre, Jean-Jacques; Le Bail, Pierre-Yves; Labbe, Catherine

    2015-07-01

    The development of fin primary cell cultures for in vitro cellular and physiological studies is hampered by slow cell outgrowth, low proliferation rate, poor viability, and sparse cell characterization. Here, we investigated whether the recycling of fresh explants after a first conventional culture could improve physiological stability and sustainability of the culture. The recycled explants were able to give a supplementary cell culture showing faster outgrowth, cleaner cell layers and higher net cell production. The cells exhibited a highly stabilized profile for marker gene expression including a low cytokeratin 49 (epithelial marker) and a high collagen 1a1 (mesenchymal marker) expression. Added to the cell spindle-shaped morphology, motility behavior, and actin organization, this suggests that the cells bore stable mesenchymal characteristics. This contrast with the time-evolving expression pattern observed in the control fresh explants during the first 2 weeks of culture: a sharp decrease in cytokeratin 49 expression was concomitant with a gradual increase in col1a1. We surmise that such loss of epithelial features for the benefit of mesenchymal ones was triggered by an epithelial to mesenchymal transition (EMT) process or by way of a progressive population replacement process. Overall, our findings provide a comprehensive characterization of this new primary culture model bearing mesenchymal features and whose stability over culture time makes those cells good candidates for cell reprogramming prior to nuclear transfer, in a context of fish genome preservation.

  7. Double-bottom chaotic map particle swarm optimization based on chi-square test to determine gene-gene interactions.

    Science.gov (United States)

    Yang, Cheng-Hong; Lin, Yu-Da; Chuang, Li-Yeh; Chang, Hsueh-Wei

    2014-01-01

    Gene-gene interaction studies focus on the investigation of the association between the single nucleotide polymorphisms (SNPs) of genes for disease susceptibility. Statistical methods are widely used to search for a good model of gene-gene interaction for disease analysis, and the previously determined models have successfully explained the effects between SNPs and diseases. However, the huge numbers of potential combinations of SNP genotypes limit the use of statistical methods for analysing high-order interaction, and finding an available high-order model of gene-gene interaction remains a challenge. In this study, an improved particle swarm optimization with double-bottom chaotic maps (DBM-PSO) was applied to assist statistical methods in the analysis of associated variations to disease susceptibility. A big data set was simulated using the published genotype frequencies of 26 SNPs amongst eight genes for breast cancer. Results showed that the proposed DBM-PSO successfully determined two- to six-order models of gene-gene interaction for the risk association with breast cancer (odds ratio > 1.0; P value <0.05). Analysis results supported that the proposed DBM-PSO can identify good models and provide higher chi-square values than conventional PSO. This study indicates that DBM-PSO is a robust and precise algorithm for determination of gene-gene interaction models for breast cancer.

  8. Double-Bottom Chaotic Map Particle Swarm Optimization Based on Chi-Square Test to Determine Gene-Gene Interactions

    Directory of Open Access Journals (Sweden)

    Cheng-Hong Yang

    2014-01-01

    Full Text Available Gene-gene interaction studies focus on the investigation of the association between the single nucleotide polymorphisms (SNPs of genes for disease susceptibility. Statistical methods are widely used to search for a good model of gene-gene interaction for disease analysis, and the previously determined models have successfully explained the effects between SNPs and diseases. However, the huge numbers of potential combinations of SNP genotypes limit the use of statistical methods for analysing high-order interaction, and finding an available high-order model of gene-gene interaction remains a challenge. In this study, an improved particle swarm optimization with double-bottom chaotic maps (DBM-PSO was applied to assist statistical methods in the analysis of associated variations to disease susceptibility. A big data set was simulated using the published genotype frequencies of 26 SNPs amongst eight genes for breast cancer. Results showed that the proposed DBM-PSO successfully determined two- to six-order models of gene-gene interaction for the risk association with breast cancer (odds ratio > 1.0; P value <0.05. Analysis results supported that the proposed DBM-PSO can identify good models and provide higher chi-square values than conventional PSO. This study indicates that DBM-PSO is a robust and precise algorithm for determination of gene-gene interaction models for breast cancer.

  9. Analytic theory for the determination of velocity and stability of bubbles in a Hele-Shaw cell. I - Velocity selection. II - Stability

    Science.gov (United States)

    Tanveer, S.

    1989-01-01

    An asymptotic theory is presented for the determination of velocity and linear stability of a steady symmetric bubble in a Hele-Shaw cell for small surface tension. First the bubble velocity relative to the fluid velocity at infinity is determined for small surface tension by means of a transcendentally small correction to the asymptotic series solution. In addition, a linear stability analysis shows that only the solution branch corresponding to the largest possible bubble velocity for given surface tension is stable, while all the others are unstable.

  10. Development and validation of stability-indicating HPLC method for determination of cefpirome sulfate.

    Science.gov (United States)

    Zalewski, Przemysław; Skibiński, Robert; Cielecka-Piontek, Judyta; Bednarek-Rajewska, Katarzyna

    2014-01-01

    The stability-indicating LC assay method was developed and validated for quantitative determination of cefpirome sulfate (CPS) in the presence of degradation products formed during the forced degradation studies. An isocratic HPLC method was developed with Lichrospher RP-18 column, 5 μm particle size, 125 mm x 4 mm column and 12 mM ammonium acetate-acetonitrile (90 : 10 v/v) as a mobile phase. The flow rate of the mobile phase was 1.0 mL/min. Detection wavelength was 270 nm and temperature was 30 degrees C. Cefpirome sulfate as other cephalosporins was subjected to stress conditions of degradation in aqueous solutions including hydrolysis, oxidation, photolysis and thermal degradation. The developed method was validated with regard to linearity, accuracy, precision, selectivity and robustness. The method was applied successfully for identification and determination of cefpirome sulfate in pharmaceuticals and during kinetic studies.

  11. Validated Stability Indicating RP-HPLC Method for the Determination of Silodosin in Pharmaceutical Dosage Form

    Directory of Open Access Journals (Sweden)

    Harischandran S

    2012-12-01

    Full Text Available A simple, rapid and economic stability indicating high performance liquid chromatography method was developed for the determination of Silodosin in pharmaceutical dosage form. The chromatographic system comprised of a reverse phase Phenomenex C 18, 5µ Silica (250×4mm column maintained at 25°C with mobile phase consisting of a mixture of methanol-water-acetonitrile-glacial acetic acid (60:27:10:3 % v/v at pH 3.2 ± 0.1 with a flow rate of 1 ml/min, determined at 270 nm. The method was linear in the range of 10-100µg/ml. The results were validated according to ICH guidelines. The method could effectively separate the drug from its degradation products.

  12. High-precision, high-throughput stability determinations facilitated by robotics and a semiautomated titrating fluorometer.

    Science.gov (United States)

    Edgell, Marshall Hall; Sims, Dorothy A; Pielak, Gary J; Yi, Fang

    2003-06-24

    The use of statistical modeling to test hypotheses concerning the determinants of protein structure requires stability data (e.g., the free energy of denaturation in H(2)O, DeltaG(HOH)) from hundreds of protein mutants. Fluorescence-monitored chemical denaturation provides a convenient method for high-precision, high-throughput DeltaG(HOH) determination. For eglin c we find that a throughput of about 20 min per protein can be attained in a two-channel semiautomated titrating fluorometer. We find also that the use of robotics for protein purification and preparation of the solutions for chemical denaturation gives highly precise DeltaG(HOH) values in which the standard deviation of values from multiple preparations (+/-0.051 kcal/mol) differs very little from multiple measurements from a single preparation (+/-0.040 kcal/mol). Since the variance introduced into model fitting by DeltaG(HOH) increases as the square of measurement error, there is a premium on precision. In fact, the fraction of stability behavior explicable by otherwise perfect models goes from 98% to only 50% over the error range commonly reported for chemical denaturation measurements (0.1-0.6 kcal/mol). We have found that the precision of chemical denaturation DeltaG(HOH) measurements depends most heavily on the precision of the instrument used, followed by protein purity and the capacity to precisely prepare the solutions used for titrations.

  13. Application of vacuum stability test to determine thermal decomposition kinetics of nitramines bonded by polyurethane matrix

    Science.gov (United States)

    Elbeih, Ahmed; Abd-Elghany, Mohamed; Elshenawy, Tamer

    2017-03-01

    Vacuum stability test (VST) is mainly used to study compatibility and stability of energetic materials. In this work, VST has been investigated to study thermal decomposition kinetics of four cyclic nitramines, 1,3,5-trinitro-1,3,5-triazinane (RDX) and 1,3,5,7-tetranitro-1,3,5,7-tetrazocane (HMX), cis-1,3,4,6-tetranitrooctahydroimidazo-[4,5-d]imidazole (BCHMX), 2,4,6,8,10,12-hexanitro-2,4,6,8,10,12-hexaazaisowurtzitane (ε-HNIW, CL-20), bonded by polyurethane matrix based on hydroxyl terminated polybutadiene (HTPB). Model fitting and model free (isoconversional) methods have been applied to determine the decomposition kinetics from VST results. For comparison, the decomposition kinetics were determined isothermally by ignition delay technique and non-isothermally by Advanced Kinetics and Technology Solution (AKTS) software. The activation energies for thermolysis obtained by isoconversional method based on VST technique of RDX/HTPB, HMX/HTPB, BCHMX/HTPB and CL20/HTPB were 157.1, 203.1, 190.0 and 176.8 kJ mol-1 respectively. Model fitting method proved that the mechanism of thermal decomposition of BCHMX/HTPB is controlled by the nucleation model while all the other studied PBXs are controlled by the diffusion models. A linear relationship between the ignition temperatures and the activation energies was observed. BCHMX/HTPB is interesting new PBX in the research stage.

  14. STABILITY-INDICATING HPLC METHOD DEVELOPMENT AND VALIDATION FOR THE DETERMINATION OF LACIDIPINE IN TABLETS

    Directory of Open Access Journals (Sweden)

    Rajavel A

    2011-05-01

    Full Text Available A stability-indicating HPLC method development and validation for the determination of Lacidipine in tablets. The determination was done for an active pharmaceutical ingredient, its pharmaceutical dosage form in the presence of degradation products impurities. The drug was subjected to stress conditions of hydrolysis (acid and base, oxidation, photolysis and thermal degradation as per International Conference on Harmonization (ICH prescribed stress conditions to show the stability-indicating method. Degradation was observed during acid, base hydrolysis, thermal, peroxide and light stressed sample and degradant was identified by LC–MS, FTIR and 1H/13C NMR spectral analysis. The generated samples were used for forced degradation studies. In the developed HPLC method, the resolution between Lacidipine and, its related impurities (namely impurity-A, impurity-B, impurity-C, impurity-LC1 and impurity-LC2 was found. The chromatographic separation was achieved on a phenomenex Luna C18, 250 mm × 4.6 mm, 5 µm column. The LC method employed an isocratic elution, and the detection wavelength was set at 240 nm. The stress samples were assayed against a qualified reference standard and the mass balance was found to be close to 99.6 %. The developed RP-LC method was validated with respect to linearity, accuracy, precision and robustness.

  15. DNA stabilization by the upregulation of estrogen signaling in BRCA gene mutation carriers.

    Science.gov (United States)

    Suba, Zsuzsanna

    2015-01-01

    Currently available scientific evidence erroneously suggests that mutagenic weakness or loss of the BRCA1/2 genes may liberate the proliferative effects of estrogen signaling, which provokes DNA damage and genomic instability. Conversely, BRCA mutation seems to be an imbalanced defect, crudely inhibiting the upregulation of estrogen receptor expression and liganded transcriptional activity, whereas estrogen receptor-repressor functions become predominant. In BRCA-proficient cases, estrogen signaling orchestrates the activity of cell proliferation and differentiation with high safety, while upregulating the expression and DNA-stabilizing impact of BRCA genes. In turn, BRCA proteins promote estrogen signaling by proper estrogen synthesis via CYP19 gene regulation and by induction of the appropriate expression and transcriptional activity of estrogen receptors. In this exquisitely organized regulatory system, the dysfunction of each player may jeopardize genome stability and lead to severe chronic diseases, such as cancer development. Female organs, such as breast, endometrium, and ovary, exhibiting regular cyclic proliferative activity are particularly vulnerable in case of disturbances in either estrogen signaling or BRCA-mediated DNA repair. BRCA mutation carrier women may apparently be healthy or exhibit clinical signs of deficient estrogen signaling in spite of hyperestrogenism. Even women who enjoy sufficient compensatory DNA-defending activities are at risk of tumor development because many endogenous and environmental factors may jeopardize the mechanisms of extreme compensatory processes. Natural estrogens have numerous benefits in tumor prevention and therapy even in BRCA mutation carriers. There are no toxic effects even in sky-high doses and all physiologic cellular functions are strongly upregulated, while malignant tumor cells are recognized and killed in a Janus-faced manner.

  16. DNA replication factor C1 mediates genomic stability and transcriptional gene silencing in Arabidopsis

    KAUST Repository

    Liu, Qian

    2010-07-01

    Genetic screening identified a suppressor of ros1-1, a mutant of REPRESSOR OF SILENCING1 (ROS1; encoding a DNA demethylation protein). The suppressor is a mutation in the gene encoding the largest subunit of replication factor C (RFC1). This mutation of RFC1 reactivates the unlinked 35S-NPTII transgene, which is silenced in ros1 and also increases expression of the pericentromeric Athila retrotransposons named transcriptional silent information in a DNA methylationindependent manner. rfc1 is more sensitive than the wild type to the DNA-damaging agent methylmethane sulphonate and to the DNA inter- and intra- cross-linking agent cisplatin. The rfc1 mutant constitutively expresses the G2/M-specific cyclin CycB1;1 and other DNA repair-related genes. Treatment with DNA-damaging agents mimics the rfc1 mutation in releasing the silenced 35S-NPTII, suggesting that spontaneously induced genomic instability caused by the rfc1 mutation might partially contribute to the released transcriptional gene silencing (TGS). The frequency of somatic homologous recombination is significantly increased in the rfc1 mutant. Interestingly, ros1 mutants show increased telomere length, but rfc1 mutants show decreased telomere length and reduced expression of telomerase. Our results suggest that RFC1 helps mediate genomic stability and TGS in Arabidopsis thaliana. © 2010 American Society of Plant Biologists.

  17. Prevalent role of gene features in determining evolutionary fates of whole-genome duplication duplicated genes in flowering plants.

    Science.gov (United States)

    Jiang, Wen-kai; Liu, Yun-long; Xia, En-hua; Gao, Li-zhi

    2013-04-01

    The evolution of genes and genomes after polyploidization has been the subject of extensive studies in evolutionary biology and plant sciences. While a significant number of duplicated genes are rapidly removed during a process called fractionation, which operates after the whole-genome duplication (WGD), another considerable number of genes are retained preferentially, leading to the phenomenon of biased gene retention. However, the evolutionary mechanisms underlying gene retention after WGD remain largely unknown. Through genome-wide analyses of sequence and functional data, we comprehensively investigated the relationships between gene features and the retention probability of duplicated genes after WGDs in six plant genomes, Arabidopsis (Arabidopsis thaliana), poplar (Populus trichocarpa), soybean (Glycine max), rice (Oryza sativa), sorghum (Sorghum bicolor), and maize (Zea mays). The results showed that multiple gene features were correlated with the probability of gene retention. Using a logistic regression model based on principal component analysis, we resolved evolutionary rate, structural complexity, and GC3 content as the three major contributors to gene retention. Cluster analysis of these features further classified retained genes into three distinct groups in terms of gene features and evolutionary behaviors. Type I genes are more prone to be selected by dosage balance; type II genes are possibly subject to subfunctionalization; and type III genes may serve as potential targets for neofunctionalization. This study highlights that gene features are able to act jointly as primary forces when determining the retention and evolution of WGD-derived duplicated genes in flowering plants. These findings thus may help to provide a resolution to the debate on different evolutionary models of gene fates after WGDs.

  18. Stability of ceftazidime in 1% and 5% buffered eye drops determined with HPLC method.

    Science.gov (United States)

    Kodym, Anna; Hapka-Zmich, Dominika; Gołab, Marta; Gwizdala, Magdalena

    2011-01-01

    The aim of the studies was to determine with HPLC method the stability of ceftazidime in buffered 1% and 5% eye drops of proposed formulary composition, which were stored for 30 days at the temperature of 4 degrees C and 20 degrees C and protected from light. The 1% and 5% eye drops were prepared under aseptic conditions by dissolving Biotum (ceftazidimum), dry injection formulation, in citrate buffer of pH 6.10-6.24. The viscosity of the eye drops was increased with polyvinyl alcohol, phenylmercuric borate combined with 2-phenylethanol was used to preserve the eye drops. The eye drops were stored for 30 days in sterile glass bottles at the temperature of 4 degrees C and 20 degrees C and protected from light. The concentration of ceftazidime and pyridine was analyzed simultaneously with HPLC method every three days; pH, osmotic pressure and viscosity were examined as well as the organoleptic analysis of the eye drops (clarity, color, odor). Storage temperature had the biggest impact on ceftazidime stability in the eye drops. The stability of the drops depended also on ceftazidime concentration in the eye drops, the presence of preservatives and polyvinyl alcohol. The time of 10% ceftazidime degradation in buffered 1% and 5% eye drops, stored at the temperature of 4 degrees C, was from 27 to 18 days in 1% eye drops and from 21 to 12 days in 5% eye drops, depending on their composition. In the eye drops which were stored at the temperature of 20 degrees C 10% ceftazidime degradation occurred on the 3rd day of storage in all 1% and 5% formulary versions.

  19. Stability-indicating RP-HPLC method for the simultaneous determination of escitalopram oxalate and clonazepam.

    Science.gov (United States)

    Kakde, Rajendra B; Satone, Dinesh D; Gadapayale, Kamalesh K; Kakde, Megha G

    2013-07-01

    The objective of the current study was to develop a validated, specific stability-indicating reversed-phase liquid chromatographic (LC) method for the quantitative determination of escitalopram oxalate and clonazepam and their related substances in bulk drugs and pharmaceutical dosage forms in the presence of degradation products. Forced degradation studies were performed on the pure drugs of escitalopram oxalate and clonazepam, as per the stress conditions prescribed by the International Conference on Harmonization (ICH) using acid, base, oxidation, thermal stress and photolytic degradation to show the stability-indicating power of the method. Significant degradation was observed during acid and alkaline hydrolysis and no degradation was observed in other stress conditions. The chromatographic method was optimized using the samples generated from forced degradation studies. Good resolution between the peaks corresponded to the active pharmaceutical ingredients, escitalopram oxalate and clonazepam, and degradation products from the analyte were achieved on an ODS Hypersil C18 column (250 × 4.6 mm) using a mobile phase consisting of a mixture of acetonitrile-50 mM phosphate buffer + 10 mM triethylamine (70:30, v/v). The detection was conducted at 268 nm. The limit of detection and the limit of quantitation for escitalopram oxalate and clonazepam were established. The stress test solutions were assayed against the qualified working standards of escitalopram oxalate and clonazepam, which indicated that the developed LC method was stability-indicating. Validation of the developed LC method was conducted as per ICH requirements. The developed LC method was found to be suitable to check the quality of bulk samples of escitalopram oxalate and clonazepam.

  20. Influence of heat stress, sex and genetic groups on reference genes stability in muscle tissue of chicken.

    Science.gov (United States)

    Cedraz de Oliveira, Haniel; Pinto Garcia, Antonio Amandio; Gonzaga Gromboni, Juliana Gracielle; Vasconcelos Farias Filho, Ronaldo; Souza do Nascimento, Carlos; Arias Wenceslau, Amauri

    2017-01-01

    Quantitative RT-PCR is an important technique for assessing gene expression. However, a proper normalization of reference genes prior to expression analyses of target genes is necessary. The best normalizer is that gene which remains stable in all samples from different treatments. The aim of this study was to identify stable reference genes for normalization of target genes in muscle tissue from three genetically divergent chickens groups (Peloco, Cobb 500® and Caneluda) under environmental (heat stress and comfort) and sex influence. Expressions of ten reference genes were tested for stability in breast muscular tissue (Pectoralis major muscle). Samples were obtained from 36 males and females of two backyard breeds (Caneluda and Peloco) and one commercial line (Cobb 500®) under two environments. The heat stress and comfort temperature were 39 and 23°C, respectively. Animals were housed in the Animal Science Department at Universidade Estadual do Sudoeste da Bahia. We analyzed the expression data by four statistical tools (SLqPCR, NormFinder, Bestkeeper and Comparative CT). According to these tools, genes stability varied according to sex, genetic group and environment, however, some genes remained stable in all analyzes. There was no difference between the most stable genes for sex effect, being MRPS27 more stable for both males and females. In general, MRPS27 was the most stable gene. Within the three genetic groups, the most stable genes were RPL5, HMBS and EEF1 to Cobb 500®, Peloco and Caneluda, respectively. Within the environment, the most stable gene under comfort and heat stress conditions was HMBS and MRPS27, respectively. BestKeeper and Comparative Ct were less correlated (28%) and SLqPCR and NormFinder were the most correlated (98%). MRPS27, RPL5 and MRPS30 genes were considered stable according the overall ranking and can be used as normalizer of relative expression of target genes in muscle tissue of chickens under heat stress.

  1. Stability-indicating HPLC method for the determination of the stability of oxytocin parenteral solutions prepared in polyolefin bags.

    Science.gov (United States)

    Kaushal, G; Sayre, B E; Prettyman, T

    2012-02-01

    Oxytocin is very commonly used in clinical settings and is a nonapeptide hormone that stimulates the contraction of uterine smooth muscles. In this study the stability of extemporaneously compounded oxytocin solutions was investigated in polyolefin bags. The sterile preparations of oxytocin were compounded to the strength of 0.02 U/mL in accordance with United States Pharmacopeia (USP) standards. In order to carry out the stability testing of these parenteral products, the solutions were stored under three different temperature conditions of -20°C (frozen), 2-6°C (refrigerated), and 22-25°C (room temperature). Three solutions from each temperature were withdrawn and were assessed for stability on days 0, 7, 15, 21, and 30 as per the USP guidelines. The assay of oxytocin was examined by an HPLC method at each time point. No precipitation, cloudiness or color change was observed during this study at all temperatures. The assay content by HPLC revealed that oxytocin retains greater than at least 90% of the initial concentrations for 21 days. There was no significant change in pH and absorbance values for 21 days under all the conditions of storage. Oxytocin parenteral solutions in the final concentration of 0.02 U/mL and diluted in normal saline are stable for at least 30 days under frozen and refrigerated conditions for 30 days. At the room temperature, the oxytocin solutions were stable for at least 21 days. The stability analysis results show that the shelf-life of 21 days observed in this study was far better than their recommended expiration dates.

  2. Stability of cefuroxime in 1% and 5% buffered eye drops determined with HPLC method.

    Science.gov (United States)

    Kodym, Anna; Wiśniewski, Andrzej; Knioła, Dawid; Olejniczak, Monika

    2011-01-01

    The aim of the studies was to develop formulary technologies of 1% and 5% eye drops containing cefuroxime with stability of at least 10-12 days. The stability was defined as the time required to reach the cutoff value of 10% degradation of cefuroxime in the drops, as determined using an HPLC assay. The drops should have such properties as optical clarity, pH in the range of 3.5 to 8.5 and osmotic pressure not lower than 280 mOsm/L. Additionally, drops of enhanced viscosity within the range 7-9 mPaxs were developed. Drops (1% and 5%) were prepared under aseptic conditions by dissolving Biofuroksym (Cefuroxime natricum) IBA Bioton--the form of the drug for dry injections--in citrate buffer of pH 6.05-6.28. Polyvinyl alcohol was used to increase the viscosity of the drops. Phenylmercuric borate at the final concentration of 0.001% was used together with beta-phenylethyl alcohol at the final concentration of 0.4% to preserve the drops. The drops were stored for 30 days in tightly closed glass bottles at the temperature of 4 degrees C and 20 degrees C, protected from light. As the course of the infection may differ in intensity, location and the area of the infection in the eye, the composition of the drops was developed at two concentrations (1% and 5%), and five formulary versions for each concentration were prepared. The concentration of cefuroxime in the drops was determined every three days using HPLC. Such properties as pH, osmotic pressure and viscosity were also examined. Additionally, organoleptic analysis (clarity, color, odor) was performed. Physical and chemical properties of all formulations of 1% and 5% drops containing cefuroxime prepared in citrate buffer of pH 6.05-6.28 met the standards set in the objective of the work. The stability of cefuroxime in buffered drops stored at the temperature of 4 degrees C, determined with HPLC as the time of 10% degradation of cefuroxime, was 15 days for 1% and 5% drops. In the drops, which were buffered and of

  3. A genomewide RNAi screen for genes that affect the stability, distribution and function of P granules in Caenorhabditis elegans.

    Science.gov (United States)

    Updike, Dustin L; Strome, Susan

    2009-12-01

    P granules are non-membrane-bound organelles found in the germ-line cytoplasm throughout Caenorhabditis elegans development. Like their "germ granule" counterparts in other animals, P granules are thought to act as determinants of the identity and special properties of germ cells, properties that include the unique ability to give rise to all tissues of future generations of an organism. Therefore, understanding how P granules work is critical to understanding how cellular immortality and totipotency are retained, gained, and lost. Here we report on a genomewide RNAi screen in C. elegans, which identified 173 genes that affect the stability, localization, and function of P granules. Many of these genes fall into specific classes with shared P-granule phenotypes, allowing us to better understand how cellular processes such as protein degradation, translation, splicing, nuclear transport, and mRNA homeostasis converge on P-granule assembly and function. One of the more striking phenotypes is caused by the depletion of CSR-1, an Argonaute associated with an endogenous siRNA pathway that functions in the germ line. We show that CSR-1 and two other endo-siRNA pathway members, the RNA-dependent RNA polymerase EGO-1 and the helicase DRH-3, act to antagonize RNA and P-granule accumulation in the germ line. Our findings strengthen the emerging view that germ granules are involved in numerous aspects of RNA metabolism, including an endo-siRNA pathway in germ cells.

  4. Elongation Factor-Tu (EF-Tu) proteins structural stability and bioinformatics in ancestral gene reconstruction

    Science.gov (United States)

    Dehipawala, Sunil; Nguyen, A.; Tremberger, G.; Cheung, E.; Schneider, P.; Lieberman, D.; Holden, T.; Cheung, T.

    2013-09-01

    A paleo-experimental evolution report on elongation factor EF-Tu structural stability results has provided an opportunity to rewind the tape of life using the ancestral protein sequence reconstruction modeling approach; consistent with the book of life dogma in current biology and being an important component in the astrobiology community. Fractal dimension via the Higuchi fractal method and Shannon entropy of the DNA sequence classification could be used in a diagram that serves as a simple summary. Results from biomedical gene research provide examples on the diagram methodology. Comparisons between biomedical genes such as EEF2 (elongation factor 2 human, mouse, etc), WDR85 in epigenetics, HAR1 in human specificity, DLG1 in cognitive skill, and HLA-C in mosquito bite immunology with EF Tu DNA sequences have accounted for the reported circular dichroism thermo-stability data systematically; the results also infer a relatively less volatility geologic time period from 2 to 3 Gyr from adaptation viewpoint. Comparison to Thermotoga maritima MSB8 and Psychrobacter shows that Thermus thermophilus HB8 EF-Tu calibration sequence could be an outlier, consistent with free energy calculation by NUPACK. Diagram methodology allows computer simulation studies and HAR1 shows about 0.5% probability from chimp to human in terms of diagram location, and SNP simulation results such as amoebic meningoencephalitis NAF1 suggest correlation. Extensions to the studies of the translation and transcription elongation factor sequences in Megavirus Chiliensis, Megavirus Lba and Pandoravirus show that the studied Pandoravirus sequence could be an outlier with the highest fractal dimension and lowest entropy, as compared to chicken as a deviant in the DNMT3A DNA methylation gene sequences from zebrafish to human and to the less than one percent probability in computer simulation using the HAR1 0.5% probability as reference. The diagram methodology would be useful in ancestral gene

  5. Simultaneous Determination and Stability Studies on Diminazene Diaceturate and Phenazone Using Developed Derivative Spectrophotometric Method

    Science.gov (United States)

    Ahmed Gadkariem, Elrasheed; Awadalla Mohamed, Magdi

    2017-01-01

    This work presents UV first derivative spectrophotometry as a precise, accurate, and feasible method for simultaneous determination of diminazene diaceturate and phenazone in bulk and dosage forms. The absorbance values of diminazene diaceturate and phenazone aqueous mixture were obtained at 398 nm and 273 nm, respectively. The developed method was proved to be linear over the concentration ranges (2–10) μg/mL and (2.496–12.48) μg/mL for diminazene diaceturate and phenazone, respectively, with good correlation coefficients (not less than 0.997). The detection and quantitation limits were found to be (LOD = 0.63 and 0.48 μg/mL; LOQ = 1.92 and 1.47 μg/mL, resp.). The developed method was employed for stability studies of both drugs under different stress conditions. Diminazene diaceturate was prone to degrade at acidic pH via first-order kinetics. The degradation process was found to be temperature dependent with an activation energy of 7.48 kcal/mole. Photo-stability was also investigated for this drug.

  6. Replicative Stress and the FHIT Gene: Roles in Tumor Suppression, Genome Stability and Prevention of Carcinogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Karras, Jenna R.; Paisie, Carolyn A.; Huebner, Kay, E-mail: kay.huebner@osumc.edu [Department of Molecular Virology, Immunology and Medical Genetics, The Ohio State University Wexner Medical Center, Columbus, OH 43210 (United States)

    2014-06-04

    The fragile FHIT gene, encompassing the chromosomal fragile site FRA3B, is an early target of DNA damage in precancerous cells. While vulnerable to DNA damage itself, FHIT protein expression is essential to protect from DNA damage-induced cancer initiation and progression by modulating genome stability, oxidative stress and levels of accumulating DNA damage. Thus, FHIT, whose expression is lost or reduced in many human cancers, is a tumor suppressor and genome caretaker whose loss initiates genome instability in preneoplastic lesions. Ongoing studies are seeking more detailed understanding of the role of FHIT in the cellular response to oxidative damage. This review discusses the relationship between FHIT, reactive oxygen species production, and DNA damage in the context of cancer initiation and progression.

  7. Independent evolutionary origin of fem paralogous genes and complementary sex determination in hymenopteran insects.

    Science.gov (United States)

    Koch, Vasco; Nissen, Inga; Schmitt, Björn D; Beye, Martin

    2014-01-01

    The primary signal of sex determination in the honeybee, the complementary sex determiner (csd) gene, evolved from a gene duplication event from an ancestral copy of the fem gene. Recently, other paralogs of the fem gene have been identified in several ant and bumblebee genomes. This discovery and the close phylogenetic relationship of the paralogous gene sequences led to the hypothesis of a single ancestry of the csd genetic system of complementary sex determination in the Hymenopteran insects, in which the fem and csd gene copies evolved as a unit in concert with the mutual transfers of sequences (concerted evolution). Here, we show that the paralogous gene copies evolved repeatedly through independent gene duplication events in the honeybee, bumblebee, and ant lineage. We detected no sequence tracts that would indicate a DNA transfer between the fem and the fem1/csd genes between different ant and bee species. Instead, we found tracts of duplication events in other genomic locations, suggesting that gene duplication was a frequent event in the evolution of these genes. These and other evidences suggest that the fem1/csd gene originated repeatedly through gene duplications in the bumblebee, honeybee, and ant lineages in the last 100 million years. Signatures of concerted evolution were not detectable, implicating that the gene tree based on neutral synonymous sites represents the phylogenetic relationships and origins of the fem and fem1/csd genes. Our results further imply that the fem1 and csd gene in bumblebees, honeybees, and ants are not orthologs, because they originated independently from the fem gene. Hence, the widely shared and conserved complementary sex determination mechanism in Hymenopteran insects is controlled by different genes and molecular processes. These findings highlight the limits of comparative genomics and emphasize the requirement to study gene functions in different species and major hymenopteran lineages.

  8. A pipeline to determine RT-QPCR control genes for evolutionary studies: application to primate gene expression across multiple tissues.

    Directory of Open Access Journals (Sweden)

    Olivier Fedrigo

    Full Text Available Because many species-specific phenotypic differences are assumed to be caused by differential regulation of gene expression, many recent investigations have focused on measuring transcript abundance. Despite the availability of high-throughput platforms, quantitative real-time polymerase chain reaction (RT-QPCR is often the method of choice because of its low cost and wider dynamic range. However, the accuracy of this technique heavily relies on the use of multiple valid control genes for normalization. We created a pipeline for choosing genes potentially useful as RT-QPCR control genes for measuring expression between human and chimpanzee samples across multiple tissues, using published microarrays and a measure of tissue-specificity. We identified 13 genes from the pipeline and from commonly used control genes: ACTB, USP49, ARGHGEF2, GSK3A, TBP, SDHA, EIF2B2, GPDH, YWHAZ, HPTR1, RPL13A, HMBS, and EEF2. We then tested these candidate genes and validated their expression stability across species. We established the rank order of the most preferable set of genes for single and combined tissues. Our results suggest that for at least three tissues (cerebral cortex, liver, and skeletal muscle, EIF2B2, EEF2, HMBS, and SDHA are useful genes for normalizing human and chimpanzee expression using RT-QPCR. Interestingly, other commonly used control genes, including TBP, GAPDH, and, especially ACTB do not perform as well. This pipeline could be easily adapted to other species for which expression data exist, providing taxonomically appropriate control genes for comparisons of gene expression among species.

  9. Stabilization of gene expression and cell morphology after explant recycling during fin explant culture in goldfish

    Energy Technology Data Exchange (ETDEWEB)

    Chenais, Nathalie; Lareyre, Jean-Jacques; Le Bail, Pierre-Yves; Labbe, Catherine, E-mail: catherine.labbe@rennes.inra.fr

    2015-07-01

    The development of fin primary cell cultures for in vitro cellular and physiological studies is hampered by slow cell outgrowth, low proliferation rate, poor viability, and sparse cell characterization. Here, we investigated whether the recycling of fresh explants after a first conventional culture could improve physiological stability and sustainability of the culture. The recycled explants were able to give a supplementary cell culture showing faster outgrowth, cleaner cell layers and higher net cell production. The cells exhibited a highly stabilized profile for marker gene expression including a low cytokeratin 49 (epithelial marker) and a high collagen 1a1 (mesenchymal marker) expression. Added to the cell spindle-shaped morphology, motility behavior, and actin organization, this suggests that the cells bore stable mesenchymal characteristics. This contrast with the time-evolving expression pattern observed in the control fresh explants during the first 2 weeks of culture: a sharp decrease in cytokeratin 49 expression was concomitant with a gradual increase in col1a1. We surmise that such loss of epithelial features for the benefit of mesenchymal ones was triggered by an epithelial to mesenchymal transition (EMT) process or by way of a progressive population replacement process. Overall, our findings provide a comprehensive characterization of this new primary culture model bearing mesenchymal features and whose stability over culture time makes those cells good candidates for cell reprogramming prior to nuclear transfer, in a context of fish genome preservation. - Highlights: • Recycled fin explants outgrow cells bearing stable mesenchymal traits. • Cell production and quality is enhanced in the recycled explant culture system. • Fresh fin primary culture is highly variable and loose epithelial traits over time.

  10. The Or Gene Enhances Carotenoid Accumulation and Stability During Post-Harvest Storage of Potato Tubers

    Institute of Scientific and Technical Information of China (English)

    Li Li; Mark Failla; Theodore W.Thannhauser; Yong Yang; Qiang Xu; Katherine Owsiany; Ralf Welsch; Chureeporn Chitchumroonchokchai; Shan Lu; Joyce Van Eck; Xiu-Xin Deng

    2012-01-01

    Provitamin A carotenoids in staple crops are not very stable during storage and their loss compromises nutritional quality.To elucidate the fundamental mechanisms underlying carotenoid accumulation and stability,we investigated transgenic potato tubers that expressed the cauliflower Orange (Or) gene.We found that the Or transgene not only promoted retention of β-carotene level,but also continuously stimulated its accumulation during 5 months of cold storage.In contrast,no increased levels of carotenoids were observed in the tubers of vector-only controls or a yellowflesh variety during the same period of storage.The increased carotenoid accumulation was found to be associated with the formation of lipoprotein-carotenoid sequestering structures,as well as with the enhanced abundance of phytoene synthase,a key enzyme in the carotenoid biosynthetic pathway.Furthermore,the provitamin A carotenoids stored were shown to be stable during simulated digestion and accessible for uptake by human intestinal absorptive cells.Proteomic analysis identified three major functional groups of proteins (i.e.heat shock proteins,glutathione-S-transferases,and carbohydrate metabolic proteins) that are potentially important in the Or-regulated carotenoid accumulation.Our results show that regulation of carotenoid sequestration capacity is an important mechanism by which carotenoid stability is regulated.Our findings suggest that induction of a proper sink structure formation in staple crops may provide the crops with a unique ability to promote and/or stabilize provitamin A accumulation during plant growth and post-harvest storage.

  11. Stability-indicating comparative methods using mekc and lc for determination of olmesartan medoxomil

    Directory of Open Access Journals (Sweden)

    Lisiane Bajerski

    2013-01-01

    Full Text Available A stability-indicating method using MEKC was validated for the analysis of olmesartan medoxomil in tablets. Successful separation was achieved using a fused silica capillary (40 cm x 50 µm i.d.; background electrolyte consisted of a combination of 10 mmol L-1 borate buffer and 5 mmol L-1 anionic detergent sodium dodecyl sulfate (95:5; v/v pH 6.5; hydrodynamic mode at 50 mBar for 5 s; 25 kV separation voltage at 25 ºC; and column temperature 25 ºC with detection at 257 nm. The proposed method, validated following ICH guidelines, was applied to the determination of this antihypertensive with good results compared with an LC method.

  12. Stability-indicating UPLC method for determining related substances and degradants in dronedarone.

    Science.gov (United States)

    Pydimarry, Surya Prakash Rao; Cholleti, Vijay Kumar; Vangala, Ranga Reddy

    2014-08-01

    A simple, sensitive and reproducible method was developed on ultra-performance liquid chromatography coupled with photodiode array detection for the quantitative determination of dronedarone hydrochloride (DRO) in drug substance and pharmaceutical dosage forms. The method is applicable for the quantification of related substances and assays of drug substances. Chromatographic separation was achieved on Acquity UPLC BEH C8 100 mm, 2.1 mm and 1.7 µm columns, using gradient elution within a short run time of 10.0 min. The eluted compounds were monitored at 288 nm, the flow rate was 0.5 mL/min and the column oven temperature was maintained at 40°C. The resolution of DRO and 11 impurities (potentials and by-products) was greater than 2.0 for all pairs of components. The high correlation coefficient value (>0.9995) indicates the clear correlations between the concentrations of investigated compound and their peak areas within the test ranges. The repeatability and intermediate precision, expressed by the relative standard deviation, were less than 2.5%. The accuracy and validity of the method were further ascertained by performing recovery studies via a spike method. The accuracy of the method, expressed as relative error, was satisfactory. No interference was observed from concomitant substances normally added to the tablets. DRO was subjected to the stress conditions of oxidative, acid, base, hydrolytic, thermal and photolytic degradation. DRO was found to degrade significantly in acid and base stress conditions and to remain stable in thermal, photolytic degradation, oxidative and hydrolytic conditions. The degradation products were well resolved from primary peak and its impurities, proving that the method is stability indicating. The developed method was validated as per International Conference on Harmonization guidelines with respect to specificity, limit of detection, limit of quantification, linearity, accuracy, precision, solution stability and robustness

  13. Stability-indicating HPTLC determination of ambroxol hydrochloride in bulk drug and pharmaceutical dosage form.

    Science.gov (United States)

    Jain, P S

    2010-01-01

    A simple, selective, precise, and stability-indicating high-performance thin-layer chromatographic (HPTLC) method for the analysis of ambroxol hydrochloride both as a bulk drug and in formulations was developed and validated. The method employed HPTLC aluminium plates precoated with silica gel 60F-254 as the stationary phase. The solvent system consisted of methanol-triethylamine (4:6 v/v). The system was found to give a compact spot for ambroxol hydrochloride (R(f) value of 0.53 +/- 0.02). Densitometric analysis of ambroxol hydrochloride was carried out in the absorbance mode at 254 nm. The linear regression analysis data for the calibration plots showed good linear relationship with r(2) = 0.9966 +/- 0.0013 with respect to peak area in the concentration range 100-1000 ng/spot. The mean value +/- standard deviation of slope and intercept were 164.85 +/- 0.72 and 1168.3 +/- 8.26 with respect to peak area. The method was validated for precision, recovery, and robustness. The limits of detection and quantitation were 10 and 30 ng/spot, respectively. Ambroxol hydrochloride was subjected to oxidation and thermal degradation. The drug undergoes degradation under oxidation and heat conditions. This indicates that the drug is susceptible to oxidation and heat. Statistical analysis proves that the method is repeatable, selective, and accurate for the estimation of said drug. Stability indicating of new chemical entities is an important part for the drug development of ambroxol hydrochloride and for its estimation in plasma and other biological fluids; the novel Statistical analysis proves that the method is repeatable and selective for the analysis of ambroxol hydrochloride as bulk drug and in pharmaceutical formulations. The proposed developed HPTLC method can be applied for identification and quantitative determination of ambroxol hydrochloride in bulk drug and dosage forms. This work is to determine the purity of the drug available from the various sources by detecting

  14. A validated stability-indicating HPLC method for determination of varenicline in its bulk and tablets

    Directory of Open Access Journals (Sweden)

    Kassem Mohamed G

    2011-06-01

    Full Text Available Abstract A simple, sensitive and accurate stability-indicating HPLC method has been developed and validated for determination of varenicline (VRC in its bulk form and pharmaceutical tablets. Chromatographic separation was achieved on a Zorbax Eclipse XDB-C8 column (150 mm × 4.6 mm i.d., particle size 5 μm, maintained at ambient temperature by a mobile phase consisted of acetonitrile and 50 mM potassium dihydrogen phosphate buffer (10:90, v/v with apparent pH of 3.5 ± 0.1 and a flow rate of 1.0 ml/min. The detection wavelength was set at 235 nm. VRC was subjected to different accelerated stress conditions. The degradation products, when any, were well resolved from the pure drug with significantly different retention time values. The method was linear (r = 0.9998 at a concentration range of 2 - 14 μg/ml. The limit of detection and limit of quantitation were 0.38 and 1.11 μg/ml, respectively. The intra- and inter-assay precisions were satisfactory; the relative standard deviations did not exceed 2%. The accuracy of the method was proved; the mean recovery of VRC was 100.10 ± 1.08%. The proposed method has high throughput as the analysis involved short run-time (~ 6 min. The method met the ICH/FDA regulatory requirements. The proposed method was successfully applied for the determination of VRC in bulk and tablets with acceptable accuracy and precisions; the label claim percentages were 99.65 ± 0.32%. The results demonstrated that the method would have a great value when applied in quality control and stability studies for VRC.

  15. Stress determination and geomechanical stability analysis of an oil well of Iran

    Indian Academy of Sciences (India)

    Ayub Elyasi; Kamran Goshtasbi; Omid Saeidi; Seyed Rahman Torabi

    2014-02-01

    In this paper, a numerical model using three-dimensional finite difference code FLAC3D is proposed for analysing the stability of an oil well drilled in four formations. Normalized Yielded Zone Area (NYZA, i.e., the ratio of surrounding yielded cross-sectional area to initial area of well) has been determined for different mud pressures. In each formation, by interpolating of obtained NYZA equal to one, the optimized mud pressure was determined using MATLAB software. Practical data including geomechanical parameters along with drilling data from one of Iranian oilfields, Mansouri-54 well have been utilized in this analysis. in situ stress was determined using stress polygon method and conducting hydraulic fracturing data in the field. Analytical solution using the Mogi–Coulomb and the Hoek–Brown failure criteria has been carried out and results are compared with the presented model. The results demonstrated that the NYZA and Hoek–Brown criteria might underestimate and overestimate the drilling mud pressure, respectively, and should be used cautiously. In the inclined section of the well, plastic zone showed more extension in the lower part than upper part of the well because of the high stress concentration.

  16. Smart stability-indicating spectrophotometric methods for determination of binary mixtures without prior separation.

    Science.gov (United States)

    El-Bardicy, Mohammad G; Lotfy, Hayam M; El-Sayed, Mohammad A; El-Tarras, Mohammad F

    2008-01-01

    Ratio subtraction and isosbestic point methods are 2 innovating spectrophotometric methods used to determine vincamine in the presence of its acid degradation product and a mixture of cinnarizine (CN) and nicergoline (NIC). Linear correlations were obtained in the concentration range from 8-40 microg/mL for vincamine (I), 6-22 microg/mL for CN (II), and 6-36 microg/mL for NIC (III), with mean accuracies 99.72 +/- 0.917% for I, 99.91 +/- 0.703% for II, and 99.58 +/- 0.847 and 99.83 +/- 1.039% for III. The ratio subtraction method was utilized for the analysis of laboratory-prepared mixtures containing different ratios of vincamine and its degradation product, and it was valid in the presence of up to 80% degradation product. CN and NIC in synthetic mixtures were analyzed by the 2 proposed methods with the total content of the mixture determined at their respective isosbestic points of 270.2 and 235.8 nm, and the content of CN was determined by the ratio subtraction method. The proposed method was validated and found to be suitable as a stability-indicating assay method for vincamine in pharmaceutical formulations. The standard addition technique was applied to validate the results and to ensure the specificity of the proposed methods.

  17. Stability of and Associations Between Social-Cognitive Determinants Over Time

    Directory of Open Access Journals (Sweden)

    Barbara Sassen

    2015-08-01

    Full Text Available This study reports on the stability of social-cognitive determinants, and on associations between social-cognitive determinants to show insight in the theory of planned behavior (TPB. In all, 278 health professionals who encourage patients to become physically active completed online TPB-based surveys at baseline (Time 1 [T1] and six months later (Time 2 [T2]. No intervention took place. No differences were found for all social-cognitive determinants measured at T1 compared with T2 (6 months later, except for intention (t test = 5.18, p < .001. Structural equation modeling—χ2(5, N = 278 = 2.35, p = .80, root mean square error of approximation = 0.00—showed that behavior T1 and attitude T1 predicted intention T1 (R2 = .57, p = <.001; that behavior T1 and barriers T1 predicted behavior T2 (R2 = .38, p = <.001; and that behavior T2, intention T1, and attitude T1 predicted intention T2 (R2 =.60, p = <.001. Intention T1 did not predict behavior T2. The model achieved a good fit with the data. Findings revealed that social-cognitive determinants remained stable over time, with intention being instable. Without intervention, the intention decreased, while the social-cognitive determinants (attitudes, perceived behavioral control, and subjective norms for intention and the corresponding behavior remained unchanged. For intervention development it seems important to value health professionals’ previous or past encouraging behavior (T1, this to change intention and behavior, or to initiate new behavior. Behavior T1 showed a predictive variable and predicted attitude T1, intention T1, barriers T1, and behavior T2. Barriers that obstruct health professionals’ encouraging behavior are encountered, and barriers influence attitudes T1 and the behavior T2 to encourage patients.

  18. Amphiphilic, low molecular weight poly(ethylene imine) derivatives with enhanced stability for efficient pulmonary gene delivery.

    Science.gov (United States)

    Roesler, Susanne; Koch, Felix P V; Schmehl, Thomas; Weissmann, Norbert; Seeger, Werner; Gessler, Tobias; Kissel, Thomas

    2011-02-01

    Poly(ethylene imine) (PEI) is a widely used transfection reagent for mammalian cells, but in vivo application of PEI 25 kDa is restricted by its toxicity. Low molecular weight (LMW) PEI is less toxic, but also less efficient than its high molecular weight equivalent, and prone to aggregation. A set of polymers was synthesized by coupling poly(ethylene glycol) (PEG) that contained either C(16/18) -chains (Cx-EO) or butyl-poly(propylene oxide)-co-poly(ethylene glycol) (ButPP). Critical micelle concentration (CMC) was determined for copolymers. Polyplexes were characterized by DNA binding ability, polyplex size and aggregation, hemolysis, and cytotoxicity. Transfection efficiency was tested in vitro and in vivo in mouse lungs. Copolymers formed stable complexes with DNA, and showed enhanced complex stability in isotonic solution for at least 1 h. CMC was determined for Cx-EO-PEI 4.7 and 8.3 at 0.0019 and 0.0037 mM, respectively; membrane activity in a haemolysis assay was demonstrated for ButPP-PEI: both factors possibly enhance endosomal escape effect after PEGylation. IC(50) values of all synthesized polymers were in the range 6-33 ng/ml. Transfection efficiency of unmodified LMW-PEIs was equivalent or better than that of PEI 25 as a result of aggregation in vitro. Cells treated with polyplexes of amphiphilic polymers showed reduced transfection compared to PEI 25. After instillation in mouse lungs, highest transfection efficiency was demonstrated with Cx-EO copolymer of lowest molecular weight PEI. A new set of polymers with low toxicity and high stability was synthesized, which contains promising candidates for pulmonary gene transfer, as documented by in vivo experiments in mice. Copyright © 2011 John Wiley & Sons, Ltd.

  19. Candidate qRT-PCR reference genes for barley that demonstrate better stability than traditional housekeeping genes

    Science.gov (United States)

    Gene transcript expression analysis is a useful tool for correlating gene activity with plant phenotype. For these studies, an appropriate reference gene is necessary to quantify the expression of target genes. Classic housekeeping genes have often been used for this purpose, but may not be consis...

  20. Determination of the stability limit of a thermoacoustic engine by means of finite elements

    NARCIS (Netherlands)

    de Jong, Anne; Wijnant, Ysbrand H.; de Boer, Andries

    2013-01-01

    A finite element model is presented to obtain the stability limit of, as an example, 2D standing wave thermoacoustic engine. The stability limit is the required heating to obtain self-sustained (thermo)acoustic oscillations. The method used to obtain the stability limit is not restricted to the exam

  1. Determination of reference genes for circadian studies in different tissues and mouse strains

    Directory of Open Access Journals (Sweden)

    Kosir Rok

    2010-08-01

    Full Text Available Abstract Background Circadian rhythms have a profound effect on human health. Their disruption can lead to serious pathologies, such as cancer and obesity. Gene expression studies in these pathologies are often studied in different mouse strains by quantitative real time polymerase chain reaction (qPCR. Selection of reference genes is a crucial step of qPCR experiments. Recent studies show that reference gene stability can vary between species and tissues, but none has taken circadian experiments into consideration. Results In the present study the expression of ten candidate reference genes (Actb, Eif2a, Gapdh, Hmbs, Hprt1, Ppib, Rn18s, Rplp0, Tbcc and Utp6c was measured in 131 liver and 97 adrenal gland samples taken from three mouse strains (C57BL/6JOlaHsd, 129Pas plus C57BL/6J and Crem KO on 129Pas plus C57BL/6J background every 4 h in a 24 h period. Expression stability was evaluated by geNorm and NormFinder programs. Differences in ranking of the most stable reference genes were observed both between individual mouse strains as well as between tissues within each mouse strain. We show that selection of reference gene (Actb that is often used for analyses in individual mouse strains leads to errors if used for normalization when different mouse strains are compared. We identified alternative reference genes that are stable in these comparisons. Conclusions Genetic background and circadian time influence the expression stability of reference genes. Differences between mouse strains and tissues should be taken into consideration to avoid false interpretations. We show that the use of a single reference gene can lead to false biological conclusions. This manuscript provides a useful reference point for researchers that search for stable reference genes in the field of circadian biology.

  2. Functional cohesion of gene sets determined by latent semantic indexing of PubMed abstracts.

    Directory of Open Access Journals (Sweden)

    Lijing Xu

    Full Text Available High-throughput genomic technologies enable researchers to identify genes that are co-regulated with respect to specific experimental conditions. Numerous statistical approaches have been developed to identify differentially expressed genes. Because each approach can produce distinct gene sets, it is difficult for biologists to determine which statistical approach yields biologically relevant gene sets and is appropriate for their study. To address this issue, we implemented Latent Semantic Indexing (LSI to determine the functional coherence of gene sets. An LSI model was built using over 1 million Medline abstracts for over 20,000 mouse and human genes annotated in Entrez Gene. The gene-to-gene LSI-derived similarities were used to calculate a literature cohesion p-value (LPv for a given gene set using a Fisher's exact test. We tested this method against genes in more than 6,000 functional pathways annotated in Gene Ontology (GO and found that approximately 75% of gene sets in GO biological process category and 90% of the gene sets in GO molecular function and cellular component categories were functionally cohesive (LPv<0.05. These results indicate that the LPv methodology is both robust and accurate. Application of this method to previously published microarray datasets demonstrated that LPv can be helpful in selecting the appropriate feature extraction methods. To enable real-time calculation of LPv for mouse or human gene sets, we developed a web tool called Gene-set Cohesion Analysis Tool (GCAT. GCAT can complement other gene set enrichment approaches by determining the overall functional cohesion of data sets, taking into account both explicit and implicit gene interactions reported in the biomedical literature.GCAT is freely available at http://binf1.memphis.edu/gcat.

  3. Stability-Indicating RP-HPLC Method for Determination of Tamsulosin HCL in Pharmaceutical Dosage Form.

    Science.gov (United States)

    Kumar, G S; Kumar, B Sai Pavan

    2012-03-01

    A selective, specific and sensitive stability-indicating high-performance liquid chromatographic method was developed and validated for the determination of Tamsulosin in in pharmaceutical dosage forms. Celecoxib was used as Internal Standard (IS). The chromatographic conditions comprised of a reversed-phase Lichrocart / Lichrosphere C18 column (250 × 4.0 mm packed with 5) with mobile phase consisting of a mixture of Acetonitrile: T.D.W. in the ratio (40: 60). Flow rate was 0.8 mL / min. Detection was carried out at 275 nm. The retention time of Tamsulosin HCl and Celecoxib were found to be 1.608 and 2.767min respectively and the linear regression analysis data for the calibration plots showed good linear relationship in the concentration range 1 - 200 g/mL. The value of correlation coefficient, slope and intercept were, 0.9995, 0.7453 and 0.4584, respectively. Tamsulosin HCl was subjected to stress conditions of degradation in aqueous solutions including acidic, alkaline, oxidation, photolysis and thermal degradation. The developed method was validated with regard to linearity, accuracy, precision, selectivity and robustness and the method was found to be precise, accurate, linear and specific. The method was employed successfully for identification and determination of Tamsulosin in pharmaceutical preparations.

  4. Patterns of proliferative activity in the colonic crypt determine crypt stability and rates of somatic evolution.

    Directory of Open Access Journals (Sweden)

    Rui Zhao

    Full Text Available Epithelial cells in the colon are arranged in cylindrical structures called crypts in which cellular proliferation and migration are tightly regulated. We hypothesized that the proliferation patterns of cells may determine the stability of crypts as well as the rates of somatic evolution towards colorectal tumorigenesis. Here, we propose a linear process model of colonic epithelial cells that explicitly takes into account the proliferation kinetics of cells as a function of cell position within the crypt. Our results indicate that proliferation kinetics has significant influence on the speed of cell movement, kinetics of mutation propagation, and sensitivity of the system to selective effects of mutated cells. We found that, of all proliferation curves tested, those with mitotic activities concentrated near the stem cell, including the actual proliferation kinetics determined in in vivo labeling experiments, have a greater ability of delaying the rate of mutation accumulation in colonic stem cells compared to hypothetical proliferation curves with mitotic activities focused near the top of the crypt column. Our model can be used to investigate the dynamics of proliferation and mutation accumulation in spatially arranged tissues.

  5. A stability-indicating high performance liquid chromatography method to determine apocynin in nanoparticles

    Directory of Open Access Journals (Sweden)

    Juliana Kovalczuk de Oliveira

    2017-04-01

    Full Text Available In this study, we developed and validated a fast, specific, sensitive, precise and stability-indicating high performance liquid chromatography (HPLC method to determine the drug apocynin in bovine serum albumin (BSA nanoparticles. Chromatographic analyses were performed on an RP C18 column and using a photodiode array detector at a wavelength of 276 nm. Mobile phase consisted of a mixture of acetonitrile and 1% acetic acid (60:40, v/v, and it was eluted isocratically at a flow rate of 0.8 mL/min. The retention time of apocynin chromatographic peak was 1.65 min. The method was linear, precise, accurate and specific in the range of 5–100 μg/mL. The intra- and inter-day precisions presented relative standard deviation (RSD values lower than 2%. The method was robust regarding changes in mobile phase proportion, but not for flow rate. Limits of detection and quantitation were 78 ng/mL and 238 ng/mL, respectively. Apocynin was exposed to acid and alkali hydrolysis, oxidation and visible light. The drug suffered mild degradation under acid and oxidation conditions and great degradation under alkali conditions. Light exposure did not degrade the drug. The method was successfully applied to determine the encapsulation efficiency of apocynin in BSA nanoparticles.

  6. The determinants and stability of money demand in the Republic of Macedonia

    Directory of Open Access Journals (Sweden)

    Jordan Kjosevski

    2013-06-01

    Full Text Available The goal of this paper is to examine the long and short-run determinants, and stability of money demand (M1 in the Republic of Macedonia using monthly data from January 2005 to October 2012. The Johansen cointegration technique and VECM model were used to fi nd the long-run and short-term dynamic relationships in money demand model. Empirical results provide the evidence that exchange rate and interest rate payable on denar time deposits up to one month explains the most variations of money demand in the long-run, while interest rate is signifi cant only in short-run. Long-run money demand function is estimated to indicate slow speed of adjustment of removing the disequilibrium. Our fi nding shows that real money demand M1 in the Republic of Macedonia is stable in the analyzed period. The results obtained in this study suggest that the National Bank should carefully monitor the exchange rate and infl ation as two most important indicators of monetary policy, because these two determinants are the main drivers of demand for money in the short and long term.

  7. Stability-Indicating HPLC Method for the Determination of Cefcapene Pivoxil.

    Science.gov (United States)

    Zalewski, Przemysław; Cielecka-Piontek, Judyta; Garbacki, Piotr; Jelińska, Anna; Karaźniewicz-Łada, Marta

    2013-04-01

    The stability-indicating LC assay method was developed and validated for quantitative determination of cefcapene pivoxil in the presence of degradation products formed during forced degradation studies. An isocratic RP-HPLC method was developed with a Lichrospher RP-18 (250 mm × 4.6 mm, 5 μm) column and the mobile phase composed of 45 volumes of acetonitrile and 55 volumes of mixture composed of citric acid 10 mmol L(-1) and potassium chloride 18 mmol L(-1). The flow rate of the mobile phase was 1 mL min(-1). Detection wavelength was 270 nm and temperature was 30 °C. Cefcapene pivoxil, similar to other cephalosporins, was subjected to stress conditions of degradation in aqueous solutions including hydrolysis, oxidation, and thermal degradation. The method was validated with regard to linearity, accuracy, precision, selectivity, and robustness. The method was applied successfully for the determination of cefcapene pivoxil during kinetic studies in aqueous solutions (pH and thermal degradation) and in solid state (oxidative, thermal, and radiolytic degradation).

  8. Influences of terrestrial determinants on the stability of surface water quality response to climate drivers

    Science.gov (United States)

    Jiang, Jiping; Kahn, Afed; Wang, Peng

    2017-04-01

    Water quality mainly depends upon the terrestrial characteristics of landscape. The current study was conducted to unveil relationships between climate elasticity of water quality (CEWQ) and terrestrial determinants based on 13 monitoring sites from three typical watersheds of Yukon, Mekong and Murray. Anthropogenic biomes and surficial geological composition was computed at basin scale. It was found that the response pattern of (T, TN-UF) and (T, water temperature) are exclusively characterized by temperature. Temperature elasticity is variable in space as compared to precipitation elasticity. The results implied that anthropogenic biomes are stronger determinants as compared to surficial geology when analyzing their relationships with CEWQ. Some important association was found between CEWQ and anthropogenic biomes which includes: positive association of dense settlements with (P, NOX-F) and (P, P-F), positive linkage of croplands with (P, NOX-F) and (P, NH4-F), negative relationship of rangelands with (P, NOX-F) and (P, DOC), and negative linkage of rangelands with (T, P-UF) and (T, water temperature). Similarly some important association was found between CEWQ and surficial geology which includes: negative linkage of clay with (P, P-F) and negative relationships of gravel and clay with (T, P-UF). This indicates that dense settlements and croplands are the main factors influencing the stability of CEWQ, and limiting high flow volume during rain will be critical for enhancing water quality.

  9. Sensitive Determination of Sertraline in Commercial Drugs and Its Stability Check in Simulated Gastric Juice.

    Science.gov (United States)

    Koçoğlu, Elif Seda; Bakırdere, Sezgin; Keyf, Seyfullah

    2016-11-01

    A sensitive analytical method was developed for the determination of sertraline in commercial drug samples by using GC-MS. The selected-ion monitoring mode was used at the most sensitive m/z 274 to obtain a lower detection limit. LOD/LOQ values were obtained as 1.6/5.4 ng/mL for sertraline under the optimum conditions. The calibration plot was linear between 5.0 and 2000 ng/mL with the correlation coefficient of 0.9999. The validated method was successfully applied to three different brands of drug samples for both qualitative and quantitative measurement of sertraline. In this experiment, four replicate extractions were performed for each brand, and the results were compared to the values written on the labels of the drug brands. Spiking experiments were also performed to check the effect of the matrixes on the determination, and it was observed that there was no shift in the retention time of the analyte. In addition, simulated gastric juice experiments were performed to check the stability of sertraline in the stomach for 240 min, and it was observed that there was no change in the structure of the analyte.

  10. Stability indicating RP HPLC method for determination of levitiracetam in pharmaceutical formulation

    Directory of Open Access Journals (Sweden)

    M. Krishna Chaitanya Prasad*

    2013-12-01

    Full Text Available The article reports on a development of RP-HPLC method for the quantitative determination of Levetiracetam in tablet dosage forms. The chromatographic separations were performed using Phenomenex C18 (250 mm x 4.6 mm i.d, 5 µm particle size column at 40 ºC temperatures. The optimum mobile phase consisted of methanol, water and acetonitrile in the ratio of 30:10:60. Auto sampler 20 µl was used and kept at 15 ºC temperature. Analysis was done with flow rate of 1.0 ml/min at 212 nm (λ max of Levetiracetam wavelength by using photodiode array (PDA detector. The drug was analyzed for acid, alkaline, oxidative, hydrolytic, photolytic and thermal degradation studies. The standard calibration curve was plotted for the drug and results showed that the drug was linear (r2 = 0.999 in the concentration range between 0.01 – 1.5 µg/ml. The results of stress testing undertaken according to the International Conference on Harmonization (ICH guidelines reveal that the selected method is selective and stability-indicating for determination of levitiracetam in pharmaceutical formualtion.

  11. Noise and Stochasticity in Gene Expression : A Pathogenic Fate Determinant

    NARCIS (Netherlands)

    Jørgensen, Mikkel Girke; Raaphorst, Renske van; Veening, Jan-Willem; Harwood, Colin; Wipat, Anil

    2013-01-01

    Not all cells in a bacterial population exhibit exactly the same phenotype, even though they grow in the same environment and are genetically identical. This phenomenon is known as phenotypic variation. The major source of phenotypic variation is noise or stochasticity in gene expression networks,

  12. Stability indicating LC method for simultaneous determination of irbesartan and hydrochlorothiazide in pharmaceutical preparations.

    Science.gov (United States)

    Rane, V P; Patil, K R; Sangshetti, J N; Yeole, R D; Shinde, D B

    2010-08-01

    A simple and precise stability-indicating liquid chromatography method is developed and validated for the quantitative simultaneous estimation of irbesartan (IRB) and hydrochlorothiazide (HCTZ) in combined pharmaceutical dosage form. A chromatographic separation of the two drugs was achieved with an Ace5 C(18) 25-cm analytical column using buffer-acetonitrile (70:30 v/v). The buffer used in mobile phase contains 50 mM ammonium acetate pH adjusted 5.5 with acetic acid. The instrumental settings are flow rate of 1.5 mL/min, column temperature at 30 degrees C, and detector wavelength of 235 nm using a photodiode array detector. IRB, HCTZ, and their combination drug products were exposed to thermal, photolytic, hydrolytic, and oxidative stress conditions, and the stressed samples were analyzed by the proposed method. Peak homogeneity data of IRB and HCTZ is obtained using photodiode array detector. In the stressed sample chromatograms, it demonstrated the specificity of the assay method for their estimation in presence of degradation products. The described method shows excellent linearity over a range of 10-200 microg/mL for IRB and 5-100 microg/mL for HCTZ. Methylparaben was used as internal standard. The correlation coefficient for IRB and HCTZ are 0.998 and 0.999. The mean recovery values for IRB and HCTZ ranged from 100.45% to 101.25%. The limit of detection for IRB and HCTZ were 0.019 and 0.023 microg/mL, respectively, and the limit of quantification were 0.053 and 0.070 microg/mL, respectively. The proposed method was suitable for quantitative determination and stability study of IRB and HCTZ in pharmaceutical preparations and also can be used in the quality control of bulk manufacturing and pharmaceutical dosage forms.

  13. Determination of optimal dead sea salt content in a cosmetic emulsion using rheology and stability measurements.

    Science.gov (United States)

    Abu-Jdayil, Basim; Mohameed, Hazim A; Bsoul, Abeer

    2008-01-01

    Dead Sea mud and salts are known for their therapeutic and cosmetic properties. The presence of Dead Sea (DS) salts in different types of cosmetics has affected the stability and the flow properties of the finished products. In this study, an attempt was made to find the optimum Dead Sea salt content in a cosmetic emulsion (model of body cream) using both rheology and stability measurements. The rheological properties were tested during a four-month storage period at three different storage temperatures: 8 degrees C, room temperature, and 45 degrees C. In addition to rheological measurements and centrifuge tests, the conductivities of the emulsion samples were also determined. The centrifuge tests showed that the cream samples containing more than 0.25 wt% of DS salt showed phase separation. The addition of DS salt to the cosmetic emulsion led to two maxima in the emulsion viscosity at salt contents of 0.07 wt% and 0.15 wt%. However, the emulsion samples containing 0.15% of DS salt was considered the optimum sample since it contained the maximum amount of salt and exhibited the maximum viscosity at all tested conditions. It was found that the viscosity of the emulsion is increased with storage time and storage temperature. This behavior was accompanied by a decrease in conductivity. This behavior was explained by water evaporation from the emulsion. However, it has been shown that the presence of DS salt in the cosmetic emulsion significantly reduces the rate of water evaporation. The conductivity measurements reflect the rate of water evaporation, and the presence of DS salt reduces the rate of conductivity. Conductivity is observed to decrease with storage time and temperature.

  14. O-GlcNAcylation determines the solubility, filament organization, and stability of keratins 8 and 18.

    Science.gov (United States)

    Srikanth, Budnar; Vaidya, Milind M; Kalraiya, Rajiv D

    2010-10-29

    Keratins 8 and 18 (K8/18) are intermediate filament proteins expressed specifically in simple epithelial tissues. Dynamic equilibrium of these phosphoglycoproteins in the soluble and filament pool is an important determinant of their cellular functions, and it is known to be regulated by site-specific phosphorylation. However, little is known about the role of dynamic O-GlcNAcylation on this keratin pair. Here, by comparing immortalized (Chang) and transformed hepatocyte (HepG2) cell lines, we have demonstrated that O-GlcNAcylation of K8/18 exhibits a positive correlation with their solubility (Nonidet P-40 extractability). Heat stress, which increases K8/18 solubility, resulted in a simultaneous increase in O-GlcNAc on these proteins. Conversely, increasing O-GlcNAc levels were associated with a concurrent increase in their solubility. This was also associated with a notable decrease in total cellular levels of K8/18. Unaltered levels of transcripts and the reduced half-life of K8 and K18 indicated their decreased stability on increasing O-GlcNAcylation. On the contrary, the K18 glycosylation mutant (K18 S29A/S30A/S48A) was notably more stable than the wild type K18 in Chang cells. The K18-O-GlcNAc mutant accumulated as aggregates upon stable expression, which possibly altered endogenous filament architecture. These results strongly indicate the involvement of O-GlcNAc on K8/18 in regulating their solubility and stability, which may have a bearing on the functions of these keratins.

  15. Validated stability-indicating TLC method for the determination of noscapine.

    Science.gov (United States)

    Ashour, Ahmed; Hegazy, Maha Abdel Monem; Moustafa, Azza Aziz; Kelani, Khadiga Omar; Fattah, Laila Elsayed Abdel

    2009-07-01

    A sensitive, selective, precise and stability-indicating thin-layer chromatographic (TLC) method was developed and validated for the analysis of noscapine, both as a bulk drug and in its formulation. The method employed TLC aluminium plates precoated with silica gel 60F-254 as the stationary phase. The solvent system consisted of chloroform-methanol (10:0.5 v/v). Densitometric analysis of noscapine and its degradation products was carried out in the absorbance mode at 254 nm. This system was found to give compact symmetrical spots for noscapine (R(f) value 0.85 +/- 0.04). Noscapine was subjected to acid and alkali hydrolysis, oxidation and photo degradation. The drug undergoes photo degradation and also degrades under acidic and basic conditions. The prepared degradation products were identified and verified through infrared (IR) and mass spectral analyses. The degraded products were also well resolved from the pure drug with significantly different R(f) values and they were quantitatively determined. The method was validated for linearity, precision, robustness, limit of detection (LOD), limit of quantitation (LOQ), specificity and accuracy. Linearity was found to be in the 1.0-10.0 microg, 0.4-3.2 microg, 1.0-9.0 microg and 0.5-5.0 microg/band ranges for noscapine, cotarnine, meconine and opionic acid, respectively. The polynomial regression analysis for the calibration plots showed a good polynomial relationship with r(2) of 0.9998, 9989, 9996 and 0.9997 for noscapine and its three degradation products, cotarnine, meconine and opionic acid, respectively. Statistical analysis proves that the method is repeatable and specific for the estimation of noscapine. As this approach could effectively separate the drug from its degradation products it can be employed as a stability-indicating method in Quality Control laboratories.

  16. Genetic Determinants for Promoter Hypermethylation in the Lungs of Smokers: A Candidate Gene-Based Study

    OpenAIRE

    Leng, Shuguang; Stidley, Christine A.; Liu, Yushi; Edlund, Christopher K.; Willink, Randall P.; Han, Younghun; Landi, Maria Teresa; Thun, Michael; Picchi, Maria A.; Bruse, Shannon E.; Crowell, Richard E.; Van Den Berg, David; Neil E Caporaso; Amos, Christopher I.; Siegfried, Jill M.

    2011-01-01

    The detection of tumor suppressor gene promoter methylation in sputum-derived exfoliated cells predicts early lung cancer. Here we identified genetic determinants for this epigenetic process and examined their biological effects on gene regulation. A two-stage approach involving discovery and replication was employed to assess the association between promoter hypermethylation of a 12-gene panel and common variation in 40 genes involved in carcinogen metabolism, regulation of methylation, and ...

  17. Stability-indicating HPLC Method for Simultaneous Determination of Terbutaline Sulphate, Bromhexine Hydrochloride and Guaifenesin.

    Science.gov (United States)

    Porel, A; Haty, Sanjukta; Kundu, A

    2011-01-01

    The aim of the present study was the development and subsequent validation of a simple, precise and stability-indicating reversed phase HPLC method for the simultaneous determination of guaifenesin, terbutaline sulphate and bromhexine hydrochloride in the presence of their potential impurities in a single run. The photolytic as well as hydrolytic impurities were detected as 3,5-dihydroxybenzoic acid, 3,5-dihydroxybenzaldehyde, 1-(3,5-dihydroxyphenyl)-2-[(1,1-dimethylethyl) amino]-ethanone from terbutaline, 2-methoxyphenol and an unknown impurity identified as (2RS)-3-(2-hydroxyphenoxy)-propane-1,2-diol from guaifenesin. The chromatographic separation of all the three active components and their impurities was achieved on Wakosil II column, using phosphate buffer (pH 3.0) and acetonitrile as mobile phase which was delivered initially in the ratio of 80:20 (v/v) for 18 min, then changed to 60:40 (v/v) for next 12 min, and finally equilibrated back to 80:20 (v/v) for 10 min. Other HPLC parameters were: Flow rate at 1.0 ml/min, detection wavelengths 248 and 280 nm, injection volume 10 μl. The calibration graphs plotted with five concentrations of each component were linear with a regression coefficient R(2) >0.9999. The limit of detection and limit of quantitation were estimated for all the five impurities. The established method was then validated for linearity, precision, accuracy, and specificity and demonstrated to be applicable to the determination of the active ingredients in commercial and model cough syrup. No interference from the formulation excipients was observed. These results suggest that this LC method can be used for the determination of multiple active ingredients and their impurities in a cough and cold syrup.

  18. The Csr system regulates genome-wide mRNA stability and transcription and thus gene expression in Escherichia coli.

    Science.gov (United States)

    Esquerré, Thomas; Bouvier, Marie; Turlan, Catherine; Carpousis, Agamemnon J; Girbal, Laurence; Cocaign-Bousquet, Muriel

    2016-04-26

    Bacterial adaptation requires large-scale regulation of gene expression. We have performed a genome-wide analysis of the Csr system, which regulates many important cellular functions. The Csr system is involved in post-transcriptional regulation, but a role in transcriptional regulation has also been suggested. Two proteins, an RNA-binding protein CsrA and an atypical signaling protein CsrD, participate in the Csr system. Genome-wide transcript stabilities and levels were compared in wildtype E. coli (MG1655) and isogenic mutant strains deficient in CsrA or CsrD activity demonstrating for the first time that CsrA and CsrD are global negative and positive regulators of transcription, respectively. The role of CsrA in transcription regulation may be indirect due to the 4.6-fold increase in csrD mRNA concentration in the CsrA deficient strain. Transcriptional action of CsrA and CsrD on a few genes was validated by transcriptional fusions. In addition to an effect on transcription, CsrA stabilizes thousands of mRNAs. This is the first demonstration that CsrA is a global positive regulator of mRNA stability. For one hundred genes, we predict that direct control of mRNA stability by CsrA might contribute to metabolic adaptation by regulating expression of genes involved in carbon metabolism and transport independently of transcriptional regulation.

  19. Macrophages and Adipocytes in Human Obesity Adipose Tissue Gene Expression and Insulin Sensitivity During Calorie Restriction and Weight Stabilization

    DEFF Research Database (Denmark)

    Capel, F.; Klimcakova, E.; Viguerie, N.

    2009-01-01

    phase with a 4-week very-low-calorie diet and a weight stabilization period composed of a 2-month low-calorie diet followed by 3-4 months of a weight maintenance diet. At each time point, a euglycemic-hyperinsulinemic clamp and subcutaneous adipose tissue biopsies were performed. Adipose tissue gene...

  20. Self-assembled ternary complexes stabilized with hyaluronic acid-green tea catechin conjugates for targeted gene delivery.

    Science.gov (United States)

    Liang, Kun; Bae, Ki Hyun; Lee, Fan; Xu, Keming; Chung, Joo Eun; Gao, Shu Jun; Kurisawa, Motoichi

    2016-03-28

    Nanosized polyelectrolyte complexes are attractive delivery vehicles for the transfer of therapeutic genes to diseased cells. Here we report the application of self-assembled ternary complexes constructed with plasmid DNA, branched polyethylenimine and hyaluronic acid-green tea catechin conjugates for targeted gene delivery. These conjugates not only stabilize plasmid DNA/polyethylenimine complexes via the strong DNA-binding affinity of green tea catechin, but also facilitate their transport into CD44-overexpressing cells via receptor-mediated endocytosis. The hydrodynamic size, surface charge and physical stability of the complexes are characterized. We demonstrate that the stabilized ternary complexes display enhanced resistance to nuclease attack and polyanion-induced dissociation. Moreover, the ternary complexes can efficiently transfect the difficult-to-transfect HCT-116 colon cancer cell line even in serum-supplemented media due to their enhanced stability and CD44-targeting ability. Confocal microscopic analysis demonstrates that the stabilized ternary complexes are able to promote the nuclear transport of plasmid DNA more effectively than binary complexes and hyaluronic acid-coated ternary complexes. The present study suggests that the ternary complexes stabilized with hyaluronic acid-green tea catechin conjugates can be widely utilized for CD44-targeted delivery of nucleic acid-based therapeutics.

  1. Oscillating Gene Expression Determines Competence for Periodic Arabidopsis Root Branching

    Science.gov (United States)

    Moreno-Risueno, Miguel A.; Van Norman, Jaimie M.; Moreno, Antonio; Zhang, Jingyuan; Ahnert, Sebastian E.; Benfey, Philip N.

    2010-01-01

    Plants and animals produce modular developmental units in a periodic fashion. In plants, lateral roots form as repeating units along the root primary axis; however, the developmental mechanism regulating this process is unknown. We found that cyclic expression pulses of a reporter gene mark the position of future lateral roots by establishing prebranch sites and that prebranch site production and root bending are periodic. Microarray and promoter-luciferase studies revealed two sets of genes oscillating in opposite phases at the root tip. Genetic studies show that some oscillating transcriptional regulators are required for periodicity in one or both developmental processes. This molecular mechanism has characteristics that resemble molecular clock–driven activities in animal species. PMID:20829477

  2. Genomic imprinting and maternal effect genes in haplodiploid sex determination

    NARCIS (Netherlands)

    van de Zande, L.; Verhulst, E. C.

    2014-01-01

    The research into the Drosophila melanogaster sex-determining system has been at the basis of all further research on insect sex determination. This further research has made it clear that, for most insect species, the presence of sufficient functional Transformer (TRA) protein in the early

  3. Genomic imprinting and maternal effect genes in haplodiploid sex determination

    NARCIS (Netherlands)

    van de Zande, L.; Verhulst, E. C.

    2014-01-01

    The research into the Drosophila melanogaster sex-determining system has been at the basis of all further research on insect sex determination. This further research has made it clear that, for most insect species, the presence of sufficient functional Transformer (TRA) protein in the early embryoni

  4. The autoregulatory loop: A common mechanism of regulation of key sex determining genes in insects

    Indian Academy of Sciences (India)

    Suresh Kumar Kumar; Gajula Gopinath; Nagraj Sambrani; Kallare P Arunkumar

    2016-06-01

    Sex determination in most insects is structured as a gene cascade, wherein a primary signal is passed through a series of sex-determining genes, culminating in a downstream double-switch known as doublesex that decides the sexual fate of the embryo. From the literature available on sex determination cascades, it becomes apparent that sex determination mechanisms have evolved rapidly. The primary signal that provides the cue to determine the sex of the embryo varies remarkably, not only among taxa, but also within taxa. Furthermore, the upstream key gene in the cascade also varies between species and even among closely related species. The order Insecta alone provides examples of astoundingly complex diversity of upstream key genes in sex determination mechanisms. Besides, unlike key upstream genes, the downstream double-switch gene is alternatively spliced to form functional sex-specific isoforms. This sex-specific splicing is conserved across insect taxa. The genes involved in the sex determination cascade such as Sex-lethal (Sxl) in Drosophila melanogaster, transformer (tra) in many other dipterans, coleopterans and hymenopterans, Feminizer (fem) in Apis mellifera, and IGF-II mRNA-binding protein (Bmimp) in Bombyx mori are reported to be regulated by an autoregulatory positive feedback loop. In this review, by taking examples from various insects, we propose the hypothesis that autoregulatory loop mechanisms of sex determination might be a general strategy. We also discuss the possible reasons for the evolution of autoregulatory loops in sex determination cascades and their impact on binary developmental choices.

  5. The autoregulatory loop: A common mechanism of regulation of key sex determining genes in insects.

    Science.gov (United States)

    Sawanth, Suresh Kumar; Gopinath, Gajula; Sambrani, Nagraj; Arunkumar, Kallare P

    2016-06-01

    Sex determination in most insects is structured as a gene cascade, wherein a primary signal is passed through a series of sex-determining genes, culminating in a downstream double-switch known as doublesex that decides the sexual fate of the embryo. From the literature available on sex determination cascades, it becomes apparent that sex determination mechanisms have evolved rapidly. The primary signal that provides the cue to determine the sex of the embryo varies remarkably, not only among taxa, but also within taxa. Furthermore, the upstream key gene in the cascade also varies between species and even among closely related species. The order Insecta alone provides examples of astoundingly complex diversity of upstream key genes in sex determination mechanisms. Besides, unlike key upstream genes, the downstream double-switch gene is alternatively spliced to form functional sex-specific isoforms. This sex-specific splicing is conserved across insect taxa. The genes involved in the sex determination cascade such as Sex-lethal (Sxl) in Drosophila melanogaster, transformer (tra) in many other dipterans, coleopterans and hymenopterans, Feminizer (fem) in Apis mellifera, and IGF-II mRNA-binding protein (Bmimp) in Bombyx mori are reported to be regulated by an autoregulatory positive feedback loop. In this review, by taking examples from various insects, we propose the hypothesis that autoregulatory loop mechanisms of sex determination might be a general strategy. We also discuss the possible reasons for the evolution of autoregulatory loops in sex determination cascades and their impact on binary developmental choices.

  6. Cell penetrating peptide-based polyplexes shelled with polysaccharide to improve stability and gene transfection

    Science.gov (United States)

    Li, Wenyu; Liu, Yajie; Du, Jianwei; Ren, Kefeng; Wang, Youxiang

    2015-04-01

    Cell-penetrating peptides (CPP) have been widely developed as a strategy to enhance cell penetrating ability and transfection. In this work, octa-arginine modified dextran gene vector with pH-sensitivity was developed via host-guest interactions. α-Cyclodextrin was modified with octa-arginine (CDR), which had excellent cell penetrating ability. Dextran was selected as a backbone and modified with azobenzene as guest units by acid-labile imine bonds (Az-I-Dex). The supramolecular polymer CDR/Az-I-Dex with high a C/A molar ratio (molar ratio of CD on CDR to Az on Az-I-Dex) was unfavorable for DNA condensation. The dextran shell of CDR/Az-I-Dex/DNA polyplexes improved the stability under physiological conditions. However, once treated with acetate buffer (pH 5.4) for 3 h, large aggregates formed rapidly due to the cleavage of the dextran shell. As expected, the vector had cell viability of 80% even when the CDR concentration increased to 100 μg mL-1. Moreover, due to the effective cellular uptake efficiency, CDR/Az-I-Dex/DNA polyplexes had 6-300 times higher transfection efficiency than CDR/DNA polyplexes. It was even higher than high molecular weight PLL-based polyplexes of HEK293 T cells. Importantly, chloroquine as an endosomal escape agent could not improve the transfection of CDR/Az-I-Dex/DNA polyplexes, which indicated that the CDR/Az-I-Dex supramolecular polymer had its own ability for endosomal escape. These results suggested that the CPP-based polyplexes shelled with polysaccharide can be promising non-viral gene delivery carriers.Cell-penetrating peptides (CPP) have been widely developed as a strategy to enhance cell penetrating ability and transfection. In this work, octa-arginine modified dextran gene vector with pH-sensitivity was developed via host-guest interactions. α-Cyclodextrin was modified with octa-arginine (CDR), which had excellent cell penetrating ability. Dextran was selected as a backbone and modified with azobenzene as guest units by acid

  7. Variation in stability of endogenous reference genes in fallopian tubes and endometrium from healthy and ectopic pregnant women.

    Science.gov (United States)

    Gebeh, Alpha K; Marczylo, Emma L; Amoako, Akwasi A; Willets, Jonathon M; Konje, Justin C

    2012-01-01

    RT-qPCR is commonly employed in gene expression studies in ectopic pregnancy. Most use RN18S1, β-actin or GAPDH as internal controls without validation of their suitability as reference genes. A systematic study of the suitability of endogenous reference genes for gene expression studies in ectopic pregnancy is lacking. The aims of this study were therefore to evaluate the stability of 12 reference genes and suggest those that are stable for use as internal control genes in fallopian tubes and endometrium from ectopic pregnancy and healthy non-pregnant controls. Analysis of the results showed that the genes consistently ranked in the top six by geNorm and NormFinder algorithms, were UBC, GAPDH, CYC1 and EIF4A2 (fallopian tubes) and UBC and ATP5B (endometrium). mRNA expression of NAPE-PLD as a test gene of interest varied between the groups depending on which of the 12 reference genes was used as internal controls. This study demonstrates that arbitrary selection of reference genes for normalisation in RT-qPCR studies in ectopic pregnancy without validation, risk producing inaccurate data and should therefore be discouraged.

  8. Determination of thermal stability of specific biomarker lipids of the freshwater fern Azolla through hydrous pyrolysis

    Science.gov (United States)

    Sap, Merel; Speelman, Eveline N.; Lewan, Michael D.; Sinninghe Damsté, Jaap S.; Reichart, Gert-Jan

    2010-05-01

    Enormous blooms of the free-floating freshwater fern Azolla occurred within the Arctic Basin during an extended period of ~1.2 Ma during the middle Eocene (Brinkhuis et al. 2006; Speelman et al., GB, 2009). The sustained growth of Azolla, currently ranking among the fastest growing plants on Earth, in a major anoxic basin may have substantially contributed to decreasing atmospheric CO2 levels by burial of Azolla-derived organic matter. Speelman et al. (OG, 2009) reported biomarkers for Azolla (1,w20 C32 - C36 diols, structurally related C29 ω20,ω21 diols, C29 1,20,21 triols, C29 dihydroxy fatty acids as well as a series of wax esters containing these mono- and dihydroxy lipids), which can be used to reconstruct palaeo-environmental conditions. Here we assess the thermal stability of these compounds, to extend their biomarker potential. We specifically focused on the thermal stability of the Azolla biomarkers using hydrous pyrolysis in order to determine which burial conditions allow reconstruction of past occurrences of Azolla. In addition, hydrous pyrolysis was also performed on samples from the Eocene Arctic Ocean (ACEX core), to test if and how the biomarkers change under higher temperatures and pressures in situ. During hydrous pyrolysis, the biomass was heated under high pressure at temperatures ranging between 220 and 365°C for 72 hours. Four experiments were also run using different durations to explore the kinetics of biomarker degradation at specific temperatures. First results indicate that the Azolla specific diols are still present at 220°C, while the corresponding wax esters are already absent. At 300°C all Azolla specific biomarkers are destroyed. More specific determination of the different biomarkers' stability and kinetics would potentially allow the reconstruction of the temperature and pressure history of Azolla deposits. Literature: • Brinkhuis, H., Schouten, S., Collinson, M. E., Sluijs, A., Sinninghe Damste, J. S., Dickens, G. R., Huber

  9. Stability of the HER2 gene after primary chemotherapy in advanced breast cancer.

    Science.gov (United States)

    Varga, Zsuzsanna; Caduff, Rosmarie; Pestalozzi, Bernhard

    2005-02-01

    We investigated whether alterations of the Her2 gene could be detected in breast cancer samples following primary chemotherapy in advanced breast cancer. The prospective study involved 23 patients with stage-II, -III or -IV breast cancer. All patients were treated with two to six cycles of fluorouracil-epirubicin and/or cyclophosphamid/epi-docetaxel. The Her2 protein and gene were assessed both on core needle biopsies prior to and on surgical specimens after completing chemotherapy using immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) methods. Estrogen and progesterone receptors (ER/PR) were also determined on both samples using IHC. Her2 status was modified in eight patients using IHC (35%) and in three patients using FISH (13%). Changes in ER/PR expression were detected in seven patients (30%). Our data suggest that alterations of the Her2 gene can occur, although not usually after primary or neoadjuvant chemotherapy. However, changes in ER/PR status seem to be a more common event; thus, both can lead to different therapeutic options. Intratumoral heterogeneity as well as sampling variations can contribute to modification of the Her2 status after primary chemotherapy.

  10. Stability - Indicating LC Method for the Simultaneous Determination of Olmesartan and Ramipril in Dosage Form

    Directory of Open Access Journals (Sweden)

    Kiran R. Patil

    2011-04-01

    Full Text Available A simple, rapid, and precise method is developed for the quantitative simultaneous estimation of Olmesartan (OL and Ramipril (RAM in combined pharmaceutical dosage form. A chromatographic separation was achieved with YMC Pack ODS A (250 x 4.6 mm analytical column. The mobile phase composed of buffer, acetonitrile and methanol (50:40:10 v/v/v. The buffer used in the mobile phase is 0.1M sodium perchlorate monohydrate in double distilled water and pH adjusted 3.0 with trifluoroacetic acid. The instrumental settings are flow rate of 1.0 mL/min, column oven temperature at 30°C and detector wavelength of 210 nm using a photodiode array detector. The resolution between OL and RAM founds to be more than 4. Theoretical plates for OL and RAM were 16599 and 15900 respectively. Tailing factor for OL and RAM was 1.11 and 1.14 respectively. Capacity factor for OL and RAM was 3.14 and 3.60 respectively. OL, RAM and combination drug product were exposed to thermal, light, hydrolytic and oxidative stress conditions, and the stressed samples were analysed by the proposed method. The proposed method was found to be suitable and accurate for quantitative determination and stability study of OL and RAM in pharmaceutical preparations.

  11. Photo-stability of Super-hydrogenated PAHs Determined by Action Spectroscopy Experiments

    Science.gov (United States)

    Wolf, M.; Kiefer, H. V.; Langeland, J.; Andersen, L. H.; Zettergren, H.; Schmidt, H. T.; Cederquist, H.; Stockett, M. H.

    2016-11-01

    We have investigated the photo-stability of pristine and super-hydrogenated pyrene cations ({{{C}}}16{{{H}}}10+m+,m=0,6,{{}} {{or}} {{}}16) by means of gas-phase action spectroscopy. Optical absorption spectra and photo-induced dissociation mass spectra are presented. By measuring the yield of mass-selected photo-fragment ions as a function of laser pulse intensity, the number of photons (and hence the energy) needed for fragmentation of the carbon backbone was determined. Backbone fragmentation of pristine pyrene ions ({{{C}}}16{{{H}}}10+) requires absorption of three photons of energy just below 3 eV, whereas super-hydrogenated hexahydropyrene (C16H{}16+) must absorb two such photons and fully hydrogenated hexadecahydropyrene (C16H{}26+) only a single photon. These results are consistent with previously reported dissociation energies for these ions. Our experiments clearly demonstrate that the increased heat capacity from the additional hydrogen atoms does not compensate for the weakening of the carbon backbone when pyrene is hydrogenated. In photodissociation regions, super-hydrogenated Polycyclic Aromatic Hydrocarbons (PAHs) have been proposed to serve as catalysts for H2 formation. Our results indicate that carbon backbone fragmentation may be a serious competitor to H2 formation at least for small hydrogenated PAHs like pyrene.

  12. Stability-Indicating LC Method for the Determination of Prasugrel Hydrochloride in Pharmaceutical Dosage Form.

    Science.gov (United States)

    Ahirrao, Vinod K; Patil, Chabutai S; Bembalkar, Saroj B; Ubale, Sanjay B; Marathe, Rajendra P; Nawale, Rajesh B; Landge, Mahadev G; Pawar, Rajendra P

    2012-01-01

    A simple, rapid and precise method was developed for the quantitative estimation of prasugrel hydrochloride in pharmaceutical dosage form. A chromatographic separation of prasugrel and its degradants was achieved with Zorbax XDB C(8), 150 × 4.6 mm, 3.5μm analytical column using aqueous solution of 0.05 M ammonium acetate pH 4.5 with acetic acid-acetonitrile (40:60 v/v). The instrumental settings include flow rate of 1.0 ml/min, column temperature at 30°C and detector wavelength of 254 nm using a photodiode array detector. Theoretical plates for prasugrel were 7023. Tailing factor for prasugrel was 1.11. Prasugrel was exposed to thermal, photolytic, hydrolytic and oxidative stress conditions, and the stressed samples were analyzed by the proposed method. Peak homogeneity data of prasugrel was obtained using photodiode array detector in the stressed sample chromatograms, which demonstrated the specificity of the method for the estimation in presence of degradants. The described method showed excellent linearity over a range of 10-300 μg/ml for prasugrel. The correlation coefficient is 0.999. The relative standard deviation of peak area for six measurements is always less than 2%. Overall, the proposed method was found to be suitable and accurate for quantitative determination and stability study of prasugrel in pharmaceutical dosage form.

  13. Synthesis of water soluble chitosan stabilized gold nanoparticles and determination of uric acid

    Science.gov (United States)

    Lanh Le, Thi; Khieu Dinh, Quang; Hoa Tran, Thai; Nguyen, Hai Phong; Le Hien Hoang, Thi; Hien Nguyen, Quoc

    2014-06-01

    Gold nanoparticles (Au-NPs) have been successfully synthesized by utilizing water soluble chitosan as reducing and stabilizing agent. The colloidal Au-NPs were characterized by UV-Vis spectroscopy and transmission electron microscopy (TEM). The results showed that the colloidal Au-NPs had a plasmon absorption band with maximum wavelength in the range of 520-526 nm and the diameters were about 8-15 nm. In addition, a new Au-NPs-modified electrode was fabricated by self-assembling Au-NPs to the surface of the L-cysteine-modified glassy carbon electrode (Au-NPs/L-Cys/GCE). The Au-NPs-modified electrode showed an excellent character for electro-catalytic oxidization of uric acid (UA) in 0.1 mol L-1 phosphate buffer solution (pH 3.2). Using differential pulse anodic stripping voltammetry (DP-ASV), a high selectivity for determination of UA has been explored for the Au-NPs-modified electrode. DP-ASV peak currents of UA increased linearly with their concentration at the range of 2.0 × 10-6 to 4.0 × 10-5 mol L-1 with the detection limit of 2.7 × 10-6 mol L-1 for UA. The proposed method was applied for the detection of UA in human urine and serum samples with satisfactory results.

  14. Colorimetric determination of hypochlorite with unmodified gold nanoparticles through the oxidation of a stabilizer thiol compound.

    Science.gov (United States)

    Zhang, Jia; Wang, Xiaolei; Yang, Xiurong

    2012-06-21

    In this article, we report a colorimetric approach for the determination of hypochlorite (OCl(-)) with gold nanoparticles (Au NPs). The test proceeds as two individual steps and selectivity is developed based on the strong oxidizing ability of hypochlorite. In concentrated phosphate buffer (PB), the red solution of citrate-capped Au NPs could be stabilized by the chemisorption of 11-mercaptoundecanoic acid (MUA), without which the colloidal suspension turned blue because of salt-induced particles aggregation. However, by its oxidizing power, OCl(-) converted the alkanethiol to a sulfonate derivative, which could not protect Au NPs from aggregation, thereby a blue solution was observed after the subsequent addition of Au suspension. With this method and under the optimal conditions (28 nm Au NP, 50 mM PB, pH 7.0, and 10 min for the colorimetric response), 1.5 μM of OCl(-) can be easily visualized by the naked eye. This sensitive and selective colorimetric assay opens up a fresh insight of facile, rapid, and reliable detection of OCl(-), and may find its future application in the monitoring of OCl(-)/HOCl in waters sanitized by chlorine or hypochlorite compounds.

  15. Photo-stability of super-hydrogenated PAHs determined by action spectroscopy experiments

    CERN Document Server

    Wolf, M; Langeland, J; Andersen, L H; Zettergren, H; Schmidt, H T; Cederquist, H; Stockett, M H

    2016-01-01

    We have investigated the photo-stability of pristine and super-hydrogenated pyrene cations C$_{16}$H$_{10+m}^+, m = 0,6, \\mathrm{\\ or\\ } 16$) by means of gas-phase action spectroscopy. Optical absorption spectra and photo-induced dissociation mass spectra are presented. By measuring the yield of mass-selected photo-fragment ions as a function of laser pulse intensity, the number of photons (and hence the energy) needed for fragmentation of the carbon backbone was determined. Backbone fragmentation of pristine pyrene ions (C$_{16}$H$_{10}^+$) requires absorption of three photons of energy just below 3 eV, whereas super-hydrogenated hexahydropyrene (C$_{16}$H$_{16}^+$) must absorb two such photons and fully hydrogenated hexadecahydropyrene (C$_{16}$H$_{26}^+$) only a single photon. These results are consistent with previously reported dissociation energies for these ions. Our experiments clearly demonstrate that the increased heat capacity from the additional hydrogen atoms does not compensate for the weakening...

  16. Stability-indicating HPLC method for simultaneous determination of montelukast and fexofenadine hydrochloride

    Directory of Open Access Journals (Sweden)

    Mona Pankhaniya

    2013-01-01

    Full Text Available A simple, specific, accurate, and stability-indicating reversed-phase high-performance liquid chromatographic method was developed for the simultaneous determination of montelukast and fexofenadine hydrochloride, using a Lichrospher ® 100, RP-18e column and a mobile phase composed of methanol:0.1% o-phosphoric acid (90:10 v/v, pH 6.8. The retention times of montelukast and fexofenadine hydrochloride were found to be 10.16 and 12.03 min, respectively. Linearity was established for montelukast and fexofenadine hydrochloride in the range of 2-10 μg/ml and 24-120 μg/ml, respectively. The percentage recoveries of montelukast and fexofenadine hydrochloride were found to be in the range of 99.09 and 99.81%, respectively. Both the drugs were subjected to acid and base hydrolysis, oxidation, photolytic, and thermal degradation conditions. The degradation products of montelukast and fexofenadine hydrochloride were well resolved from the pure drug with significant differences in their retention time values. This method can be successfully employed for simultaneous quantitative analysis of montelukast and fexofenadine hydrochloride in bulk drugs and formulations.

  17. The distribution of fat in dried dairy particles determines flavor release and flavor stability.

    Science.gov (United States)

    Park, C W; Drake, M A

    2014-04-01

    Dried dairy ingredients are utilized in various food and beverage applications for their nutritional, functional, and sensory properties. Dried dairy ingredients include milk powders of varying fat content and heat treatment and buttermilk powder, along with both milk and whey proteins of varying protein contents. The flavor of these ingredients is the most important characteristic that determines consumer acceptance of the ingredient applications. Lipid oxidation is the main mechanism for off-flavor development in dried dairy ingredients. The effects of various unit operations on the flavor of dried dairy ingredients have been investigated. Recent research documented that increased surface free fat in spray dried WPC80 was associated with increased lipid oxidation and off-flavors. Surface free fat in spray-dried products is fat on the surface of the powder that is not emulsified. The most common emulsifiers present in dried dairy ingredients are proteins and phospholipids. Currently, only an association between surface free fat and lipid oxidation has been presented. The link between surface free fat in dried dairy ingredients and flavor and flavor stability has not been investigated. In this review, some hypotheses for the role of surface free fat on the flavor of dried dairy ingredients are presented along with proposed mechanisms.

  18. Stability-Indicating TLC-densitometric determination of nebivolol hydrochloride in bulk and pharmaceutical dosage form.

    Science.gov (United States)

    Shirkhedkar, Atul A; Bugdane, Prasad M; Surana, Sanjay J

    2010-02-01

    A simple, selective, precise, and stability-indicating high-performance thin-layer chromatographic (HPTLC) method for densitometric determination of nebivolol hydrochloride both as a bulk drug and in formulation was developed and validated as per the international conference on harmonization guidelines (ICH). The method employed TLC aluminium plates precoated with silica gel 60F254 as the stationary phase. The solvent system consisted of toluene-methanol-triethylamine (3.8:1.2:0.2 v/v/v). Densitometry analysis of nebivolol hydrochloride was carried out in the absorbance mode at 281 nm. The system was found to give compact spot for nebivolol hydrochloride (R(f) value of 0.33 +/- 0.02). The linear regression analysis data for the calibration plots showed good relationship with r(2)= 0.9994 +/- 0.0002 in the concentration range 500-3000 ng/spot. The mean value +/- SD of slope and intercept were 3.761 +/- 0.017 and 127.39 +/- 19.53 with respect to peak area. The limits of detection (LOD) and limit of quantitation (LOQ) were 63.10 ng/spot and 191.23 ng/ spot, respectively. Nebivolol hydrochloride was subjected to acid and alkali hydrolysis, oxidation, thermal degradation, and photodegradation. All the peaks of degradation products were well-resolved from the standard drug with significantly different R(f) values. Statistical analysis proves that the method is repeatable, selective and accurate for the estimation of said drug. The proposed developed HPTLC method can be applied for identification and quantitative determination of nebivolol hydrochloride in the bulk and pharmaceutical dosage form.

  19. Possible role of the WDR3 gene on genome stability in thyroid cancer patients.

    Directory of Open Access Journals (Sweden)

    Wilser Andrés García-Quispes

    Full Text Available The role of the WDR3 gene on genomic instability has been evaluated in a group of 115 differentiated thyroid cancer (DTC patients. Genomic instability has been measured according to the response of peripheral blood lymphocytes to ionizing radiation (0.5 Gy. The response has been measured with the micronucleus (MN test evaluating the frequency of binucleated cells with MN (BNMN, both before and after the irradiation. No differences between genotypes, for the BNMN frequencies previous the irradiation, were observed. Nevertheless significant decreases in DNA damage after irradiation were observed in individuals carrying the variant alleles for each of the three genotyped SNPs: rs3754127 [-8.85 (-15.01 to -2.70, P<0.01]; rs3765501 [-8.98 (-15.61 to -2.36, P<0.01]; rs4658973 [-8.70 (-14.94 to -2.46, P<0.01]. These values correspond to those obtained assuming a dominant model. This study shows for the first time that WDR3 can modulate genome stability.

  20. The Use of Amelogenin Gene in Sex Determination from Human Skeletal Fragments and Teeth Specimens

    Directory of Open Access Journals (Sweden)

    Abdullahi Daudu Zagga

    2014-08-01

    Full Text Available Alternative approaches to sex determination of DNA samples involve investigation of regions of the amelogenin gene. This is the gene that encodes tooth enamel and is present on both the X and Y chromosomes. A review composed via Medline Internet search of literature and contributions from our experiences as well as experiences from colleagues. The rareness of failures in sex determination provides confidence in current techniques, but amelogenin gene method (singly of sex determination is not without failures. Amelogenin PCR method/system of sex determination should not, at the moment, completely replace traditional methods of sex identification. Hence, sex identification with amelogenin gene, of subjects for forensic purposes should be conducted as much as possible through a multiple morphological-molecular combined methods to avoid fallibility of amelogenin gene. [Archives Medical Review Journal 2014; 23(4.000: 605-622

  1. Factors Determining the Stability of a Gas Cell in an Elastic Medium

    NARCIS (Netherlands)

    Fyrillas, M.M.; Kloek, W.; van Vliet, T.; Mellema, J.

    1999-01-01

    In this paper we consider the stability of a gas cell embedded in an infinite elastic medium. The stability criterion obtained extends the classical result by Gibbs, y < 2E, to include the shear modulus of the elastic material. Interestingly, besides the shear modulus another parameter appears which

  2. Factors Determining the Stability of a Gas Cell in an Elastic Medium

    NARCIS (Netherlands)

    Fyrillas, M.M.; Kloek, W.; Vliet, van T.; Mellema, J.

    1999-01-01

    In this paper we consider the stability of a gas cell embedded in an infinite elastic medium. The stability criterion obtained extends the classical result by Gibbs, y < 2E, to include the shear modulus of the elastic material. Interestingly, besides the shear modulus another parameter appears whic

  3. Fast determination of operational stability of the soluble acetylacetone-cleaving enzyme Dke1 in an enzyme membrane reactor.

    Science.gov (United States)

    Hofer, Hannes; Steiner, Walter

    2005-11-01

    The main aim of this study was the determination of the operational stability of soluble Dke1 (EC 1.13.11.50) in an enzyme membrane reactor. In order to calculate the half-life of soluble Dke1, the K (M) of oxygen must be known. The determination of this constant was done using progress curve analysis (K (M) = 260 micromol l(-1)). In a next step, the reactor system was studied by building a mathematical model for calculation of the reactor system, using Berkeley Madonna ver. 8.0.1 software. After that, the determination of the half-life of Dke1 under operational conditions at different temperatures (5, 10, 15, 25, 30, 35 degrees C) was performed. The quantitative criterion for stability was the value of the first-order rate constant of monomolecular inactivation. The experiments showed that soluble Dke1 is poorly stable. The half-life ranged from 308 min at 5 degrees C to 9 min at 35 degrees C. This method for determining the half-life is quite applicable for enzymes which are poorly stable. In addition, both the storage stability and the operational stability can be determined.

  4. Can experts in acetabular fracture care determine hip stability after posterior wall fractures using plain radiographs and computed tomography?

    Science.gov (United States)

    Davis, Adrian T; Moed, Berton R

    2013-10-01

    Hip stability status after a posterior wall acetabular fracture involving 20%-50% of the posterior wall is difficult to determine. However, noted experts have professed that hip stability can be accurately determined by careful review of high-quality anteroposterior and oblique plain radiographs and a computed tomography scan. The objective of this investigation was to evaluate the interobserver and intraobserver reliabilities and accuracies in determining hip stability status by fellowship-trained orthopedic traumatologists expert in acetabular fracture care using these studies. Reliability and validation study. Level 1 trauma center. Fifteen patients with isolated unilateral posterior wall (OTA 62-A1) acetabular fractures involving 20%-50% of the posterior acetabular wall and known clinical outcome had undergone dynamic stress fluoroscopy under anesthesia to determine hip stability. High-quality anteroposterior and oblique plain radiographs and axial computed tomography images of 15 fractures involving 20%-50% of the posterior acetabular wall were reviewed in random order by 4 fellowship-trained orthopedic traumatologists specializing in acetabular fracture care in 2 separate sessions. The second session occurred after a minimum 1-month washout period. Determination of hip stability status was made for each fracture at the 2 time points based on the images along with any history of dislocation of the hip at the time of injury. These determinations were compared with the findings of examination under anesthesia, which served as the gold standard. Measurement of agreement using the Kappa statistic. Although intraobserver reliability was good (0.65), interobserver reliability was poor (0.12). In addition, percent correct was only 53% (32/60) for the initial reading and only 52% (31/60) for the second. For the initial reading, sensitivity and specificity were 100% (28/28) and 13% (4/32), respectively. For the second reading, the sensitivity and specificity were 57

  5. Digital transcriptome analysis of putative sex-determination genes in papaya (Carica papaya.

    Directory of Open Access Journals (Sweden)

    Naoya Urasaki

    Full Text Available Papaya (Carica papaya is a trioecious plant species that has male, female and hermaphrodite flowers on different plants. The primitive sex chromosomes genetically determine the sex of the papaya. Although draft sequences of the papaya genome are already available, the genes for sex determination have not been identified, likely due to the complicated structure of its sex-chromosome sequences. To identify the candidate genes for sex determination, we conducted a transcriptome analysis of flower samples from male, female and hermaphrodite plants using high-throughput SuperSAGE for digital gene expression analysis. Among the short sequence tags obtained from the transcripts, 312 unique tags were specifically mapped to the primitive sex chromosome (X or Y(h sequences. An annotation analysis revealed that retroelements are the most abundant sequences observed in the genes corresponding to these tags. The majority of tags on the sex chromosomes were located on the X chromosome, and only 30 tags were commonly mapped to both the X and Y(h chromosome, implying a loss of many genes on the Y(h chromosome. Nevertheless, candidate Y(h chromosome-specific female determination genes, including a MADS-box gene, were identified. Information on these sex chromosome-specific expressed genes will help elucidating sex determination in the papaya.

  6. Digital transcriptome analysis of putative sex-determination genes in papaya (Carica papaya).

    Science.gov (United States)

    Urasaki, Naoya; Tarora, Kazuhiko; Shudo, Ayano; Ueno, Hiroki; Tamaki, Moritoshi; Miyagi, Norimichi; Adaniya, Shinichi; Matsumura, Hideo

    2012-01-01

    Papaya (Carica papaya) is a trioecious plant species that has male, female and hermaphrodite flowers on different plants. The primitive sex chromosomes genetically determine the sex of the papaya. Although draft sequences of the papaya genome are already available, the genes for sex determination have not been identified, likely due to the complicated structure of its sex-chromosome sequences. To identify the candidate genes for sex determination, we conducted a transcriptome analysis of flower samples from male, female and hermaphrodite plants using high-throughput SuperSAGE for digital gene expression analysis. Among the short sequence tags obtained from the transcripts, 312 unique tags were specifically mapped to the primitive sex chromosome (X or Y(h)) sequences. An annotation analysis revealed that retroelements are the most abundant sequences observed in the genes corresponding to these tags. The majority of tags on the sex chromosomes were located on the X chromosome, and only 30 tags were commonly mapped to both the X and Y(h) chromosome, implying a loss of many genes on the Y(h) chromosome. Nevertheless, candidate Y(h) chromosome-specific female determination genes, including a MADS-box gene, were identified. Information on these sex chromosome-specific expressed genes will help elucidating sex determination in the papaya.

  7. Housekeeping gene stability influences the quantification of osteogenic markers during stem cell differentiation to the osteogenic lineage.

    Science.gov (United States)

    Quiroz, Felipe Garcia; Posada, Olga M; Gallego-Perez, Daniel; Higuita-Castro, Natalia; Sarassa, Carlos; Hansford, Derek J; Agudelo-Florez, Piedad; López, Luis E

    2010-04-01

    Real-time reverse transcription PCR (RT-qPCR) relies on a housekeeping or normalizer gene whose expression remains constant throughout the experiment. RT-qPCR is commonly used for characterization of human bone marrow mesenchymal stem cells (hBMSCs). However, to the best of our knowledge, there are no studies validating the expression stability of the genes used as normalizers during hBMSCs differentiation. This work aimed to study the stability of the housekeeping genes beta-actin, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and ribosomal protein L13A (RPL13A) during the osteogenic differentiation of hBMSCs. Their stability was evaluated via RT-qPCR in 14 and 20 day differentiation assays to the osteogenic lineage. Different normalization strategies were evaluated to quantify the osteogenic markers collagen type I, bone sialoprotein and osteonectin. Cell differentiation was confirmed via alizarin red staining. The results demonstrated up-regulation of beta-actin with maximum fold changes (MFC) of 4.38. GAPDH and RPL13A were not regulated by osteogenic media after 14 days and presented average fold changes lower than 2 in 20 day cultures. RPL13A (MFC < 2) had a greater stability when normalizing as a function of culture time compared with GAPDH (MFC gene in osteogenic differentiation studies of hBMSCs. This work highlights the importance of validating the normalizer genes used for stem cells characterization via RT-qPCR.

  8. Stability-indicating HPTLC determination of imatinib mesylate in bulk drug and pharmaceutical dosage form.

    Science.gov (United States)

    Vadera, N; Subramanian, G; Musmade, P

    2007-01-17

    A simple, selective, precise and stability-indicating high-performance thin-layer chromatographic method of analysis of imatinib mesylate both as a bulk drug and in formulations was developed and validated. The method employed HPTLC aluminium plates precoated with silica gel 60F-254 as the stationary phase. The solvent system consisted of chloroform:methanol (6:4, v/v). The system was found to give compact spot for imatinib mesylate (R(f) value of 0.53+/-0.02). Densitometric analysis of imatinib mesylate was carried out in the absorbance mode at 276 nm. The linear regression analysis data for the calibration plots showed good linear relationship with r(2)=0.9966+/-0.0013 with respect to peak area in the concentration range 100-1000 ng per spot. The mean value+/-S.D. of slope and intercept were 164.85+/-0.72 and 1168.3+/-8.26 with respect to peak area. The method was validated for precision, recovery and robustness. The limits of detection and quantitation were 10 and 30 ng per spot, respectively. Imatinib mesylate was subjected to acid and alkali hydrolysis, oxidation and thermal degradation. The drug undergoes degradation under acidic, basic, oxidation and heat conditions. This indicates that the drug is susceptible to acid, base hydrolysis, oxidation and heat. Statistical analysis proves that the method is repeatable, selective and accurate for the estimation of said drug. The proposed developed HPTLC method can be applied for identification and quantitative determination of imatinib mesylate in bulk drug and dosage forms.

  9. Stability-Indicating HPTLC Determination of Imatinib Mesylate in Bulk Drug and Pharmaceutical Dosage

    Science.gov (United States)

    Musmade, P.; Vadera, N.; Subramanian, G.

    A simple, selective, precise and stability-indicating high-performance thin-layer chromatographic method of analysis of imatinib mesylate both as a bulk drug and in formulations was developed and validated. The method employed HPTLC aluminum plates precoated with silica gel 60F-254 as the stationary phase. The solvent system consisted of chloroform:methanol (6:4, v/v). The system was found to give compact spot for imatinib mesylate (R f value of 0.53 ± 0.02). Densitometric analysis of imatinib mesylate was carried out in the absorbance mode at 276 nm. The linear regression analysis data for the calibration plots showed good linear relationship with r 2 = 0.9966 ± 0.0013 with respect to peak area in the concentration range 100-1,000 ng per spot. The mean value ± SD of slope and intercept were 164.85 ± 0.72 and 1168.3 ± 8.26, respectively, with respect to peak area. The method was validated for precision, recovery, and robustness. The limits of detection and quantitation were 10 and 30 ng per spot, respectively. Imatinib mesylate was subjected to acid and alkali hydrolysis, and oxidation and thermal degradation. The drug undergoes degradation under acidic, basic, oxidation, and heat conditions. This indicates that the drug is susceptible to acid, base hydrolysis, oxidation, and heat. Statistical analysis proves that the method is repeatable, selective, and accurate for the estimation of the said drug. The proposed developed HPTLC method can be applied for identification and quantitative determination of imatinib mesylate in bulk drug and dosage forms.

  10. A Graphical Procedure for the Simultaneous Determination of the Distribution Constant of Iodine and the Stability Constants of Trihalide Anions.

    Science.gov (United States)

    Kahwa, I. A.

    1984-01-01

    Discusses a graphical procedure which allows the distribution constant of iodine to be determined simultaneously with the trihalide anion stability constant. In addition, the procedure extends the experimental chemistry from distribution equilibria to important thermodynamic and bonding features. Advantages of using the procedure are also…

  11. Thickness determination of large-area films of yttria-stabilized zirconia produced by pulsed laser deposition

    DEFF Research Database (Denmark)

    Pryds, N.; Christensen, Bo Toftmann; Bilde-Sørensen, Jørgen;

    2006-01-01

    Films of yuria-stabilized zirconia (YSZ) on a polished silicon substrate of diameter up to 125 mm have been produced in a large-area pulsed laser deposition (PLD) setup under typical PLD conditions. The film thickness over the full film area has been determined by energy-dispersive Xray spectrome...

  12. Expression stabilities of candidate reference genes for RT-qPCR under different stress conditions in soybean.

    Directory of Open Access Journals (Sweden)

    Shuhua Ma

    Full Text Available Due to its accuracy, sensitivity and high throughput, real time quantitative PCR (RT-qPCR has been widely used in analysing gene expression. The quality of data from such analyses is affected by the quality of reference genes used. Expression stabilities for nine candidate reference genes widely used in soybean were evaluated under different stresses in this study. Our results showed that EF1A and ACT11 were the best under salinity stress, TUB4, TUA5 and EF1A were the best under drought stress, ACT11 and UKN2 were the best under dark treatment, and EF1B and UKN2 were the best under virus infection. EF1B and UKN2 were the top two genes which can be reliably used in all of the stress conditions assessed.

  13. Sry and SoxE genes: How they participate in mammalian sex determination and gonadal development?

    Science.gov (United States)

    She, Zhen-Yu; Yang, Wan-Xi

    2017-03-01

    In mammals, sex determination defines the differentiation of the bipotential genital ridge into either testes or ovaries. Sry, the mammalian Y-chromosomal testis-determining gene, is a master regulator of male sex determination. It acts to switch the undifferentiated genital ridge towards testis development, triggering the adoption of a male fate. Sry initiates a cascade of gene networks through the direct regulation of Sox9 expression and promotes supporting cell differentiation, Leydig cell specification, vasculature formation and testis cord development. In the absence of Sry, alternative genetic cascades, including female sex-determining genes RSPO1, Wnt4/β-catenin and Foxl2, are involved in the formation of female genitalia and the maintenance of female ovarian development. The mutual antagonisms between male and female sex-determining pathways are crucial in not just the initiation but also the maintenance of the somatic sex of the gonad throughout the organism's lifetime. Any imbalances in above sex-determining genes can cause disorders of sex development in humans and mice. In this review, we provide a detailed summary of the expression profiles, biochemical properties and developmental functions of Sry and SoxE genes in embryonic testis development and adult gonadal development. We also briefly summarize the dedicate balances between male and female sex-determining genes in mammalian sex development, with particular highlights on the molecular actions of Sry and Sox9 transcription factors.

  14. Diet-gene interactions between dietary fat intake and common polymorphisms in determining lipid metabolism

    Energy Technology Data Exchange (ETDEWEB)

    Corella, D.

    2009-07-01

    Current dietary guidelines for fat intake have not taken into consideration the possible genetic differences underlying the individual variability in responsiveness to dietary components. Genetic variability has been identified in humans for all the known lipid metabolism-related genes resulting in a plethora of candidate genes and genetic variants to examine in diet-gene interaction studies focused on fat consumption. Some examples of fat-gene interaction are reviewed. These include: the interaction between total intake and the 14C/T in the hepatic lipase gene promoter in determining high-density lipoprotein cholesterol (HDL-C) metabolism; the interaction between polyunsaturated fatty acids (PUFA) and the 5G/A polymorphism in the APOA1 gene plasma HDL-C concentrations; the interaction between PUFA and the L162V polymorphism in the PPARA gene in determining triglycerides and APOC3 concentrations; and the interaction between PUFA intake and the -1131T>C in the APOA5 gene in determining triglyceride metabolism. Although hundreds of diet-gene interaction studies in lipid metabolism have been published, the level of evidence to make specific nutritional recommendations to the population is still low and more research in nutrigenetics has to be undertaken. (Author) 31 refs.

  15. Phylogenetic distribution and evolutionary dynamics of the sex determination genes doublesex and transformer in insects

    NARCIS (Netherlands)

    Geuverink, E.; Beukeboom, L. W.

    2014-01-01

    Sex determination in insects is characterized by a gene cascade that is conserved at the bottom but contains diverse primary signals at the top. The bottom master switch gene doublesex is found in all insects. Its upstream regulator transformer is present in the orders Hymenoptera, Coleoptera and

  16. Phylogenetic distribution and evolutionary dynamics of the sex determination genes doublesex and transformer in insects

    NARCIS (Netherlands)

    Geuverink, E.; Beukeboom, L. W.

    2014-01-01

    Sex determination in insects is characterized by a gene cascade that is conserved at the bottom but contains diverse primary signals at the top. The bottom master switch gene doublesex is found in all insects. Its upstream regulator transformer is present in the orders Hymenoptera, Coleoptera and Di

  17. Stability-indicating assay method for determination of actarit, its process related impurities and degradation products: Insight into stability profile and degradation pathways☆

    Directory of Open Access Journals (Sweden)

    A. Abiramasundari

    2014-12-01

    Full Text Available The stability of the drug actarit was studied under different stress conditions like hydrolysis (acid, alkaline and neutral, oxidation, photolysis and thermal degradation as recommended by International Conference on Harmonization (ICH guidelines. Drug was found to be unstable in acidic, basic and photolytic conditions and produced a common degradation product while oxidative stress condition produced three additional degradation products. Drug was impassive to neutral hydrolysis, dry thermal and accelerated stability conditions. Degradation products were identified, isolated and characterized by different spectroscopic analyses. Drug and the degradation products were synthesized by a new route using green chemistry. The chromatographic separation of the drug and its impurities was achieved in a phenomenex luna C18 column employing a step gradient elution by high performance liquid chromatography coupled to photodiode array and mass spectrometry detectors (HPLC–PDA–MS. A specific and sensitive stability-indicating assay method for the simultaneous determination of the drug actarit, its process related impurities and degradation products was developed and validated.

  18. Single molecule force spectroscopy reveals critical roles of hydrophobic core packing in determining the mechanical stability of protein GB1.

    Science.gov (United States)

    Bu, Tianjia; Wang, Hui-Chuan Eileen; Li, Hongbin

    2012-08-21

    Understanding molecular determinants of protein mechanical stability is important not only for elucidating how elastomeric proteins are designed and functioning in biological systems but also for designing protein building blocks with defined nanomechanical properties for constructing novel biomaterials. GB1 is a small α/β protein and exhibits significant mechanical stability. It is thought that the shear topology of GB1 plays an important role in determining its mechanical stability. Here, we combine single molecule atomic force microscopy and protein engineering techniques to investigate the effect of side chain reduction and hydrophobic core packing on the mechanical stability of GB1. We engineered seven point mutants and carried out mechanical φ-value analysis of the mechanical unfolding of GB1. We found that three mutations, which are across the surfaces of two subdomains that are to be sheared by the applied stretching force, in the hydrophobic core (F30L, Y45L, and F52L) result in significant decrease in mechanical unfolding force of GB1. The mechanical unfolding force of these mutants drop by 50-90 pN compared with wild-type GB1, which unfolds at around 180 pN at a pulling speed of 400 nm/s. These results indicate that hydrophobic core packing plays an important role in determining the mechanical stability of GB1 and suggest that optimizing hydrophobic interactions across the surfaces that are to be sheared will likely be an efficient method to enhance the mechanical stability of GB1 and GB1 homologues.

  19. DETERMINATION OF HYDRO-ECOLOGICAL FACTORS OF THE VOLGA-CASPIAN AQUATIC ECOSYSTEMS STABILITY IN DESIGNING THEIR PROTECTION

    Directory of Open Access Journals (Sweden)

    Natalia Mitina

    2010-01-01

    Full Text Available This paper discusses possible consequences of changes in the Volga-Caspian aquatic ecosystems resulting from climate change according to the data scenarios of the Worldwide Meteorological Organization. Main hydro-ecological factors of stability of the Northern Caspian Sea ecosystems have been determined. It appears that ecosystem stability in the Northern Caspian Sea is primarily affected by natural conditions. It is essential to model riparian ecosystem processes and winter regime of reservoirs to develop strategies for mitigation of negative impacts of climate change. Such measures may include improvement of existing dams in the Volga region.

  20. Evaluation of the stability of acetaminophen in pluronic lecithin organogel and the determination of an appropriate beyond-use date.

    Science.gov (United States)

    Peacock, Gina F; Sauvageot, Jurgita

    2012-01-01

    Transdermal acetaminophen in Pluronic lecithin organogel (APAP-PLO) has been anecdotally reported as beneficial when used in cancer patients in the hospice setting. However, there is currently no published information regarding the stability of APAP-PLO. The objective of this study was to identify an appropriate formulation of APAP-PLO and to evaluate the stability of that formulation in order to determine an appropriate beyond-use date. APAP-PLO 50% was prepared by a local compounding pharmacy and analyzed at 0, 7, 14, 28, 45, 60, 90, and 180 days using a stability-indicating high-performance liquid chromatographic method. The mean concentrations and standard deviations were determined for each time point. Physical stability was also assessed by visual observation at each time point. The beyond-use date was determined as the time period that the samples maintained at least 90 percent of the initial concentration. At 180 days, the APAP-PLO was physically stable as noted by visual observation, and the concentration was 102 +/- 4.8 percent of initial concentration indicating that a beyond-use date of 180 days would be appropriate for this formulation.

  1. Estrogen Receptor beta 2 Induces Hypoxia Signature of Gene Expression by Stabilizing HIF-1 alpha in Prostate Cancer

    OpenAIRE

    Prasenjit Dey; Velazquez-Villegas, Laura A.; Michelle Faria; Anthony Turner; Philp Jonsson; Paul Webb; Cecilia Williams; Jan-Åke Gustafsson; Ström, Anders M.

    2015-01-01

    The estrogen receptor (ER) beta variant ER beta 2 is expressed in aggressive castration-resistant prostate cancer and has been shown to correlate with decreased overall survival. Genome-wide expression analysis after ER beta 2 expression in prostate cancer cells revealed that hypoxia was an overrepresented theme. Here we show that ER beta 2 interacts with and stabilizes HIF-1 alpha protein in normoxia, thereby inducing a hypoxic gene expression signature. HIF-1 alpha is known to stimulate met...

  2. The Csr system regulates genome-wide mRNA stability and transcription and thus gene expression in Escherichia coli

    OpenAIRE

    Thomas Esquerré; Marie Bouvier; Catherine Turlan; Carpousis, Agamemnon J.; Laurence Girbal; Muriel Cocaign-Bousquet

    2016-01-01

    Bacterial adaptation requires large-scale regulation of gene expression. We have performed a genome-wide analysis of the Csr system, which regulates many important cellular functions. The Csr system is involved in post-transcriptional regulation, but a role in transcriptional regulation has also been suggested. Two proteins, an RNA-binding protein CsrA and an atypical signaling protein CsrD, participate in the Csr system. Genome-wide transcript stabilities and levels were compared in wildtype...

  3. Determination of the elastic modulus of fly ash-based stabilizer applied in the trackbed

    Science.gov (United States)

    Lojda, Vít; Lidmila, Martin; Pýcha, Marek

    2017-09-01

    This paper describes a unique application of a fly ash-based stabilizer in the trackbed of a railway main line. The key goals of the stabilizer application are to protect the subgrade against the ingress of rain water, to increase the frost resistance and to remediate the natural ground constituted of weathered rock. The stabilizer was designed as a mixture of fly ash, generated as a waste material from coal plants, gypsum, calcium oxide and water. The mixture recipe was developed in a laboratory over several years. In 2005, a trial section of a railway line with subgrade consisting of clay limestone (weathered marlite) was built in the municipality of Smiřice. Since then, periodical measurements including collection of samples for laboratory evaluation of the fly ash-based stabilizer have taken place. Over the time span of the measurements, changes in mineral composition and development of fly ash transforming structures leading to the formation of C-A-S-H gel were detected. This paper describes the experimental laboratory investigation of the influence of dynamic loading on the elastic modulus of fly ash stabilizer samples and the development of permanent deformation of the samples with increasing number of loading cycles.

  4. Individual Differences in Exercise Behavior: Stability and Change in Genetic and Environmental Determinants From Age 7 to 18.

    Science.gov (United States)

    Huppertz, Charlotte; Bartels, Meike; de Zeeuw, Eveline L; van Beijsterveldt, Catharina E M; Hudziak, James J; Willemsen, Gonneke; Boomsma, Dorret I; de Geus, Eco J C

    2016-09-01

    Exercise behavior during leisure time is a major source of health-promoting physical activity and moderately tracks across childhood and adolescence. This study aims to investigate the absolute and relative contribution of genes and the environment to variance in exercise behavior from age 7 to 18, and to elucidate the stability and change of genetic and shared environmental factors that underlie this behavior. The Netherlands Twin Register collected data on exercise behavior in twins aged approximately 7, 10, 12, 14, 16 and 18 years (N = 27,332 twins; 48 % males; 47 % with longitudinal assessments). Three exercise categories (low, middle, high) were analyzed by means of liability threshold models. First, a univariate model was fitted using the largest available cross-sectional dataset with linear and quadratic effects of age as modifiers on the means and variance components. Second, a simplex model was fitted on the longitudinal dataset. Heritability was low in 7-year-olds (14 % in males and 12 % in females), but gradually increased up to age 18 (79 % in males and 49 % in females), whereas the initially substantial relative influence of the shared environment decreased with age (from 80 to 4 % in males and from 80 to 19 % in females). This decrease was due to a large increase in the genetic variance. The longitudinal model showed the genetic effects in males to be largely stable and to accumulate from childhood to late adolescence, whereas in females, they were marked by both transmission and innovation at all ages. The shared environmental effects tended to be less stable in both males and females. In sum, the clear age-moderation of exercise behavior implies that family-based interventions might be useful to increase this behavior in children, whereas individual-based interventions might be better suited for adolescents. We showed that some determinants of individual differences in exercise behavior are stable across childhood and youth, whereas

  5. STABILITY INDICATING RP-LC METHOD FOR DETERMINATION OF RASAGILINE MESYLATE IN BULK AND PHARMACEUTICAL DOSAGE FORMS

    Directory of Open Access Journals (Sweden)

    R. Narendra Kumar

    2010-10-01

    Full Text Available An isocratic stability indicating liquid chromatographic method has been developed and validated for the determination of Rasagiline in bulk drug and its pharmaceutical dosage forms. Separation of the drug with degradation products was achieved using Puroshere Star, C18, 150 x 4.6mm; 5μm column as stationary phase and pH 7.0(±0.05 buffer: Acetonitrile (40:60,v/v as mobile phase at a flow rate of 1.0 mL/min. UV detection was performed at 210 nm. The method is linear over the range of 4.8 – 150.5 μg/mL. The percent recovery of drug in dosage forms was ranged from 98.0 to 102.1. The method is simple, rapid, precise, selective and stability indicating and can be used for the assay in quality control and stability studies samples.

  6. Transition state trajectory stability determines barrier crossing rates in chemical reactions induced by time-dependent oscillating fields

    CERN Document Server

    Craven, Galen T; Hernandez, Rigoberto

    2015-01-01

    When a chemical reaction is driven by an external field, the transition state that the system must pass through as it changes from reactant to product -for example, an energy barrier- becomes time-dependent. We show that for periodic forcing the rate of barrier crossing can be determined through stability analysis of the non-autonomous transition state. Specifically, strong agreement is observed between the difference in the Floquet exponents describing stability of the transition state trajectory, which defines a recrossing-free dividing surface [G. T. Craven, T. Bartsch, and R. Hernandez, Phys. Rev. E 89, 040801(R) (2014)], and the rates calculated by simulation of ensembles of trajectories. This result opens the possibility to extract rates directly from the intrinsic stability of the transition state, even when it is time-dependent, without requiring a numerically-expensive simulation of the long-time dynamics of a large ensemble of trajectories.

  7. THE DETERMINATION OF A CRITICAL VALUE FOR DYNAMIC STABILITY OF SEMICONDUCTOR LASER DIODE WITH EXTERNAL OPTICAL FEEDBACK

    Directory of Open Access Journals (Sweden)

    Remzi YILDIRIM

    1998-01-01

    Full Text Available In this study, dynamic stability analysis of semiconductor laser diodes with external optical feedback has been realized. In the analysis the frequency response of the transfer function of laser diode H jw( , the transfer m function of laser diode with external optical feedback TF jw( , and optical feedback transfer function m K jw( obtained from small signal equations has been m accomplished using Nyquist stability analysis in complex domain. The effect of optical feedback on the stability of the system has been introduced and to bring the laser diode to stable condition the working critical boundary range of dampig frequency and reflection power constant (R has been determined. In the study the reflection power has been taken as ( .

  8. Beyond initiation-limited translational bursting: the effects of burst size distributions on the stability of gene expression

    KAUST Repository

    Kuwahara, Hiroyuki

    2015-11-04

    A main source of gene expression noise in prokaryotes is translational bursting. It arises from efficient translation of mRNAs with low copy numbers, which makes the production of protein copies highly variable and pulsatile. To obtain analytical solutions, previous models to capture this noise source had to assume translation to be initiation-limited, representing the burst size by a specific type of a long-tail distribution. However, there is increasing evidence suggesting that the initiation is not the rate-limiting step in certain settings, for example, under stress conditions. Here, to overcome the limitations imposed by the initiation-limited assumption, we present a new analytical approach that can evaluate biological consequences of the protein burst size with a general distribution. Since our new model can capture the contribution of other factors to the translational noise, it can be used to analyze the effects of gene expression noise in more general settings. We used this new model to analytically analyze the connection between the burst size and the stability of gene expression processes in various settings. We found that the burst size with different distributions can lead to quantitatively and qualitatively different stability characteristics of protein abundance and can have non-intuitive effects. By allowing analysis of how the stability of gene expression processes changes based on various distributions of translational noise, our analytical approach is expected to enable deeper insights into the control of cell fate decision-making, the evolution of cryptic genetic variations, and fine-tuning of gene circuits.

  9. Mobile antibiotic resistance - the spread of genes determining the resistance of bacteria through food products.

    Science.gov (United States)

    Godziszewska, Jolanta; Guzek, Dominika; Głąbski, Krzysztof; Wierzbicka, Agnieszka

    2016-07-07

    In recent years, more and more antibiotics have become ineffective in the treatment of bacterial nfections. The acquisition of antibiotic resistance by bacteria is associated with circulation of genes in the environment. Determinants of antibiotic resistance may be transferred to pathogenic bacteria. It has been shown that conjugation is one of the key mechanisms responsible for spread of antibiotic resistance genes, which is highly efficient and allows the barrier to restrictions and modifications to be avoided. Some conjugative modules enable the transfer of plasmids even between phylogenetically distant bacterial species. Many scientific reports indicate that food is one of the main reservoirs of these genes. Antibiotic resistance genes have been identified in meat products, milk, fruits and vegetables. The reason for such a wide spread of antibiotic resistance genes is the overuse of antibiotics by breeders of plants and animals, as well as by horizontal gene transfer. It was shown, that resistance determinants located on mobile genetic elements, which are isolated from food products, can easily be transferred to another niche. The antibiotic resistance genes have been in the environment for 30 000 years. Their removal from food products is not possible, but the risks associated with the emergence of multiresistant pathogenic strains are very large. The only option is to control the emergence, selection and spread of these genes. Therefore measures are sought to prevent horizontal transfer of genes. Promising concepts involve the combination of developmental biology, evolution and ecology in the fight against the spread of antibiotic resistance.

  10. The patterns of sex determination and differentiation genes in green sea turtle (Chelonia mydas

    Directory of Open Access Journals (Sweden)

    Anggraini Barlian

    2015-08-01

    Full Text Available Green sea turtle (C. mydas is one of TSD (Temperature-dependent Sex Determination, TSD animals which mean that their sex is determined by the egg’s incubation temperature. Genotypic Sex Determination (GSD homologous genes play a role in TSD process. Until now, research on the pattern of sex determination genes in C.mydas has not been conducted yet. The aim of this research is to reveal sex determination and differentiation genes expression in Mesonephros-Gonad (MG complexes of C. mydas embryos which incubated in masculinizing temperature (MT and feminizing temperature (FT. C. mydas eggs were incubated in 3 different stage of TSP (Thermosensitive Period at masculinizing temperature (26±10C, MT and feminizing temperature (31±10C FT. Mesonefros-gonad complexes were isolated at Pre-TSP stage (FT at 14th day, MT at 24th day, TSP stage (FT at 24th day, MT at 36th day and differentiated stage (FT at 40th day, MT at 58th day. RNA from mesonephros-gonad (MG complexes were converted into cDNA by RT-PCR process. Pattern of Sf1, Wt1, Aromatase, FoxL2, Sox9, Wnt4, Fgf9 and Rspo1 genes expression were analyzed by quantitative Real Time PCR (qPCR method with ?-actin gene as an internal control. Result of this study shown that expression pattern of Sf1, Wt1, Aromatase, FoxL2, Sox9, Wnt4, Fgf9, and Rspo1genes in gonadal embryo of C. mydas were increased during gonadal development stage. Four genes expression patterns (Wnt4, Fgf9, Rspo1, and FoxL2 have shown that these genes have role in sexual differentiation rather than in sexual determination.

  11. Size and molecular flexibility of sugars determine the storage stability of freeze-dried proteins

    NARCIS (Netherlands)

    Tonnis, W. F.; Mensink, M. A.; de Jager, A.; Maarschalk, K. van der Voort; Frijlink, H. W.; Hinrichs, W. L. J.

    2015-01-01

    Protein-based biopharmaceuticals are generally produced as aqueous solutions and stored refrigerated to obtain sufficient shelf life. Alternatively, proteins may be freeze-dried in the presence of sugars to allow storage stability at ambient conditions for prolonged periods. However, to act as a sta

  12. Qualitative community stability determines parasite establishment and richness in estuarine marshes

    Directory of Open Access Journals (Sweden)

    Tavis K. Anderson

    2013-06-01

    Full Text Available The establishment of parasites with complex life cycles is generally thought to be regulated by free-living species richness and the stability of local ecological interactions. In this study, we test the prediction that stable host communities are prerequisite for the establishment of complex multi-host parasite life cycles. The colonization of naïve killifish, Fundulus heteroclitus, by parasites was investigated in 4 salt marsh sites that differed in time since major ecological restoration, and which provided a gradient in free-living species richness. The richness of the parasite community, and the rate at which parasite species accumulated in the killifish, were similar between the low diversity unrestored site and the two high diversity (10- and 20-year restored marsh sites. The parasite community in the newly restored marsh (0 year included only directly-transmitted parasite species. To explain the paradox of a low diversity, highly invaded salt marsh (unrestored having the same parasite community as highly diverse restored marsh sites (10 and 20 yrs we assessed qualitative community stability. We find a significant correlation between system stability and parasite species richness. These data suggest a role for local stability in parasite community assembly, and support the idea that stable trophic relationships are required for the persistence of complex parasite life cycles.

  13. Determination of Myoglobin Stability by Circular Dichroism Spectroscopy: Classic and Modern Data Analysis

    Science.gov (United States)

    Mehl, Andrew F.; Crawford, Mary A.; Zhang, Lei

    2009-01-01

    Few laboratory procedures describe the use of circular dichroism (CD) at the undergraduate level. To increase the number of laboratory exercises using CD, a thermal denaturation study of myoglobin using CD is described to assess protein stability. Values obtained from a more classic linear data analysis approach are consistent with data analyzed…

  14. Determination of the stability constants for cobalt, nickel and palladium homogeneous catalyst complexes containing triphenylphosphine ligands

    NARCIS (Netherlands)

    Djekic, T.; Zivkovic, Z.; van der Ham, A.G.J.; de Haan, A.B.

    2006-01-01

    Homogeneous catalysts are complex compounds that are always in equilibrium with their free metal, free ligand and other forms of complexes. The ratios between different species are defined by the stability constants, which are influenced by different parameters such as the type of metal, ligand, cou

  15. [Diversity and genetic stability of yeast flocculation caused by variation of tandem repeats in yeast flocculin genes].

    Science.gov (United States)

    Yue, Feng; Guo, Xuena; He, Xiuping; Zhang, Borun

    2013-07-01

    Yeast flocculation is described as a reversible, asexual and calcium dependent process, in which cells adhere to form flocs by interaction of specific cell surface proteins named flocculins on yeast cells with mannose residues present on the cell wall of adjacent yeast cells. Yeast flocculation provides a very economical and convenient pathway for separation of yeast cells from the fermentation broth or removal of heavy metal ions from effluent. A large number of tandem repeats have been found in genes encoding flocculins, which not only have great regulatory effect on the structure and function of flocculins, generating the diversity of flocculation characteristics, but lead to genetic instability in flocculation as well for driving slippage and recombination reactions within and between FLO genes. Here, the research progress in effect of variation of tandem repeats in FLO genes on flocculation characteristics and genetic stability were reviewed to direct and promote the controllable application of flocculation in industrial fermentation process and environmental remediation.

  16. Stability-indicating RP-HPLC method for simultaneous determination of gatifloxacin and flurbiprofen in binary combination

    Directory of Open Access Journals (Sweden)

    Islam Ullah Khan

    2014-04-01

    Full Text Available A stability-indicating RP-HPLC method is presented for determination of gatifloxacin and flurbiprofen in binary combination. Gatifloxacin, flurbiprofen and their degradation products were detected at 254 nm using a BDS Hypersil C8 (250 X 4.6 mm, 5 µm column and mixture of 20 mM phosphate buffer (pH 3.0 and methanol 30:70 v/v as mobile phase. Response was linear over the range of 15-105 mg mL-1 for gatifloxacin (r² > 0.998 and of 1.5-10.5 mg mL-1 for flurbiprofen (r² > 0.999. The developed method efficiently separated the analytical peaks from degradation products (peak purity index > 0.9999. The method developed can be applied successfully for determination of gatifloxacin and flurbiprofen in human serum, urine, pharmaceutical formulations, and their stability studies.

  17. Application of droplet digital PCR to determine copy number of endogenous genes and transgenes in sugarcane.

    Science.gov (United States)

    Sun, Yue; Joyce, Priya Aiyar

    2017-08-28

    Droplet digital PCR combined with the low copy ACT allele as endogenous reference gene, makes accurate and rapid estimation of gene copy number in Q208 (A) and Q240 (A) attainable. Sugarcane is an important cultivated crop with both high polyploidy and aneuploidy in its 10 Gb genome. Without a known copy number reference gene, it is difficult to accurately estimate the copy number of any gene of interest by PCR-based methods in sugarcane. Recently, a new technology, known as droplet digital PCR (ddPCR) has been developed which can measure the absolute amount of the target DNA in a given sample. In this study, we deduced the true copy number of three endogenous genes, actin depolymerizing factor (ADF), adenine phosphoribosyltransferase (APRT) and actin (ACT) in three Australian sugarcane varieties, using ddPCR by comparing the absolute amounts of the above genes with a transgene of known copy number. A single copy of the ACT allele was detected in Q208 (A) , two copies in Q240 (A) , but was absent in Q117. Copy number variation was also observed for both APRT and ADF, and ranged from 9 to 11 in the three tested varieties. Using this newly developed ddPCR method, transgene copy number was successfully determined in 19 transgenic Q208 (A) and Q240 (A) events using ACT as the reference endogenous gene. Our study demonstrates that ddPCR can be used for high-throughput genetic analysis and is a quick, accurate and reliable alternative method for gene copy number determination in sugarcane. This discovered ACT allele would be a suitable endogenous reference gene for future gene copy number variation and dosage studies of functional genes in Q208 (A) and Q240 (A) .

  18. A fragile lattice: replacing bacteriophage lambda's head stability gene D with the shp gene of phage 21 generates the Mg2+-dependent virus, lambda shp.

    Science.gov (United States)

    Wendt, Jennifer L; Feiss, Michael

    2004-08-15

    Phage lambda DNA packaging is accompanied by prohead expansion, due to structural changes in gpE, the major capsid protein. Rearrangement of the gpE lattice creates binding sites for trimers of gpD, the head stabilization protein. lambda-Like phage 21's shp gene is homologous to lambda's D gene. gpD and gpShp share 49% amino acid identity. To ask whether gpShp could stabilize the lambda head shell, we replaced lambda's D gene with shp, creating lambda shp. Unlike lambda or 21, lambda shp was strictly dependent on the presence of 10(-2) M Mg2+, and lambda shp virions were very sensitive to chelating agents. Density gradient studies indicated that the lambda gpE lattice was underpopulated with gpShp. gpD's N-terminus has been proposed to contact gpE, and we found that lambda D/shp, which produces a chimeric protein with the N-terminus of gpD and the C-terminus of gpShp, was Mg2+-independent and more stable than lambda shp.

  19. Identification of two key genes controlling chill haze stability of beer in barley (Hordeum vulgare L)

    OpenAIRE

    Ye, Lingzhen; Huang, Yuqing; Dai, Fei; Ning, Huajiang; Li, Chengdao; Zhou, Meixue; Zhang, Guoping

    2015-01-01

    Background In bright beer, haze formation is a serious quality problem, degrading beer quality and reducing its shelf life. The quality of barley (Hordeum vulgare L) malt, as the main raw material for beer brewing, largely affects the colloidal stability of beer. Results In this study, the genetic mechanism of the factors affecting beer haze stability in barley was studied. Quantitative trait loci (QTL) analysis of alcohol chill haze (ACH) in beer was carried out using a Franklin/Yerong doubl...

  20. Potential flux landscapes determine the global stability of a Lorenz chaotic attractor under intrinsic fluctuations.

    Science.gov (United States)

    Li, Chunhe; Wang, Erkang; Wang, Jin

    2012-05-21

    We developed a potential flux landscape theory to investigate the dynamics and the global stability of a chemical Lorenz chaotic strange attractor under intrinsic fluctuations. Landscape was uncovered to have a butterfly shape. For chaotic systems, both landscape and probabilistic flux are crucial to the dynamics of chaotic oscillations. Landscape attracts the system down to the chaotic attractor, while flux drives the coherent motions along the chaotic attractors. Barrier heights from the landscape topography provide a quantitative measure for the robustness of chaotic attractor. We also found that the entropy production rate and phase coherence increase as the molecular numbers increase. Power spectrum analysis of autocorrelation function provides another way to quantify the global stability of chaotic attractor. We further found that limit cycle requires more flux and energy to sustain than the chaotic strange attractor. Finally, by detailed analysis we found that the curl probabilistic flux may provide the origin of the chaotic attractor.

  1. Determination of stability of epimetamorphic rock slope using Minimax Probability Machine

    Directory of Open Access Journals (Sweden)

    Manoj Kumar

    2016-01-01

    Full Text Available The article employs Minimax Probability Machine (MPM for the prediction of the stability status of epimetamorphic rock slope. The MPM gives a worst-case bound on the probability of misclassification of future data points. Bulk density (d, height (H, inclination (β, cohesion (c and internal friction angle (φ have been used as input of the MPM. This study uses the MPM as a classification technique. Two models {Linear Minimax Probability Machine (LMPM and Kernelized Minimax Probability Machine (KMPM} have been developed. The generalization capability of the developed models has been checked by a case study. The experimental results demonstrate that MPM-based approaches are promising tools for the prediction of the stability status of epimetamorphic rock slope.

  2. Habitat area and climate stability determine geographical variation in plant species range sizes.

    Science.gov (United States)

    Morueta-Holme, Naia; Enquist, Brian J; McGill, Brian J; Boyle, Brad; Jørgensen, Peter M; Ott, Jeffrey E; Peet, Robert K; Símová, Irena; Sloat, Lindsey L; Thiers, Barbara; Violle, Cyrille; Wiser, Susan K; Dolins, Steven; Donoghue, John C; Kraft, Nathan J B; Regetz, Jim; Schildhauer, Mark; Spencer, Nick; Svenning, Jens-Christian

    2013-12-01

    Despite being a fundamental aspect of biodiversity, little is known about what controls species range sizes. This is especially the case for hyperdiverse organisms such as plants. We use the largest botanical data set assembled to date to quantify geographical variation in range size for ~ 85 000 plant species across the New World. We assess prominent hypothesised range-size controls, finding that plant range sizes are codetermined by habitat area and long- and short-term climate stability. Strong short- and long-term climate instability in large parts of North America, including past glaciations, are associated with broad-ranged species. In contrast, small habitat areas and a stable climate characterise areas with high concentrations of small-ranged species in the Andes, Central America and the Brazilian Atlantic Rainforest region. The joint roles of area and climate stability strengthen concerns over the potential effects of future climate change and habitat loss on biodiversity.

  3. Habitat area and climate stability determine geographical variation in plant species range sizes

    DEFF Research Database (Denmark)

    Morueta-Holme, Naia; Enquist, Brian J.; McGill, Brian J.

    2013-01-01

    Despite being a fundamental aspect of biodiversity, little is known about what controls species range sizes. This is especially the case for hyperdiverse organisms such as plants. We use the largest botanical data set assembled to date to quantify geographical variation in range size for ~85,000 ...... concerns over the potential effects of future climate change and habitat loss on biodiversity.......,000 plant species across the New World. We assess prominent hypothesised range-size controls, finding that plant range sizes are codetermined by habitat area and long- and short-term climate stability. Strong short- and long-term climate instability in large parts of North America, including past...... glaciations, are associated with broad-ranged species. In contrast, small habitat areas and a stable climate characterise areas with high concentrations of small-ranged species in the Andes, Central America and the Brazilian Atlantic Rainforest region. The joint roles of area and climate stability strengthen...

  4. Determination of physical and chemical stability in pressurised metered dose inhalers: potential new techniques.

    Science.gov (United States)

    Ooi, Jesslynn; Traini, Daniela; Boyd, Ben J; Gaisford, Simon; Young, Paul M

    2015-01-01

    Pressurised metered dose inhalers (pMDIs) are subject to rigorous physical and chemical stability tests during formulation. Due to the time and cost associated with product development studies, there is a need for online techniques to fast screen new formulations in terms of physical and chemical (physico-chemical) stability. The problem with achieving this is that pMDIs are by their definition, pressurised, making the direct observation of physico-chemical properties in situ difficult. This review highlights the characterisation tools that can enhance the product development process for pMDIs. Techniques investigated include: laser diffraction, Raman spectroscopy, isothermal ampoule calorimetry, titration calorimetry and gas perfusion calorimetry. The operational principles behind each technique are discussed and complemented with examples from the literature. Laser diffraction is well placed to analyse real-time physical stability as a function of particle size; however, its use is restricted to suspension pMDIs. Raman spectroscopy can be potentially used to attain both suspension and solution pMDI spectra in real time; however, the majority of experiments are ex-valve chemical composition mapping. Calorimetry is an effective technique in capturing both chemical and physical degradations of APIs in real time but requires redevelopment to withstand pressure for the purposes of pMDI screening.

  5. Sex Determination by CHDW and CHDZ Genes of Avian Sex Chromosomes in Nymphicus hollandicus

    OpenAIRE

    CERİT, Harun; Kozet AVANUS

    2007-01-01

    The aim of this study was sex determination in Nymphicus hollandicus without giving it any harm and obtaining accurate results by DNA analysis. CHD genes are preserved within avian Z and W sex chromosomes. The intron regions of the CHDW and CHDZ genes vary between male (ZZ) and female (ZW) individuals. The method used in this study was based on this difference. DNA was extracted from feathers instead of blood. The intron regions of CHDW and CHDZ genes were amplified by sex specific primers (P...

  6. Determinants of cell- and gene-specific transcriptional regulation by the glucocorticoid receptor.

    Science.gov (United States)

    So, Alex Yick-Lun; Chaivorapol, Christina; Bolton, Eric C; Li, Hao; Yamamoto, Keith R

    2007-06-01

    The glucocorticoid receptor (GR) associates with glucocorticoid response elements (GREs) and regulates selective gene transcription in a cell-specific manner. Native GREs are typically thought to be composite elements that recruit GR as well as other regulatory factors into functional complexes. We assessed whether GR occupancy is commonly a limiting determinant of GRE function as well as the extent to which core GR binding sequences and GRE architecture are conserved at functional loci. We surveyed 100-kb regions surrounding each of 548 known or potentially glucocorticoid-responsive genes in A549 human lung cells for GR-occupied GREs. We found that GR was bound in A549 cells predominately near genes responsive to glucocorticoids in those cells and not at genes regulated by GR in other cells. The GREs were positionally conserved at each responsive gene but across the set of responsive genes were distributed equally upstream and downstream of the transcription start sites, with 63% of them >10 kb from those sites. Strikingly, although the core GR binding sequences across the set of GREs varied extensively around a consensus, the precise sequence at an individual GRE was conserved across four mammalian species. Similarly, sequences flanking the core GR binding sites also varied among GREs but were conserved at individual GREs. We conclude that GR occupancy is a primary determinant of glucocorticoid responsiveness in A549 cells and that core GR binding sequences as well as GRE architecture likely harbor gene-specific regulatory information.

  7. Determinants of cell- and gene-specific transcriptional regulation by the glucocorticoid receptor.

    Directory of Open Access Journals (Sweden)

    Alex Yick-Lun So

    2007-06-01

    Full Text Available The glucocorticoid receptor (GR associates with glucocorticoid response elements (GREs and regulates selective gene transcription in a cell-specific manner. Native GREs are typically thought to be composite elements that recruit GR as well as other regulatory factors into functional complexes. We assessed whether GR occupancy is commonly a limiting determinant of GRE function as well as the extent to which core GR binding sequences and GRE architecture are conserved at functional loci. We surveyed 100-kb regions surrounding each of 548 known or potentially glucocorticoid-responsive genes in A549 human lung cells for GR-occupied GREs. We found that GR was bound in A549 cells predominately near genes responsive to glucocorticoids in those cells and not at genes regulated by GR in other cells. The GREs were positionally conserved at each responsive gene but across the set of responsive genes were distributed equally upstream and downstream of the transcription start sites, with 63% of them >10 kb from those sites. Strikingly, although the core GR binding sequences across the set of GREs varied extensively around a consensus, the precise sequence at an individual GRE was conserved across four mammalian species. Similarly, sequences flanking the core GR binding sites also varied among GREs but were conserved at individual GREs. We conclude that GR occupancy is a primary determinant of glucocorticoid responsiveness in A549 cells and that core GR binding sequences as well as GRE architecture likely harbor gene-specific regulatory information.

  8. Antibiotic resistance, efflux pump genes and virulence determinants in Enterococcus spp. from surface water systems.

    Science.gov (United States)

    Molale, L G; Bezuidenhout, Cornelius Carlos

    2016-11-01

    The aim of this study was to report on antibiotic susceptibility patterns as well as highlight the presence of efflux pump genes and virulence genetic determinants in Enterococcus spp. isolated from South African surface water systems. One hundred and twenty-four Enterococcus isolates consisting of seven species were identified. Antimicrobial susceptibility testing revealed a high percentage of isolates was resistant to β-lactams and vancomycin. Many were also resistant to other antibiotic groups. These isolates were screened by PCR, for the presence of four efflux pump genes (mefA, tetK, tetL and msrC). Efflux genes mefA and tetK were not detected in any of the Enterococcus spp. However, tetL and msrC were detected in 17 % of the Enterococcus spp. The presence of virulence factors in the Enterococcus spp. harbouring efflux pump genes was determined. Virulence determinants were detected in 86 % of the Enterococcus spp. harbouring efflux pump genes. Four (asa1, cylA, gel and hyl) of the five virulence factors were detected. The findings of this study have demonstrated that Enterococcus from South African surface water systems are resistant to multiple antibiotics, some of which are frequently used for therapy. Furthermore, these isolates harbour efflux pump genes coding for resistance to antibiotics and virulence factors which enhance their pathogenic potential.

  9. De novo transcriptome sequencing to identify the sex-determination genes in Hyriopsis schlegelii.

    Science.gov (United States)

    Shi, Jianwu; Hong, Yijiang; Sheng, Junqing; Peng, Kou; Wang, Junhua

    2015-01-01

    This study presents the first analysis of expressed transcripts in the spermary and ovary of Hyriopsis schlegelii (H. schlegelii). A total of 132,055 unigenes were obtained and 31,781 of these genes were annotated. In addition, 19,511 upregulated and 25,911 downregulated unigenes were identified in the spermary. Ten sex-determination genes were selected and further analyzed by real-time PCR. In addition, mammalian genes reported to govern sex-determination pathways, including Sry, Dmrt1, Dmrt2, Sox9, GATA4, and WT1 in males and Wnt4, Rspo1, Foxl2, and β-catenin in females, were also identified in H. schlegelii. These results suggest that H. schlegelii and mammals use similar gene regulatory mechanisms to control sex determination. Moreover, genes associated with dosage compensation mechanisms, such as Msl1, Msl2, and Msl3, and hermaphrodite phenotypes, such as Tra-1, Tra-2α, Tra-2β, Fem1A, Fem1B, and Fem1C, were also identified in H. schlegelii. The identification of these genes indicates that diverse regulatory mechanisms regulate sexual polymorphism in H. schlegelii.

  10. Spectrophotometric determination of L-cysteine by using polyvinylpyrrolidone-stabilized silver nanoparticles in the presence of barium ions

    Science.gov (United States)

    Bamdad, Farzad; Khorram, Fateme; Samet, Maryam; Bamdad, Kourosh; Sangi, Mohammad Reza; Allahbakhshi, Fateme

    2016-05-01

    In this article a simple and selective colorimetric probe for cysteine determination using silver nano particles (AgNPS) is described. The determination process was based upon the surface plasmon resonance properties of polyvinylpyrrolidone-stabilized AgNPS. Interaction of AgNPS with cysteine molecules in the presence of barium ions induced a red shift in the surface plasmon resonance (SPR) maximum of AgNPs, as a result of nanoparticle aggregation. Consequently, yellow color of AgNP solution was changed to pink. The linear range for the determination of cysteine was 3.2-8.2 μM (R = 0.9965) with a limit of detection equal to 2.8 μM (3σ). The proposed method was successfully applied to the determination of cysteine in human plasma samples. Acceptable recovery results of the spiked samples confirmed the validity of the proposed method.

  11. Simpler Evaluation of Predictions and Signature Stability for Gene Expression Data

    Directory of Open Access Journals (Sweden)

    Yvonne E. Pittelkow

    2009-01-01

    Full Text Available Scientific advances are raising expectations that patient-tailored treatment will soon be available. The development of resulting clinical approaches needs to be based on well-designed experimental and observational procedures that provide data to which proper biostatistical analyses are applied. Gene expression microarray and related technology are rapidly evolving. It is providing extremely large gene expression profiles containing many thousands of measurements. Choosing a subset from these gene expression measurements to include in a gene expression signature is one of the many challenges needing to be met. Choice of this signature depends on many factors, including the selection of patients in the training set. So the reliability and reproducibility of the resultant prognostic gene signature needs to be evaluated, in such a way as to be relevant to the clinical setting. A relatively straightforward approach is based on cross validation, with separate selection of genes at each iteration to avoid selection bias. Within this approach we developed two different methods, one based on forward selection, the other on genes that were statistically significant in all training blocks of data. We demonstrate our approach to gene signature evaluation with a well-known breast cancer data set.

  12. Rapid and tunable method to temporally control gene editing based on conditional Cas9 stabilization.

    Science.gov (United States)

    Senturk, Serif; Shirole, Nitin H; Nowak, Dawid G; Corbo, Vincenzo; Pal, Debjani; Vaughan, Alexander; Tuveson, David A; Trotman, Lloyd C; Kinney, Justin B; Sordella, Raffaella

    2017-02-22

    The CRISPR/Cas9 system is a powerful tool for studying gene function. Here, we describe a method that allows temporal control of CRISPR/Cas9 activity based on conditional Cas9 destabilization. We demonstrate that fusing an FKBP12-derived destabilizing domain to Cas9 (DD-Cas9) enables conditional Cas9 expression and temporal control of gene editing in the presence of an FKBP12 synthetic ligand. This system can be easily adapted to co-express, from the same promoter, DD-Cas9 with any other gene of interest without co-modulation of the latter. In particular, when co-expressed with inducible Cre-ER(T2), our system enables parallel, independent manipulation of alleles targeted by Cas9 and traditional recombinase with single-cell specificity. We anticipate this platform will be used for the systematic characterization and identification of essential genes, as well as the investigation of the interactions between functional genes.

  13. A framework for list representation, enabling list stabilization through incorporation of gene exchangeabilities

    CERN Document Server

    Soneson, Charlotte

    2011-01-01

    Analysis of multivariate data sets from e.g. microarray studies frequently results in lists of genes which are associated with some response of interest. The biological interpretation is often complicated by the statistical instability of the obtained gene lists with respect to sampling variations, which may partly be due to the functional redundancy among genes, implying that multiple genes can play exchangeable roles in the cell. In this paper we use the concept of exchangeability of random variables to model this functional redundancy and thereby account for the instability attributable to sampling variations. We present a flexible framework to incorporate the exchangeability into the representation of lists. The proposed framework supports straightforward robust comparison between any two lists. It can also be used to generate new, more stable gene rankings incorporating more information from the experimental data. Using a microarray data set from lung cancer patients we show that the proposed method prov...

  14. Stability-indicating comparative methods using mekc and lc for determination of olmesartan medoxomil

    OpenAIRE

    Lisiane Bajerski; Paim,Clésio S.; Pereira,Andrea G.; Carolina L. Dias; Rocheli C. Rossi; Vítor Todeschini; Martin Steppe; Ana Maria Bergold; Pedro E. Fröehlich

    2013-01-01

    A stability-indicating method using MEKC was validated for the analysis of olmesartan medoxomil in tablets. Successful separation was achieved using a fused silica capillary (40 cm x 50 µm i.d.); background electrolyte consisted of a combination of 10 mmol L-1 borate buffer and 5 mmol L-1 anionic detergent sodium dodecyl sulfate (95:5; v/v) pH 6.5; hydrodynamic mode at 50 mBar for 5 s; 25 kV separation voltage at 25 ºC; and column temperature 25 ºC with detection at 257 nm. The proposed metho...

  15. Demyelination determinants map to the spike glycoprotein gene of coronavirus mouse hepatitis virus.

    Science.gov (United States)

    Das Sarma, J; Fu, L; Tsai, J C; Weiss, S R; Lavi, E

    2000-10-01

    Demyelination is the pathologic hallmark of the human immune-mediated neurologic disease multiple sclerosis, which may be triggered or exacerbated by viral infections. Several experimental animal models have been developed to study the mechanism of virus-induced demyelination, including coronavirus mouse hepatitis virus (MHV) infection in mice. The envelope spike (S) glycoprotein of MHV contains determinants of properties essential for virus-host interactions. However, the molecular determinants of MHV-induced demyelination are still unknown. To investigate the mechanism of MHV-induced demyelination, we examined whether the S gene of MHV contains determinants of demyelination and whether demyelination is linked to viral persistence. Using targeted RNA recombination, we replaced the S gene of a demyelinating virus (MHV-A59) with the S gene of a closely related, nondemyelinating virus (MHV-2). Recombinant viruses containing an S gene derived from MHV-2 in an MHV-A59 background (Penn98-1 and Penn98-2) exhibited a persistence-positive, demyelination-negative phenotype. Thus, determinants of demyelination map to the S gene of MHV. Furthermore, viral persistence is insufficient to induce demyelination, although it may be a prerequisite for the development of demyelination.

  16. Transcriptional expression of type I interferon response genes and stability of housekeeping genes in the human endometrium and endometriosis

    DEFF Research Database (Denmark)

    Vestergaard, Anna L; Knudsen, Ulla B; Munk, Torben

    2011-01-01

    Endometriosis is a painful chronic female disease defined by the presence of endometrial tissue implants in ectopic locations. The pathogenesis is much debated, and type I interferons could be involved. The expression of genes of the type I interferon response were profiled by a specific PCR Array...... was suitable for normalization of qRT-PCR studies of eutopic vs. ectopic endometrium. In the endometrial cell lines HEC1A, HEC1B, Ishikawa, and RL95-2, HMBS and HPRT1 were most stably expressed. The interferon-specific PCR Array indicated significantly different expression of the genes BST2, COL16A1, HOXB2......, and ISG20 between the endometrial tissue types. However, by correctly normalized qRT-PCR, levels of BST2, COL16A1, and the highly type I IFN-stimulated genes ISG12A and 6-16 displayed insignificant variations. Conversely, HOXB2 and ISG20 transcriptions were significantly reduced in endometriosis lesions...

  17. Recruitment of septin cytoskeletal proteins by botulinum toxin A protease determines its remarkable stability.

    Science.gov (United States)

    Vagin, Olga; Tokhtaeva, Elmira; Garay, Patton E; Souda, Puneet; Bassilian, Sara; Whitelegge, Julian P; Lewis, Ramilla; Sachs, George; Wheeler, Larry; Aoki, Roger; Fernandez-Salas, Ester

    2014-08-01

    Proteolytic cleavage of synaptosomal-associated protein 25 by the light chain of botulinum neurotoxin type A (LCA) results in a blockade of neurotransmitter release that persists for several months in motor neurons. The L428A/L429A mutation in LCA is known to significantly shorten both the proteolytic and neuroparalytic effects of the neurotoxin in mice. To elucidate the cellular mechanism for LCA longevity, we studied the effects of L428A/L429A mutation on the interactome, localization and stability of LCA expressed in cultured neuronal cells. Mass spectrometry analysis of the LCA interactome showed that the mutation prevented the interaction of LCA with septins. The wild-type LCA was concentrated in plasma-membrane-associated clusters, colocalizing with septins-2 and septin-7, which accumulated in these clusters only in the presence of LCA. The L428A/L429A mutation decreased co-clustering of LCA and septins and accelerated proteasomal and non-proteasomal degradation of LCA. Similarly, the impairment of septin oligomerization by forchlorfenuron or silencing of septin-2 prevented LCA interaction and clustering with septins and increased LCA degradation. Therefore, the dileucine-mediated LCA-septin co-clustering is crucial for the long-lasting stabilization of LCA-related proteolytic and presumably neuroparalytic activity. © 2014. Published by The Company of Biologists Ltd.

  18. Stability Indicating HPLC Determination of Risperidone in Bulk Drug and Pharmaceutical Formulations

    Directory of Open Access Journals (Sweden)

    Zarna R. Dedania

    2011-01-01

    Full Text Available The objective of the current study was to develop a validated stability-indicating assay method (SIAM for risperidone after subjecting it to forced decomposition under hydrolysis, oxidation, photolysis, and thermal stress conditions. The liquid chromatographic separation was achieved isocratically on a symmetry C18 column (5 μm size, 250 mm × 4.6 mm i.d. using a mobile phase containing methanol: acetonitrile (80 : 20, v/v at a flow rate of 1 mL/min and UV detection at 280 nm. Retention time of risperidone was found to be 3.35±0.01. The method was linear over the concentration range of 10–60 μg/mL(2=0.998 with a limit of detection and quantitation of 1.79 and 5.44 μg/mL, respectively. The method has the requisite accuracy, specificity, sensitivity, and precision to assay risperidone in bulk form and pharmaceutical dosage forms. Degradation products resulting from the stress studies did not interfere with the detection of Risperidone, and the assay is thus stability indicating.

  19. Same-strand overlapping genes in bacteria: compositional determinants of phase bias

    Directory of Open Access Journals (Sweden)

    Landan Giddy

    2008-08-01

    Full Text Available Abstract Background Same-strand overlapping genes may occur in frameshifts of one (phase 1 or two nucleotides (phase 2. In previous studies of bacterial genomes, long phase-1 overlaps were found to be more numerous than long phase-2 overlaps. This bias was explained by either genomic location or an unspecified selection advantage. Models that focused on the ability of the two genes to evolve independently did not predict this phase bias. Here, we propose that a purely compositional model explains the phase bias in a more parsimonious manner. Same-strand overlapping genes may arise through either a mutation at the termination codon of the upstream gene or a mutation at the initiation codon of the downstream gene. We hypothesized that given these two scenarios, the frequencies of initiation and termination codons in the two phases may determine the number for overlapping genes. Results We examined the frequencies of initiation- and termination-codons in the two phases, and found that termination codons do not significantly differ between the two phases, whereas initiation codons are more abundant in phase 1. We found that the primary factors explaining the phase inequality are the frequencies of amino acids whose codons may combine to form start codons in the two phases. We show that the frequencies of start codons in each of the two phases, and, hence, the potential for the creation of overlapping genes, are determined by a universal amino-acid frequency and species-specific codon usage, leading to a correlation between long phase-1 overlaps and genomic GC content. Conclusion Our model explains the phase bias in same-strand overlapping genes by compositional factors without invoking selection. Therefore, it can be used as a null model of neutral evolution to test selection hypotheses concerning the evolution of overlapping genes. Reviewers This article was reviewed by Bill Martin, Itai Yanai, and Mikhail Gelfand.

  20. Posttranscriptional regulation of GAP-43 gene expression in PC12 cells through protein kinase C-dependent stabilization of the mRNA.

    Science.gov (United States)

    Perrone-Bizzozero, N I; Cansino, V V; Kohn, D T

    1993-03-01

    We have previously shown that nerve growth factor (NGF) selectively stabilizes the GAP-43 mRNA in PC12 cells. To study the cellular mechanisms for this post-transcriptional control and to determine the contribution of mRNA stability to GAP-43 gene expression, we examined the effects of several agents that affect PC12 cell differentiation on the level of induction and rate of degradation of the GAP-43 mRNA. The NGF-mediated increase in GAP-43 mRNA levels and neurite outgrowth was mimicked by the phorbol ester TPA, but not by dibutyryl cAMP or the calcium ionophore A12783. Downregulation of protein kinase C (PKC) by high doses of phorbol esters or selective PKC inhibitors prevented the induction of this mRNA by NGF, suggesting that NGF and TPA act through a common PKC-dependent pathway. In mRNA decay studies, phorbol esters caused a selective 6-fold increase in the half-life of the GAP-43 mRNA, which accounts for most of the induction of this mRNA by TPA. The phorbol ester-induced stabilization of GAP-43 mRNA was blocked by the protein kinase inhibitor polymyxin B and was partially inhibited by dexamethasone, an agent that blocks GAP-43 expression and neuronal differentiation in PC12 cells. In contrast, the rates of degradation and the levels of the GAP-43 mRNA in control and TPA-treated cells were not affected by cycloheximide treatment. Thus, changes in GAP-43 mRNA turnover do not appear to require continuous protein synthesis. In conclusion, these data suggest that PKC activity regulates the levels of the GAP-43 mRNA in PC12 cells through a novel, translation-independent mRNA stabilization mechanism.

  1. Regulation of Nitrogenase Gene Expression by Transcript Stability in the Cyanobacterium Anabaena variabilis

    OpenAIRE

    Pratte, Brenda S.; Thiel, Teresa

    2014-01-01

    The nitrogenase gene cluster in cyanobacteria has been thought to comprise multiple operons; however, in Anabaena variabilis, the promoter for the first gene in the cluster, nifB1, appeared to be the primary promoter for the entire nif cluster. The structural genes nifHDK1 were the most abundant transcripts; however, their abundance was not controlled by an independent nifH1 promoter, but rather, by RNA processing, which produced a very stable nifH1 transcript and a moderately stable nifD1 tr...

  2. Evaluation of Sample Stability and Automated DNA Extraction for Fetal Sex Determination Using Cell-Free Fetal DNA in Maternal Plasma

    Directory of Open Access Journals (Sweden)

    Elena Ordoñez

    2013-01-01

    Full Text Available Objective. The detection of paternally inherited sequences in maternal plasma, such as the SRY gene for fetal sexing or RHD for fetal blood group genotyping, is becoming part of daily routine in diagnostic laboratories. Due to the low percentage of fetal DNA, it is crucial to ensure sample stability and the efficiency of DNA extraction. We evaluated blood stability at 4°C for at least 24 hours and automated DNA extraction, for fetal sex determination in maternal plasma. Methods. A total of 158 blood samples were collected, using EDTA-K tubes, from women in their 1st trimester of pregnancy. Samples were kept at 4°C for at least 24 hours before processing. An automated DNA extraction was evaluated, and its efficiency was compared with a standard manual procedure. The SRY marker was used to quantify cfDNA by real-time PCR. Results. Although lower cfDNA amounts were obtained by automated DNA extraction (mean 107,35 GE/mL versus 259,43 GE/mL, the SRY sequence was successfully detected in all 108 samples from pregnancies with male fetuses. Conclusion. We successfully evaluated the suitability of standard blood tubes for the collection of maternal blood and assessed samples to be suitable for analysis at least 24 hours later. This would allow shipping to a central reference laboratory almost from anywhere in Europe.

  3. Is a sex-determining gene(s) necessary for sex-determination in amphibians? Steroid hormones may be the key factor.

    Science.gov (United States)

    Nakamura, M

    2013-01-01

    Amphibians have 2 genetic sex-determining systems, one with male (XX/XY) and one with female (ZZ/ZW) heterogamety. While the ancestral state of sex-determination is thought to be female heterogamety, male and female heterogametic types were probably once interchangeable. The Japanese frog Rana rugosa has both XX/XY and ZZ/ZW systems within a single species in certain local populations. However, steroid hormones can alter the phenotypic sex epigenetically. In R. rugosa, steroidogenic enzyme expression starts before sex-determination in the indifferent gonad, and these enzymes become active in both male and female tadpoles. Androgens are produced in the indifferent gonad of male tadpoles at high levels, whereas estrogens are synthesized in females. In this regard, the observed enhanced expression of the hormone-metabolizing genes, CYP19 in the female gonad and CYP17 in males, may be crucial for sex-determination. Moreover, with FSH known to increase estrogen synthesis in the vertebrate ovary, observed upregulation of FSH receptor (FSHR) expression in the indifferent gonad of female tadpoles is intriguing. These data suggest that steroid hormones could be crucial for sex-determination in R. rugosa, with the consequence that upregulation of CYP19 and FSHR expression is necessary for female and CYP17 for male sex-determination.

  4. Stabilization of cancer-specific gene carrier via hydrophobic interaction for a clear-cut response to cancer signaling.

    Science.gov (United States)

    Kim, Chan Woo; Toita, Riki; Kang, Jeong-Hun; Li, Kai; Lee, Eun Kyung; Zhao, Guo Xi; Funamoto, Daiki; Nobori, Takanobu; Nakamura, Yuta; Mori, Takeshi; Niidome, Takuro; Katayama, Yoshiki

    2013-09-28

    Here, we developed a new gene carrier, comprising a linear polyethylenimine (LPEI) grafted with a hydrophobically modified cationic peptide containing a long alkyl chain, for use in cancer-specific gene delivery. The cationic peptide is a substrate of protein kinase Cα (PKCα), which is known to be activated specifically in cancer cells. The hydrophobically modified LPEI-peptide conjugate (LPEI-C10-peptide) could form a polyplex with DNA through electrostatic and hydrophobic interactions between the anionic DNA strands and the cationic peptide substrate. The hydrophobic modification of the peptide did not affect the reactivity of the peptide toward PKCα, while the polyplex showed improved intracellular uptake. Because of the efficient endosomal escape and enhanced stability, the polyplex significantly improved the transgene regulation responding to intracellular PKCα activity.

  5. Functional analysis of sex-determination genes by gene silencing with LNA-DNA gapmers in the silkworm, Bombyx mori.

    Science.gov (United States)

    Sakai, Hiroki; Sakaguchi, Honami; Aoki, Fugaku; Suzuki, Masataka G

    2015-08-01

    The sexual fate of B. mori is determined genetically; ZW, female and ZZ, male. Recently, we successfully identified a strong candidate gene at the top of the sex determination cascade in B. mori. This gene was termed Feminizer (Fem) and revealed to be a source of Fem-piRNA. Further, we found that B. mori doublesex (Bmdsx) splicing was markedly altered to produce the male-type isoform when a Fem-piRNA inhibitor was injected into ZW embryos. Moreover, knockdown of Masculinizer (Masc), a Fem-piRNA target gene, altered to produce the female-type isoform of Bmdsx in male embryos. However, it remains unclear as to whether Masc directly regulates the sex-specific expression of Bmdsx. In previous studies, we determined that the male-specific isoform of the Bombyx homolog of IGF-II mRNA-binding protein (Imp(M)) was involved in the male-specific splicing of Bmdsx. In an attempt to clarify the genetic relationship between Fem, Masc, Imp(M), and Bmdsx, knockdown experiments were performed. Knockdown of Fem shifted into male-type Bmdsx, Imp(M) and Masc in female embryos. Knockdown of Masc led to the production of the female-type Bmdsx and a dramatic reduction in Imp(M) expression in male embryos. Knockdown of Imp(M) shifted Bmdsx splice mode from the male-type into the female-type. Our results suggest that: (1) Fem reduces Masc expression, (2) Masc dramatically induces Imp(M) expression, and (3) Imp(M) shifting Bmdsx splice mode from the female-type into the male-type. Based on these findings, we propose a possible genetic cascade regulating sex determination in B. mori.

  6. Observability and confidence of stability and control derivatives determined in real time

    Science.gov (United States)

    Noriega, Alfonso

    Stability and control derivatives of an aircraft were estimated from real flight test data in real time. A higher language block diagram library was developed for this purpose. Parameter identification techniques and requirements were used to detect and rate maneuvers present in the data. These ratings were used to blend newly calculated derivatives with previously known values by means of a Kalman filter. The Kalman filter output was used to identify the health of control surfaces actuators. Statistical and measured data were used to predict the probability that an actuator failure has occurred at any given time during the flight. Sweeps of all the tuning parameters of the system were performed, and it was demonstrated that these tuning parameters can be used to obtain the desired performance based on requirements.

  7. A validated HPLC stability-indicating method for the determination of diacerhein in bulk drug substance.

    Science.gov (United States)

    Giannellini, Valerio; Salvatore, Francesco; Bartolucci, Gianluca; Coran, Silvia A; Bambagiotti-Alberti, Massimo

    2005-09-15

    A novel stability-indicating high-performance liquid chromatographic (HPLC) method was developed and validated for the assay of diacerhein in bulk forms. Diacerhein was found to degrade in alkaline and acidic conditions and also under oxidative stress. The drug was stable to dry heat and in presence of light. Resolution of drug, its potential impurities and degradation products were achieved on a RP18 (endcapped) column utilizing 0.1 M phosphoric acid and methanol (40:60, v/v) as eluent at the detection wavelength of 254 nm. The validation studies were carried out fulfilling International Conference on Harmonisation (ICH) requirements. The procedure was found to be specific, linear, precise (including intermediate precision), accurate and robust.

  8. Systematic mapping of occluded genes by cell fusion reveals prevalence and stability of cis-mediated silencing in somatic cells

    Science.gov (United States)

    Looney, Timothy J.; Zhang, Li; Chen, Chih-Hsin; Lee, Jae Hyun; Chari, Sheila; Mao, Frank Fuxiang; Pelizzola, Mattia; Zhang, Lu; Lister, Ryan; Baker, Samuel W.; Fernandes, Croydon J.; Gaetz, Jedidiah; Foshay, Kara M.; Clift, Kayla L.; Zhang, Zhenyu; Li, Wei-Qiang; Vallender, Eric J.; Wagner, Ulrich; Qin, Jane Yuxia; Michelini, Katelyn J.; Bugarija, Branimir; Park, Donghyun; Aryee, Emmanuel; Stricker, Thomas; Zhou, Jie; White, Kevin P.; Ren, Bing; Schroth, Gary P.; Ecker, Joseph R.; Xiang, Andy Peng; Lahn, Bruce T.

    2014-01-01

    Both diffusible factors acting in trans and chromatin components acting in cis are implicated in gene regulation, but the extent to which either process causally determines a cell's transcriptional identity is unclear. We recently used cell fusion to define a class of silent genes termed “cis-silenced” (or “occluded”) genes, which remain silent even in the presence of trans-acting transcriptional activators. We further showed that occlusion of lineage-inappropriate genes plays a critical role in maintaining the transcriptional identities of somatic cells. Here, we present, for the first time, a comprehensive map of occluded genes in somatic cells. Specifically, we mapped occluded genes in mouse fibroblasts via fusion to a dozen different rat cell types followed by whole-transcriptome profiling. We found that occluded genes are highly prevalent and stable in somatic cells, representing a sizeable fraction of silent genes. Occluded genes are also highly enriched for important developmental regulators of alternative lineages, consistent with the role of occlusion in safeguarding cell identities. Alongside this map, we also present whole-genome maps of DNA methylation and eight other chromatin marks. These maps uncover a complex relationship between chromatin state and occlusion. Furthermore, we found that DNA methylation functions as the memory of occlusion in a subset of occluded genes, while histone deacetylation contributes to the implementation but not memory of occlusion. Our data suggest that the identities of individual cell types are defined largely by the occlusion status of their genomes. The comprehensive reference maps reported here provide the foundation for future studies aimed at understanding the role of occlusion in development and disease. PMID:24310002

  9. Application of stability-indicating HPTLC method for quantitative determination of metadoxine in pharmaceutical dosage form.

    Science.gov (United States)

    Kaul, Neeraj; Agrawal, Himani; Patil, Bharat; Kakad, Abhijit; Dhaneshwar, S R

    2005-04-01

    A sensitive, selective, precise and stability-indicating high-performance thin-layer chromatographic method for analysis of metadoxine both as a bulk drug and in formulations was developed and validated. The method employed TLC aluminium plates precoated with silica gel 60F-254 as the stationary phase. The solvent system consisted of acetone-chloroform-methanol-ammonia (7.0:4.0:3.0:1.2, v/v/v/v). Densitometric analysis of metadoxine was carried out in the absorbance mode at 315 nm. This system was found to give compact spots for metadoxine (Rf value of 0.45+/-0.02, for six replicates). Metadoxine was subjected to acid, alkali and neutral hydrolysis, oxidation, dry and wet heat treatment and photo and UV degradation. The drug undergoes degradation under all stress conditions. Also, the degraded products were well resolved from the pure drug with significantly different Rf values. The method was validated for linearity, precision, robustness, LOD, LOQ, specificity and accuracy. Linearity was found to be in the range of 100-1500 ng/spot with significantly high value of correlation coefficient r2=0.9997+/-1.02. The linear regression analysis data for the calibration plots showed good linear relationship with r2=0.9999+/-0.58 in the working concentration range of 200-700 ng/spot. The mean value of slope and intercept were 0.11+/-0.04 and 18.73+/-1.89, respectively. The limits of detection and quantitation were 50 and 100 ng/spot, respectively. Statistical analysis proves that the method is repeatable and specific for the estimation of the said drug. As the method could effectively separate the drug from its degradation products, it can be employed as a stability-indicating one. Moreover, the proposed HPTLC method was utilized to investigate the kinetics of acid and base degradation process. Arrhenius plot was constructed and activation energy was calculated respectively for acid and base degradation process.

  10. Determination of ivermectin stability by high-performance thin-layer chromatography

    Directory of Open Access Journals (Sweden)

    Mohd. Ali

    2011-06-01

    Full Text Available A rapid, sensi tive and stabi l i ty-signi fying high-performance thin-layer chromatographic (HPTLC method was developed and validated for the quantitative estimation of ivermectin (IVM as a bulk drug and in pharmaceutical formulations. The separation was achieved on Lichrospher TLC aluminum plates pre-coated with silica gel 60F-254 (20cm×10cm×200 :m using n-hexane: acetone: ethylacetate (6.5: 3.5: 0.1 v/v/v as mobi le phase. The densitometric analysis was carried out at 247 nm wavelength. Compact spots of IVM were found at Rf = 26±0.02. For proposed procedure, linearity (r2 = 0.9989, limit of quantification (24.9 ng spot−1, limit of detection (8.22 ng spot−1 recovery (98.25–100.16%, and inter as well intra-day precision (≤2.21 was found to be satisfactory. We have synthesized polymeric nanoparticles encapsulated formulat ion of ivermectin (IVM-NPs ; ut i l izing micel lar aggregates of cross-l inked random copolymer Nisopropylacrylamide (NIPAAM with N-vinyl-2-pyrrolidone (VP and polyethyleneglycol monoacrylate (PEG-A for lymphat ic targeting and i t was also quanti f ied by the developed method. IVM and formulations were subjected to acid and alkali hydrolysis, oxidation and photo-degradation. The drug undergoes degradation under acidic, basic, light and oxidation conditions. This indicates that the drug is susceptible to acid- base hydrolysis, oxidation and photo-oxidation and the developed method is selective for quantifying IVM even in the presence of degradatnts. The method was applicable for routine analysis and stability testing of IVM in pharmaceutical drug delivery systems. As the method could effectively separate the said drug from its degradation products, it can be employed as a stability indicating one.

  11. The Acinar Cage: Basement Membranes Determine Molecule Exchange and Mechanical Stability of Human Breast Cell Acini.

    Directory of Open Access Journals (Sweden)

    Aljona Gaiko-Shcherbak

    Full Text Available The biophysical properties of the basement membrane that surrounds human breast glands are poorly understood, but are thought to be decisive for normal organ function and malignancy. Here, we characterize the breast gland basement membrane with a focus on molecule permeation and mechanical stability, both crucial for organ function. We used well-established and nature-mimicking MCF10A acini as 3D cell model for human breast glands, with ether low- or highly-developed basement membrane scaffolds. Semi-quantitative dextran tracer (3 to 40 kDa experiments allowed us to investigate the basement membrane scaffold as a molecule diffusion barrier in human breast acini in vitro. We demonstrated that molecule permeation correlated positively with macromolecule size and intriguingly also with basement membrane development state, revealing a pore size of at least 9 nm. Notably, an intact collagen IV mesh proved to be essential for this permeation function. Furthermore, we performed ultra-sensitive atomic force microscopy to quantify the response of native breast acini and of decellularized basement membrane shells against mechanical indentation. We found a clear correlation between increasing acinar force resistance and basement membrane formation stage. Most important native acini with highly-developed basement membranes as well as cell-free basement membrane shells could both withstand physiologically relevant loads (≤ 20 nN without loss of structural integrity. In contrast, low-developed basement membranes were significantly softer and more fragile. In conclusion, our study emphasizes the key role of the basement membrane as conductor of acinar molecule influx and mechanical stability of human breast glands, which are fundamental for normal organ function.

  12. The lysosomal stability as a biomarker for the determination of pollution in aquatic environments

    Directory of Open Access Journals (Sweden)

    Maria Loreto Nazar

    2008-10-01

    Full Text Available This work studied the effects caused by five different formulae of gasoline on the stability of the lysosomes isolated from the liver of the tilapia fish (Oreochromis niloticus. The functional integrity of the lysosomal membranes was evaluated via the acid phosphatase activity. The results showed that there were significant changes in the stability of the lysosomes exposed to the presence of the hydrocarbons in the environment. Therefore, considering the method's simplicity, the sensitivity of the responses and its low cost the assessment of the lysosomal activity could be an important tool for the study of the effects of pollution in the aquatic environments.A procura de biomarcadores de agentes poluidores, mais simples e menos custosos, tem levado ao estudo dos lisossomos, isolados de animais componentes da biota nos ambientes contaminados, principalmente por poluentes com características lipofílicas, a exemplo dos hidrocarbonetos policíclicos e seus derivados. Este trabalho estudou os efeitos provocados por 05 diferentes formulações de gasolina sobre a estabilidade de lisossomos, isolados de fígado de tilápia (Oreochromis niloticus. A integridade funcional das membranas lisossômicas foi avaliada através da atividade da fosfatase ácida, expressa em mU/mg de proteínas totais. Os resultados obtidos mostraram que existem alterações significativas na estabilidade dos lisossomos isolados de fígado de tilápias submetidas aos efeitos de hidrocarbonetos presentes no meio ambiente. Portanto, levando em conta a simplicidade, a sensibilidade de resposta e o baixo custo, os autores recomendam a avaliação da atividade lisossômica, como uma importante ferramenta para o estudo dos efeitos da poluição dos meios aquáticos.

  13. Identification of floral genes for sex determination in Calamus palustris Griff. by using suppression subtractive hybridization.

    Science.gov (United States)

    Ng, C Y; Wickneswari, R; Choong, C Y

    2014-08-07

    Calamus palustris Griff. is an economically important dioecious rattan species in Southeast Asia. However, dioecy and onset of flowering at 3-4 years old render uncertainties in desired female:male seedling ratios to establish a productive seed orchard for this rattan species. We constructed a subtractive library for male floral tissue to understand the genetic mechanism for gender determination in C. palustris. The subtractive library produced 1536 clones with 1419 clones of high quality. Reverse Northern screening showed 313 clones with differential expression, and sequence analyses clustered them into 205 unigenes, including 32 contigs and 173 singletons. The subtractive library was further validated with reverse transcription-quantitative polymerase chain reaction analysis. Homology identification classified the unigenes into 12 putative functional proteins with 83% unigenes showing significant match to proteins in databases. Functional annotations of these unigenes revealed genes involved in male flower development, including MADS-box genes, pollen-related genes, phytohormones for flower development, and male flower organ development. Our results showed that the male floral genes may play a vital role in sex determination in C. palustris. The identified genes can be exploited to understand the molecular basis of sex determination in C. palustris.

  14. A Toolbox for Quantitative Gene Expression in Varroa destructor: RNA Degradation in Field Samples and Systematic Analysis of Reference Gene Stability.

    Science.gov (United States)

    Campbell, Ewan M; McIntosh, Catriona H; Bowman, Alan S

    2016-01-01

    Varroa destructor is the major pest of Apis mellifera and contributes to the global honey bee health crisis threatening food security. Developing new control strategies to combat Varroa will require the application of molecular biology, including gene expression studies by quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR). Both high quality RNA samples and suitable stable internal reference genes are required for accurate gene expression studies. In this study, ten candidate genes (succinate dehydrogenase (SDHA), NADH dehydrogenase (NADH), large ribsosmal subunit, TATA-binding protein, glyceraldehyde-3-phosphate dehydrogenase, 18S rRNA (18S), heat-shock protein 90 (HSP90), cyclophilin, α-tubulin, actin), were evaluated for their suitability as normalization genes using the geNorm, Normfinder, BestKeeper, and comparative ΔCq algorithims. Our study proposes the use of no more than two of the four most stable reference genes (NADH, 18S, SDHA and HSP90) in Varroa gene expression studies. These four genes remain stable in phoretic and reproductive stage Varroa and are unaffected by Deformed wing virus load. When used for determining changes in vitellogenin gene expression, the signal-to-noise ratio (SNR) for the relatively unstable genes actin and α-tubulin was much lower than for the stable gene combinations (NADH + HSP90 +18S; NADH + HSP90; or NADH). Using both electropherograms and RT-qPCR for short and long amplicons as quality controls, we demonstrate that high quality RNA can be recovered from Varroa up to 10 days later stored at ambient temperature if collected into RNAlater and provided the body is pierced. This protocol allows the exchange of Varroa samples between international collaborators and field sample collectors without requiring frozen collection or shipping. Our results make important contributions to gene expression studies in Varroa by proposing a validated sampling protocol to obtain high quality Varroa RNA and the

  15. A Toolbox for Quantitative Gene Expression in Varroa destructor: RNA Degradation in Field Samples and Systematic Analysis of Reference Gene Stability

    Science.gov (United States)

    Campbell, Ewan M.; McIntosh, Catriona H.; Bowman, Alan S.

    2016-01-01

    Varroa destructor is the major pest of Apis mellifera and contributes to the global honey bee health crisis threatening food security. Developing new control strategies to combat Varroa will require the application of molecular biology, including gene expression studies by quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR). Both high quality RNA samples and suitable stable internal reference genes are required for accurate gene expression studies. In this study, ten candidate genes (succinate dehydrogenase (SDHA), NADH dehydrogenase (NADH), large ribsosmal subunit, TATA-binding protein, glyceraldehyde-3-phosphate dehydrogenase, 18S rRNA (18S), heat-shock protein 90 (HSP90), cyclophilin, α-tubulin, actin), were evaluated for their suitability as normalization genes using the geNorm, Normfinder, BestKeeper, and comparative ΔCq algorithims. Our study proposes the use of no more than two of the four most stable reference genes (NADH, 18S, SDHA and HSP90) in Varroa gene expression studies. These four genes remain stable in phoretic and reproductive stage Varroa and are unaffected by Deformed wing virus load. When used for determining changes in vitellogenin gene expression, the signal-to-noise ratio (SNR) for the relatively unstable genes actin and α-tubulin was much lower than for the stable gene combinations (NADH + HSP90 +18S; NADH + HSP90; or NADH). Using both electropherograms and RT-qPCR for short and long amplicons as quality controls, we demonstrate that high quality RNA can be recovered from Varroa up to 10 days later stored at ambient temperature if collected into RNAlater and provided the body is pierced. This protocol allows the exchange of Varroa samples between international collaborators and field sample collectors without requiring frozen collection or shipping. Our results make important contributions to gene expression studies in Varroa by proposing a validated sampling protocol to obtain high quality Varroa RNA and the

  16. A Toolbox for Quantitative Gene Expression in Varroa destructor: RNA Degradation in Field Samples and Systematic Analysis of Reference Gene Stability.

    Directory of Open Access Journals (Sweden)

    Ewan M Campbell

    Full Text Available Varroa destructor is the major pest of Apis mellifera and contributes to the global honey bee health crisis threatening food security. Developing new control strategies to combat Varroa will require the application of molecular biology, including gene expression studies by quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR. Both high quality RNA samples and suitable stable internal reference genes are required for accurate gene expression studies. In this study, ten candidate genes (succinate dehydrogenase (SDHA, NADH dehydrogenase (NADH, large ribsosmal subunit, TATA-binding protein, glyceraldehyde-3-phosphate dehydrogenase, 18S rRNA (18S, heat-shock protein 90 (HSP90, cyclophilin, α-tubulin, actin, were evaluated for their suitability as normalization genes using the geNorm, Normfinder, BestKeeper, and comparative ΔCq algorithims. Our study proposes the use of no more than two of the four most stable reference genes (NADH, 18S, SDHA and HSP90 in Varroa gene expression studies. These four genes remain stable in phoretic and reproductive stage Varroa and are unaffected by Deformed wing virus load. When used for determining changes in vitellogenin gene expression, the signal-to-noise ratio (SNR for the relatively unstable genes actin and α-tubulin was much lower than for the stable gene combinations (NADH + HSP90 +18S; NADH + HSP90; or NADH. Using both electropherograms and RT-qPCR for short and long amplicons as quality controls, we demonstrate that high quality RNA can be recovered from Varroa up to 10 days later stored at ambient temperature if collected into RNAlater and provided the body is pierced. This protocol allows the exchange of Varroa samples between international collaborators and field sample collectors without requiring frozen collection or shipping. Our results make important contributions to gene expression studies in Varroa by proposing a validated sampling protocol to obtain high quality Varroa

  17. Redox regulation of genome stability by effects on gene expression, epigenetic pathways and DNA damage/repair.

    Science.gov (United States)

    Mikhed, Yuliya; Görlach, Agnes; Knaus, Ulla G; Daiber, Andreas

    2015-08-01

    Reactive oxygen and nitrogen species (e.g. H2O2, nitric oxide) confer redox regulation of essential cellular signaling pathways such as cell differentiation, proliferation, migration and apoptosis. In addition, classical regulation of gene expression or activity, including gene transcription to RNA followed by translation to the protein level, by transcription factors (e.g. NF-κB, HIF-1α) and mRNA binding proteins (e.g. GAPDH, HuR) is subject to redox regulation. This review will give an update of recent discoveries in this field, and specifically highlight the impact of reactive oxygen and nitrogen species on DNA repair systems that contribute to genomic stability. Emphasis will be placed on the emerging role of redox mechanisms regulating epigenetic pathways (e.g. miRNA, DNA methylation and histone modifications). By providing clinical correlations we discuss how oxidative stress can impact on gene regulation/activity and vise versa, how epigenetic processes, other gene regulatory mechanisms and DNA repair can influence the cellular redox state and contribute or prevent development or progression of disease.

  18. Redox regulation of genome stability by effects on gene expression, epigenetic pathways and DNA damage/repair

    Directory of Open Access Journals (Sweden)

    Yuliya Mikhed

    2015-08-01

    Full Text Available Reactive oxygen and nitrogen species (e.g. H2O2, nitric oxide confer redox regulation of essential cellular signaling pathways such as cell differentiation, proliferation, migration and apoptosis. In addition, classical regulation of gene expression or activity, including gene transcription to RNA followed by translation to the protein level, by transcription factors (e.g. NF-κB, HIF-1α and mRNA binding proteins (e.g. GAPDH, HuR is subject to redox regulation. This review will give an update of recent discoveries in this field, and specifically highlight the impact of reactive oxygen and nitrogen species on DNA repair systems that contribute to genomic stability. Emphasis will be placed on the emerging role of redox mechanisms regulating epigenetic pathways (e.g. miRNA, DNA methylation and histone modifications. By providing clinical correlations we discuss how oxidative stress can impact on gene regulation/activity and vise versa, how epigenetic processes, other gene regulatory mechanisms and DNA repair can influence the cellular redox state and contribute or prevent development or progression of disease.

  19. Mice lacking pituitary tumor transforming gene show elevated exposure of DGalNAc carbohydrate determinants

    Directory of Open Access Journals (Sweden)

    Lutsyk A. D.

    2012-04-01

    Full Text Available Aim. To investigate the influence of pituitary tumor transforming gene (pttg-1 knockout on glycome of parenchimal organs by means of lectin histochemistry. Methods. DGalNAc, DGlcNAc, NeuNAc carbohydrate determinants were labelled with soybean agglutinin (SBA and wheat germ agglutinin (WGA, conjugated to peroxidase, with subsequent visualization of the lectin-binding sites with diaminobenzidine. The testes and kidneys of murine strain BL6/C57 with the pttg-1 gene knockout (PTTG-KO were compared to the wild type (PTTG-WT animals, both groups 1 month of age. Results. Knockout of the pttg-1 gene was accompanied by enhanced exposure of the DGalNAc sugar residues within the Golgi complex of secondary spermatocytes, in a brush border of renal tubules and on the lumenal surface of collecting ducts. Conclusions. This study suggests that knockout of the pttg-1 gene may lead to the changes in carbohydrate processing in mammalian organism.

  20. Defining genes using "blueprint" versus "instruction" metaphors: effects for genetic determinism, response efficacy, and perceived control.

    Science.gov (United States)

    Parrott, Roxanne; Smith, Rachel A

    2014-01-01

    Evidence supports mixed attributions aligned with personal and/or clinical control and gene expression for health in this era of genomic science and health care. We consider variance in these attributions and possible relationships to individual mind sets associated with essentialist beliefs that genes determine health versus threat beliefs that genes increase susceptibility for disease and severity linked to gene-environment interactions. Further, we contribute to theory and empirical research to evaluate the use of metaphors to define genes. Participants (N = 324) read a message that varied the introduction by providing a definition of genes that used either an "instruction" metaphor or a "blueprint" metaphor. The "instruction" metaphor compared to the "blueprint" metaphor promoted stronger threat perceptions, which aligned with both belief in the response efficacy of genetic research for health and perceived behavioral control linked to genes and health. The "blueprint" metaphor compared to the "instruction" metaphor promoted stronger essentialist beliefs, which aligned with more intense positive regard for the efficacy of genetic research and human health. Implications for health communicators include societal effects aligned with stigma and discrimination that such findings portend.

  1. Detection of Genes that Determine Maize Grain Quality Characteristics and Resistance to Stress Factors

    Directory of Open Access Journals (Sweden)

    Markovskyi, O.V.

    2014-01-01

    Full Text Available 200 experimental maize samples (Maize Company were examined for the presence of genes that determine the quality characteristics of grain (wx and fl-2 genes, herbicide (bar (pat, epsps genes and insect (cry-genes resistance. The total DNA was extracted from maize living plant tissue. Primers to detect wx, fl-2, bar (pat, mepsps, CP4 epsps, cry1A(b, cry1F, cry1A.105, mcry3A, cry2Ab2, cry3Bb1, cry34Ab1, cry35Ab1 genes were designed and selected. Multiplex and Touchdown PCR were worked out. PCR amplification of certain sequences was carried out. No transgenes (bar (pat, mepsps, CP4 epsps, cry1A(b, cry1F, cry1A.105, mcry3A, cry2Ab2, cry3Bb1, cry34Ab1, cry35Ab1 were found among 200 analyzed experimental maize samples. At the same time, fl-2 gene was found in 41 samples, wx gene was found in 192 analyzed samples.

  2. Far-red fluorescence gene reporter tomography for determination of placement and viability of cell-based gene therapies.

    Science.gov (United States)

    Lu, Yujie; Darne, Chinmay D; Tan, I-Chih; Zhu, Banghe; Hall, Mary A; Lazard, Zawaunyka W; Davis, Alan R; Simpson, Lashan; Sevick-Muraca, Eva M; Olmsted-Davis, Elizabeth A

    2013-10-01

    Non-invasive injectable cellular therapeutic strategies based on sustained delivery of physiological levels of BMP-2 for spinal fusion are emerging as promising alternatives, which could provide sufficient fusion without the associated surgical risks. However, these injectable therapies are dependent on bone formation occurring only at the specific target region. In this study, we developed and deployed fluorescence gene reporter tomography (FGRT) to provide information on in vivo cell localization and viability. This information is sought to confirm the ideal placement of the materials with respect to the area where early bone reaction is required, ultimately providing three dimensional data about the future fusion. However, because almost all conventional fluorescence gene reporters require visible excitation wavelengths, current in vivo imaging of fluorescent proteins is limited by high tissue absorption and confounding autofluorescence. We previously administered fibroblasts engineered to produce BMP-2, but is difficult to determine 3-D information of placement prior to bone formation. Herein we used the far-red fluorescence gene reporter, IFP1.4 to report the position and viability of fibroblasts and developed 3-D tomography to provide placement information. A custom small animal, far-red fluorescence tomography system integrated into a commercial CT scanner was used to assess IFP1.4 fluorescence and to demark 3-D placement of encapsulated fibroblasts with respect to the vertebrae and early bone formation as assessed from CT. The results from three experiments showed that the placement of the materials within the spine could be detected. This work shows that in vivo fluorescence gene reporter tomography of cell-based gene therapy is feasible and could help guide cell-based therapies in preclinical models.

  3. Functional conservation of the fruitless male sex-determination gene across 250 Myr of insect evolution

    NARCIS (Netherlands)

    Gailey, Donald A; Billeter, Jean-Christophe; Liu, Jim H; Bauzon, Frederick; Allendorfer, Jane B; Goodwin, Stephen F

    2006-01-01

    Male sexual behavior in the fruit fly Drosophila melanogaster is regulated by fruitless (fru), a sex-determination gene specifying the synthesis of BTB-Zn finger proteins that likely function as male-specific transcriptional regulators. Expression of fru in the nervous system specifies male sexual b

  4. Functional conservation of the fruitless male sex-determination gene across 250 Myr of insect evolution

    NARCIS (Netherlands)

    Gailey, Donald A; Billeter, Jean-Christophe; Liu, Jim H; Bauzon, Frederick; Allendorfer, Jane B; Goodwin, Stephen F

    2006-01-01

    Male sexual behavior in the fruit fly Drosophila melanogaster is regulated by fruitless (fru), a sex-determination gene specifying the synthesis of BTB-Zn finger proteins that likely function as male-specific transcriptional regulators. Expression of fru in the nervous system specifies male sexual b

  5. Functional conservation of the fruitless male sex-determination gene across 250 Myr of insect evolution

    NARCIS (Netherlands)

    Gailey, Donald A; Billeter, Jean-Christophe; Liu, Jim H; Bauzon, Frederick; Allendorfer, Jane B; Goodwin, Stephen F

    Male sexual behavior in the fruit fly Drosophila melanogaster is regulated by fruitless (fru), a sex-determination gene specifying the synthesis of BTB-Zn finger proteins that likely function as male-specific transcriptional regulators. Expression of fru in the nervous system specifies male sexual

  6. CLONING AND SEQUENCING OF PSEUDOMONAS GENES DETERMINING SODIUM DODECYL-SULFATE BIODEGRADATION

    NARCIS (Netherlands)

    DAVISON, J; BRUNEL, F; PHANOPOULOS, A; PROZZI, D; TERPSTRA, P

    1992-01-01

    The nucleotide sequences of two genes involved in sodium dodecyl sulfate (SDS) degradation, by Pseudomonas, have been determined. One of these, sdsA, codes for an alkyl sulfatase (58 957 Da) and has similarity (31.8% identity over a 201-amino acid stretch) to the N terminus of a predicted protein of

  7. Oligogalacturonide-auxin antagonism does not require posttranscriptional gene silencing or stabilization of auxin response repressors in Arabidopsis.

    Science.gov (United States)

    Savatin, Daniel V; Ferrari, Simone; Sicilia, Francesca; De Lorenzo, Giulia

    2011-11-01

    α-1-4-Linked oligogalacturonides (OGs) derived from plant cell walls are a class of damage-associated molecular patterns and well-known elicitors of the plant immune response. Early transcript changes induced by OGs largely overlap those induced by flg22, a peptide derived from bacterial flagellin, a well-characterized microbe-associated molecular pattern, although responses diverge over time. OGs also regulate growth and development of plant cells and organs, due to an auxin-antagonistic activity. The molecular basis of this antagonism is still unknown. Here we show that, in Arabidopsis (Arabidopsis thaliana), OGs inhibit adventitious root formation induced by auxin in leaf explants as well as the expression of several auxin-responsive genes. Genetic, biochemical, and pharmacological experiments indicate that inhibition of auxin responses by OGs does not require ethylene, jasmonic acid, and salicylic acid signaling and is independent of RESPIRATORY BURST OXIDASE HOMOLOGUE D-mediated reactive oxygen species production. Free indole-3-acetic acid levels are not noticeably altered by OGs. Notably, OG- as well as flg22-auxin antagonism does not involve any of the following mechanisms: (1) stabilization of auxin-response repressors; (2) decreased levels of auxin receptor transcripts through the action of microRNAs. Our results suggest that OGs and flg22 antagonize auxin responses independently of Aux/Indole-3-Acetic Acid repressor stabilization and of posttranscriptional gene silencing.

  8. STABILITY INDICATING RP-LC METHOD FOR DETERMINATION OF RASAGILINE MESYLATE IN BULK AND PHARMACEUTICAL DOSAGE FORMS

    OpenAIRE

    R. Narendra Kumar; Nageswara Rao, G.; P.Y. Naidu

    2010-01-01

    An isocratic stability indicating liquid chromatographic method has been developed and validated for the determination of Rasagiline in bulk drug and its pharmaceutical dosage forms. Separation of the drug with degradation products was achieved using Puroshere Star, C18, 150 x 4.6mm; 5μm column as stationary phase and pH 7.0(±0.05) buffer: Acetonitrile (40:60,v/v) as mobile phase at a flow rate of 1.0 mL/min. UV detection was performed at 210 nm. The method is linear over the range of 4.8 – 1...

  9. Stability-indicating high-performance thin-layer chromatographic method for quantitative determination of omeprazole in capsule dosage form.

    Science.gov (United States)

    Jha, Preeta; Parveen, Rabea; Khan, Suroor A; Alam, Ozair; Ahmad, Sayeed

    2010-01-01

    A novel HPTLC method has been developed and validated for quantitative determination of omeprazole (OPZ) in capsule dosage form. The method was validated according to the International Conference on Harmonization guidelines for accuracy, precision, linearity, specificity, and robustness. HPTLC aluminum sheets precoated with silica gel 60F24 were used as the stationary phase and chloroform-methanol (9 + 1) as the mobile phase. The mobile phase was found to give compact bands for OPZ (Rf value of 0.39 +/- 0.12) in densitometric analysis in the absorbance mode at 302 nm. The linear regression analysis data for the calibration plots showed good linearity (r2 = 0.997) with respect to peak area in the concentration range 50-3000 ng/band. The mean values of the slope and intercept were 9.896 +/- 0.0753 and 1870.761 +/- 16.866, respectively. The method was also applied for stability testing of OPZ in different stress conditions and found to be accurate, linear, precise, robust, specific, and stability indicating. The method proposed can be used for QC and stability testing of different dosage forms such as tablets and capsules, as well as for bulk drug analysis of OPZ.

  10. Noxa determines localization and stability of MCL-1 and consequently ABT-737 sensitivity in small cell lung cancer.

    Science.gov (United States)

    Nakajima, W; Hicks, M A; Tanaka, N; Krystal, G W; Harada, H

    2014-02-13

    The sensitivity to ABT-737, a prototype BH3 mimetic drug, varies in a broad range in small cell lung cancer (SCLC) cells. We have previously shown that the expression of Noxa, a BH3-only pro-apoptotic BCL-2 family protein, is the critical determinant of ABT-737 sensitivity. We show here that Noxa regulates the localization and stability of MCL-1, an anti-apoptotic member, which results in modulating ABT-737 sensitivity. Mutations in Noxa within the BH3 domain, the carboxyl terminus mitochondrial targeting domain, or of ubiquitinated lysines not only change the localization and stability of Noxa itself but also affect the mitochondrial localization and phosphorylation/ubiquitination status of MCL-1 and consequently modulate sensitivity to ABT-737. Results of studies utilizing these mutant proteins indicate that Noxa recruits MCL-1 from the cytosol to the mitochondria. Translocation of MCL-1 initiates its phosphorylation and subsequent ubiquitination, which triggers proteasome-mediated degradation. The precise regulatory mechanisms of Noxa/MCL-1 expression and stability could provide alternative targets to modulate apoptosis induced by BH3 mimetic drugs or other chemotherapeutic reagents.

  11. Stability-Indicating HPLC-UV Method for Vitamin D3 Determination in Solutions, Nutritional Supplements and Pharmaceuticals.

    Science.gov (United States)

    Temova, Žane; Roškar, Robert

    2016-08-01

    A simple and fast high-performance liquid chromatography method with UV detection for determination of vitamin D3 in stability studies as well as in solutions, nutritional supplements and pharmaceuticals was developed. Successful separation of vitamin D3 from its degradation products was achieved on a Gemini C18 100 × 3.0 mm column using a mixture of acetonitrile and water (99:1, v/v) as а mobile phase. The method was successfully validated according to the ICH guidelines. The described reversed-phase HPLC method is favorable compared with other published HPLC-UV methods because of its stability-indicating nature, short run time (3.3 min) and wide analytical range with outstanding linearity, accuracy and precision. The method was further applied for quantification of vitamin D3 in selected liquid and solid nutritional supplements and prescription medicines, confirming its suitability for routine analysis. Degradation products, formed under stress conditions (hydrolysis, oxidation, photolysis and thermal degradation), were additionally elucidated by suitable equipment (LC-DAD-MS) to confirm the stability-indicating nature of the developed method.

  12. Communication: Transition state trajectory stability determines barrier crossing rates in chemical reactions induced by time-dependent oscillating fields.

    Science.gov (United States)

    Craven, Galen T; Bartsch, Thomas; Hernandez, Rigoberto

    2014-07-28

    When a chemical reaction is driven by an external field, the transition state that the system must pass through as it changes from reactant to product--for example, an energy barrier--becomes time-dependent. We show that for periodic forcing the rate of barrier crossing can be determined through stability analysis of the non-autonomous transition state. Specifically, strong agreement is observed between the difference in the Floquet exponents describing stability of the transition state trajectory, which defines a recrossing-free dividing surface [G. T. Craven, T. Bartsch, and R. Hernandez, "Persistence of transition state structure in chemical reactions driven by fields oscillating in time," Phys. Rev. E 89, 040801(R) (2014)], and the rates calculated by simulation of ensembles of trajectories. This result opens the possibility to extract rates directly from the intrinsic stability of the transition state, even when it is time-dependent, without requiring a numerically expensive simulation of the long-time dynamics of a large ensemble of trajectories.

  13. Glutamine synthetase stabilizes the binding of GlnR to nitrogen fixation gene operators.

    Science.gov (United States)

    Fernandes, Gabriela de C; Hauf, Ksenia; Sant'Anna, Fernando H; Forchhammer, Karl; Passaglia, Luciane M P

    2017-03-01

    Biological nitrogen fixation (BNF) is a high energy demanding process carried out by diazotrophic microorganisms that supply combined nitrogen to the biosphere. The genes related to BNF are strictly regulated, but these mechanisms are poorly understood in gram-positive bacteria. The transcription factor GlnR was proposed to regulate nitrogen fixation-related genes based on Paenibacillus comparative genomics. In order to validate this proposal, we investigated BNF regulatory sequences in Paenibacillus riograndensis SBR5(T) genome. We identified GlnR-binding sites flanking σ(A) -binding sites upstream from BNF-related genes. GlnR binding to these sites was demonstrated by surface plasmon resonance spectroscopy. GlnR-DNA affinity is greatly enhanced when GlnR is in complex with feedback-inhibited (glutamine-occupied) glutamine synthetase (GS). GlnR-GS complex formation is also modulated by ATP and AMP. Thereby, gene repression exerted by the GlnR-GS complex is coupled with nitrogen (glutamine levels) and energetic status (ATP and AMP). Finally, we propose a DNA-looping model based on multiple operator sites that represents a strong and strict regulation for these genes. © 2017 Federation of European Biochemical Societies.

  14. Stability of Barley stripe mosaic virus induced gene silencing in barley

    DEFF Research Database (Denmark)

    Bruun-Rasmussen, Marianne; Madsen, Christian Toft; Jessing, Stine

    2007-01-01

    Virus-induced gene silencing (VIGS) can be used as a powerful tool for functional genomics studies in plants. With this approach, it is possible to target most genes and downregulate the messenger (m)RNA in a sequence-specific manner. Barley stripe mosaic virus (BSMV) is an established VIGS vector...... for barley and wheat; however, silencing using this vector is generally transient, with efficient silencing often being confined to the first two or three systemically infected leaves. To investigate this further, part of the barley Phytoene desaturase (PDS) gene was inserted into BSMV and the resulting...... inoculation, although large parts of the insert had been lost from the virus vector. The instability of the insert, observed consistently throughout our experiments, offers an explanation for the transient nature of silencing when using BSMV as a VIGS vector....

  15. Determination of Algae and Macrophyte Species Distribution in Three Wastewater Stabilization Ponds Using Metagenomics Analysis

    Directory of Open Access Journals (Sweden)

    Jack Wallace

    2015-06-01

    Full Text Available This study involved the evaluation of algae and macrophyte species distributions in three wastewater stabilization ponds (WSPs at a wastewater treatment plant in Ontario, Canada, which has experienced high pH levels at the final effluent and excessive algae growth during the summer since 2003. From samples collected from the system, the relative abundances of specific algae and aquatic plant (macrophyte taxa were assessed and correlated to water chemistry data. A strong shift from the dominance of green algae, chlorophyceae, in WSP#2, to the dominance of aquatic macrophytes, embryophyta, in WSP#4, was observed and corresponded to field observations. Correlation of the abundances to nutrient parameters suggested that the macronutrient rich conditions in WSP#2 allowed floating green algae to proliferate against macrophytes. In WSP#1 and WSP#4, macrophytes competed against algae and thrived, due to their adaptability to lower nutrient conditions. The pH increases occurred primarily in WSP#2 and were not buffered or reduced in WSP#1 and WSP#4. Two alternatives strategies for pH control were recommended for the system: decreasing algae growth in WSP#2 through duckweed seeding or macronutrient loading reduction; or designing and implementing a constructed wetland (CW in WSP#4 with soil and vegetation to buffer pH prior to release.

  16. Role of terrestrial ecosystems in determining CO{sub 2} stabilization and recovery behaviour

    Energy Technology Data Exchange (ETDEWEB)

    Jones, Chris; Liddicoat, Spencer (Met Office Hadley Centre, Exeter (United Kingdom)); Lowe, Jason (Met Office Hadley Centre, Dept. of Meteorology, Univ. of Reading, Reading (United Kingdom))

    2010-11-15

    Terrestrial ecosystems are sensitive to climate and can also influence it through both biophysical and biogeochemical feedbacks. Natural carbon uptake by ecosystems will control future evolution of CO{sub 2} and climate, but the ecosystems themselves may be committed to long-term changes. Here we use a coupled climate-carbon cycle GCM with dynamic vegetation to investigate the policy-relevance of these feedbacks in several idealized scenarios. Our results show that the natural carbon cycle in the ocean and on land controls the recovery of atmospheric CO{sub 2} following emissions reductions at three action points during the 21st century. Initial rates of recovery are similar but for different reasons. Ocean carbon uptake exceeds terrestrial uptake, with higher CO{sub 2} levels leading to increased ocean uptake whereas on land greater climate change at higher CO{sub 2} leads to decreased carbon storage. There are long-term committed changes to terrestrial ecosystems which vary in sign regionally and create a complex dynamic response of terrestrial carbon storage as it slowly approaches a new steady state. Neither stabilization nor CO{sub 2} recovery allows ecosystems to recover back to their initial state and the ecosystems continue to respond for decades or even centuries following emissions reductions. These long-term committed changes, in addition to realized, transient changes, must be considered when defining dangerous climate change and identifying emission-pathways to avoid it.

  17. Physical determinants of bipolar mitotic spindle assembly and stability in fission yeast

    Science.gov (United States)

    Blackwell, Robert; Edelmaier, Christopher; Sweezy-Schindler, Oliver; Lamson, Adam; Gergely, Zachary R.; O’Toole, Eileen; Crapo, Ammon; Hough, Loren E.; McIntosh, J. Richard; Glaser, Matthew A.; Betterton, Meredith D.

    2017-01-01

    Mitotic spindles use an elegant bipolar architecture to segregate duplicated chromosomes with high fidelity. Bipolar spindles form from a monopolar initial condition; this is the most fundamental construction problem that the spindle must solve. Microtubules, motors, and cross-linkers are important for bipolarity, but the mechanisms necessary and sufficient for spindle assembly remain unknown. We describe a physical model that exhibits de novo bipolar spindle formation. We began with physical properties of fission-yeast spindle pole body size and microtubule number, kinesin-5 motors, kinesin-14 motors, and passive cross-linkers. Our model results agree quantitatively with our experiments in fission yeast, thereby establishing a minimal system with which to interrogate collective self-assembly. By varying the features of our model, we identify a set of functions essential for the generation and stability of spindle bipolarity. When kinesin-5 motors are present, their bidirectionality is essential, but spindles can form in the presence of passive cross-linkers alone. We also identify characteristic failed states of spindle assembly—the persistent monopole, X spindle, separated asters, and short spindle, which are avoided by the creation and maintenance of antiparallel microtubule overlaps. Our model can guide the identification of new, multifaceted strategies to induce mitotic catastrophes; these would constitute novel strategies for cancer chemotherapy. PMID:28116355

  18. Role of In vivo reflectance confocal microscopy in determining stability in vitiligo: A preliminary study

    Directory of Open Access Journals (Sweden)

    Wei LI

    2013-01-01

    Full Text Available Background: Vitiligo is an acquired pigmentary disorder. In vivo reflectance confocal microscopy (RCM reproducible imaging technique has already been reported to be useful in the diagnosis of other skin diseases. Objective: To define RCM features of vitiligo on different clinical stages. Materials and Methods: A total of 125 patients with a clinical diagnosis of vitiligo were included in this study. After informed consent, lesional skins of those vitiligo patients were characterized by using RCM. Five patients with inflammatory cell infiltration observed at the edge of skin lesions and another 5 patients without inflammatory cell infiltration were selected. Biopsies were performed at same sites of the RCM examination areas for histological and immune-histological analysis. Results: In the active stage of vitiligo, the RCM examination revealed that the bright dermal papillary rings presented at the dermoepidermal junction level in normal skin lost their integrity or totally disappeared, border between vitiligo lesion and normal skin became unclear, and highly refractile cells that referred to infiltrated inflammatory cells could be seen within the papillary dermis at the edge of the lesions. In the stable stage of vitiligo, the RCM showed a complete loss of melanin in lesional skin and a clear border between lesional and normal skin. Conclusion: A simple clinical examination with RCM may reliably and efficiently allow evaluation of the stability status of vitiligo lesions.

  19. Determination of genetic stability in long-term somatic embryogenic cultures and derived plantlets of cork oak using microsatellite markers.

    Science.gov (United States)

    Lopes, Tina; Pinto, Glória; Loureiro, João; Costa, Armando; Santos, Conceição

    2006-09-01

    Microsatellites were used to test genetic stability in somatic embryos (SE) of Quercus suber L. The SE were obtained by a simple somatic embryogenesis protocol: leaf explants from two adult plants (QsG0, QsG5) and from two juvenile plants (QsGM1, QsGM2) were inoculated on Murashige and Skoog (MS) medium with 2,4-dichlorophenoxyacetic acid and zeatin. Calluses with primary embryogenic structures were transferred to MSWH (MS medium without growth regulators) and SE proliferated by secondary somatic embryogenesis. High morphological heterogeneity was found among cotyledonary SE. However, converted plants looked morphologically normal with well-developed rooting systems and shoots. The genetic stability of the plant material during the somatic embryogenesis process was evaluated by using six to eight nuclear microsatellites transferred from Q. myrsinifolia Blume, Q. petraea (Matts.) Liebl. and Q. robur L. Five of eight microsatellites distinguished among the genotypes analyzed, and for QsG0, QsGM1 and QsGM2, uniform microsatellite patterns were generally observed within and between SE and the respective donor genotypes. For genotype QsG5, the same pattern was observed in all samples analyzed except one, where the mutation percentage was 2.5%. We conclude that microsatellite markers can be used to assess genetic stability of clonal materials and to determine genetic stability throughout the process of somatic embryogenesis. The simple somatic embryogenesis protocol described has potential for the commercial propagation of Q. suber because it results in a low percentage of mutations.

  20. Polarographic Determination of Composition and Thermodynamic Stability Constant of a Complex Metal Ion.

    Science.gov (United States)

    Marin, Dolores; Mendicuti, Francisco

    1988-01-01

    Describes a laboratory experiment designed to encourage laboratory cooperation among individual undergraduate students or groups. Notes each student contributes results individually and the exchange of data is essential to obtain final results. Uses the polarographic method for determining complex metal ions. (MVL)

  1. Determine the need to research the time-related stability decay of bord and pillar systems

    CSIR Research Space (South Africa)

    Oberholzer, JW

    1997-07-01

    Full Text Available This report summarizes the findings of the work conducted to determine the need to research the time dependent decay of coal mine bord and pillar workings. It is intended for use by the relevant SIMRAC committee as background information to assist...

  2. Immediate postarousal sleep dynamics: an important determinant of sleep stability in obstructive sleep apnea.

    Science.gov (United States)

    Younes, Magdy; Hanly, Patrick J

    2016-04-01

    Arousability from sleep is increasingly recognized as an important determinant of the clinical spectrum of sleep disordered breathing (SDB). Patients with SDB display a wide range of arousability. The reason for these differences is not known. We hypothesized that differences in the speed with which sleep deepens following arousals/awakenings (postarousal sleep dynamics) is a major determinant of these differences in arousability in patients with SDB. We analyzed 40 preexisting clinical polysomnography records from patients with a range of SDB severity (apnea-hypopnea index 5-135/h). Sleep depth was determined every 3 s using the odds ratio product (ORP) method, a continuous index of sleep depth (0 = deep sleep, 2.5 = full wakefulness) that correlates strongly (r = 0.98) with arousability (Younes M, Ostrowski M, Soiferman M, Younes H, Younes M, Raneri J, and Hanly P. Sleep 38: 641-654, 2015). Time course of ORP was determined from end of arousal until the next arousal. All arousals were analyzed (142 ± 65/polysomnogram). ORP increased from 0.58 ± 0.32 during sleep to 1.67 ± 0.35 during arousals. ORP immediately (first 9 s) following arousals/awakenings (ORP-9) ranged from 0.21(very deep sleep) to 1.71 (highly arousable state) in different patients. In patients with high ORP-9, sleep deepened slowly (over minutes) beyond 9 s but only if no arousals/awakenings recurred. ORP-9 correlated strongly with average non-rapid eye movement sleep depth (r = 0.87, P sleep architecture. We conclude that postarousal sleep dynamics are highly variable among patients with sleep-disordered breathing and largely determine average sleep depth and continuity.

  3. Developmental expression of "germline"- and "sex determination"-related genes in the ctenophore Mnemiopsis leidyi.

    Science.gov (United States)

    Reitzel, Adam M; Pang, Kevin; Martindale, Mark Q

    2016-01-01

    An essential developmental pathway in sexually reproducing animals is the specification of germ cells and the differentiation of mature gametes, sperm and oocytes. The "germline" genes vasa, nanos and piwi are commonly identified in primordial germ cells, suggesting a molecular signature for the germline throughout animals. However, these genes are also expressed in a diverse set of somatic stem cells throughout the animal kingdom leaving open significant questions for whether they are required for germline specification. Similarly, members of the Dmrt gene family are essential components regulating sex determination and differentiation in bilaterian animals, but the functions of these transcription factors, including potential roles in sex determination, in early diverging animals remain unknown. The phylogenetic position of ctenophores and the genome sequence of the lobate Mnemiopsis leidyi motivated us to determine the compliment of these gene families in this species and determine expression patterns during development. Our phylogenetic analyses of the vasa, piwi and nanos gene families show that Mnemiopsis has multiple genes in each family with multiple lineage-specific paralogs. Expression domains of Mnemiopsis nanos, vasa and piwi, during embryogenesis from fertilization to the cydippid stage, were diverse, with little overlapping expression and no or little expression in what we think are the germ cells or gametogenic regions. piwi paralogs in Mnemiopsis had distinct expression domains in the ectoderm during development. We observed overlapping expression domains in the apical organ and tentacle apparatus of the cydippid for a subset of "germline genes," which are areas of high cell proliferation, suggesting that these genes are involved with "stem cell" specification and maintenance. Similarly, the five Dmrt genes show diverse non-overlapping expression domains, with no clear evidence for expression in future gametogenic regions of the adult. We also

  4. Development and validation of stability indicating method for determination of sertraline following ICH guidlines and its determination in pharmaceuticals and biological fluids

    Directory of Open Access Journals (Sweden)

    El-Enany Nahed M

    2011-10-01

    Full Text Available Abstract Background Sertraline is a well known antidepressant drug which belongs to a class called selective serotonin reuptake inhibitor. Most published methods do not enable studying the stability of this drug in different stress conditions. Results Two new methods were developed for the determination of sertraline (SER. Both methods are based on coupling with 4-chloro-7-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl in borate buffer of pH 7.8 and measuring the reaction product spectrophotometrically at 395 nm (Method I or spectrofluorimetrically at 530 nm upon excitation at 480 nm (Method II. The response-concentration plots were rectilinear over the range 2-24 μg/mL and 0.25-5 μg/mL for methods I and II respectively with LOD of 0.18 μg/mL and 0.07 μg/mL, and LOQ of 0.56 μg/mL and 0.21 μg/mL for methods I and II, respectively. Conclusion Both methods were applied to the analysis of commercial tablets and the results were in good agreement with those obtained using a reference method. The fluorimetric method was further applied to the in vivo determination of SER in human plasma. A proposal of the reaction pathway was presented. The spectrophotometric method was extended to stability study of SER. The drug was exposed to alkaline, acidic, oxidative and photolytic degradation according to ICH guidelines. Moreover, the method was utilized to investigate the kinetics of oxidative degradation of the drug. The apparent first order rate constant and t1/2 of the degradation reaction were determined.

  5. Transcriptional rewiring of the sex determining dmrt1 gene duplicate by transposable elements.

    Directory of Open Access Journals (Sweden)

    Amaury Herpin

    2010-02-01

    Full Text Available Control and coordination of eukaryotic gene expression rely on transcriptional and posttranscriptional regulatory networks. Evolutionary innovations and adaptations often require rapid changes of such networks. It has long been hypothesized that transposable elements (TE might contribute to the rewiring of regulatory interactions. More recently it emerged that TEs might bring in ready-to-use transcription factor binding sites to create alterations to the promoters by which they were captured. A process where the gene regulatory architecture is of remarkable plasticity is sex determination. While the more downstream components of the sex determination cascades are evolutionary conserved, the master regulators can switch between groups of organisms even on the interspecies level or between populations. In the medaka fish (Oryzias latipes a duplicated copy of dmrt1, designated dmrt1bY or DMY, on the Y chromosome was shown to be the master regulator of male development, similar to Sry in mammals. We found that the dmrt1bY gene has acquired a new feedback downregulation of its expression. Additionally, the autosomal dmrt1a gene is also able to regulate transcription of its duplicated paralog by binding to a unique target Dmrt1 site nested within the dmrt1bY proximal promoter region. We could trace back this novel regulatory element to a highly conserved sequence within a new type of TE that inserted into the upstream region of dmrt1bY shortly after the duplication event. Our data provide functional evidence for a role of TEs in transcriptional network rewiring for sub- and/or neo-functionalization of duplicated genes. In the particular case of dmrt1bY, this contributed to create new hierarchies of sex-determining genes.

  6. A validated stability-indicating liquid chromatographic method for the determination of retapamulin in topical dosage form.

    Science.gov (United States)

    Nalwade, Santaji; Reddy, Vangala Ranga

    2014-03-01

    A sensitive, stability-indicating reversed-phase high-performance liquid chromatographic method is developed and validated for the quantitative determination of retapamulin in topical dosage form. The chromatographic separation is achieved by using a C18 column (XTerra RP 18 250 × 4.6 mm, 5 µm) at 30°C. The mobile phase comprises a mixture of 0.05M potassium dihydrogen phosphate buffer (pH 6.1), acetonitrile and methanol in the ratio of 35:50:15 (v/v/v). The flow rate is set at 1.0 mL/min and chromatograms are extracted at 243 nm using a photodiode array detector. The method is validated with respect to linearity, accuracy, precision, robustness and forced degradation studies, which further prove the stability-indicating supremacy of the method. During forced degradation studies, retapamulin is observed to be labile to oxidative and base hydrolysis stress and stable in thermal, photolytic and acid hydrolysis stress. The degradation products are well separated from the retapamulin peak, thus proving the stability-indicating superiority of the method. The method is found to be sensitive for retapamulin, with a detection limit of 25 ng/mL and a quantification limit of 80 ng/mL. The proposed method is found to be very sensitive and accurate for the determination of retapamulin in topical dosage form. The method is also demonstrated to be robust, because it is resistant to small variations of chromatographic variables such as pH, mobile phase composition, flow rate and column temperature.

  7. Evaluation of the stability of fluoxetine in pluronic lecithin organogel and the determination of an appropriate beyond-use date.

    Science.gov (United States)

    Peacock, Gina F; Sauvageot, Jurgita

    2014-01-01

    Fluoxetine is a commonly prescribed psychotropic medication for a variety of behavioral diagnoses in veterinary practice, and fluoxetine in Pluronic lecithin organogel has been used successfully in treating inappropriate urine spraying in felines. Historically, pharmacists have assigned a variety of beyond-use dates to extemporaneously compound drugs in Pluronic lecithin organogel. The objective of this study was to evaluate the stability of fluoxetine in Pluronic lecithin organogel over a period of six months and to determine an appropriate beyond-use date. A stability-indicating high-performance liquid chromatography method for fluoxetine in Pluronic lecithin organogel was validated in our laboratory. Fluoxetine-Pluronic lecithin organogel 50 mg/mL was prepared by a local compounding pharmacy and analyzed by high-performance liquid chromatograph at 0, 7, 14, 21, 28, 45, 60, 90, and 180 days. Physical stability was also assessed by visual observation. At each time point percent of initial concentration was calculated. The beyond-use date was determined as the time period that the samples maintained at least 90 percent of the initial concentration. At 180 days, the mean percent of initial concentration was 99 +/- 1.5 and, visually, the fluoxetine-Pluronic lecithin organogel retained the original color and consistency, without detectable separation of the different phases of Pluronic lecithin organogel. Since fluoxetine was physically stable and retained greater than 90 percent of initial concentration in Pluronic lecithin organogel for 180 days when stored at room temperature and protected from light, a beyond-use date of 180 days is appropriate.

  8. Determining suitable lego-structures to estimate stability of larger peptide nanostructures using computational methods.

    Science.gov (United States)

    Beke, Tamás; Czajlik, András; Csizmadia, Imre G; Perczel, András

    2006-02-02

    Nanofibers, nanofilms and nanotubes constructed of one to four strands of oligo-alpha- and oligo-beta-peptides were obtained by using carefully selected building units. Lego-type approaches based on thermoneutral isodesmic reactions can be used to reconstruct the total energies of both linear and tubular periodic nanostructures with acceptable accuracy. Total energies of several different nanostructures were accurately determined with errors typically falling in the subchemical range. Thus, attention will be focused on the description of suitable isodesmic reactions that have enabled the determination of the total energy of polypeptides and therefore offer a very fast, efficient and accurate method to obtain energetic information on large and even very large nanosystems.

  9. Development and validation of stability-indicating HPLC method for simultaneous determination of meropenem and potassium clavulanate.

    Science.gov (United States)

    Zalewski, Przemysław; Cielecka-Piontek, Judyta; Paczkowska, Magdalena

    2014-01-01

    A stability-indicating LC assay method was developed and validated for a simultaneous determination of meropenem and potassium clavulanate in the presence of degradation products formed during acid-base hydrolysis, oxidation and thermolysis. The isocratic RP-HPLC method was developed with a LiChrospher RP-18 (250 mm x 4.6 mm, 5 microm) column and gradient elution of 12 mmol/L ammonium acetate and acetonitrile. The flow rate of the mobile phase was 1.0 mL/min, the detection wavelength 220 nm and the temperature 303 K. The method was validated with regard to linearity, accuracy, precision, selectivity and robustness, and was applied successfully for the determination of meropenem and potassium clavulanate separately as well as jointly in pharmaceutical formulations.

  10. DEVELOPMENT AND VALIDATION OF THE STABILITY-INDICATING LC-UV METHOD FOR DETERMINATION OF CEFOZOPRAN HYDROCHLORIDE.

    Science.gov (United States)

    Zalewski, Przemysław; Garbacki, Piotr; Cielecka-Piontek, Judyta; Bednarek-Rajewska, Katarzyna; Krause, Anna

    2015-01-01

    The stability-indicating LC assay method was developed and validated for quantitative determination of cefozopran hydrochloride (CZH) in the presence of degradation products formed during the forced degradation studies. An isocratic, RP-HPLC method was developed with C-18 (250 mm x 4.6 mm, 5 µm) column and 12 mM ammonium acetate-acetonitrile (92:8, v/v) as a mobile phase. The flow rate of the mobile phase was 1.0 mL/min. Detection wavelength was 260 not and temperature was 30°C. Cefozopran hydrochloride as other cephalosporins was subjected to stress conditions of degradation in aqueous solutions including hydrolysis, oxidation, photolysis and thermal degradation. The developed method was validated with regard to linearity, accuracy, precision, selectivity and robustness. The method was applied successfully for identification and determination of cefozopran hydrochloride in pharmaceuticals and during kinetic studies.

  11. Influence of the Rancimat parameters on the determination of oxidative stability index of Sesamum Indicum L. Oil

    Directory of Open Access Journals (Sweden)

    Eudes Villanueva López

    2013-09-01

    Full Text Available The objective of this research was to determine the oxidative stability index (OSI in virgin oil seed Sesamum indicum L. (Sesame previously extracted by cold pressing, clarified by centrifugation and stored under nitrogen atmosphere and cooling on. The OSI was determined by accelerated Rancimat test, it was used for 3.0 ± 0.1 g of sample temperature parameters (110, 130 and 150 °C and air flow (15, 20 and 25 L/h. Applying the Rancimat test, it was found by ANOVA (p < 0.05 that the influence of temperature on the OSI was highly significant, whereas the only air flow was significant. By extrapolation method, values were calculated at usual temperatures OSI oil storage (25 °C to give stability times 214, 242 and 222 days, also the activation energy of the oxidation reaction of sesame oil for different air flows, is 97.28, 98.79 and 96.86 kJ / mol for 15, 20 and 25 L/h respectively.

  12. Physical and chemical properties and stability of sodium cefazolin in buffered eye drops determined with HPLC method.

    Science.gov (United States)

    Kodym, Anna; Bilski, Piotr; Domańska, Agata; Hełminiak, Łukasz; Jabłońska, Maria; Jachymska, Anna

    2012-01-01

    The aim of the studies was to analyze the stability of 1% and 5% eye drops containing sodium cefazolin, prepared in citrate buffer of pH 6.11-6.27, which were stored at the temperature of 4 degrees C and 20 degrees C with light protection. The drops were prepared under aseptic conditions by dissolving sodium cefazolin (Biofazolin, IBA Bioton), dry injection form of the drug, in citrate buffer. The viscosity of the drops was increased using polyvinyl alcohol. The drops were preserved with phenylmercuric borate of 0.001% concentration mixed with beta-phenylethyl alcohol of 0.4% concentration in the drops. The concentration of cefazolin was determined at every three days using HPLC method. Besides, the measurements of pH, osmotic pressure and viscosity were performed as well as the organoleptic analysis of the drops (clarity, color, odor). The concentration of cefazolin in 1% drops after the 30-day-storage at the temperature of 4 degrees C, depending on their composition, decreased in the range of 2.17-6.02%. In 5% drops the decrease in cefazolin concentration was similar, i.e., after 30-day-storage at the temperature of 4 degrees C it was 1.62-6.76%. In 1% and 5% drops stored at the temperature of 20 degrees C the stability of the drops determined as the 10% degradation time of cefazolin was in the range of 9-15 days.

  13. Ultraviolet absorbance titration for the determination of conditional stability constants of Hg(II) and dissolved organic matter

    Institute of Scientific and Technical Information of China (English)

    BAI Yingchen; WU Fengchang; WAN Guojiang; LIU Congqiang; FU Pingqing; LI Wen

    2008-01-01

    Strong interaction between natural dissolved organic matter (DOM) and Hg(II) may influence the transport, conversion, toxicity and bio-validity of mercury in the environment. In this paper ultraviolet (UV) absorbance titration was employed for the first time for the determination of the conditional stability constants of Hg(II) and (DOM). With increasing Hg(II) concentrations, the UV absorbance of fulvic acid, humic acid, and DOM in river increases progressively. By linear and non-linear model fitting, the conditional stability constants (lgK) of Hg(II) and DOM were worked out to be 3.54-4.93 and 3.64-4.85, respectively. The results are consistent with those acquired by the typical fluorescence quenching titration method, with the maximum relative error of lgK being 2.6% and the average relative error being 0.2%. The UV absorbance titration method has the advantages of rapid determination, simple performance, and it will probably become a new approach to studying interactions between DOM and trace metallic ions.

  14. Evaluation of the stability of ketoprofen in pluronic lecithin organogel and the determination of an appropriate beyond-use date.

    Science.gov (United States)

    Peacock, Gina F; Sauvageot, Jurgita; Addo, Richard T

    2014-01-01

    Previous reports indicate that pharmacists are assigning a wide variety of beyond-use dates to extemporaneously compounded medications in topical Pluronic lecithin organogel. The objective of this study was to evaluate the stability of ketoprofen in Pluronic lecithin organogel over a period of six months and to determine an appropriate beyond-use date for this formulation. A stability-indicating high-performance liquid chromatography method for ketoprofen in Pluronic lecithin organogel was validated in our laboratory. Samples of the formulation were analyzed by high-performance liquid chromatography at 0, 7, 14, 21, 28, 45, 60, 90, and 180 days. At each time point, the average concentration and average percent of initial concentration were calculated. The beyond-use date was determined as the time period that the samples were physically stable and maintained at least 90% of the initial concentration. Ketoprofen in Pluronic lecithin organogel was chemically and physically stable for six months when stored at room temperature and protected from light. Therefore, a beyond-use date of six months is appropriate for this preparation.

  15. Potentiometric titration for determining the composition and stability of metal(II) alginates and pectinates in aqueous solutions

    Science.gov (United States)

    Kaisheva, N. Sh.; Kaishev, A. Sh.

    2015-07-01

    The compositions and stabilities of Cu2+, Mn2+, Pb2+, Ca2+, Zn2+, Cd2+, Co2+, and Ni2+ alginates and pectinates are determined in aqueous solutions via titrimetry and potentiometry with calculations performed using Bjerrum's method, the curve intersection technique, and the equilibrium shift method. It is found that the interaction between Cu2+ and polyuronides is a stepwise process and, depending on the ligand concentration and the method of determination, Cu2+ alginate can be characterized by its ML, ML2, and ML3 compositions (where M is the metal ion and L is the structural unit of polyuronide) and stability constants logβ = 2.65, 5.00-5.70, and 7.18-7.80, respectively. The compositions of Cu2+ pectinates are ML and ML2 with logβ = 3.00 and 7.64-7.94, respectively. It is concluded that Pb2+, Ca2+, Mn2+, Zn2+, Cd2+, Co2+, and Ni2+ ions form only alginates and pectinates of ML2 composition with logβ values of 3.45 (Pb2+ alginate), 2.20 (Ca2+ alginate), 1.06 (Mn2+ alginate), 3.51 (Pb2+ pectinate), 2.35 (Ca2+ pectinate), and 1.24 (Mn2+ pectinate). The pectinates are shown to be more stable than the alginates, the most stable compounds being those formed by polyuronides and Cu2+. The least stable are those with Mn2+.

  16. An approach to determining the local boundaries of voltage stability region with wind farms in power injection space

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    A new approach to determining the local boundaries of voltage stability region in power injection space (IVSR) for power system with wind farms is presented. It can be used for power system voltage stability analysis and on-line security assessment with considering the output uncertainty of the wind farms. Firstly, power flow tracing and double dispatching are used to determine the generators that are closely related to the wind farms, in order to balance the power fluctuations caused by the wind speed variation. Then, modal analysis is used to get the key generators to achieve an effective dimension reduction for IVSR. Finally, the forecasting output power (or wind speed) of wind farms is divided into several intervals. For each interval, the corresponding local IVSR boundaries can be calculated by the method based on small disturbance. Moreover, parallel process is used to accelerate the computing speed. The presented approach is validated by several power systems. It can be revealed that the approach can give the local IVSR boundaries at different wind speeds and has a good engineering application prospect.

  17. Thermal stability of photovoltaic a-Si:H determined by neutron reflectometry

    Energy Technology Data Exchange (ETDEWEB)

    Qviller, A. J., E-mail: atlejq@ife.no; Haug, H.; You, C. C.; Hasle, I. M.; Marstein, E. S.; Frommen, C.; Hauback, B. C. [Institute for Energy Technology, P.O. Box 40, NO-2027 Kjeller (Norway); Dennison, A. J. C.; Vorobiev, A. [Division for Materials Physics, Department of Physics and Astronomy, Uppsala University, Box 516, SE-751 20 Uppsala (Sweden); Institut Laue-Langevin, 71 Avenue des Martyrs, 38000 Grenoble (France); Østreng, E.; Fjellvåg, H. [Department of Chemistry, Centre for Materials Science and Nanotechnology, University of Oslo, P.O. Box 1033, Blindern, NO-0315 Oslo (Norway); Hjörvarsson, B. [Division for Materials Physics, Department of Physics and Astronomy, Uppsala University, Box 516, SE-751 20 Uppsala (Sweden)

    2014-12-08

    Neutron and X-ray reflectometry were used to determine the layer structure and hydrogen content of thin films of amorphous silicon (a-Si:H) deposited onto crystalline silicon (Si) wafers for surface passivation in solar cells. The combination of these two reflectometry techniques is well suited for non-destructive probing of the structure of a-Si:H due to being able to probe buried interfaces and having sub-nanometer resolution. Neutron reflectometry is also unique in its ability to allow determination of density gradients of light elements such as hydrogen (H). The neutron scattering contrast between Si and H is strong, making it possible to determine the H concentration in the deposited a-Si:H. In order to correlate the surface passivation properties supplied by the a-Si:H thin films, as quantified by obtainable effective minority carrier lifetime, photoconductance measurements were also performed. It is shown that the minority carrier lifetime falls sharply when H has been desorbed from a-Si:H by annealing.

  18. Ratio manipulating spectrophotometry versus chemometry as stability indicating methods for cefquinome sulfate determination.

    Science.gov (United States)

    Yehia, Ali M; Arafa, Reham M; Abbas, Samah S; Amer, Sawsan M

    2016-01-15

    Spectral resolution of cefquinome sulfate (CFQ) in the presence of its degradation products was studied. Three selective, accurate and rapid spectrophotometric methods were performed for the determination of CFQ in the presence of either its hydrolytic, oxidative or photo-degradation products. The proposed ratio difference, derivative ratio and mean centering are ratio manipulating spectrophotometric methods that were satisfactorily applied for selective determination of CFQ within linear range of 5.0-40.0 μg mL(-1). Concentration Residuals Augmented Classical Least Squares was applied and evaluated for the determination of the cited drug in the presence of its all degradation products. Traditional Partial Least Squares regression was also applied and benchmarked against the proposed advanced multivariate calibration. Experimentally designed 25 synthetic mixtures of three factors at five levels were used to calibrate and validate the multivariate models. Advanced chemometrics succeeded in quantitative and qualitative analyses of CFQ along with its hydrolytic, oxidative and photo-degradation products. The proposed methods were applied successfully for different pharmaceutical formulations analyses. These developed methods were simple and cost-effective compared with the manufacturer's RP-HPLC method.

  19. Ratio manipulating spectrophotometry versus chemometry as stability indicating methods for cefquinome sulfate determination

    Science.gov (United States)

    Yehia, Ali M.; Arafa, Reham M.; Abbas, Samah S.; Amer, Sawsan M.

    2016-01-01

    Spectral resolution of cefquinome sulfate (CFQ) in the presence of its degradation products was studied. Three selective, accurate and rapid spectrophotometric methods were performed for the determination of CFQ in the presence of either its hydrolytic, oxidative or photo-degradation products. The proposed ratio difference, derivative ratio and mean centering are ratio manipulating spectrophotometric methods that were satisfactorily applied for selective determination of CFQ within linear range of 5.0-40.0 μg mL- 1. Concentration Residuals Augmented Classical Least Squares was applied and evaluated for the determination of the cited drug in the presence of its all degradation products. Traditional Partial Least Squares regression was also applied and benchmarked against the proposed advanced multivariate calibration. Experimentally designed 25 synthetic mixtures of three factors at five levels were used to calibrate and validate the multivariate models. Advanced chemometrics succeeded in quantitative and qualitative analyses of CFQ along with its hydrolytic, oxidative and photo-degradation products. The proposed methods were applied successfully for different pharmaceutical formulations analyses. These developed methods were simple and cost-effective compared with the manufacturer's RP-HPLC method.

  20. Development and validation of stability indicating TLC densitometric and spectrophotometric methods for determination of Clobetasol propionate

    Directory of Open Access Journals (Sweden)

    Yasmine Farouk Bassuoni

    2016-12-01

    Full Text Available Two simple analytical techniques that manipulate the inherent spectroscopic properties of the drug differently were developed for Clobetasol propionate (CP determination in the presence of its alkaline hydrolytic degradation products. The first method depends on TLC-densitometric determination of the UV–visualized bands after TLC separation of CP in the presence of its alkaline degradation products in its bulk and pharmaceutical formulation. Separation was performed on preactivated silica gel 60 F254 TLC plates using ethyl acetate:hexane:ammonia (5:5:0.2, by volume as a developing system followed by scanning at 240 nm. Linear correlation was obtained in the range of 0.10–0.50 μg/band. The second method was ratio difference spectrophotometry. It was applied by measuring the difference in peak amplitude of the ratio spectra at 243.40 and 256.40 nm. The selectivity of both methods was checked by analyzing laboratory prepared mixtures containing different concentrations of CP and its alkaline degradation products. The methods were validated in compliance with ICH guidelines. The methods determined CP in its bulk powder with average percentage recoveries of 99.60% ± 1.09 and 99.44% ± 1.60 for densitometry and ratio difference, respectively. Both methods were successfully applied for quantification of CP in its commercial cream.

  1. Trichostatin A stabilizes the expression of pluripotent genes in human mesenchymal stem cells during ex vivo expansion.

    Directory of Open Access Journals (Sweden)

    Bing Han

    Full Text Available BACKGROUND: Mesenchymal stem cells (MSCs have been considered as ideal cells for the treatment of a variety of diseases. However, aging and spontaneous differentiation of MSCs during culture expansion dampen their effectiveness. Previous studies suggest that ex vivo aging of MSCs is largely caused by epigenetic changes particularly a decline of histone H3 acetylation levels in promoter regions of pluripotent genes due to inappropriate growth environment. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we examined whether histone deacetylase inhibitor trichostatin A (TSA could suppress the histone H3 deacetylation thus maintaining the primitive property of MSCs. We found that in regular adherent culture, human MSCs became flatter and larger upon successive passaging, while the expression of pluripotent genes such as Oct4, Sox2, Nanog, Rex-1, CD133 and TERT decreased markedly. Administration of low concentrations of TSA in culture significantly suppressed the morphological changes in MSCs otherwise occurred during culture expansion, increased their proliferation while retaining their cell contact growth inhibition property and multipotent differentiation ability. Moreover, TSA stabilized the expression of the above pluripotent genes and histone H3 acetylation levels in K9 and K14 in promoter regions of Oct4, Sox2 and TERT. CONCLUSIONS/SIGNIFICANCE: Our results suggest that TSA may serve as an effective culture additive to maintain the primitive feature of MSCs during culture expansion.

  2. Exploration of structural stability in deleterious nsSNPs of the XPA gene: A molecular dynamics approach

    Directory of Open Access Journals (Sweden)

    N NagaSundaram

    2011-01-01

    Full Text Available Background: Distinguishing the deleterious from the massive number of non-functional nsSNPs that occur within a single genome is a considerable challenge in mutation research. In this approach, we have used the existing in silico methods to explore the mutation-structure-function relationship in the XPA gene. Materials and Methods: We used the Sorting Intolerant From Tolerant (SIFT, Polymorphism Phenotyping (PolyPhen, I-Mutant 2.0, and the Protein Analysis THrough Evolutionary Relationships methods to predict the effects of deleterious nsSNPs on protein function and evaluated the impact of mutation on protein stability by Molecular Dynamics simulations. Results: By comparing the scores of all the four in silico methods, nsSNP with an ID rs104894131 at position C108F was predicted to be highly deleterious. We extended our Molecular dynamics approach to gain insight into the impact of this non-synonymous polymorphism on structural changes that may affect the activity of the XPA gene. Conclusion: Based on the in silico methods score, potential energy, root-mean-square deviation, and root-mean-square fluctuation, we predict that deleterious nsSNP at position C108F would play a significant role in causing disease by the XPA gene. Our approach would present the application of in silico tools in understanding the functional variation from the perspective of structure, evolution, and phenotype.

  3. Native and Reconstituted Plasma Lipoproteins in Nanomedicine: Physicochemical Determinants of Nanoparticle Structure, Stability, and Metabolism.

    Science.gov (United States)

    Pownall, Henry J; Rosales, Corina; Gillard, Baiba K; Ferrari, Mauro

    2016-09-01

    Although many acute and chronic diseases are managed via pharmacological means, challenges remain regarding appropriate drug targeting and maintenance of therapeutic levels within target tissues. Advances in nanotechnology will overcome these challenges through the development of lipidic particles, including liposomes, lipoproteins, and reconstituted high-density lipoproteins (rHDL) that are potential carriers of water-soluble, hydrophobic, and amphiphilic molecules. Herein we summarize the properties of human plasma lipoproteins and rHDL, identify the physicochemical determinants of lipid transfer between phospholipid surfaces, and discuss strategies for increasing the plasma half-life of lipoprotein- and liposome-associated molecules.

  4. Amelogenin Gene - The Pioneer in Gender Determination from Forensic Dental Samples

    Science.gov (United States)

    Bhosale, Satish; Singh, Rajeshwar; Gubrellay, Priyanka; Patil, Jitendra; Sehdev, Bhumika; Bhagat, Sachin; Bansal, Tajinder

    2017-01-01

    Introduction In the event of any mass fatality incident, DNA analysis plays a vital role in disaster victim identification. Teeth are one of the most resistant structures in the human body that resist decomposition hence making them prime choice for extracting DNA for identification of individuals. Polymerase Chain Reaction (PCR) analysis that target regions of Amelogenin gene have become the method of choice for sex determination of biological samples. Aim Determining the sex of a given DNA sample from either dental pulp or dentin of tooth and help in identification of missing persons and disaster victims. Materials and Methods In our study 50 teeth samples were studied and they were subjected to various environmental conditions along with freshly extracted teeth taken as control for duration of one month and three months. Pulpal tissue was retrieved from the teeth specimens by access opening of root canals and for incinerated samples, the specimens were crushed. From the DNA that was extracted from the dental pulp sample Amelogenin gene locus was used for sex determination by amplifying a segment of X-Y homologous gene locus through PCR analysis. Results ANOVA test and t-test proved to be statistically significant and 100% retrieval rate was observed in samples. Conclusion Pulpal tissue along with degenerating odontoblastic processes yield sufficient amount of DNA for gender determination when subjected to various forensic conditions with maximum accuracy. PMID:28384982

  5. Validated stability-indicating spectrofluorimetric methods for the determination of ebastine in pharmaceutical preparations

    Directory of Open Access Journals (Sweden)

    Eid Manal

    2011-03-01

    Full Text Available Abstract Two sensitive, selective, economic, and validated spectrofluorimetric methods were developed for the determination of ebastine (EBS in pharmaceutical preparations depending on reaction with its tertiary amino group. Method I involves condensation of the drug with mixed anhydrides (citric and acetic anhydrides producing a product with intense fluorescence, which was measured at 496 nm after excitation at 388 nm. Method (IIA describes quantitative fluorescence quenching of eosin upon addition of the studied drug where the decrease in the fluorescence intensity was directly proportional to the concentration of ebastine; the fluorescence quenching was measured at 553 nm after excitation at 457 nm. This method was extended to (Method IIB to apply first and second derivative synchronous spectrofluorimetric method (FDSFS & SDSFS for the simultaneous analysis of EBS in presence of its alkaline, acidic, and UV degradation products. The proposed methods were successfully applied for the determination of the studied compound in its dosage forms. The results obtained were in good agreement with those obtained by a comparison method. Both methods were utilized to investigate the kinetics of the degradation of the drug.

  6. Determination of Electrochemical Performance and Thermo-Mechanical-Chemical Stability of SOFCs from Defect Modeling

    Energy Technology Data Exchange (ETDEWEB)

    Eric Wachsman; Keith L. Duncan

    2006-09-30

    This research was focused on two distinct but related issues. The first issue concerned using defect modeling to understand the relationship between point defect concentration and the electrochemical, thermo-chemical and mechano-chemical properties of typical solid oxide fuel cell (SOFC) materials. The second concerned developing relationships between the microstructural features of SOFC materials and their electrochemical performance. To understand the role point defects play in ceramics, a coherent analytical framework was used to develop expressions for the dependence of thermal expansion and elastic modulus on point defect concentration in ceramics. These models, collectively termed the continuum-level electrochemical model (CLEM), were validated through fits to experimental data from electrical conductivity, I-V characteristics, elastic modulus and thermo-chemical expansion experiments for (nominally pure) ceria, gadolinia-doped ceria (GDC) and yttria-stabilized zirconia (YSZ) with consistently good fits. The same values for the material constants were used in all of the fits, further validating our approach. As predicted by the continuum-level electrochemical model, the results reveal that the concentration of defects has a significant effect on the physical properties of ceramic materials and related devices. Specifically, for pure ceria and GDC, the elastic modulus decreased while the chemical expansion increased considerably in low partial pressures of oxygen. Conversely, the physical properties of YSZ remained insensitive to changes in oxygen partial pressure within the studied range. Again, the findings concurred exactly with the predictions of our analytical model. Indeed, further analysis of the results suggests that an increase in the point defect content weakens the attractive forces between atoms in fluorite-structured oxides. The reduction treatment effects on the flexural strength and the fracture toughness of pure ceria were also evaluated at

  7. Stabilized determination of geopotential coefficients by the mixed hom-BLUP approach

    Science.gov (United States)

    Middel, B.; Schaffrin, B.

    1989-01-01

    For the determination of geopotential coefficients, data can be used from rather different sources, e.g., satellite tracking, gravimetry, or altimetry. As each data type is particularly sensitive to certain wavelengths of the spherical harmonic coefficients it is of essential importance how they are treated in a combination solution. For example the longer wavelengths are well described by the coefficients of a model derived by satellite tracking, while other observation types such as gravity anomalies, delta g, and geoid heights, N, from altimetry contain only poor information for these long wavelengths. Therefore, the lower coefficients of the satellite model should be treated as being superior in the combination. In the combination a new method is presented which turns out to be highly suitable for this purpose due to its great flexibility combined with robustness.

  8. [Stability of the structure and antigenic determinants of adenovirus type 1 native hexon to proteases].

    Science.gov (United States)

    Kiseleva, E K; Khil'ko, S N; Grigor'ev, V G; Diachenko, N S; Vantsak, N P

    1986-08-01

    Hexon capsomers of human adenovirus type 1 (h1) labeled by iodine 125 were digested in a native state (trimers) by trypsin, chymotrypsin or papain, and the resulting hydrolysates were analyzed by SDS-PAGE. In each case, a discrete and temporally stable pattern of relatively large fragments was revealed. The degree of hexon polypeptide hydrolysis was maximal for papain, intermediate for chymotrypsin and minimal for trypsin, the largest fragments in the digest being 32, 40 and 80 kD, respectively. At room temperature, all the electrophoretically discernible hexon proteolytical fragments were held together in structures resembling intact hexon trimers and could be regarded as "hexon cores", of which papain hexon cores were the most stable during SDS-PAGE. Radioimmunoprecipitation analysis revealed a complete absence of native hexon antigenicity in thermodenaturated fragments of hexon protease digests, while native trypsin, chymotrypsin and papain hexon cores could be precipitated by hexon-specific antibodies. The immunoprecipitated material contained all of the hexon fragments found in appropriate hexon cores and retained the structure of the original cores. Trypsin, chymotrypsin and papain hexon cores were shown to possess at least part of native Ad h1 hexon antigenic determinants of each of the following specificities: species-specific (epsilon), cross-reactive with hexon of human adenoviruses (h3 and h6), simian adenovirus (sim 16), bovine adenoviruses (bos 3 and bos 7) and avian adenovirus (Aviadenovirus gal 1 or CELO). Thus, the full spectrum of known hexon antigenic determinants (species-specific to intergenus-crossreactive) is at least portly stable against protease attack of native hexon capsomers.

  9. Determination of quercetin in pharmaceutical formations via its reaction with potassium titanyloxalate. Determination of the stability constants of the quercetin titanyloxalato complex

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    DUSAN MALESEV

    2005-05-01

    Full Text Available Asimple, rapid and accurate procedure for the quantitative determination of quercetin in its pure form and in formulations has been developed. The method is based on the spectrophotometric determination of a complex formed between quercetin and potassium titanyloxalate in 50 % ethanolic solutions. To characterize the quercetin titanyloxalato complex, the stability constants of the complex were determinated potentiometrically and spectrophotometrically at different temperatures (T = 26.0 oC, 34 oC and 39.0 oC, as well as at different ionic strengths (I = 5.0×10-4 mol dm-3, 3.0×10-2 mol dm-3 and 6.0×10-2 mol dm-3 and the thermodynamic parameters were calculated. As quercetin is usually conjugated to vitamin C in pharmaceutical formulations, two procedures for the quantitative determination of quercetin by this complexing reaction were tested both in the absence and presence of ascorbic acid. In both procedures, the Beer law was obeyed over the same concentration range of quercetin, i.e., 0.85 mg mL-1×16.9 mg mL-1. In the first procedure in the absence of ascobic acid the molar absorptivity coefficient of the quercetin-titanyloxalate complex is a = 2.49×104 mol-1 dm3 cm-1, Sandells sensitivity of the method is S = 1.35×10-2 mg cm-2 and the detection limit is d = 0.67 mg mL-1. Whereas, in the presence of ascorbic acid (second procedure a = 3.04×104 mol-1 dm3 cm-1, S = 1.11×10-2 ug mL-1. The proposed method was verified for the determination of quercetin in pharmaceutical dosage forms.

  10. Regeneration of roots from callus reveals stability of the developmental program for determinate root growth in Sonoran Desert Cactaceae.

    Science.gov (United States)

    Shishkova, Svetlana; García-Mendoza, Edith; Castillo-Díaz, Vicente; Moreno, Norma E; Arellano, Jesús; Dubrovsky, Joseph G

    2007-05-01

    In some Sonoran Desert Cactaceae the primary root has a determinate root growth: the cells of the root apical meristem undergo only a few cell division cycles and then differentiate. The determinate growth of primary roots in Cactaceae was found in plants cultivated under various growth conditions, and could not be reverted by any treatment tested. The mechanisms involved in root meristem maintenance and determinate root growth in plants remain poorly understood. In this study, we have shown that roots regenerated from the callus of two Cactaceae species, Stenocereus gummosus and Ferocactus peninsulae, have a determinate growth pattern, similar to that of the primary root. To demonstrate this, a protocol for root regeneration from callus was established. The determinate growth pattern of roots regenerated from callus suggests that the program of root development is very stable in these species. These findings will permit future analysis of the role of certain Cactaceae genes in the determinate pattern of root growth via the regeneration of transgenic roots from transformed calli.

  11. Development and evaluation of a rapid strategy to determine enterotoxin gene content in Staphylococcus aureus.

    Science.gov (United States)

    Fischer, Adrien; Francois, Patrice; Holtfreter, Silva; Broeker, Barbara; Schrenzel, Jacques

    2009-05-01

    Enterotoxins of S. aureus are important molecules displaying superantigenic properties. To date no less than 18 enterotoxins have been identified in S. aureus and their role has been documented in very diverse diseases. Using available nucleotide sequence information, we developed a rapid and automated PCR-based approach to evaluate enterotoxin content in S aureus. We studied a collection of S. aureus strains previously analyzed for enterotoxins gene content and report a perfect correlation between simplex and multiplex PCR assays for the presence of all enterotoxin genes described so far. The determination of enterotoxin content relies on 4 multiplex PCR tubes whose amplification products are resolved by a rapid microcapillary electrophoresis. Automated analysis of the PCR profiles evaluates for the presence of the 18 enterotoxin genes in less than 3 h and at moderate cost. Finally, the use of enterotoxin gene content for genotyping purpose was compared to multi-locus variable number of tandem repeat assay and spa genotyping. Analysis revealed an important homogeneity of the genetic backgrounds for strains harboring the egc cluster as well as a large diversity for strains harboring other enterotoxins but lacking the egc cluster. A combined genotyping method that includes rapid enterotoxin content determination appears informative for various epidemiological survey purposes.

  12. The sex determination gene shows no founder effect in the giant honey bee, Apis dorsata.

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    Zhi Yong Liu

    Full Text Available BACKGROUND: All honey bee species (Apis spp share the same sex determination mechanism using the complementary sex determination (csd gene. Only individuals heterogeneous at the csd allele develop into females, and the homozygous develop into diploid males, which do not survive. The honeybees are therefore under selection pressure to generate new csd alleles. Previous studies have shown that the csd gene is under balancing selection. We hypothesize that due to the long separation from the mainland of Hainan Island, China, that the giant honey bees (Apis dorsata should show a founder effect for the csd gene, with many different alleles clustered together, and these would be absent on the mainland. METHODOLOGY/PRINCIPAL FINDINGS: We sampled A. dorsata workers from both Hainan and Guangxi Provinces and then cloned and sequenced region 3 of the csd gene and constructed phylogenetic trees. We failed to find any clustering of the csd alleles according to their geographical origin, i.e. the Hainan and Guangxi samples did not form separate clades. Further analysis by including previously published csd sequences also failed to show any clade-forming in both the Philippines and Malaysia. CONCLUSIONS/SIGNIFICANCE: Results from this study and those from previous studies did not support the expectations of a founder effect. We conclude that because of the extremely high mating frequency of A. dorsata queens, a founder effect does not apply in this species.

  13. Molecular characterization of genes encoding the quinolone resistance determining regions of Malaysian Streptococcus pneumoniae strains

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    Kumari N

    2008-01-01

    Full Text Available Genes encoding the quinolones resistance determining regions (QRDRs in Streptococcus pneumoniae were detected by PCR and the sequence analysis was carried out to identify point mutations within these regions. The study was carried out to observe mutation patterns among S. pneumoniae strains in Malaysia. Antimicrobial susceptibility testing of 100 isolates was determined against various antibiotics, out of which 56 strains were categorised to have reduced susceptibility to ciprofloxacin (≥2 μg/mL. These strains were subjected to PCR amplification for presence of the gyrA, parC , gyrB and parE genes. Eight representative strains with various susceptibilities to fluoroquinolones were sequenced. Two out of the eight isolates that were sequenced were shown to have a point mutation in the gyrA gene at position Ser81. The detection of mutation at codon Ser81 of the gyrA gene suggested the potential of developing fluoroquinolone resistance among S. pneumoniae isolates in Malaysia. However, further experimental work is required to confirm the involvement of this mutation in the development of fluoroquinolone resistance in Malaysia.

  14. Metabolic stability and determination of cytochrome P450 isoenzymes' contribution to the metabolism of medetomidine in dog liver microsomes.

    Science.gov (United States)

    Duhamel, Marie-Claude; Troncy, Eric; Beaudry, Francis

    2010-08-01

    Medetomidine is a potent and selective alpha2-adrenergic agonist. The activation of alpha2-adrenergic receptor mediates a variety of effects including sedation, analgesia, relief of anxiety, vasoconstriction and bradycardia. However, our main interest is the sedative effects of medetomidine when used as a premedicant prior surgery in companion animals, especially in dogs. Recently, data suggested that following intravenous infusion at six dosing regiments non-linear pharmacokinetics was observed. Major causes of non-linear pharmacokinetics are the elimination of the drug not following a simple first-order kinetics and/or the elimination half-life changing due to saturation of an enzyme system. The goal of this study was to establish the metabolic stability and determine the metabolic pathway of medetomidine in dog liver microsomes. Consequently, Michaelis-Menten parameters (V(max), K(m)), T(1/2) and CL(i) were determined. The incubations were performed in a microcentrifuge tube and containing various concentrations of medetomidine (10-5000 nM), 1 mg/mL of microsomal proteins suspended in 0.1 M phosphate buffer, pH 7.4. Microsomal suspensions were preincubated with NADPH (1 mM) for 5 min at 37 degrees C prior to fortification with medetomidine. Samples were taken at various time points for kinetic information and the initial velocity (v(i)) was determined after 10 min incubation. The reaction was stopped by the addition of an internal standard solution (100 ng/mL of dextrometorphan in acetone). Medetomidine concentrations were determined using a selective and sensitive HPLC-ESI/MS/MS method. Using non-linear regression, we determined a K(m) value of 577 nM, indicating relatively low threshold enzyme saturation consistent with previous in vivo observation. The metabolic stability was determined at a concentration of 100 nm (dog liver microsomes, also consistent with previous in vivo data. Moreover, results suggest that principally medetomidine is metabolized by the

  15. A boolean model of the cardiac gene regulatory network determining first and second heart field identity.

    Directory of Open Access Journals (Sweden)

    Franziska Herrmann

    Full Text Available Two types of distinct cardiac progenitor cell populations can be identified during early heart development: the first heart field (FHF and second heart field (SHF lineage that later form the mature heart. They can be characterized by differential expression of transcription and signaling factors. These regulatory factors influence each other forming a gene regulatory network. Here, we present a core gene regulatory network for early cardiac development based on published temporal and spatial expression data of genes and their interactions. This gene regulatory network was implemented in a Boolean computational model. Simulations reveal stable states within the network model, which correspond to the regulatory states of the FHF and the SHF lineages. Furthermore, we are able to reproduce the expected temporal expression patterns of early cardiac factors mimicking developmental progression. Additionally, simulations of knock-down experiments within our model resemble published phenotypes of mutant mice. Consequently, this gene regulatory network retraces the early steps and requirements of cardiogenic mesoderm determination in a way appropriate to enhance the understanding of heart development.

  16. A Boolean Model of the Cardiac Gene Regulatory Network Determining First and Second Heart Field Identity

    Science.gov (United States)

    Zhou, Dao; Kestler, Hans A.; Kühl, Michael

    2012-01-01

    Two types of distinct cardiac progenitor cell populations can be identified during early heart development: the first heart field (FHF) and second heart field (SHF) lineage that later form the mature heart. They can be characterized by differential expression of transcription and signaling factors. These regulatory factors influence each other forming a gene regulatory network. Here, we present a core gene regulatory network for early cardiac development based on published temporal and spatial expression data of genes and their interactions. This gene regulatory network was implemented in a Boolean computational model. Simulations reveal stable states within the network model, which correspond to the regulatory states of the FHF and the SHF lineages. Furthermore, we are able to reproduce the expected temporal expression patterns of early cardiac factors mimicking developmental progression. Additionally, simulations of knock-down experiments within our model resemble published phenotypes of mutant mice. Consequently, this gene regulatory network retraces the early steps and requirements of cardiogenic mesoderm determination in a way appropriate to enhance the understanding of heart development. PMID:23056457

  17. A rough set based rational clustering framework for determining correlated genes.

    Science.gov (United States)

    Jeyaswamidoss, Jeba Emilyn; Thangaraj, Kesavan; Ramar, Kadarkarai; Chitra, Muthusamy

    2016-06-01

    Cluster analysis plays a foremost role in identifying groups of genes that show similar behavior under a set of experimental conditions. Several clustering algorithms have been proposed for identifying gene behaviors and to understand their significance. The principal aim of this work is to develop an intelligent rough clustering technique, which will efficiently remove the irrelevant dimensions in a high-dimensional space and obtain appropriate meaningful clusters. This paper proposes a novel biclustering technique that is based on rough set theory. The proposed algorithm uses correlation coefficient as a similarity measure to simultaneously cluster both the rows and columns of a gene expression data matrix and mean squared residue to generate the initial biclusters. Furthermore, the biclusters are refined to form the lower and upper boundaries by determining the membership of the genes in the clusters using mean squared residue. The algorithm is illustrated with yeast gene expression data and the experiment proves the effectiveness of the method. The main advantage is that it overcomes the problem of selection of initial clusters and also the restriction of one object belonging to only one cluster by allowing overlapping of biclusters.

  18. Natural resource landscapes of a marine bacterium reveal distinct fitness-determining genes across the genome.

    Science.gov (United States)

    Takemura, Alison F; Corzett, Christopher H; Hussain, Fatima; Arevalo, Philip; Datta, Manoshi; Yu, Xiaoqian; Le Roux, Frederique; Polz, Martin F

    2017-06-01

    Heterotrophic bacteria exploit diverse microhabitats in the ocean, from particles to transient gradients. Yet the degree to which genes and pathways can contribute to an organism's fitness on such complex and variable natural resource landscapes remains poorly understood. Here, we determine the gene-by-gene fitness of a generalist saprophytic marine bacterium (Vibrio sp. F13 9CS106) on complex resources derived from its natural habitats - copepods (Apocyclops royi) and brown algae (Fucus vesiculosus) - and as reference substrates, glucose and the polysaccharide alginate, derived from brown algal cell walls. We find that resource complexity strongly buffers fitness costs of mutations, and that anabolic rather than catabolic pathways are more stringently required, likely due to functional redundancy in the latter. Moreover, while carbohydrate-rich algae requires several synthesis pathways, protein-rich Apocyclops does not, suggesting this ancestral habitat for Vibrios is a replete medium with metabolically redundant substrates. We also identify a candidate fitness trade-off for algal colonization: deletion of mshA increases mutant fitness. Our results demonstrate that gene fitness depends on habitat composition, and suggest that this generalist uses distinct resources in different natural habitats. The results further indicate that substrate replete conditions may lead to relatively relaxed selection on catabolic genes. © 2017 Society for Applied Microbiology and John Wiley & Sons Ltd.

  19. A Validated Stability-Indicating and Stereoselective HPLC Method for the Determination of Lenalidomide Enantiomers in Bulk Form and Capsules

    Science.gov (United States)

    Alzoman, Nourah Z.

    2016-01-01

    A simple, rapid and stability-indicating chiral HPLC (CHR-HPLC) method was designed for the enantiomeric separation of lenalidomide (LDM) in the presence of its degradation products. LDM was exposed to different accelerated stress factors. The degradation products were well resolved from the pure drug enantiomers. Separation of the LDM enantiomers was achieved on a LUX 5U cellulose-2 chiral column (250 × 4.6 mm i.d.) with a mobile phase consisting of methanol : glacial acetic acid : triethyl amine (100 : 0.01 : 0.01, v/v/v) at a flow rate of 1.2 mL/min. The detection wavelength was 220 nm, and ornidazole was the internal standard. The chiral method was validated in terms of its specificity, linearity, range, precision and accuracy as well as solution stability, robustness, limit of detection and limit of quantification. The calibration curve was linear for concentrations ranging from 2 to 1,000 ng/mL (r = 0.9999) for both LDM enantiomers. The proposed method, which met International Conference on Harmonization/Food and Drug Administration regulatory requirements, was utilized successfully for the determination of LDM in bulk and in capsules with acceptable accuracy and precision; the label demand percentages were 100.09 ± 0.80 and 99.97 ± 0.93 for the S-(−) and R-(+)-LDM enantiomers, respectively. Based on these results, this method should have great value when applied to quality control and stability studies of LDM. PMID:26850732

  20. A rapid stability indicating LC-method for determination of praziquantel in presence of its pharmacopoeial impurities

    Directory of Open Access Journals (Sweden)

    Hisham Hashem

    2017-02-01

    Full Text Available This study reports for the first time about a stability indicating RP-HPLC method for quantitative determination of Praziquantel (PZQ in bulk powder and dosage form and in presence of its pharmacopoeial impurities. The chromatographic separation was carried out on (Caltrex AI® calixarene column, a relatively new packing material. Chromatography was done using an isocratic binary mobile phase consisting of ACN and 25 mM ammonium acetate (NH4Ac in the ratio of 40:60 at flow rate of 1 mL min−1, 30 °C and 210 nm wavelength for detection. The elution time of PZQ was found to be 6.15 ± 0.03 min. The method was validated for system suitability, linearity, precision, limits of detection and quantitation, specificity, stability and robustness. The robustness study was done for small changes in temperature, flow rate, wavelength of detection and % of ACN in mobile phase. Stability tests were done through exposure of the analyte solution to five different stress conditions: Reflux with 1 N HCl, reflux with 1 N NaOH, reflux with 30% H2O2, thermal degradation of powder and exposure to UV radiation. Limits of detection and quantification were found to be 0.56 and 1.70 μg mL−1, respectively. The recovery value of this method was 100.30% ± 1.10 and the reproducibility was within 1.31.

  1. Simultaneous, stability indicating, HPLC-DAD determination of guaifenesin and methyl and propyl-parabens in cough syrup.

    Science.gov (United States)

    Grosa, Giorgio; Del Grosso, Erika; Russo, Roberta; Allegrone, Gianna

    2006-06-07

    A stability indicating high performance liquid chromatography procedure has been developed for the simultaneous determination of guaifenesin (GUA), methyl p-hydroxybenzoate (MHB) and propyl p-hydroxybenzoate (PHB) in a commercial cough syrup dosage form. The method was specific and stability indicating as chromatographic conditions were selected to provide adequate separation of GUA, MHB and PHB from the putative degradation products guaiacol (GUAI) and p-hydroxybenzoic acid (HBA) as well as from excipients. The isocratic separation and quantitation were achieved within 17 min on a 150-mm column with an ether-linked phenyl stationary phase and a hydrophilic endcapping. The mobile phase was constituted of eluant A: aqueous phosphate buffer (pH 3.0, 10 mM)/acetonitrile 25/75 (v/v) and eluant B:methanol; the A:B ratio was 85:15 (v/v) with a flow rate 1 ml min-1 and detection of analytes at 254 and 276 nm. The method showed good linearity for the GUA-MHB-PHB mixture in the 95-285, 4-12, and 1-3 microg ml-1 ranges, respectively, being all the square of the correlation coefficients greater than 0.999. The interday R.S.D.s were 1.17, 1.14, and 0.91%, for GUA, MHB, and PHP, respectively. The method demonstrated also to be accurate; indeed the average recoveries, at 100% of the target assay concentration, were 100.5, 100.3, and 100.7% with relative standard deviations of 0.8, 0.7, and 0.4% for GUA, MHB, and PHB, respectively, from laboratory prepared samples. The applicability of the method was evaluated in commercial dosage form analysis as well as in stability studies.

  2. Estrogen Receptor β2 Induces Hypoxia Signature of Gene Expression by Stabilizing HIF-1α in Prostate Cancer.

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    Prasenjit Dey

    Full Text Available The estrogen receptor (ER β variant ERβ2 is expressed in aggressive castration-resistant prostate cancer and has been shown to correlate with decreased overall survival. Genome-wide expression analysis after ERβ2 expression in prostate cancer cells revealed that hypoxia was an overrepresented theme. Here we show that ERβ2 interacts with and stabilizes HIF-1α protein in normoxia, thereby inducing a hypoxic gene expression signature. HIF-1α is known to stimulate metastasis by increasing expression of Twist1 and increasing vascularization by directly activating VEGF expression. We found that ERβ2 interacts with HIF-1α and piggybacks to the HIF-1α response element present on the proximal Twist1 and VEGF promoters. These findings suggest that at least part of the oncogenic effects of ERβ2 is mediated by HIF-1α and that targeting of this ERβ2 - HIF-1α interaction may be a strategy to treat prostate cancer.

  3. Estrogen Receptor β2 Induces Hypoxia Signature of Gene Expression by Stabilizing HIF-1α in Prostate Cancer

    Science.gov (United States)

    Dey, Prasenjit; Velazquez-Villegas, Laura A.; Faria, Michelle; Turner, Anthony; Jonsson, Philp; Webb, Paul; Williams, Cecilia; Gustafsson, Jan-Åke; Ström, Anders M.

    2015-01-01

    The estrogen receptor (ER) β variant ERβ2 is expressed in aggressive castration-resistant prostate cancer and has been shown to correlate with decreased overall survival. Genome-wide expression analysis after ERβ2 expression in prostate cancer cells revealed that hypoxia was an overrepresented theme. Here we show that ERβ2 interacts with and stabilizes HIF-1α protein in normoxia, thereby inducing a hypoxic gene expression signature. HIF-1α is known to stimulate metastasis by increasing expression of Twist1 and increasing vascularization by directly activating VEGF expression. We found that ERβ2 interacts with HIF-1α and piggybacks to the HIF-1α response element present on the proximal Twist1 and VEGF promoters. These findings suggest that at least part of the oncogenic effects of ERβ2 is mediated by HIF-1α and that targeting of this ERβ2 – HIF-1α interaction may be a strategy to treat prostate cancer. PMID:26010887

  4. Identification of Multiple Cryptococcal Fungicidal Drug Targets by Combined Gene Dosing and Drug Affinity Responsive Target Stability Screening

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    Yoon-Dong Park

    2016-08-01

    Full Text Available Cryptococcus neoformans is a pathogenic fungus that is responsible for up to half a million cases of meningitis globally, especially in immunocompromised individuals. Common fungistatic drugs, such as fluconazole, are less toxic for patients but have low efficacy for initial therapy of the disease. Effective therapy against the disease is provided by the fungicidal drug amphotericin B; however, due to its high toxicity and the difficulty in administering its intravenous formulation, it is imperative to find new therapies targeting the fungus. The antiparasitic drug bithionol has been recently identified as having potent fungicidal activity. In this study, we used a combined gene dosing and drug affinity responsive target stability (GD-DARTS screen as well as protein modeling to identify a common drug binding site of bithionol within multiple NAD-dependent dehydrogenase drug targets. This combination genetic and proteomic method thus provides a powerful method for identifying novel fungicidal drug targets for further development.

  5. Automatic Sieve-Shaker for Determining Soil Aggregate Stability and Dimensional Distribution Using a Vertical Oscillation System

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    Rosario Dell’Aquila

    Full Text Available The soil aggregate stability is determined generally by sifting the soil samples in water using a sieve-shaker (wet sieving. The Author has developed an original model of automatic sieve-shaker using a vertical oscillation system to the aim of an its possible use to determine the soil aggregate stability and dimensional distribution. The purpose of this note is to describe the construction and performance of the prototype currently used in the Laboratory for the Soil Structure Study of the ISAFOM – CNR. The proposed sieve-shaker, with the introduction of some innovations (protected by Italy Patent 0001332102, realizes the submersion and levelling of the soil samples using a lifter to support the containers with the water. With 6 workplaces it allows to process simultaneously up to 6 soil samples according to different test cycles. By means of the control panel it is possible to set up various determinations with the stroke of 3 cm and the oscillation frequency from 4 up to 80 oscillations per minute. The performance of the proposed sieve-shaker was verified with a technical test to verify the performance of the 6 workplaces to oscillation speed increasing up to 60 oscillations per minute and an agronomic test. The results have been submitted to analysis of variance considering the plots of the field from which have been taken the samples for repetitions and the six workplaces of the proposed sieve-shaker for experimental theses. The differences between the various workplaces have not been significant. This demonstrates that the behavior of the various workplaces is uniform. The dispersion in water at constant shaking time and increasing oscillation speed has evidenced a very significant inverse relation between the index of aggregate stability in water (IASW and number of oscillations per minute. This result demonstrates a constant performance of the proposed sieve-shaker to varying of the oscillation speed. The agnonomic test has demonstrated

  6. Degradable terpolymers with alkyl side chains demonstrate enhanced gene delivery potency and nanoparticle stability.

    Science.gov (United States)

    Eltoukhy, Ahmed A; Chen, Delai; Alabi, Christopher A; Langer, Robert; Anderson, Daniel G

    2013-03-13

    Degradable, cationic poly(β-amino ester)s (PBAEs) with alkyl side chains are developed for non-viral gene delivery. Nanoparticles formed from these PBAE terpolymers exhibit significantly enhanced DNA transfection potency and resistance to aggregation. These hydrophobic PBAE terpolymers, but not PBAEs lacking alkyl side chains, support interaction with PEG-lipid conjugates, facilitating their functionalization with shielding and targeting moieties and accelerating the in vivo translation of these materials. Copyright © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. Prenatal sex determination in suspicious cases of X-linked recessive diseases by the amelogenin gene

    Directory of Open Access Journals (Sweden)

    Amir Abbas Rahimi

    2014-02-01

    Conclusion: Sex detection of fetus before delivery in the first trimester of pregnancy, will prevent babies with abnormalities being born. It can also be used in detection of recessive sex related diseases in In Vitro Fertilization cases for sex detection and to transfer female fetus to the mother. Our optimized molecular detection system was designed on the basis of amelogenin gene, which can determine the sex in blood, chorionic villi, and single cell in vitro fertilization with high sensitivity and specificity.

  8. Stability indicating RP-HPLC method for simultaneous determination of piroxicam and ofloxacin in binary combination.

    Science.gov (United States)

    John, Peter; Azeem, Waqar; Ashfaq, Muhammad; Khan, Islam Ullah; Razzaq, Syed Naeem

    2015-09-01

    A simple and precise RP-HPLC method was developed for simultaneous determination of piroxicam and ofloxacin in pharmaceutical formulations and human serum. Optimum separations of piroxicam, ofloxacin and stress-induced degradation products were achieved by use of Hypersil BDS C8 column (250 x 4.6mm, 5 μm). The mobile phase was a mixture of acetonitrile: 0.012M K2HPO4: 0.008M sodium citrate (both buffers mixed and pH adjusted to 2.8) (50:25:25 v/v/v) delivered at flow rate of 1.5 mL min⁻¹ using DAD at 254 nm. Response was linear function of concentration over the ranges of 70-130 mg mL⁻¹ for piroxicam and ofloxacin (r² ≥ 0.999). The method efficiently separated the analytical peaks from degradation products with acceptable tailing and resolution. The developed method was successfully used for concurrent analysis of piroxicam and ofloxacin in pharmaceutical formulations, human serum and in vitro drug interaction studies.

  9. Intrinsic karyotype stability and gene copy number variations may have laid the foundation for tetraploid wheat formation.

    Science.gov (United States)

    Zhang, Huakun; Bian, Yao; Gou, Xiaowan; Dong, Yuzhu; Rustgi, Sachin; Zhang, Bangjiao; Xu, Chunming; Li, Ning; Qi, Bao; Han, Fangpu; von Wettstein, Diter; Liu, Bao

    2013-11-26

    Polyploidy or whole-genome duplication is recurrent in plant evolution, yet only a small fraction of whole-genome duplications has led to successful speciation. A major challenge in the establishment of nascent polyploids is sustained karyotype instability, which compromises fitness. The three putative diploid progenitors of bread wheat, with AA, SS (S ∼ B), and DD genomes occurred sympatrically, and their cross-fertilization in different combinations may have resulted in fertile allotetraploids with various genomic constitutions. However, only SSAA or closely related genome combinations have led to the speciation of tetraploid wheats like Triticum turgidum and Triticum timopheevii. We analyzed early generations of four newly synthesized allotetraploid wheats with genome compositions S(sh)S(sh)A(m)A(m), S(l)S(l)AA, S(b)S(b)DD, and AADD by combined fluorescence and genomic in situ hybridization-based karyotyping. Results of karyotype analyses showed that although S(sh)S(sh)A(m)A(m) and S(l)S(l)AA are characterized by immediate and persistent karyotype stability, massive aneuploidy and extensive chromosome restructuring are associated with S(b)S(b)DD and AADD in which parental subgenomes showed markedly different propensities for chromosome gain/loss and rearrangements. Although compensating aneuploidy and reciprocal translocation between homeologs prevailed, reproductive fitness was substantially compromised due to chromosome instability. Strikingly, localized genomic changes in repetitive DNA and copy-number variations in gene homologs occurred in both chromosome stable lines, S(sh)S(sh)A(m)A(m) and S(l)S(l)AA. Our data demonstrated that immediate and persistent karyotype stability is intrinsic to newly formed allotetraploid wheat with genome combinations analogous to natural tetraploid wheats. This property, coupled with rapid gene copy-number variations, may have laid the foundation of tetraploid wheat establishment.

  10. Application of a stability-indicating thin-layer chromatographic method to the determination of tenatoprazole in pharmaceutical dosage forms.

    Science.gov (United States)

    Dhaneshwar, Sunil R; Bhusari, Vidhya K; Mahadik, Mahadeo V; Santakumari, B

    2009-01-01

    A sensitive, selective, precise, and stability-indicating thin-layer chromatographic (TLC) method was developed and validated for the determination of tenatoprazole both as a bulk drug and in formulation. The method uses TLC aluminum plates precoated with Silica Gel 60F-254 as the stationary phase and the solvent system toluene-ethyl acetate-methanol (6 + 4 + 1, v/v/v). This system gave compact spots for tenatoprazole (Rf value of 0.34 +/- 0.02). Tenatoprazole was subjected to acid and alkali hydrolysis, oxidation, and photodegradation. The peaks of the degradation products were well-resolved from that of the pure drug and had significantly different Rf values. Densitometric analysis of tenatoprazole was performed in the absorbance mode at 306 nm. The linear regression analysis data for the calibration plots showed a good linear relationship over the concentration range of 100-1500 ng/spot. The mean values of the correlation coefficient, slope, and intercept were 0.9989 +/- 1.42, 10.27 +/- 0.965, and 4894.2 +/- 1.24, respectively. The method was validated for precision, robustness, and recovery. The limit of detection and limit of quantitation were 50 and 100 ng/spot, respectively. Statistical analysis showed that the method is repeatable and selective for estimation of tenatoprazole. Because the method can separate the drug from its degradation products, it can be used to monitor stability.

  11. Determination of arsenic in diesel, gasoline and naphtha by graphite furnace atomic absorption spectrometry using microemulsion medium for sample stabilization.

    Science.gov (United States)

    Brandão, Geisamanda Pedrini; de Campos, Reinaldo Calixto; Luna, Aderval Severino; de Castro, Eustáquio Vinicius Ribeiro; de Jesus, Honério Coutinho

    2006-08-01

    A procedure for the determination of As in diesel, gasoline and naphtha at microg L(-1) levels by GFAAS is proposed. Sample stabilization was achieved by the formation of three component solutions prepared by mixing appropriate volumes of the samples propan-1-ol and nitric acid aqueous solution. This mixture resulted in a one-phase medium, which was indefinitely stable. No changes in the analyte signals were observed over several days in spiked samples, proving long-term stabilization ability. The use of conventional (Pd) and permanent (Ir) modification was investigated and the former was preferred. Central composite design multivariate optimization defined the optimum microemulsion composition as well as the temperature program. In this way, calibration using aqueous analytical solutions was possible, since the same sensitivity was observed in the investigated microemulsion media and in 0.2% v/v HNO(3). Coefficients of correlation larger than 0.999 and an As characteristic mass of 22 pg were observed. Recoveries (n=4) obtained from spiked samples were 98+/-4, 99+/-3 and 103+/-5%, and the limits of detection in the original samples were 1.8, 1.2 and 1.5 microg L(-1) for diesel, gasoline and naphtha, respectively. Validation was performed by the analysis of a set of commercial samples by independent comparative procedures. No significant difference (Student's t-test, pnaphtha, equivalent to a sample throughput of 7 h(-1) for diesel and 10 h(-1) for gasoline and naphtha.

  12. Simultaneous determination of aliskiren and hydrochlorothiazide from their pharmaceutical preparations using a validated stability-indicating MEKC method.

    Science.gov (United States)

    Sangoi, Maximiliano S; Wrasse-Sangoi, Micheli; Oliveira, Paulo R; Rolim, Clarice M B; Steppe, Martin

    2011-08-01

    A stability-indicating MEKC method was developed and validated for the simultaneous determination of aliskiren (ALI) and hydrochlorothiazide (HCTZ) in pharmaceutical formulations using ranitidine as an internal standard (IS). Optimal conditions for the separation of ALI, HCTZ and its major impurity chlorothiazide (CTZ), IS and degradation products were investigated. The method employed 47 mM Tris buffer and 47 mM anionic detergent SDS solution at pH 10.2 as the background electrolyte. MEKC method was performed on a fused-silica capillary (40 cm) at 28°C. Applied voltage was 26 kV (positive polarity) and photodiode array (PDA) detector was set at 217 nm. The method was validated in accordance with the ICH requirements. The method was linear over the concentration range of 5-100 and 60-1200 μg/mL for HCTZ and ALI, respectively (r(2) >0.9997). The stability-indicating capability of the method was established by enforced degradation studies combined with peak purity assessment using the PDA detection. Precision and accuracy evaluated by RSD were lower than 2%. The method proved to be robust by a fractional factorial design evaluation. The proposed MEKC method was successfully applied for the quantitative analysis of ALI and HCTZ both individually and in a combined dosage tablet formulation to support the quality control.

  13. Development of stability-indicating UHPLC method for the quantitative determination of silodosin and its related substances.

    Science.gov (United States)

    Shaik, Jafer Vali; Saladi, Shantikumar; Sait, Shakil S

    2014-08-01

    A novel, specific and stability-indicating reversed-phase (RP) ultra-high-performance liquid chromatography (UHPLC) method, which is mass compatible, was developed and validated for the quantitative determination of silodosin and its related substances. Silodosin was subjected to stress conditions like hydrolysis (acid and basic), oxidation, photolysis and thermal degradation, as per the guidelines of the International Conference Harmonization, to show that the method is stability-indicating. The proposed UHPLC method has a resolution of greater than 2.0 between silodosin and its process-related impurities. The chromatographic separation was achieved on an Agilent Poroshell 120 EC-C18 column (50 × 4.6mm i.d.; particle size, 2.7 µm). The method employed a linear gradient elution using a mobile phase consisting of acetonitrile and 10 mM ammonium acetate buffer with 0.1% triethyl amine, with pH adjusted to 6.0, monitored at 273 nm. The developed RP-LC method was validated with respect to linearity, accuracy, precision and robustness. The known process impurities were separated and their structure was confirmed by using liquid chromatography-mass spectrometry and direct mass analysis.

  14. Development, Optimization, and Validation of a Green and Stability-Indicating HPLC Method for Determination of Daptomycin in Lyophilized Powder.

    Science.gov (United States)

    Tótoli, Eliane Gandolpho; Salgado, Hérida Regina Nunes

    2015-01-01

    Daptomycin is an antimicrobial that plays an important role in clinical practice today because it is considered a promising drug to combat resistant strains, such as methicilin and vancomycin-resistant Gram-positive bacteria. Considering the analysis of daptomycin in a pharmaceutical dosage form, the only method found in literature uses potentially toxic organic solvents. Therefore, the objective of this work was to develop a green and stability-indicating HPLC method for determination of daptomycin in lyophilized powder. The mobile phase was ethanol-water (55+45, v/v) at pH 4.5 pumped at a flow rate of 0.6 mL/min. A C18 column was used, and UV detection was performed at 221 nm. Stress degradation studies were conducted in order to demonstrate the specificity and stability-indicating capability of the method. The method was validated according to International Conference on Harmonization guidelines, proving to be linear (r=0.9996), precise, accurate, robust (demonstrated by the Plackett-Burman model), and specific within the range 20-70 μg/mL. The retention time of daptomycin was 5.8 min. It can be concluded that the validated method can be a fast, safe, and environmentally friendly alternative for the analysis of daptomycin.

  15. Stability analysis and determination of rock pillar between two adjacent caverns in different regions of Asmari formation in Iran

    Institute of Scientific and Technical Information of China (English)

    Abdollahipour Abolfazl a; ⇑; Ghannadshirazi Hossein b

    2014-01-01

    Large underground caverns are commonly used in variety of applications. In many cases, because of the geomechanical limitations of dimensions and requirement of high volume, several parallel caverns are used. Plastic zone integration requires a larger rock pillar distance of theses adjacent caverns while eco-nomic and access reasons require smaller distance. In Iran many underground projects are located in West and South West. Asmari formation covers a large part of these regions. The stability of underground spaces that are constructed or will be constructed in this formation has been investigated. A proper cross section based on plastic analysis and a stability criterion has been proposed for each region. Finally, in each case, allowable rock pillar between adjacent caverns with similar dimension was determined with two methods (numerical analysis and fire service law). Results show that Fire Service Law uses a very con-servative safety factor and it was proposed to use a correction factor for allowable distance based on application of underground space.

  16. A stability-indicating high performance liquid chromatographic assay for the simultaneous determination of atenolol and lercanidipine hydrochloride in tablets

    Directory of Open Access Journals (Sweden)

    H O Kaila

    2011-01-01

    Full Text Available A simple, rapid, precise and accurate isocratic reversed phase stability indicating HPLC method was developed and validated for the simultaneous determination of atenolol and lercanidipine hydrochloride in commercial tablets. The chromatographic separation was achieved on phenomenex Gemini C18 (250×4.6 mm, 5 μm column using a mobile phase consisting of acetonitrile and buffer (20 mM potassium dihydrogen phosphate pH 3.5 in the ratio of (55:45, v/v at a flow rate of 1.0 ml/min and UV detection at 235 nm. The linearity of the proposed method was investigated in the range of 40-160 μg/ml (r 2 =0.9995 for atenolol and 8-32 μg/ml (r 2 =0.9993 for lercanidipine. Degradation products produced as a result of stress studies did not interfere with the detection of atenolol and lercanidipine and the assay can thus be considered stability-indicating.

  17. Phylogenetic distribution and evolutionary dynamics of the sex determination genes doublesex and transformer in insects.

    Science.gov (United States)

    Geuverink, E; Beukeboom, L W

    2014-01-01

    Sex determination in insects is characterized by a gene cascade that is conserved at the bottom but contains diverse primary signals at the top. The bottom master switch gene doublesex is found in all insects. Its upstream regulator transformer is present in the orders Hymenoptera, Coleoptera and Diptera, but has thus far not been found in Lepidoptera and in the basal lineages of Diptera. transformer is presumed to be ancestral to the holometabolous insects based on its shared domains and conserved features of autoregulation and sex-specific splicing. We interpret that its absence in basal lineages of Diptera and its order-specific conserved domains indicate multiple independent losses or recruitments into the sex determination cascade. Duplications of transformer are found in derived families within the Hymenoptera, characterized by their complementary sex determination mechanism. As duplications are not found in any other insect order, they appear linked to the haplodiploid reproduction of the Hymenoptera. Further phylogenetic analyses combined with functional studies are needed to understand the evolutionary history of the transformer gene among insects. © 2013 S. Karger AG, Basel.

  18. Brain transcriptional stability upon prion protein-encoding gene invalidation in zygotic or adult mouse

    Directory of Open Access Journals (Sweden)

    Béringue Vincent

    2010-07-01

    Full Text Available Abstract Background The physiological function of the prion protein remains largely elusive while its key role in prion infection has been expansively documented. To potentially assess this conundrum, we performed a comparative transcriptomic analysis of the brain of wild-type mice with that of transgenic mice invalidated at this locus either at the zygotic or at the adult stages. Results Only subtle transcriptomic differences resulting from the Prnp knockout could be evidenced, beside Prnp itself, in the analyzed adult brains following microarray analysis of 24 109 mouse genes and QPCR assessment of some of the putatively marginally modulated loci. When performed at the adult stage, neuronal Prnp disruption appeared to sequentially induce a response to an oxidative stress and a remodeling of the nervous system. However, these events involved only a limited number of genes, expression levels of which were only slightly modified and not always confirmed by RT-qPCR. If not, the qPCR obtained data suggested even less pronounced differences. Conclusions These results suggest that the physiological function of PrP is redundant at the adult stage or important for only a small subset of the brain cell population under classical breeding conditions. Following its early reported embryonic developmental regulation, this lack of response could also imply that PrP has a more detrimental role during mouse embryogenesis and that potential transient compensatory mechanisms have to be searched for at the time this locus becomes transcriptionally activated.

  19. Ionic modulation of QPX stability as a nano-switch regulating gene expression in neurons

    Science.gov (United States)

    Baghaee Ravari, Soodeh

    G-quadruplexes (G-QPX) have been the subject of intense research due to their unique structural configuration and potential applications, particularly their functionality in biological process as a novel type of nano--switch. They have been found in critical regions of the human genome such as telomeres, promoter regions, and untranslated regions of RNA. About 50% of human DNA in promoters has G-rich regions with the potential to form G-QPX structures. A G-QPX might act mechanistically as an ON/OFF switch, regulating gene expression, meaning that the formation of G-QPX in a single strand of DNA disrupts double stranded DNA, prevents the binding of transcription factors (TF) to their recognition sites, resulting in gene down-regulation. Although there are numerous studies on biological roles of G-QPXs in oncogenes, their potential formation in neuronal cells, in particular upstream of transcription start sites, is poorly investigated. The main focus of this research is to identify stable G-QPXs in the 97bp active promoter region of the choline acetyltransferase (ChAT) gene, the terminal enzyme involved in synthesis of the neurotransmitter acetylcholine, and to clarify ionic modulation of G-QPX nanostructures through the mechanism of neural action potentials. Different bioinformatics analyses (in silico), including the QGRS, quadparser and G4-Calculator programs, have been used to predict stable G-QPX in the active promoter region of the human ChAT gene, located 1000bp upstream from the TATA box. The results of computational studies (using those three different algorithms) led to the identification of three consecutive intramolecular G-QPX structures in the negative strand (ChAT G17-2, ChAT G17, and ChAT G29) and one intramolecular G-QPX structure in the positive strand (ChAT G30). Also, the results suggest the possibility that nearby G-runs in opposed DNA strands with a short distance of each other may be able to form a stable intermolecular G-QPX involving two DNA

  20. Stability evaluation of reference genes for real-time PCR in zebrafish (Danio rerio) exposed to cadmium chloride and subsequently infected by bacteria Aeromonas hydrophila

    Energy Technology Data Exchange (ETDEWEB)

    Lang, Xingping; Wang, Lan, E-mail: lanwang@sxu.edu.cn; Zhang, Zuobing, E-mail: zbzhang@sxu.edu.cn

    2016-01-15

    Highlights: • Cd exposure affects the stability of reference genes for real-time PCR in zebrafish. • Reference genes present different stability in the five tissues (spleen, kidney, liver, gills and intestine) of zebrafish after Cd exposure. • Bacterial infection further affects the stability of reference genes in Cd-treated zebrafish. - Abstract: Environmental and occupational cadmium (Cd) toxicity is a global concern, and the model organism zebrafish is an ideal species to investigate Cd toxicity. Among various detecting techniques, quantitative real-time PCR (qPCR) is a sensitive and efficient tool. Stable reference genes are critical for relative qPCR analysis. However, accumulated evidence shows that conventional reference genes can vary significantly under different experimental setups. Here we evaluated the stability of eight candidate reference genes of zebrafish with or without exposure to different concentrations of Cd. The results showed that the best four suitable reference genes in the five selected organs were: (1) spleen: β-actin > gapdh > ef1α > rpl13α; (2) kidney: rplp2 > rpl7 > β-actin > ef1α; (3) liver: rpl7 > rpl13α > β-actin > ef1α; (4) gills: rplp2 > gapdh > rnf7 > ef1α; (5) intestine: ef1α > rnf7 > rplp2 > rpl13α. Moreover, we further assessed the expression stability of the four reference genes for Cd immunotoxicology studies in zebrafish. The expression profiles showed that ef1α in spleen and kidney, rpl13a in liver and rplp2 in intestine were the most suitable reference genes at 12 h and 9 days after the injection with Aeromonas hydrophila following Cd exposure. In gills, the expression of gapdh was more stable than ef1α after 9 days of bacteria challenge while ef1α showed a higher stability than gapdh at 12 h after bacteria injection. In conclusion, this study has demonstrated that different tissues of zebrafish have different suitable reference genes after Cd exposure and the subsequently pathogenic insults for q

  1. Diet-gene interactions between dietary fat intake and common polymorphisms in determining lipid metabolism

    Directory of Open Access Journals (Sweden)

    Corella, Dolores

    2009-03-01

    Full Text Available Current dietary guidelines for fat intake have not taken into consideration the possible genetic differences underlying the individual variability in responsiveness to dietary components. Genetic variability has been identified in humans for all the known lipid metabolim-related genes resulting in a plethora of candidate genes and genetic variants to examine in diet-gene interaction studies focused on fat consumption. Some examples of fat-gene interaction are reviewed. These include: the interaction between total intake and the 514C/T in the hepatic lipase gene promoter in determining high-density lipoprotein cholesterol (HDL-C metabolism; the interaction between polyunsaturated fatty acids (PUFA and the 75G/A polymorphism in the APOA1 gene plasma HDL-C concentrations; the interaction between PUFA and the L162V polymorphism in the PPARA gene in determining triglycerides and APOC3 concentrations; and the interaction between PUFA intake and the 1131TC in the APOA5 gene in determining triglyceride metabolism. Although hundreds of diet-gene interaction studies in lipid metabolism have been published, the level of evidence to make specific nutritional recommendations to the population is still low and more research in nutrigenetics has to be undertaken.Las recomendaciones dietéticas actuales referentes al consumo de grasas en la dieta han sido realizadas sin tener en cuenta las posibles diferencias genéticas de las personas que podrían ser las responsables de las diferentes respuestas interindividuales que frecuentemente se observan ante la misma dieta. La presencia de variabilidad genética ha sido puesta de manifiesto para todos los genes relacionados con el metabolismo lipídico, por lo que existe un ingente número de genes y de variantes genéticas para ser incluidas en los estudios sobre interacciones dieta-genotipo en el ámbito específico del consumo de grasas y aceites. Se revisarán algunos ejemplos sobre interacciones grasa

  2. Evaluation of stability and simultaneous determination of fimasartan and amlodipine by a HPLC method in combination tablets

    Directory of Open Access Journals (Sweden)

    Hyeon Woo Moon

    2014-06-01

    Full Text Available A simple, rapid, accurate, precise and robust HPLC method was developed for the simultaneous determination of fimasartan and amlodipine in tablet dosage form. Furthermore, stability of active ingredients was evaluated under normal and stress conditions. The isocratic elution was accomplished by Nucleosil C18 column (250 mm × 4.6 mm, 5 μm at 40 °C. The mobile phase consisted of acetonitrile and 0.02 M monopotassium phosphate buffer (pH 2.2 in the ratio of 50:50 (v/v was eluted at 1.0 ml/min. The eluent was monitored by the UV detector for fimasartan and amlodipine at 237 nm for 8 min, detection time. The validation of HPLC method was carried out in accordance with the ICH guidelines.

  3. Determining the power-law wind-profile exponent under near-neutral stability conditions at sea

    Science.gov (United States)

    Hsu, S. A.; Meindl, Eric A.; Gilhousen, David B.

    1994-01-01

    On the basis of 30 samples from near-simultaneous overwater measurements by pairs of anemometers located at different heights in the Gulf of Mexico and off the Chesapeake Bay, Virginia, the mean and standard deviation for the exponent of the power-law wind profile over the ocean under near-neutral atmospheric stability conditions were determined to be 0.11 +/- 0.03. Because this mean value is obtained from both deep and shallow water environments, it is recommended for use at sea to adjust the wind speed measurements at different heights to the standard height of 10 m above the mean sea surface. An example to apply this P value to estimate the momentum flux or wind stress is provided.

  4. Performance analysis of a GPS Interferometric attitude determination system for a gravity gradient stabilized spacecraft. M.S. Thesis

    Science.gov (United States)

    Stoll, John C.

    1995-01-01

    The performance of an unaided attitude determination system based on GPS interferometry is examined using linear covariance analysis. The modelled system includes four GPS antennae onboard a gravity gradient stabilized spacecraft, specifically the Air Force's RADCAL satellite. The principal error sources are identified and modelled. The optimal system's sensitivities to these error sources are examined through an error budget and by varying system parameters. The effects of two satellite selection algorithms, Geometric and Attitude Dilution of Precision (GDOP and ADOP, respectively) are examined. The attitude performance of two optimal-suboptimal filters is also presented. Based on this analysis, the limiting factors in attitude accuracy are the knowledge of the relative antenna locations, the electrical path lengths from the antennae to the receiver, and the multipath environment. The performance of the system is found to be fairly insensitive to torque errors, orbital inclination, and the two satellite geometry figures-of-merit tested.

  5. Determining the power-law wind-profile exponent under near-neutral stability conditions at sea

    Science.gov (United States)

    Hsu, S. A.; Meindl, Eric A.; Gilhousen, David B.

    1994-01-01

    On the basis of 30 samples from near-simultaneous overwater measurements by pairs of anemometers located at different heights in the Gulf of Mexico and off the Chesapeake Bay, Virginia, the mean and standard deviation for the exponent of the power-law wind profile over the ocean under near-neutral atmospheric stability conditions were determined to be 0.11 +/- 0.03. Because this mean value is obtained from both deep and shallow water environments, it is recommended for use at sea to adjust the wind speed measurements at different heights to the standard height of 10 m above the mean sea surface. An example to apply this P value to estimate the momentum flux or wind stress is provided.

  6. Determination of Stability Constants of Triazole Ligand Carrying Naphthol Group with Heavy Metal Ions in Aqueous Solutions

    Directory of Open Access Journals (Sweden)

    Nurhan Gümrükçüoğlu

    2013-12-01

    Full Text Available Interaction of 4-(2-hydroxy-1-naphthylmethylamino-3-methyl-5-(4-tolyl-4H-1,2,4-triazole with heavy metal cations such as Cu2+, Co2+, Cd2+, Ni2+ and Pb2+ was investigated by using UV-visible spectrophotometric technique. The complex stability constants (Log β were determined in aqueous as well as in methanol: water (1:1 system at 25 ± 0.1°C by Buschmann’s method and Valeur’s methods, respectively. The ligand showed good sensitivity for Co2+ with a linear range of 210-6M to 310-5M.

  7. Stability and multiattractor dynamics of a toggle switch based on a two-stage model of stochastic gene expression.

    Science.gov (United States)

    Strasser, Michael; Theis, Fabian J; Marr, Carsten

    2012-01-04

    A toggle switch consists of two genes that mutually repress each other. This regulatory motif is active during cell differentiation and is thought to act as a memory device, being able to choose and maintain cell fate decisions. Commonly, this switch has been modeled in a deterministic framework where transcription and translation are lumped together. In this description, bistability occurs for transcription factor cooperativity, whereas autoactivation leads to a tristable system with an additional undecided state. In this contribution, we study the stability and dynamics of a two-stage gene expression switch within a probabilistic framework inspired by the properties of the Pu/Gata toggle switch in myeloid progenitor cells. We focus on low mRNA numbers, high protein abundance, and monomeric transcription-factor binding. Contrary to the expectation from a deterministic description, this switch shows complex multiattractor dynamics without autoactivation and cooperativity. Most importantly, the four attractors of the system, which only emerge in a probabilistic two-stage description, can be identified with committed and primed states in cell differentiation. To begin, we study the dynamics of the system and infer the mechanisms that move the system between attractors using both the quasipotential and the probability flux of the system. Next, we show that the residence times of the system in one of the committed attractors are geometrically distributed. We derive an analytical expression for the parameter of the geometric distribution, therefore completely describing the statistics of the switching process and elucidate the influence of the system parameters on the residence time. Moreover, we find that the mean residence time increases linearly with the mean protein level. This scaling also holds for a one-stage scenario and for autoactivation. Finally, we study the implications of this distribution for the stability of a switch and discuss the influence of the

  8. Determination of an acceptable assimilable organic carbon (AOC) level for biological stability in water distribution systems with minimized chlorine residual.

    Science.gov (United States)

    Ohkouchi, Yumiko; Ly, Bich Thuy; Ishikawa, Suguru; Kawano, Yoshihiro; Itoh, Sadahiko

    2013-02-01

    There is considerable interest in minimizing the chlorine residual in Japan because of increasing complaints about a chlorinous odor in drinking water. However, minimizing the chlorine residual causes the microbiological water quality to deteriorate, and stricter control of biodegradable organics in finished water is thus needed to maintain biological stability during water distribution. In this investigation, an acceptable level of assimilable organic carbon (AOC) for biologically stable water with minimized chlorine residual was determined based on the relationship between AOC, the chlorine residual, and bacterial regrowth. In order to prepare water samples containing lower AOC, the fractions of AOC and biodegradable organic matter (BOM) in tap water samples were reduced by converting into biomass after thermal hydrolysis of BOM at alkaline conditions. The batch-mode incubations at different conditions of AOC and chlorine residual were carried out at 20 °C, and the presence or absence of bacterial regrowth was determined. The determined curve for biologically stable water indicated that the acceptable AOC was 10.9 μg C/L at a minimized chlorine residual (0.05 mg Cl(2)/L). This result indicated that AOC removal during current water treatment processes in Japan should be significantly enhanced prior to minimization of the chlorine residual in water distribution.

  9. Thermodynamic stability of Ca{sub 3}TeO{sub 6} determined by a solid electrolyte EMF method

    Energy Technology Data Exchange (ETDEWEB)

    Sukhomlinov, D., E-mail: dmitry.sukhomlinov@aalto.fi [Aalto University School of Chemical Technology, Department of Materials Science and Engineering, Metallurgical Thermodynamics and Modeling Research Group, Vuorimiehentie 2 K, PO Box 16200, FI-00076 Aalto (Finland); Tesfaye, F. [Aalto University School of Chemical Technology, Department of Materials Science and Engineering, Metallurgical Thermodynamics and Modeling Research Group, Vuorimiehentie 2 K, PO Box 16200, FI-00076 Aalto (Finland); Åbo Akademi University, Process Chemistry Centre, Piispankatu 8, FI-20500 Turku (Finland); Taskinen, P. [Aalto University School of Chemical Technology, Department of Materials Science and Engineering, Metallurgical Thermodynamics and Modeling Research Group, Vuorimiehentie 2 K, PO Box 16200, FI-00076 Aalto (Finland)

    2015-09-10

    Highlights: • Gibbs energy of formation of Ca{sub 3}TeO{sub 6} experimentally determined for the first time. • Oxygen concentration galvanic cells based on YSZ solid electrolyte were employed. • In the Ca–Te–O system, Ca{sub 3}TeO{sub 6} coexists with CaO and Te. - Abstract: The standard thermodynamic properties of Ca{sub 3}TeO{sub 6} were determined electrochemically utilizing fast O{sup 2−} ion conducting solid electrolyte yttria-stabilized zirconia. The ternary phase was synthesized from the pure oxides CaO and TeO{sub 2} in excess of CaO. The electromotive force measurements were performed on two similar electrochemical cells of the type Te + CaO + Ca{sub 3}TeO{sub 6}|YSZ|O{sub 2}, within the temperature range from 850 to 949 K. The standard Gibbs energy of formation for the ternary compound Ca{sub 3}TeO{sub 6} was determined for the first time, based on the experimental data obtained.

  10. Nucleosome Stability Distinguishes Two Different Promoter Types at All Protein-Coding Genes in Yeast.

    Science.gov (United States)

    Kubik, Slawomir; Bruzzone, Maria Jessica; Jacquet, Philippe; Falcone, Jean-Luc; Rougemont, Jacques; Shore, David

    2015-11-01

    Previous studies indicate that eukaryotic promoters display a stereotypical chromatin landscape characterized by a well-positioned +1 nucleosome near the transcription start site and an upstream -1 nucleosome that together demarcate a nucleosome-free (or -depleted) region. Here we present evidence that there are two distinct types of promoters distinguished by the resistance of the -1 nucleosome to micrococcal nuclease digestion. These different architectures are characterized by two sequence motifs that are broadly deployed at one set of promoters where a nuclease-sensitive ("fragile") nucleosome forms, but concentrated in a narrower, nucleosome-free region at all other promoters. The RSC nucleosome remodeler acts through the motifs to establish stable +1 and -1 nucleosome positions, while binding of a small set of general regulatory (pioneer) factors at fragile nucleosome promoters plays a key role in their destabilization. We propose that the fragile nucleosome promoter architecture is adapted for regulation of highly expressed, growth-related genes.

  11. Interactions of cubilin with megalin and the product of the amnionless gene (AMN): effect on its stability.

    Science.gov (United States)

    Ahuja, Rajiv; Yammani, Raghunatha; Bauer, Joseph A; Kalra, Seema; Seetharam, Shakuntla; Seetharam, Bellur

    2008-03-01

    Cubilin, a 456 kDa multipurpose receptor lacking in both transmembrane and cytoplasmic domains is expressed in the apical BBMs (brush border membranes) of polarized epithelia. Cubilin interacts with two transmembrane proteins, AMN, a 45-50 kDa protein product of the amnionless gene, and megalin, a 600 kDa giant endocytic receptor. In vitro, three fragments of cubilin, the 113-residue N-terminus and CUB domains 12-17 and 22-27, demonstrated Ca2+-dependent binding to megalin. Immunoprecipitation and immunoblotting studies using detergent extracts of rat kidney BBMs revealed that cubilin interacts with both megalin and AMN. Ligand (intrinsic factor-cobalamin)-affinity chromatography showed that in renal BBMs, functional cubilin exists as a complex with both AMN and megalin. Cubilin and AMN levels were reduced by 80% and 55-60% respectively in total membranes and BBMs obtained from kidney of megalin antibody-producing rabbits. Immunohistochemical analysis and turnover studies for cubilin in megalin or AMN gene-silenced opossum kidney cells showed a significant reduction (85-90%) in cubilin staining and a 2-fold decrease in its half-life. Taken together, these results indicate that three distinct regions of cubilin bind to megalin and its interactions with both megalin and AMN are essential for its intracellular stability.

  12. Using data from the Microsoft Kinect 2 to determine postural stability in healthy subjects: A feasibility trial

    Science.gov (United States)

    Smeragliuolo, Anna H.; Long, John Davis; Bumanlag, Silverio Joseph; He, Victor; Lampe, Anna

    2017-01-01

    The objective of this study was to determine whether kinematic data collected by the Microsoft Kinect 2 (MK2) could be used to quantify postural stability in healthy subjects. Twelve subjects were recruited for the project, and were instructed to perform a sequence of simple postural stability tasks. The movement sequence was performed as subjects were seated on top of a force platform, and the MK2 was positioned in front of them. This sequence of tasks was performed by each subject under three different postural conditions: “both feet on the ground” (1), “One foot off the ground” (2), and “both feet off the ground” (3). We compared force platform and MK2 data to quantify the degree to which the MK2 was returning reliable data across subjects. We then applied a novel machine-learning paradigm to the MK2 data in order to determine the extent to which data from the MK2 could be used to reliably classify different postural conditions. Our initial comparison of force plate and MK2 data showed a strong agreement between the two devices, with strong Pearson correlations between the trunk centroids “Spine_Mid” (0.85 ± 0.06), “Neck” (0.86 ± 0.07) and “Head” (0.87 ± 0.07), and the center of pressure centroid inferred by the force platform. Mean accuracy for the machine learning classifier from MK2 was 97.0%, with a specific classification accuracy breakdown of 90.9%, 100%, and 100% for conditions 1 through 3, respectively. Mean accuracy for the machine learning classifier derived from the force platform data was lower at 84.4%. We conclude that data from the MK2 has sufficient information content to allow us to classify sequences of tasks being performed under different levels of postural stability. Future studies will focus on validating this protocol on large populations of individuals with actual balance impairments in order to create a toolkit that is clinically validated and available to the medical community. PMID:28196139

  13. Using data from the Microsoft Kinect 2 to determine postural stability in healthy subjects: A feasibility trial.

    Science.gov (United States)

    Dehbandi, Behdad; Barachant, Alexandre; Smeragliuolo, Anna H; Long, John Davis; Bumanlag, Silverio Joseph; He, Victor; Lampe, Anna; Putrino, David

    2017-01-01

    The objective of this study was to determine whether kinematic data collected by the Microsoft Kinect 2 (MK2) could be used to quantify postural stability in healthy subjects. Twelve subjects were recruited for the project, and were instructed to perform a sequence of simple postural stability tasks. The movement sequence was performed as subjects were seated on top of a force platform, and the MK2 was positioned in front of them. This sequence of tasks was performed by each subject under three different postural conditions: "both feet on the ground" (1), "One foot off the ground" (2), and "both feet off the ground" (3). We compared force platform and MK2 data to quantify the degree to which the MK2 was returning reliable data across subjects. We then applied a novel machine-learning paradigm to the MK2 data in order to determine the extent to which data from the MK2 could be used to reliably classify different postural conditions. Our initial comparison of force plate and MK2 data showed a strong agreement between the two devices, with strong Pearson correlations between the trunk centroids "Spine_Mid" (0.85 ± 0.06), "Neck" (0.86 ± 0.07) and "Head" (0.87 ± 0.07), and the center of pressure centroid inferred by the force platform. Mean accuracy for the machine learning classifier from MK2 was 97.0%, with a specific classification accuracy breakdown of 90.9%, 100%, and 100% for conditions 1 through 3, respectively. Mean accuracy for the machine learning classifier derived from the force platform data was lower at 84.4%. We conclude that data from the MK2 has sufficient information content to allow us to classify sequences of tasks being performed under different levels of postural stability. Future studies will focus on validating this protocol on large populations of individuals with actual balance impairments in order to create a toolkit that is clinically validated and available to the medical community.

  14. Insights on the Horizontal Gene Transfer of Carbapenemase Determinants in the Opportunistic Pathogen Acinetobacter baumannii

    Directory of Open Access Journals (Sweden)

    Gabriela Jorge Da Silva

    2016-08-01

    Full Text Available Horizontal gene transfer (HGT is a driving force to the evolution of bacteria. The fast emergence of antimicrobial resistance reflects the ability of genetic adaptation of pathogens. Acinetobacter baumannii has emerged in the last few decades as an important opportunistic nosocomial pathogen, in part due to its high capacity of acquiring resistance to diverse antibiotic families, including to the so-called last line drugs such as carbapenems. The rampant selective pressure and genetic exchange of resistance genes hinder the effective treatment of resistant infections. A. baumannii uses all the resistance mechanisms to survive against carbapenems but production of carbapenemases are the major mechanism, which may act in synergy with others. A. baumannii appears to use all the mechanisms of gene dissemination. Beyond conjugation, the mostly reported recent studies point to natural transformation, transduction and outer membrane vesicles-mediated transfer as mechanisms that may play a role in carbapenemase determinants spread. Understanding the genetic mobilization of carbapenemase genes is paramount in preventing their dissemination. Here we review the carbapenemases found in A. baumannii and present an overview of the current knowledge of contributions of the various HGT mechanisms to the molecular epidemiology of carbapenem resistance in this relevant opportunistic pathogen.

  15. Parameter estimation and determinability analysis applied to Drosophila gap gene circuits

    Directory of Open Access Journals (Sweden)

    Jaeger Johannes

    2008-09-01

    Full Text Available Abstract Background Mathematical modeling of real-life processes often requires the estimation of unknown parameters. Once the parameters are found by means of optimization, it is important to assess the quality of the parameter estimates, especially if parameter values are used to draw biological conclusions from the model. Results In this paper we describe how the quality of parameter estimates can be analyzed. We apply our methodology to assess parameter determinability for gene circuit models of the gap gene network in early Drosophila embryos. Conclusion Our analysis shows that none of the parameters of the considered model can be determined individually with reasonable accuracy due to correlations between parameters. Therefore, the model cannot be used as a tool to infer quantitative regulatory weights. On the other hand, our results show that it is still possible to draw reliable qualitative conclusions on the regulatory topology of the gene network. Moreover, it improves previous analyses of the same model by allowing us to identify those interactions for which qualitative conclusions are reliable, and those for which they are ambiguous.

  16. c-Jun N-terminal kinase phosphorylation of MARCKSL1 determines actin stability and migration in neurons and in cancer cells.

    Science.gov (United States)

    Björkblom, Benny; Padzik, Artur; Mohammad, Hasan; Westerlund, Nina; Komulainen, Emilia; Hollos, Patrik; Parviainen, Lotta; Papageorgiou, Anastassios C; Iljin, Kristiina; Kallioniemi, Olli; Kallajoki, Markku; Courtney, Michael J; Mågård, Mats; James, Peter; Coffey, Eleanor T

    2012-09-01

    Cell migration is a fundamental biological function, critical during development and regeneration, whereas deregulated migration underlies neurological birth defects and cancer metastasis. MARCKS-like protein 1 (MARCKSL1) is widely expressed in nervous tissue, where, like Jun N-terminal protein kinase (JNK), it is required for neural tube formation, though the mechanism is unknown. Here we show that MARCKSL1 is directly phosphorylated by JNK on C-terminal residues (S120, T148, and T183). This phosphorylation enables MARCKSL1 to bundle and stabilize F-actin, increase filopodium numbers and dynamics, and retard migration in neurons. Conversely, when MARCKSL1 phosphorylation is inhibited, actin mobility increases and filopodium formation is compromised whereas lamellipodium formation is enhanced, as is cell migration. We find that MARCKSL1 mRNA is upregulated in a broad range of cancer types and that MARCKSL1 protein is strongly induced in primary prostate carcinomas. Gene knockdown in prostate cancer cells or in neurons reveals a critical role for MARCKSL1 in migration that is dependent on the phosphorylation state; phosphomimetic MARCKSL1 (MARCKSL1(S120D,T148D,T183D)) inhibits whereas dephospho-MARCKSL1(S120A,T148A,T183A) induces migration. In summary, these data show that JNK phosphorylation of MARCKSL1 regulates actin homeostasis, filopodium and lamellipodium formation, and neuronal migration under physiological conditions and that, when ectopically expressed in prostate cancer cells, MARCKSL1 again determines cell movement.

  17. Determination of manganese in diesel, gasoline and naphtha by graphite furnace atomic absorption spectrometry using microemulsion medium for sample stabilization

    Science.gov (United States)

    Brandão, Geisamanda Pedrini; de Campos, Reinaldo Calixto; de Castro, Eustáquio Vinicius Ribeiro; de Jesus, Honério Coutinho

    2008-08-01

    The determination of Mn in diesel, gasoline and naphtha samples at µg L - 1 level by graphite furnace atomic absorption spectrometry, after sample stabilization in a three-component medium (microemulsion) was investigated. Microemulsions were prepared by mixing appropriate volumes of sample, propan-1-ol and nitric acid aqueous solution, and a stable system was immediately and spontaneously formed. After multivariate optimization by central composite design the optimum microemulsion composition as well as the temperature program was defined. In this way, calibration using aqueous analytical solution was possible, since the same sensitivity was observed in the optimized microemulsion media and 0.2% v/v HNO 3. The use of modifier was not necessary. Recoveries at the 3 µg L - 1 level using both inorganic and organic Mn standards spiked solutions ranged from 98 to 107% and the limits of detection were 0.6, 0.5 and 0.3 µg L - 1 in the original diesel, gasoline and naphtha samples, respectively. The Mn characteristic mass 3.4 pg. Typical relative standard deviation ( n = 5) of 8, 6 and 7% were found for the samples prepared as microemulsions at concentration levels of 1.3, 0.8, and 1.5 µg L - 1 , respectively. The total determination cycle lasted 4 min for diesel and 3 min for gasoline and naphtha, equivalent to a sample throughput of 7 h - 1 for duplicate determinations in diesel and 10 h - 1 for duplicate determinations in gasoline and naphtha. Accuracy was also assessed by using other method of analysis (ASTM D 3831-90). No statistically significant differences were found between the results obtained with the proposed method and the reference method in the analysis of real samples.

  18. Effective Surfactants Blend Concentration Determination for O/W Emulsion Stabilization by Two Nonionic Surfactants by Simple Linear Regression.

    Science.gov (United States)

    Hassan, A K

    2015-01-01

    In this work, O/W emulsion sets were prepared by using different concentrations of two nonionic surfactants. The two surfactants, tween 80(HLB=15.0) and span 80(HLB=4.3) were used in a fixed proportions equal to 0.55:0.45 respectively. HLB value of the surfactants blends were fixed at 10.185. The surfactants blend concentration is starting from 3% up to 19%. For each O/W emulsion set the conductivity was measured at room temperature (25±2°), 40, 50, 60, 70 and 80°. Applying the simple linear regression least squares method statistical analysis to the temperature-conductivity obtained data determines the effective surfactants blend concentration required for preparing the most stable O/W emulsion. These results were confirmed by applying the physical stability centrifugation testing and the phase inversion temperature range measurements. The results indicated that, the relation which represents the most stable O/W emulsion has the strongest direct linear relationship between temperature and conductivity. This relationship is linear up to 80°. This work proves that, the most stable O/W emulsion is determined via the determination of the maximum R² value by applying of the simple linear regression least squares method to the temperature-conductivity obtained data up to 80°, in addition to, the true maximum slope is represented by the equation which has the maximum R² value. Because the conditions would be changed in a more complex formulation, the method of the determination of the effective surfactants blend concentration was verified by applying it for more complex formulations of 2% O/W miconazole nitrate cream and the results indicate its reproducibility.

  19. CHD1 regulates cell fate determination by activation of differentiation-induced genes

    DEFF Research Database (Denmark)

    Baumgart, Simon J; Najafova, Zeynab; Hossan, Tareq

    2017-01-01

    . Furthermore, we observed that CHD1-dependent genes are mainly induced during osteoblast differentiation and are characterized by higher levels of CHD1 occupancy around the TSS. Interestingly, CHD1 depletion resulted in increased pausing of RNA Polymerase II (RNAPII) and decreased H2A.Z occupancy close......The coordinated temporal and spatial activation of gene expression is essential for proper stem cell differentiation. The Chromodomain Helicase DNA-binding protein 1 (CHD1) is a chromatin remodeler closely associated with transcription and nucleosome turnover downstream of the transcriptional start...... to the TSS, but not at enhancer regions. These findings reveal a novel role for CHD1 during osteoblast differentiation and provide further insights into the intricacies of epigenetic regulatory mechanisms controlling cell fate determination....

  20. STABILITY INDICATING LIQUID CHROMATOGRAPHIC METHOD FOR THE SIMULTANEOUS DETERMINATION OF OLMESARTAN MEDOXOMIL AND AZELNIDIPINE IN COMBINED TABLET DOSAGE FORM

    Directory of Open Access Journals (Sweden)

    Raveendra Babu Ganduri

    2014-06-01

    Full Text Available A stability indicating RP-HPLC method for the simultaneous determination of olmesartan medoxomil (OLM and azelnidipine from combined tablet dosage form was developed. The separation was accomplished on Inertsil 3V (4.6 mm X 100 mm; particle size 3 μm column using a mobile phase consisting of potassium dihydrogen phosphate buffer (pH adjusted to 3.0 with orthophosphoric acid and acetonitrile in gradient elution mode. The analytes were monitored by a photo diode array (PDA detector set at 255 nm and the flow rate was kept at 2.0 mL min-1. The retention time for olmesartan medoxomil and azelnidipine were 3.148 and 3.704 respectively. Linearity was observed in the concentration range of 10-60 μg/mL for olmesartan medoxomil and 4-24 μg/mL azelnidipine. Both the drugs were subjected to acid, alkali and neutral hydrolysis, oxidation, dry heat and photolytic degradation. The degradants were well resolved from the pure drugs. The method could be used for simultaneous determination of olmesartan medoxomil and azelnidipine in bulk and combined dosage form.

  1. The structure of HIV-1 genomic RNA in the gp120 gene determines a recombination hot spot in vivo.

    Science.gov (United States)

    Galetto, Román; Moumen, Abdeladim; Giacomoni, Véronique; Véron, Michel; Charneau, Pierre; Negroni, Matteo

    2004-08-27

    By frequently rearranging large regions of the genome, genetic recombination is a major determinant in the plasticity of the human immunodeficiency virus type I (HIV-1) population. In retroviruses, recombination mostly occurs by template switching during reverse transcription. The generation of retroviral vectors provides a means to study this process after a single cycle of infection of cells in culture. Using HIV-1-derived vectors, we present here the first characterization and estimate of the strength of a recombination hot spot in HIV-1 in vivo. In the hot spot region, located within the C2 portion of the gp120 envelope gene, the rate of recombination is up to ten times higher than in the surrounding regions. The hot region corresponds to a previously identified RNA hairpin structure. Although recombination breakpoints in vivo cluster in the top portion of the hairpin, the bias for template switching in this same region appears less marked in a cell-free system. By modulating the stability of this hairpin we were able to affect the local recombination rate both in vitro and in infected cells, indicating that the local folding of the genomic RNA is a major parameter in the recombination process. This characterization of reverse transcription products generated after a single cycle of infection provides insights in the understanding of the mechanism of recombination in vivo and suggests that specific regions of the genome might be prompted to yield different rates of evolution due to the presence of circumscribed recombination hot spots.

  2. Facet stability of crystals I. Factors determining the polyhedral (in)-stability of silver single crystals during electrocrystallization at high current densities

    Science.gov (United States)

    Nanev, Chr. N.; Rashkov, R. St.

    1992-06-01

    Loss of the polyhedral stability as a result of emerging depressions on crystal faces has been observed during both vapour and solution growth under diffusion control, as well as by electrocrystallization at high current densities. A difference was found only when a quantitative comparison of the stability of the crystal shapes with the existing theoretical predictions was attempted. With the growth of zinc and cadmium single crystals from the vapour phase this phenomenon appears earlier, i.e. at smaller sizes than the expected figures, while the silver single crystals are more steady — they withstand one order of magnitude higher of current densities than the calculated values before the appearance of the depressions, in spite of the fact that the presence of an (inhomogeneous) electrical field in the second case has to decrease the polyhedral stability. One possible explanation of this fact is that the electrocrystallization of silver proceeds in highly concentrated solutions, for which Seeger's equation, laying in the base of the quantitative elucidations in this case, does not hold true. Correspondingly, here (part I of the paper) we are trying a more general approach, while part II represents a new, alternative way for explaining the higher stability of the faceted forms of the silver single crystals.

  3. Simultaneous stability-indicating HPLC method for the determination of cisapride, methylparaben and propylparaben in oral suspension

    Directory of Open Access Journals (Sweden)

    Jutima Boonleang

    2010-08-01

    Full Text Available A simultaneous stability-indicating HPLC method for the determination of cisapride, methylparaben and propylparabenin oral suspensions has been developed and validated. Baseline separation was achieved on a C18 column at room temperature(25°C by gradient elution with mobile phase consisting of solvent A: 10% v/v acetonitrile in 0.13% w/v sodium-1-pentanesulfonate pH 8 and solvent B: acetonitrile. The gradient program was as follows: 0-5 min: 20 to 56% solvent B; 5-7min: 56 to 85% solvent B; 7-10 min: 85% solvent B. The flow rate of mobile phase was 1.2 mL/min. The injection volume was20 L. Detection and peak purity assessments were performed by photo-diode array detector set at 275 nm with scan modein the range of 190-400 nm. The method was selective, accurate and precise. It provided chromatograms with good peak shapeand acceptable resolutions of greater than 4.4 for all analytes including the degradation products formed in oral suspensionsin about 8.5 min. All analyte peaks were pure. The accuracy of all analytes was in the range of 99.20-100.6%. The within-runand between-run relative standard deviations were less than 1.50%. The calibration curves for cisapride, methylparaben, andpropylparaben were linear over the concentration range of 10.0-75.0 g/mL, 8.0-100.0 g/mL, and 0.8-10.0 g/mL, respectivelywith r2 greater than 0.999. This developed method was successfully applied to the stability study of cisapride, methylparabenand propylparaben in oral suspension formulations.

  4. Determination of Edoxaban in Bulk and in Tablet Dosage Form by Stability Indicating High-Performance Liquid Chromatography

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    Pasam Satyanarayana Reddy , V. Shanmukha Kumar Jagarlapudi, Chandra Bala Sekaran

    2016-03-01

    Full Text Available Background: Edoxaban is an orally active direct factor Xa inhibitor. The aim of the present study was to develop a stability indicating HPLC method for the quantification of edoxaban in bulk and in tablet dosage form. Methods: Edoxaban was separated on Hypersil BDS C18 column (250 x 4.6 mm, i.d. 5µm using 0.1M K2HPO4: Methanol (65:35, v/v as an isocratic mobile phase at a flow rate of 1.0 ml/min. Detection was performed using photodiode array detector set at 245 nm. The chromatographic conditions were optimized. The method was validated as per the guidelines given by International Conference on Harmonization guidelines. Results: Edoxaban was eluted at 3.785 min with a total run time of 6 min. The calibration curve was found to be linear over the concentration range of 5–200 μg/ml. Limit of detection and limit of quantification for edoxaban are 0.209 µg/ml and 0.698 µg/ml, respectively. The intra- and inter-day precision values were ≤0.710% and the accuracy ranged from 99.824-100.720%. Besides, all the validation results were within acceptability criteria of general assay. The stability indicating nature of the method was established by subjecting the edoxaban to stress conditions such as acid and base hydrolyses, oxidative, photo- and thermal degradations. The degraded products formed in all stress conditions were resolved successfully from the edoxaban. Conclusion: The developed and validated method is suitable for the determination of edoxaban in bulk and in commercial tablet dosage form.

  5. Stability-Indicating RP-HPLC Method for Determination of Guanfacine Hydrochloride in Bulk Drugs and in Pharmaceutical Dosage Form

    Directory of Open Access Journals (Sweden)

    Vinod K. Ahirrao

    2011-04-01

    Full Text Available A novel stability-indicating RP-HPLC method was developed and validated for quantitative determination of guanfacine hydrochloride in bulk drug and in pharmaceutical dosage form. An isocratic, reversed phase HPLC method was developed to separate the drug from the degradation products, using Apollo, C18 (250mm x 4.6mm, 5µm column with mobile phase of 50mM Ammonium acetate (volatile buffer and acetonitrile (65:35, v/v. UV detection has been done at wavelength 220 nm. The guanfacine hydrochloride was subjected to the stress conditions of hydrolysis (acid, base, oxidation, photolysis and thermal degradation. The stressed samples were analyzed by the proposed method. The analyte peak shape was excellent. The described method shows excellent linearity over a range of 30 – 450 µg/mL. The correlation coefficient for guanfacine hydrochloride was 0.999. The limit of detection for Guanfacine hydrochloride is 0.011 µg/mL and the limit of quantification is 0.038 µg/mL respectively.Degradation was observed for guanfacine hydrochloride in base, thermal and in 30% H2O2 conditions. The drug was found to be stable in the other stress conditions attempted. The degradation products were well resolved from main peak. The percentage recovery of guanfacine hydrochloride was ranged from (99.2% to 100.5% in pharmaceutical dosage form. The developed method was validated with respect to the linearity, accuracy (recovery, precision, specificity and robustness. The forced degradation studies prove the stability indicating power of the method.

  6. Quorum Sensing Coordinates Cooperative Expression of Pyruvate Metabolism Genes To Maintain a Sustainable Environment for Population Stability

    Science.gov (United States)

    Hawver, Lisa A.; Giulietti, Jennifer M.; Baleja, James D.

    2016-01-01

    ABSTRACT Quorum sensing (QS) is a microbial cell-cell communication system that regulates gene expression in response to population density to coordinate collective behaviors. Yet, the role of QS in resolving the stresses caused by the accumulation of toxic metabolic by-products at high cell density is not well defined. In response to cell density, QS could be involved in reprogramming of the metabolic network to maintain population stability. Using unbiased metabolomics, we discovered that Vibrio cholerae mutants genetically locked in a low cell density (LCD) QS state are unable to alter the pyruvate flux to convert fermentable carbon sources into neutral acetoin and 2,3-butanediol molecules to offset organic acid production. As a consequence, LCD-locked QS mutants rapidly lose viability when grown with fermentable carbon sources. This key metabolic switch relies on the QS-regulated small RNAs Qrr1-4 but is independent of known QS regulators AphA and HapR. Qrr1-4 dictate pyruvate flux by translational repression of the enzyme AlsS, which carries out the first step in acetoin and 2,3-butanediol biosynthesis. Consistent with the idea that QS facilitates the expression of a common trait in the population, AlsS needs to be expressed cooperatively in a group of cells. Heterogeneous populations with high percentages of cells not expressing AlsS are unstable. All of the cells, regardless of their respective QS states, succumb to stresses caused by toxic by-product accumulation. Our results indicate that the ability of the bacteria to cooperatively control metabolic flux through QS is critical in maintaining a sustainable environment and overall population stability. PMID:27923919

  7. Multidrug resistance 1 gene polymorphisms may determine Crohn's disease behavior in patients from Rio de Janeiro

    Directory of Open Access Journals (Sweden)

    Ana Teresa P. Carvalho

    2014-01-01

    Full Text Available OBJECTIVES: Conflicting data from studies on the potential role of multidrug resistance 1 gene polymorphisms in inflammatory bowel disease may result from the analysis of genetically and geographically distinct populations. Here, we investigated whether multidrug resistance 1 gene polymorphisms are associated with inflammatory bowel diseases in patients from Rio de Janeiro. METHODS: We analyzed 123 Crohn's disease patients and 83 ulcerative colitis patients to determine the presence of the multidrug resistance 1 gene polymorphisms C1236T, G2677T and C3435T. In particular, the genotype frequencies of Crohn's disease and ulcerative colitis patients were analyzed. Genotype-phenotype associations with major clinical characteristics were established, and estimated risks were calculated for the mutations. RESULTS: No significant difference was observed in the genotype frequencies of the multidrug resistance 1 G2677T/A and C3435T polymorphisms between Crohn's disease and ulcerative colitis patients. In contrast, the C1236T polymorphism was significantly more common in Crohn's disease than in ulcerative colitis (p = 0.047. A significant association was also found between the multidrug resistance 1 C3435T polymorphism and the stricturing form of Crohn's disease (OR: 4.13; p = 0.009, whereas no association was found with penetrating behavior (OR: 0.33; p = 0.094. In Crohn's disease, a positive association was also found between the C3435T polymorphism and corticosteroid resistance/refractoriness (OR: 4.14; p = 0.010. However, no significant association was found between multidrug resistance 1 gene polymorphisms and UC subphenotypic categories. CONCLUSION: The multidrug resistance 1 gene polymorphism C3435T is associated with the stricturing phenotype and an inappropriate response to therapy in Crohn's disease. This association with Crohn's disease may support additional pathogenic roles for the multidrug resistance 1 gene in regulating gut

  8. Determination of Asymptomatic Chlamydia Trachomatis Infections by Omp1 Gene Based -PCR

    Directory of Open Access Journals (Sweden)

    Masoumeh Nazer

    2008-01-01

    Full Text Available Objective: The objective of this research was to determine the prevalenceof genital C. trachomatis infection in asymptomatic women by using highlysensitive nested-polymerase chain reaction (PCR in urine sample.Materials and Methods: One hundred-forty asymptomatic women wererandomly selected from those who attended gynecology out patient departmentof Hazraat e Rasool Hospital in Tehran. First catch urine specimen were collectedfrom all the participants. DNA extraction was performed by means of HighPure PCR Template Preparation Kit (HPPTP according to the manufacture’sinstructions. Extracted DNA was tested by omp1 gene based nested-PCR,using sets of primers to amplify C. trachomatis omp1 gene. Visualizationof a 1027 bp fragment from omp1 gene in agarose gel electrophoresis wasconsidered as a positive result.Results: In total, 140 urines were tested for determination of C. trachomatisinfection. C. trachomatis omp-1 was detected in 22.1% of cases (31/140. Theoverall prevalence rates of C. trachomatis in the urine sample as determined byomp1 based nested-PCR were 4.3% in group I (age, 44 years. The highest prevalence of C. trachomatis infection (12.1% wasseen in women aged 25-34 years. This finding was not statistically significant(p=0.710. Also, there was not relation between C. trachomatis infection andsome probable risk factors such as young age (<25 years, STD history andmissing use of barrier contraceptive in this study.Conclusion: The prevalence of C.trachomatis infection in the women notseeking health care warrants more comprehensive study using high sensitiveomp1 based nested- PCR to identify and treat a large number of infectedwomen in Iran.

  9. Sequence length polymorphisms within primate amelogenin and amelogenin-like genes: usefulness in sex determination.

    Science.gov (United States)

    Morrill, Benson H; Rickords, Lee F; Schafstall, Heather J

    2008-10-01

    Sequence length polymorphisms between the amelogenin (AMELX) and the amelogenin-like (AMELY) genes both within and between several mammalian species have been identified and utilized for sex determination, species identification, and to elucidate evolutionary relationships. Sex determination via polymerase chain reaction (PCR) assays of the AMELX and AMELY genes has been successful in greater apes, prosimians, and two species of old world monkeys. To date, no sex determination PCR assay using AMELX and AMELY has been developed for new world monkeys. In this study, we present partial AMELX and AMELY sequences for five old world monkey species (Mandrillus sphinx, Macaca nemestrina, Macaca fuscata, Macaca mulatta, and Macaca fascicularis) along with primer sets that can be used for sex determination of these five species. In addition, we compare the sequences we generated with other primate AMELX and AMELY sequences available on GenBank and discuss sequence length polymorphisms and their usefulness in sex determination within primates. The mandrill and four species of macaque all share two similar deletion regions with each other, the human, and the chimpanzee in the region sequenced. These two deletion regions are 176-181 and 8 nucleotides in length. In analyzing existing primate sequences on GenBank, we also discovered that a separate six-nucleotide polymorphism located approximately 300 nucleotides upstream of the 177 nucleotide polymorphism in sequences of humans and chimps was also present in two species of new world monkeys (Saimiri boliviensis and Saimiri sciureus). We designed primers that incorporate this polymorphism, creating the first AMELX and AMELY PCR primer set that has been used successfully to generate two bands in a new world monkey species.

  10. Stability-indicating methods for the determination of erdosteine in the presence of its acid degradation products.

    Science.gov (United States)

    Moustafa, Nadia M; Badawey, Amr M; Lamie, Nesrine T; El-Aleem, Abd El-Aziz B Abd

    2014-01-01

    Four accurate, sensitive, and reproducible stability-indicating methods for the determination of erdosteine in the presence of its acid degradation products are presented. The first method involves processing the spectra by using a first-derivative method at 229 nm in a concentration range of 10-70 microg/mL. The mean percentage recovery was 100.43 +/- 0.977. The second method is based on ratio-spectra first derivative spectrophotometry at 227.4 and 255 nm over a concentration range of 10-70 microg/mL. The mean percentage recovery was 99.65 +/- 1.122% and 100.02 +/- 1.306% at 227.4 and 255 nm, respectively. The third method utilizes quantitative densitometric evaluation of the TLC of erdosteine in the presence of its acid degradation products, and uses methanol-chloroform-ammonia (7 + 3 +/- 0.01, v/v/v) as the mobile phase. TLC chromatograms were scanned at 235 nm. This method analyzes erdosteine in a concentration range of 2.4-5.6 microg/spot, with a mean percentage recovery of 100.03 +/- 1.015%. The fourth method is HPLC for the simultaneous determination of erdosteine in the presence of its acid degradation products. The mobile phase consists of water-methanol (65 + 35, v/v). The standard curve of erdosteine showed good linearity over a concentration range of 10-80 microg/mL, with a mean percentage recovery of 99.90 +/- 1.207%. These methods were successfully applied to the determination of erdosteine in bulk powder, laboratory-prepared mixtures containing different percentages of the degradation products, and pharmaceutical dosage forms. The validity of results was assessed by applying the standard addition technique. The results obtained agreed statistically with those obtained by a reported method, showing no significant differences with respect to accuracy and precision.

  11. The Role of Retinal Determination Gene Network (RDGN) in Hormone Signaling Transduction and Prostate Tumorigenes

    Science.gov (United States)

    2015-12-01

    crossed with Probasin-erbB2Δ (Pb-erbB2) transgenic mice to create triple transgenic mice, Dach1fl/fl/Pb-Cre/ Pb-erbB2. A). In order to determine the role...DACH1 blocked YB-1–induced mammary tumor growth and EMT in mice D). Conditional Dach1 gene knockout in the prostate demonstrates a role for...DACH1 to restrain AR negative and AR positive prostate cancer cell contact independent growth. In PC3 cells DACH1 inhibited colony formation (Fig.2a

  12. EXPRESSING HUMAN MATURED BRAIN-DERIVED NEUROTROPHIC FACTOR GENE IN E. Coli AND DETERMINING ITS BIOACTIVITY

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Objective Expressing the human matured brain-derived neurotrophic factor (mBDNF) gene in E.Coli and determining its bioactivity. Methods The resulting gene of mBDNF was subcloned into the EcoRI-BamHI site of the expression vector plasmid pBV220. The ligation products were used to transform the competent E. Coli DH5α. The proteins of mBDNF were experessed by temperature inducing. The expression products were dealed with solubilizing inclusion bodies and refolding protein. It was introduced into the embryonic chicken DRG to test whether the expressed mBDNF is a biologically active protein. Results The recombinant plasmid pBV/mBDNF was successfully constructed. By temperature inducing,under the control of the bacteriophage λ PL promoter, the experessed mBDNF protein was a 14Kd non-fusion protein,which existed in E. Coli as inclusion bodies. The size of expressed mBDNF is identical to the prediction. Bioactivity of the products was proved that it could support the cell survival and neurite growth in the primary cultures of embryonic 8-day-old chicken DRG neurons as compared to control.Conclusion The mBDNF gene can be expressed bioactively in E. Coli.

  13. Drosophila Hox and sex-determination genes control segment elimination through EGFR and extramacrochetae activity.

    Directory of Open Access Journals (Sweden)

    David Foronda

    Full Text Available The formation or suppression of particular structures is a major change occurring in development and evolution. One example of such change is the absence of the seventh abdominal segment (A7 in Drosophila males. We show here that there is a down-regulation of EGFR activity and fewer histoblasts in the male A7 in early pupae. If this activity is elevated, cell number increases and a small segment develops in the adult. At later pupal stages, the remaining precursors of the A7 are extruded under the epithelium. This extrusion requires the up-regulation of the HLH protein Extramacrochetae and correlates with high levels of spaghetti-squash, the gene encoding the regulatory light chain of the non-muscle myosin II. The Hox gene Abdominal-B controls both the down-regulation of spitz, a ligand of the EGFR pathway, and the up-regulation of extramacrochetae, and also regulates the transcription of the sex-determining gene doublesex. The male Doublesex protein, in turn, controls extramacrochetae and spaghetti-squash expression. In females, the EGFR pathway is also down-regulated in the A7 but extramacrochetae and spaghetti-squash are not up-regulated and extrusion of precursor cells is almost absent. Our results show the complex orchestration of cellular and genetic events that lead to this important sexually dimorphic character change.

  14. Adaptive Evolution of cry Genes in Bacillus thuringiensis:Implications for Their Specificity Determination

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    The cry gene family, produced during the late exponential phase of growth in Bacillus thuringiensis, is a large, still-growing family of homologous genes, in which each gene encodes a protein with strong specific activity against only one or a few insect species. Extensive studies are mostly focusing on the structural and functional relationships of Cry proteins, and have revealed several residues or domains that are important for the target recognition and receptor attachment. In this study,we have employed a maximum likelihood method to detect evidence of adaptive evolution in Cry proteins, and have identified 24 positively selected residues, which are all located in Domain Ⅱ or Ⅲ. Combined with known data from mutagenesis studies, the majority of these residues, at the molecular level, contribute much to the insect specificity determination. We postulate that the potential pressures driving the diversification of Cry proteins may be in an attempt to adapt for the "arm race" between δ-endotoxins and the targeted insects, or to enlarge their target spectra, hence result in the functional divergence. The sites identified to be under positive selection would provide targets for further structural and functional analyses on Cry proteins.

  15. Fine-mapping of genes determining extrafusal fiber properties in murine soleus muscle.

    Science.gov (United States)

    Carroll, A M; Cheng, R; Collie-Duguid, E S R; Meharg, C; Scholz, M E; Fiering, S; Fields, J L; Palmer, A A; Lionikas, A

    2017-03-01

    Muscle fiber cross-sectional area (CSA) and proportion of different fiber types are important determinants of muscle function and overall metabolism. Genetic variation plays a substantial role in phenotypic variation of these traits; however, the underlying genes remain poorly understood. This study aimed to map quantitative trait loci (QTL) affecting differences in soleus muscle fiber traits between the LG/J and SM/J mouse strains. Fiber number, CSA, and proportion of oxidative type I fibers were assessed in the soleus of 334 genotyped female and male mice of the F34 generation of advanced intercross lines (AIL) derived from the LG/J and SM/J strains. To increase the QTL detection power, these data were combined with 94 soleus samples from the F2 intercross of the same strains. Transcriptome of the soleus muscle of LG/J and SM/J females was analyzed by microarray. Genome-wide association analysis mapped four QTL (genome-wide P < 0.05) affecting the properties of muscle fibers to chromosome 2, 3, 4, and 11. A 1.5-LOD QTL support interval ranged between 2.36 and 4.67 Mb. On the basis of the genomic sequence information and functional and transcriptome data, we identified candidate genes for each of these QTL. The combination of analyses in F2 and F34 AIL populations with transcriptome and genomic sequence data in the parental strains is an effective strategy for refining QTL and nomination of the candidate genes.

  16. A Novel Candidate Gene for Temperature-Dependent Sex Determination in the Common Snapping Turtle.

    Science.gov (United States)

    Schroeder, Anthony L; Metzger, Kelsey J; Miller, Alexandra; Rhen, Turk

    2016-05-01

    Temperature-dependent sex determination (TSD) was described nearly 50 years ago. Researchers have since identified many genes that display differential expression at male- vs. female-producing temperatures. Yet, it is unclear whether these genes (1) are involved in sex determination per se, (2) are downstream effectors involved in differentiation of ovaries and testes, or (3) are thermo-sensitive but unrelated to gonad development. Here we present multiple lines of evidence linking CIRBP to sex determination in the snapping turtle, Chelydra serpentina We demonstrate significant associations between a single nucleotide polymorphism (SNP) (c63A > C) in CIRBP, transcript levels in embryonic gonads during specification of gonad fate, and sex in hatchlings from a thermal regime that produces mixed sex ratios. The A allele was induced in embryos exposed to a female-producing temperature, while expression of the C allele did not differ between female- and male-producing temperatures. In accord with this pattern of temperature-dependent, allele-specific expression, AA homozygotes were more likely to develop ovaries than AC heterozygotes, which, in turn, were more likely to develop ovaries than CC homozygotes. Multiple regression using SNPs in CIRBP and adjacent loci suggests that c63A > C may be the causal variant or closely linked to it. Differences in CIRBP allele frequencies among turtles from northern Minnesota, southern Minnesota, and Texas reflect small and large-scale latitudinal differences in TSD pattern. Finally, analysis of CIRBP protein localization reveals that CIRBP is in a position to mediate temperature effects on the developing gonads. Together, these studies strongly suggest that CIRBP is involved in determining the fate of the bipotential gonad.

  17. Gene dosage as a relevant mechanism contributing to the determination of ovarian function in Turner syndrome

    Science.gov (United States)

    Castronovo, Chiara; Rossetti, Raffaella; Rusconi, Daniela; Recalcati, Maria P.; Cacciatore, Chiara; Beccaria, Elena; Calcaterra, Valeria; Invernizzi, Pietro; Larizza, Daniela; Finelli, Palma; Persani, Luca

    2014-01-01

    STUDY QUESTION What is the burden of X chromosome mosaicism in the occurrence of spontaneous menarche (SM) in Turner syndrome (TS)? SUMMARY ANSWER SM was significantly associated with X chromosome mosaicism in the TS patients; a mosaicism with around 10% euploid cell line may predict spontaneous pubertal development when determined by molecular-cytogenetic techniques on uncultivated tissues. WHAT IS KNOWN ALREADY Spontaneous puberty can be observed in a minority of patients with TS, more frequently, but not exclusively, in those with a high level of 46,XX/45,X mosaicism at standard karyotype. The genetic mechanisms contributing to ovarian function in TS patients are still not determined. However, submicroscopic X-linked and autosomal copy number variations (CNVs) have recently emerged as an important genetic risk category for premature ovarian insufficiency and may be involved in modulating the TS ovarian phenotype. STUDY DESIGN, SIZE, DURATION A group of 40 patients with a diagnosis of TS at conventional karyotyping participated in the study; 6 patients had SM and 34 patients had primary amenorrhoea (PA). All clinical data and the patients’ DNA samples were collected over the years at a single paediatric clinic. PARTICIPANTS/MATERIALS, SETTING, METHODS The patients' samples were used to perform both genetic (Copy Number Assay) and molecular-cytogenetic (array-CGH and iFISH, interphase-FISH) analyses in order to evaluate the X chromosome mosaicism rate and to detect possible rare CNVs of genes with a known or predicted role in female fertility. MAIN RESULTS AND THE ROLE OF CHANCE All TS patients showed variable percentages of the 46,XX lineage, but these percentages were higher in the SM group (P < 0.01). A mosaicism around 10% for the euploid cell line may predict spontaneous pubertal development when determined by molecular-cytogenetic techniques performed in uncultivated tissues. A few CNVs involving autosomal and X-linked ovary-related loci were identified by

  18. Members of a large retroposon family are determinants of post-transcriptional gene expression in Leishmania.

    Directory of Open Access Journals (Sweden)

    Frédéric Bringaud

    2007-09-01

    Full Text Available Trypanosomatids are unicellular protists that include the human pathogens Leishmania spp. (leishmaniasis, Trypanosoma brucei (sleeping sickness, and Trypanosoma cruzi (Chagas disease. Analysis of their recently completed genomes confirmed the presence of non-long-terminal repeat retrotransposons, also called retroposons. Using the 79-bp signature sequence common to all trypanosomatid retroposons as bait, we identified in the Leishmania major genome two new large families of small elements--LmSIDER1 (785 copies and LmSIDER2 (1,073 copies--that fulfill all the characteristics of extinct trypanosomatid retroposons. LmSIDERs are approximately 70 times more abundant in L. major compared to T. brucei and are found almost exclusively within the 3'-untranslated regions (3'UTRs of L. major mRNAs. We provide experimental evidence that LmSIDER2 act as mRNA instability elements and that LmSIDER2-containing mRNAs are generally expressed at lower levels compared to the non-LmSIDER2 mRNAs. The considerable expansion of LmSIDERs within 3'UTRs in an organism lacking transcriptional control and their role in regulating mRNA stability indicate that Leishmania have probably recycled these short retroposons to globally modulate the expression of a number of genes. To our knowledge, this is the first example in eukaryotes of the domestication and expansion of a family of mobile elements that have evolved to fulfill a critical cellular function.

  19. Chromosomal antioxidant genes have metal ion-specific roles as determinants of bacterial metal tolerance.

    Science.gov (United States)

    Harrison, Joe J; Tremaroli, Valentina; Stan, Michelle A; Chan, Catherine S; Vacchi-Suzzi, Caterina; Heyne, Belinda J; Parsek, Matthew R; Ceri, Howard; Turner, Raymond J

    2009-10-01

    Microbiological metal toxicity involves redox reactions between metal species and cellular molecules, and therefore, we hypothesized that antioxidant systems might be chromosomal determinants affecting the susceptibility of bacteria to metal toxicity. Here, survival was quantified in metal ion-exposed planktonic cultures of several Escherichia coli strains, each bearing a mutation in a gene important for redox homeostasis. This characterized approximately 250 gene-metal combinations and identified that sodA, sodB, gor, trxA, gshA, grxA and marR have distinct roles in safeguarding or sensitizing cells to different toxic metal ions (Cr(2)O(7)(2-), Co(2+), Cu(2+), Ag(+), Zn(2+), AsO(2)(-), SeO(3)(2-) or TeO(3)(2-)). To shed light on these observations, fluorescent sensors for reactive oxygen species (ROS) and reduced thiol (RSH) quantification were used to ascertain that different metal ions exert oxidative toxicity through disparate modes-of-action. These oxidative mechanisms of metal toxicity were categorized as involving ROS and thiol-disulfide chemistry together (AsO(2)(-), SeO(3)(2-)), ROS predominantly (Cu(2+), Cr(2)O(7)(2-)) or thiol-disulfide chemistry predominantly (Ag(+), Co(2+), Zn(2+), TeO(3)(2-)). Corresponding to this, promoter-luxCDABE fusions showed that toxic doses of different metal ions up- or downregulate the transcription of gene sets marking distinct pathways of cellular oxidative stress. Altogether, our findings suggest that different metal ions are lethal to cells through discrete pathways of oxidative biochemistry, and moreover, indicate that chromosomally encoded antioxidant systems may have metal ion-specific physiological roles as determinants of bacterial metal tolerance.

  20. Use of SNPs to determine the breakpoints of complex deficiencies, facilitating gene mapping in Caenorhabditis elegans

    Directory of Open Access Journals (Sweden)

    Hoffmann Melissa

    2005-05-01

    Full Text Available Abstract Background Genetic deletions or deficiencies have been used for gene mapping and discovery in various organisms, ranging from the nematode Caenorhabditis elegans all the way to humans. One problem with large deletions is the determination of the location of their breakpoints. This is exacerbated in the case of complex deficiencies that delete a region of the genome, while retaining some of the intervening sequence. Previous methods, using genetic complementation or cytology were hampered by low marker density and were consequently not very precise at positioning the breakpoints of complex deficiencies. The identification of increasing numbers of Single Nucleotide Polymorphisms (SNPs has resulted in the use of these as genetic markers, and consequently in their utilization for defining the breakpoints of deletions using molecular biology methods. Results Here, we show that SNPs can be used to help position the breakpoints of a complex deficiency in C. elegans. The technique uses a combination of genetic crosses and molecular biology to generate robust and highly reproducible results with strong internal controls when trying to determine the breakpoints of deficiencies. The combined use of this technique and standard genetic mapping allowed us to rapidly narrow down the region of interest in our attempts to clone a gene. Conclusion Unlike previous methods used to locate deficiency breakpoints, our technique has the advantage of not being limited by the amount of starting material. It also incorporates internal controls to eliminate false positives and negatives. The technique can also easily be adapted for use in other organisms in which both genetic deficiencies and SNPs are available, thereby aiding gene discovery in these other models.

  1. Water-vortex-stabilized electric arc: III. Radial energy transport, determination of water-vapour-boundary and arc performance

    Science.gov (United States)

    Jenista, Jirí

    2003-12-01

    This paper is concerned with numerical modelling of an electric arc stabilized by a water vortex. The two-dimensional axisymmetric model presented includes the arc discharge area between the cathode and the outlet nozzle of the water plasma torch. The aims of the numerical simulations are: (1) to assess the influence of radial position of the water-vapour-boundary in the discharge chamber on arc performance and overall radial energy transport within the arc; (2) to determine the most probable mass flow rates and radii of the water-vapour-boundary in the discharge chamber for a prescribed current; (3) to demonstrate arc performance for two radiation models involved; and (4) to estimate validity of local thermodynamic equilibrium (LTE) conditions within the arc column. The rate of evaporation of water is calculated from the conduction and radiation heat fluxes at the water vapour surface for the specified mass flow rate. The behaviour of such an arc has been studied for a range of current 300-600 A. It is shown that changes of bulk magnitudes of different terms in the momentum and energy equations within the arc column as a function of arc radius enable us to reveal transitions of temperature and velocity fields from one steady state to a qualitatively different one. The best fit between experiment and numerical simulation for all currents exists for the mean arc radius ~3.3 mm. Deviations from LTE within the arc column are estimated with the criteria for kinetic equilibrium and spatial temperature gradients.

  2. Development and validation of stability indicating HPLC and HPTLC methods for determination of sulpiride and mebeverine hydrochloride in combination.

    Science.gov (United States)

    Naguib, Ibrahim A; Abdelkawy, Mohammed

    2010-09-01

    Validated sensitive and highly selective stability indicating methods are adopted for simultaneous quantitative determination of sulpiride and mebeverine hydrochloride in presence of their reported impurities and hydrolytic degradates whether in pure forms or in pharmaceutical formulation. The first method is High Performance Liquid Chromatography, where the mixture of sulpiride and mebeverine hydrochloride together with the reported interferents plus metopimazine as internal standard are separated on a reversed phase cyano column (5 microm ps, 250 mm x 4.6 id) using acetonitrile: water (70:30 v/v) adjusted to pH = 7 as a mobile phase. The drugs were detected at 221 nm over a concentration range of 5-40 microg ml(-1) and 5-60 microg ml(-1) with mean percentage recoveries 99.75% (S.D. 0.910) and 99.99% (S.D. 0.450) for sulpiride and mebeverine hydrochloride respectively. The second method is High Performance Thin Layer Chromatography, where sulpiride and mebeverine hydrochloride are separated on silica gel HPTLC F(254) plates using absolute ethanol:methylene chloride:triethyl amine (7:3:0.2 by volume) as mobile phase and scanning of the separated bands at 221 nm over a concentration range of 0.4-1.4 and 0.2-1.6 microg band(-1) with mean percentage recoveries 101.01% (S.D. 1.991) and 100.40% (S.D. 1.868) for sulpiride and mebeverine hydrochloride respectively.

  3. Validated stability-indicating HPLC method for the determination of mesna in presence of its degradation products.

    Science.gov (United States)

    Rizk, Mohamed; Taha, Elham A; Mowaka, Shereen; Abdallah, Youmna M

    2015-01-01

    A rapid and simple stability-indicating liquid chromatographic method was developed and validated for analysis of mesna in the presence of its degradation products in drug substance and drug products in a run time not >5 min. The separation was achieved on a RP amide C16 column at room temperature using methanol-phosphate buffer (10:90, v/v, pH 3.0) as mobile phase at a flow rate of 1 mL min(-1) and UV detection at 210 nm. The detector response for mesna was linear over the selected concentration range from 50 to 1000 µg mL(-1) with a correlation coefficient 0.9998. The limit of detection and the limit of quantitation were 7.5 and 22.7 µg mL(-1), respectively. The solution was stable for at least 5 days. Baseline resolution between mesna and its degradation products was achieved. Diode array detection peak purity tests showed no peak interfered with mesna peak. Moreover, the method was successfully applied for the determination of mesna in two different commercially available drug products.

  4. Stability-indicating LC-UV method for the determination of eszopiclone and degradation impurities in tablet dosage form.

    Science.gov (United States)

    Shaikh, Kabeer; Patil, Ashish; Gite, Sandeep

    2014-04-01

    A sensitive, stability-indicating reversed-phase high-performance liquid chromatographic method was developed for the determination of eszopiclone and related impurities in tablet dosage form. The chromatographic separation was achieved on an Inertsil C18 column (250 × 4.6 mm, 5 µm), using a mobile phase consisting of 0.05M monobasic sodium phosphate buffer containing 0.8% sodium lauryl sulfate (pH 3.5) and acetonitrile in the ratio of 60:40 (v/v), at a flow rate of 1.5 mL/min and temperature of 40°C. Quantification was achieved with photodiode array detection at 303 nm. The described method showed excellent linearity over a range of limits of quantification to 4.8 µg/mL (150% of specification limit; i.e., 3.2 µg/mL). The drug product was subjected to the stress conditions of oxidative, acid, base, thermal and photolytic degradation. Eszopiclone degradation was observed in acid hydrolysis, base hydrolysis and peroxide stress conditions. Eszopiclone was stable in thermal and photolytic degradation conditions. The developed method is simple, selective and accurate for the quantification of impurities and degradation products of eszopiclone in tablet dosage form.

  5. Validation of a stability-indicating RP-LC method for the determination of tigecycline in lyophilized powder.

    Science.gov (United States)

    da Silva, Lucélia Magalhães; Salgado, Hérida Regina Nunes

    2013-02-01

    A reversed-phase liquid chromatography (RP-LC) method was validated for the determination of tigecycline in lyophilized powder. The LC method was conducted on a Luna C18 column (250 × 4.6 mm i.d.), maintained at room temperature. The mobile phase consisted of buffer containing sodium phosphate monobasic (0.015M) and oxalic acid (0.015M) (pH 7.0)-acetonitrile (75:25, v/v), run at a flow rate of 1.0 mL/min and using ultraviolet detection at 280 nm. The chromatographic separation was obtained with a retention time of 8.6 min, and was linear in the range of 40-100 µg/mL (r(2) = 0.9997). The specificity and stability-indicating capability of the method was proven through forced degradation studies, which also showed no interference of the excipients. The accuracy was 99.01% with a bias lower than 1.81%. The limits of detection and quantitation were 1.67 and 5.05 µg/mL, respectively. Moreover, method validation demonstrated satisfactory results for precision and robustness. The proposed method was applied for the analysis of the lyophilized powder formulation, contributing to improve the quality control and to assure the therapeutic efficacy.

  6. Development and validation of a stability-indicating liquid chromatography method for the determination of dabigatran etexilate in capsules.

    Science.gov (United States)

    Bernardi, Raquel M; Fröehlich, Pedro E; Bergold, Ana M

    2013-01-01

    An RP-LC method was developed and validated for the determination of dabigatran etexilate in a capsule formulation. The LC method was carried out on a Zorbax C18 column (250 x 4.6 mm id). The mobile phase consisted of acetonitrile and a solution of triethylamine 0.1%, pH 6.0 adjusted with phosphoric acid (65 + 35, v/v) at a flow rate of 1.0 mL/min. The diode array detector was set at 225 nm. The chromatographic separation was obtained with retention time of 6.31 min, and linearity was in the range of 10-70 microg/mL (R2 = 0.9991). The specificity and stability-indicating capability of the method was proven through degradation studies and showing that there was no interference from the excipients. The accuracy was 100.23%, with an RSD of 1.34%. The LOD and LOQ were 0.04 and 10 microg/mL, respectively. Moreover, method validation demonstrated acceptable results for robustness and a system suitability test. The proposed method was applied for the analysis of the capsules to ensure their therapeutic efficacy.

  7. A validated stability-indicative UPLC method for nilotinib hydrochloride for the determination of process-related and degradation impurities.

    Science.gov (United States)

    Kondra, Sudhakar Babu; Madireddy, Venkataramanna; Chilukuri, Mohanareddy; Papadasu, Narayanareddy; Jonnalagadda, Latha

    2014-09-01

    A novel stability-indicating ultra performance liquid chromatographic (UPLC) method has been developed for quantitative determination of nilotinib hydrochloride in active pharmaceutical ingredients along with four impurities (imp-1, imp-2, imp-3 and imp-4). The method is applicable to the quantification of related compounds and assay of nilotinib hydrochloride drug. Efficient chromatographic separation was achieved on a Shim-pack XR-ODS II, 75 × 3.0 mm, 1.8-µm column with a gradient mobile phase combination. Quantification was carried at 260 nm at a flow rate of 0.6 mL min(-1). Stress degradation conditions were established for nilotinib hydrochloride by subjecting it to acid, base, oxidation, humidity, thermal and photolysis. The stress samples were assayed against a qualified reference standard and the mass balance was found close to 97.0%. The developed UPLC method was validated according to the present International Conference on Harmonisation guidelines for specificity, detection limit, quantitation limit, linearity, accuracy, precision, intermediate precision and robustness. The resolution between nilotinib hydrochloride and four potential impurities is found to be >2.0. Regression analysis shows as r value (correlation coefficient) of >0.999 for nilotinib hydrochloride and four potential impurities.

  8. Stability indicating spectrophotometric and spectrodensitometric methods for the determination of diatrizoate sodium in presence of its degradation product

    Science.gov (United States)

    Abd El-Rahman, Mohamed K.; Riad, Safaa M.; Abdel Gawad, Sherif A.; Fawaz, Esraa M.; Shehata, Mostafa A.

    2015-02-01

    Three sensitive, selective, and precise stability indicating methods for the determination of the X-ray contrast agent, diatrizoate sodium (DTA), in the presence of its acidic degradation product (highly cytotoxic 3,5 diamino metabolite) and in pharmaceutical formulation were developed and validated. The first method is a first derivative (D1) spectrophotometric one, which allows the determination of DTA in the presence of its degradate at 231.2 nm (corresponding to zero crossing of the degradate) over a concentration range of 2-24 μg/mL with mean percentage recovery 99.95 ± 0.97%. The second method is the first derivative of the ratio spectra (DD1) by measuring the peak amplitude at 227 nm over the same concentration range as D1 spectrophotometric method, with mean percentage recovery 99.99 ± 1.15%. The third method is a TLC-densitometric one, where DTA was separated from its degradate on silica gel plates using chloroform:methanol:ammonium hydroxide (20:10:2 by volume) as a developing system. This method depends on quantitative densitometric evaluation of thin layer chromatogram of DTA at 238 nm over a concentration range of 4-20 μg/spot, with mean percentage recovery 99.88 ± 0.89%. The selectivity of the proposed methods was tested using laboratory-prepared mixtures. The proposed methods have been successfully applied to the analysis of DTA in pharmaceutical dosage forms without interference from other dosage form additives. The results were statistically compared with the official US pharmacopeial method. No significant difference for either accuracy or precision was observed.

  9. Determination of Scattering and Absorption Coefficients for Plasma-Sprayed Yttria-Stabilized Zirconia Thermal Barrier Coatings at Elevated Temperatures

    Science.gov (United States)

    Eldridge, Jeffrey I.; Spuckler, Charles M.; Markham, James R.

    2009-01-01

    The temperature dependence of the scattering and absorption coefficients for a set of freestanding plasma-sprayed 8 wt% yttria-stabilized zirconia (8YSZ) thermal barrier coatings (TBCs) was determined at temperatures up to 1360 C in a wavelength range from 1.2 micrometers up to the 8YSZ absorption edge. The scattering and absorption coefficients were determined by fitting the directional-hemispherical reflectance and transmittance values calculated by a four-flux Kubelka Munk method to the experimentally measured hemispherical-directional reflectance and transmittance values obtained for five 8YSZ thicknesses. The scattering coefficient exhibited a continuous decrease with increasing wavelength and showed no significant temperature dependence. The scattering is primarily attributed to the relatively temperature-insensitive refractive index mismatch between the 8YSZ and its internal voids. The absorption coefficient was very low (less than 1 per centimeter) at wavelengths between 2 micrometers and the absorption edge and showed a definite temperature dependence that consisted of a shift of the absorption edge to shorter wavelengths and an increase in the weak absorption below the absorption edge with increasing temperature. The shift in the absorption edge with temperature is attributed to strongly temperature-dependent multiphonon absorption. While TBC hemispherical transmittance beyond the absorption edge can be predicted by a simple exponential decrease with thickness, below the absorption edge, typical TBC thicknesses are well below the thickness range where a simple exponential decrease in hemispherical transmittance with TBC thickness is expected. [Correction added after online publication August 11, 2009: "edge to a shorter wavelengths" has been updated as edge to shorter wavelengths."

  10. Gene expression in chromosomal Ridge domains : influence on transcription, mRNA stability, codon usage, and evolution

    NARCIS (Netherlands)

    Gierman, H.J.

    2010-01-01

    Chromosomes are the long DNA molecules that carry the genetic code of our genes. Each gene encodes a protein, but also contains the information that controls the activity of that gene. In this thesis, we find that chromosomal domains with many active genes (so-called 'Ridges'), also control gene

  11. Sex determination in femurs of modern Egyptians: A comparative study between metric measurements and SRY gene detection

    Directory of Open Access Journals (Sweden)

    Iman F. Gaballah

    2014-12-01

    Conclusion: The SRY gene detection method for sex determination is quick and simple, requiring only one PCR reaction. It corroborates the results obtained from anatomical measurements and further confirms the sex of the femur bone in question.

  12. Low-level laser irradiation alters mRNA expression from genes involved in DNA repair and genomic stabilization in myoblasts

    Science.gov (United States)

    Trajano, L. A. S. N.; Sergio, L. P. S.; Silva, C. L.; Carvalho, L.; Mencalha, A. L.; Stumbo, A. C.; Fonseca, A. S.

    2016-07-01

    Low-level lasers are used for the treatment of diseases in soft and bone tissues, but few data are available regarding their effects on genomic stability. In this study, we investigated mRNA expression from genes involved in DNA repair and genomic stabilization in myoblasts exposed to low-level infrared laser. C2C12 myoblast cultures in different fetal bovine serum concentrations were exposed to low-level infrared laser (10, 35 and 70 J cm-2), and collected for the evaluation of DNA repair gene expression. Laser exposure increased gene expression related to base excision repair (8-oxoguanine DNA glycosylase and apurinic/apyrimidinic endonuclease 1), nucleotide excision repair (excision repair cross-complementation group 1 and xeroderma pigmentosum C protein) and genomic stabilization (ATM serine/threonine kinase and tumor protein p53) in normal and low fetal bovine serum concentrations. Results suggest that genomic stability could be part of a biostimulation effect of low-level laser therapy in injured muscles.

  13. Stability of glycoprotein gene sequences of herpes simplex virus type 2 from primary to recurrent human infection, and diversity of the sequences among patients attending an STD clinic

    Science.gov (United States)

    2014-01-01

    Background Herpes simplex virus type 2 (HSV-2) is sexually transmitted, leading to blisters and ulcers in the genito-anal region. After primary infection the virus is present in a latent state in neurons in sensory ganglia. Reactivation and production of new viral particles can cause asymptomatic viral shedding or new lesions. Establishment of latency, maintenance and reactivation involve silencing of genes, continuous suppression of gene activities and finally gene activation and synthesis of viral DNA. The purpose of the present work was to study the genetic stability of the virus during these events. Methods HSV-2 was collected from 5 patients with true primary and recurrent infections, and the genes encoding glycoproteins B,G,E and I were sequenced. Results No nucleotide substitution was observed in any patient, indicating genetic stability. However, since the total number of nucleotides in these genes is only a small part of the total genome, we cannot rule out variation in other regions. Conclusions Although infections of cell cultures and animal models are useful for studies of herpes simplex virus, it is important to know how the virus behaves in the natural host. We observed that several glycoprotein gene sequences are stable from primary to recurrent infection. However, the virus isolates from the different patients were genetically different. PMID:24502528

  14. Discovery and evaluation of candidate sex-determining genes and xenobiotics in the gonads of lake sturgeon (Acipenser fulvescens).

    Science.gov (United States)

    Hale, Matthew C; Jackson, James R; Dewoody, J Andrew

    2010-07-01

    Modern pyrosequencing has the potential to uncover many interesting aspects of genome evolution, even in lineages where genomic resources are scarce. In particular, 454 pyrosequencing of nonmodel species has been used to characterize expressed sequence tags, xenobiotics, gene ontologies, and relative levels of gene expression. Herein, we use pyrosequencing to study the evolution of genes expressed in the gonads of a polyploid fish, the lake sturgeon (Acipenser fulvescens). Using 454 pyrosequencing of transcribed genes, we produced more than 125 MB of sequence data from 473,577 high-quality sequencing reads. Sequences that passed stringent quality control thresholds were assembled into 12,791 male contigs and 32,629 female contigs. Average depth of coverage was 4.2 x for the male assembly and 5.5x for the female assembly. Analytical rarefaction indicates that our assemblies include most of the genes expressed in lake sturgeon gonads. Over 86,700 sequencing reads were assigned gene ontologies, many to general housekeeping genes like protein, RNA, and ion binding genes. We searched specifically for sex determining genes and documented significant sex differences in the expression of two genes involved in animal sex determination, DMRT1 and TRA-1. DMRT1 is the master sex determining gene in birds and in medaka (Oryzias latipes) whereas TRA-1 helps direct sexual differentiation in nematodes. We also searched the lake sturgeon assembly for evidence of xenobiotic organisms that may exist as endosymbionts. Our results suggest that exogenous parasites (trematodes) and pathogens (protozoans) apparently have infected lake sturgeon gonads, and the trematodes have horizontally transferred some genes to the lake sturgeon genome.

  15. Application of spectrophotometric, densitometric, and HPLC techniques as stability indicating methods for determination of Zaleplon in pharmaceutical preparations

    Science.gov (United States)

    Metwally, Fadia H.; Abdelkawy, M.; Abdelwahab, Nada S.

    2007-12-01

    Spectrophotometric, spectrodensitometric and HPLC are stability indicating methods described for determination of Zaleplon in pure and dosage forms. As Zaleplon is easily degradable, the proposed techniques in this manuscript are adopted for its determination in presence of its alkaline degradation product, namely N-[4-(3-cyano-pyrazolo[1,5a]pyridin-7-yl)-phenyl]- N-ethyl-acetamide. These approaches are successfully applied to quantify Zaleplon using the information included in the absorption spectra of appropriate solutions. The second derivative (D 2) spectrophotometric method, allows determination of Zaleplon without interference of its degradate at 235.2 nm using 0.01N HCl as a solvent with obedience to Beer's law over a concentration range of 1-10 μg ml -1 with mean percentage recovery 100.24 ± 0.86%. The first derivative of the ratio spectra ( 1DD) based on the simultaneous use of ( 1DD) and measurement at 241.8 nm using the same solvent and over the same concentration range as (D 2) spectrophotometric method, with mean percentage recovery 99.9 ± 1.07%. The spectrodensitometric analysis allows the separation and quantitation of Zaleplon from its degradate on silica gel plates using chloroform:acetone:ammonia solution (9:1:0.2 by volume) as a mobile phase. This method depends on quantitave densitometric evaluation of thin layer chromatogram of Zaleplon at 338 nm over a concentration range of 0.2-1 μg band -1, with mean percentage recovery 99.73 ± 1.35. Also a reversed-phase liquid chromatographic method using 5-C8 (22 cm × 4.6 mm i.d. 5 μm particle size) column was described and validated for quantitation of Zaleplon using acetonitrile:deionised water (35:65, v/v) as a mobile phase using Paracetamol as internal standard and a flow rate of 1.5 ml min -1 with UV detection of the effluent at 232 nm at ambient temperature over a concentration range of 2-20 μg ml -1 with mean percentage recovery 100.19 ± 1.15%. The insignificance difference of the proposed

  16. Water-vortex-stabilized electric arc: III. Radial energy transport, determination of water-vapour-boundary and arc performance

    Energy Technology Data Exchange (ETDEWEB)

    Jenista, Jiri [Institute of Plasma Physics ASCR, Za Slovankou 3, PO Box 17, Prague 8, 182 21 (Czech Republic)

    2003-12-07

    This paper is concerned with numerical modelling of an electric arc stabilized by a water vortex. The two-dimensional axisymmetric model presented includes the arc discharge area between the cathode and the outlet nozzle of the water plasma torch. The aims of the numerical simulations are: (1) to assess the influence of radial position of the water-vapour-boundary in the discharge chamber on arc performance and overall radial energy transport within the arc; (2) to determine the most probable mass flow rates and radii of the water-vapour-boundary in the discharge chamber for a prescribed current; (3) to demonstrate arc performance for two radiation models involved; and (4) to estimate validity of local thermodynamic equilibrium (LTE) conditions within the arc column. The rate of evaporation of water is calculated from the conduction and radiation heat fluxes at the water vapour surface for the specified mass flow rate. The behaviour of such an arc has been studied for a range of current 300-600 A. It is shown that changes of bulk magnitudes of different terms in the momentum and energy equations within the arc column as a function of arc radius enable us to reveal transitions of temperature and velocity fields from one steady state to a qualitatively different one. The best fit between experiment and numerical simulation for all currents exists for the mean arc radius {approx} 3.3 mm. Deviations from LTE within the arc column are estimated with the criteria for kinetic equilibrium and spatial temperature gradients.

  17. Determination of aliskiren in tablet dosage forms by a validated stability-indicating RP-LC method.

    Science.gov (United States)

    Wrasse-Sangoi, M; Sangoi, M S; Oliveira, P R; Secretti, L T; Rolim, C M B

    2011-02-01

    A reversed-phase liquid chromatography (RP-LC) method is validated for the determination of aliskiren in tablet dosage form. The LC method is carried out on a Waters XBridge C(18) column (150 × 4.6 mm i.d.), maintained at 25°C. The mobile phase consisted of acetonitrile:water (95:5, v/v)/phosphoric acid (25 mM, pH 3.0) (40:60, v/v), run at a flow rate of 1.0 mL/min, with photodiode array detector set at 229 nm. The chromatographic separation is obtained with aliskiren retention time of 3.68 min, and it is linear in the range of 10-300 μg/mL (r = 0.9999). The limits of detection and quantitation are 2.38 and 7.93 μg/mL, respectively. The specificity and stability-indicating capability of the method are proven through degradation studies, which also showed that there is no interference of the formulation excipients, showing that peak is free from any coeluting peak. The method showed adequate precision, with a relative standard deviation (RSD) values lower than 0.92%. Good values of accuracy were also obtained, with a mean value of 99.55%. Experimental design is used during validation to calculate method robustness. The proposed method is applied for the analysis of the tablet dosage forms, contributing to improve the quality control and to assure the therapeutic efficacy.

  18. Determination of Hydrogen Bond Structure in Water versus Aprotic Environments To Test the Relationship Between Length and Stability

    Energy Technology Data Exchange (ETDEWEB)

    Sigala, Paul A.; Ruben, Eliza A.; Liu, Corey W.; Piccoli, Paula M. B.; Hohenstein, Edward G.; Martinez, Todd J.; Schultz, Arthur J.; Herschiag, Daniel

    2015-05-06

    Hydrogen bonds profoundly influence the architecture and activity of biological macromolecules. Deep appreciation of hydrogen bond contributions to biomolecular function thus requires a detailed understanding of hydrogen bond structure and energetics and the relationship between these properties. Hydrogen bond formation energies (Delta G(f)) are enormously more favorable in aprotic solvents than in water, and two classes of contributing factors have been proposed to explain this energetic difference, focusing respectively on the isolated and hydrogen-bonded species: (I) water stabilizes the dissociated donor and acceptor groups much better than aprotic solvents, thereby reducing the driving force for hydrogen bond formation; and (II) water lengthens hydrogen bonds compared to aprotic environments, thereby decreasing the potential energy within the hydrogen bond. Each model has been proposed to provide a dominant contribution to Delta G(f), but incisive tests that distinguish the importance of these contributions are lacking. Here we directly test the structural basis of model II. Neutron crystallography, NMR spectroscopy, and quantum mechanical calculations demonstrate that O-H center dot center dot center dot O hydrogen bonds in crystals, chloroform, acetone, and water have nearly identical lengths and very similar potential energy surfaces despite Delta G(f) differences >8 kcal/mol across these solvents. These results rule out a substantial contribution from solvent-dependent differences in hydrogen bond structure and potential energy after association (model II) and thus support the conclusion that differences in hydrogen bond Delta G(f) are predominantly determined by solvent interactions with the dissociated groups (model I). These findings advance our understanding of universal hydrogen-bonding interactions and have important implications for biology and engineering.

  19. Allelic variation of bile salt hydrolase genes in Lactobacillus salivarius does not determine bile resistance levels.

    LENUS (Irish Health Repository)

    Fang, Fang

    2009-09-01

    Commensal lactobacilli frequently produce bile salt hydrolase (Bsh) enzymes whose roles in intestinal survival are unclear. Twenty-six Lactobacillus salivarius strains from different sources all harbored a bsh1 allele on their respective megaplasmids. This allele was related to the plasmid-borne bsh1 gene of the probiotic strain UCC118. A second locus (bsh2) was found in the chromosomes of two strains that had higher bile resistance levels. Four Bsh1-encoding allele groups were identified, defined by truncations or deletions involving a conserved residue. In vitro analyses showed that this allelic variation was correlated with widely varying bile deconjugation phenotypes. Despite very low activity of the UCC118 Bsh1 enzyme, a mutant lacking this protein had significantly lower bile resistance, both in vitro and during intestinal transit in mice. However, the overall bile resistance phenotype of this and other strains was independent of the bsh1 allele type. Analysis of the L. salivarius transcriptome upon exposure to bile and cholate identified a multiplicity of stress response proteins and putative efflux proteins that appear to broadly compensate for, or mask, the effects of allelic variation of bsh genes. Bsh enzymes with different bile-degrading kinetics, though apparently not the primary determinants of bile resistance in L. salivarius, may have additional biological importance because of varying effects upon bile as a signaling molecule in the host.

  20. Master regulators in development: Views from the Drosophila retinal determination and mammalian pluripotency gene networks.

    Science.gov (United States)

    Davis, Trevor L; Rebay, Ilaria

    2017-01-15

    Among the mechanisms that steer cells to their correct fate during development, master regulatory networks are unique in their sufficiency to trigger a developmental program outside of its normal context. In this review we discuss the key features that underlie master regulatory potency during normal and ectopic development, focusing on two examples, the retinal determination gene network (RDGN) that directs eye development in the fruit fly and the pluripotency gene network (PGN) that maintains cell fate competency in the early mammalian embryo. In addition to the hierarchical transcriptional activation, extensive positive transcriptional feedback, and cooperative protein-protein interactions that enable master regulators to override competing cellular programs, recent evidence suggests that network topology must also be dynamic, with extensive rewiring of the interactions and feedback loops required to navigate the correct sequence of developmental transitions to reach a final fate. By synthesizing the in vivo evidence provided by the RDGN with the extensive mechanistic insight gleaned from the PGN, we highlight the unique regulatory capabilities that continual reorganization into new hierarchies confers on master control networks. We suggest that deeper understanding of such dynamics should be a priority, as accurate spatiotemporal remodeling of network topology will undoubtedly be essential for successful stem cell based therapeutic efforts.

  1. Stability of three methods for two-dimensional sociometric status determination based on the procedure of Asher, Singleton, Tinsley and Hymel

    NARCIS (Netherlands)

    Maassen, G.H; Steenbeek, H.W.; Van Geert, P. L. C.

    2004-01-01

    This study aimed at comparing the stability of three methods for two-dimensional sociometric status determination, including (1) the recently developed SSrat technique (Maassen, Akkermans, & Van der Linden, 1996), as well as (2) the procedure of Howes (1988), which is based on the algorithm and clas

  2. The Impact of Social-Cognitive Stress on Speech Variability, Determinism, and Stability in Adults Who Do and Do Not Stutter

    Science.gov (United States)

    Jackson, Eric S.; Tiede, Mark; Beal, Deryk; Whalen, D. H.

    2016-01-01

    Purpose: This study examined the impact of social-cognitive stress on sentence-level speech variability, determinism, and stability in adults who stutter (AWS) and adults who do not stutter (AWNS). We demonstrated that complementing the spatiotemporal index (STI) with recurrence quantification analysis (RQA) provides a novel approach to both…

  3. Functional analysis of the stability determinant AlfB of pBET131, a miniplasmid derivative of bacillus subtilis (natto) plasmid pLS32.

    Science.gov (United States)

    Tanaka, Teruo

    2010-03-01

    Bacillus subtilis plasmid pBET131 is a derivative of pLS32, which was isolated from a natto strain of Bacillus subtilis. The DNA region in pBET131 that confers segregational stability contains an operon consisting of three genes, of which alfA, encoding an actin-like ATPase, and alfB are essential for plasmid stability. In this work, the alfB gene product and its target DNA region were studied in detail. Transcription of the alf operon initiated from a sigma(A)-type promoter was repressed by the alfB gene product. Overproduction of AlfA was inhibitory to cell growth, suggesting that the repression of the alf operon by AlfB is important for maintaining appropriate levels of AlfA. An electrophoretic mobility shift assay and footprinting analysis with purified His-tagged AlfB showed that it bound to a DNA region containing three tandem repeats of 8-bp AT-rich sequence (here designated parN), which partially overlaps the -35 sequence of the promoter. A sequence alteration in the first or third repeat did not affect the AlfB binding and plasmid stability, whereas that in the second repeat resulted in inhibition of these phenomena. The repression of alfA-lacZ expression was observed in the constructs carrying a mutation in either the first or third repeat, but not in the second repeat, indicating a correlation between plasmid stability, AlfB binding, and repression. It was also demonstrated by the yeast two-hybrid system that AlfA and AlfB interact with each other and among themselves. From these results, it was concluded that AlfB participates in partitioning pBET131 by forming a complex with AlfA and parN, the mode of which is typified by the type II partition mechanism.

  4. NCYM, a Cis-antisense gene of MYCN, encodes a de novo evolved protein that inhibits GSK3β resulting in the stabilization of MYCN in human neuroblastomas.

    Directory of Open Access Journals (Sweden)

    Yusuke Suenaga

    2014-01-01

    Full Text Available The rearrangement of pre-existing genes has long been thought of as the major mode of new gene generation. Recently, de novo gene birth from non-genic DNA was found to be an alternative mechanism to generate novel protein-coding genes. However, its functional role in human disease remains largely unknown. Here we show that NCYM, a cis-antisense gene of the MYCN oncogene, initially thought to be a large non-coding RNA, encodes a de novo evolved protein regulating the pathogenesis of human cancers, particularly neuroblastoma. The NCYM gene is evolutionally conserved only in the taxonomic group containing humans and chimpanzees. In primary human neuroblastomas, NCYM is 100% co-amplified and co-expressed with MYCN, and NCYM mRNA expression is associated with poor clinical outcome. MYCN directly transactivates both NCYM and MYCN mRNA, whereas NCYM stabilizes MYCN protein by inhibiting the activity of GSK3β, a kinase that promotes MYCN degradation. In contrast to MYCN transgenic mice, neuroblastomas in MYCN/NCYM double transgenic mice were frequently accompanied by distant metastases, behavior reminiscent of human neuroblastomas with MYCN amplification. The NCYM protein also interacts with GSK3β, thereby stabilizing the MYCN protein in the tumors of the MYCN/NCYM double transgenic mice. Thus, these results suggest that GSK3β inhibition by NCYM stabilizes the MYCN protein both in vitro and in vivo. Furthermore, the survival of MYCN transgenic mice bearing neuroblastoma was improved by treatment with NVP-BEZ235, a dual PI3K/mTOR inhibitor shown to destabilize MYCN via GSK3β activation. In contrast, tumors caused in MYCN/NCYM double transgenic mice showed chemo-resistance to the drug. Collectively, our results show that NCYM is the first de novo evolved protein known to act as an oncopromoting factor in human cancer, and suggest that de novo evolved proteins may functionally characterize human disease.

  5. Gain-of-function mutations of fem-3, a sex-determination gene in Caenorhabditis elegans

    Energy Technology Data Exchange (ETDEWEB)

    Barton, M.K.; Schedl, T.B.; Kimble, J.

    1987-01-01

    The authors have isolated nine gain-of-function (gf) alleles of the sex-determination gene fem-3 as suppressors of feminizing mutations in fem-1 and fem-2. The wild type fem-3 gene is needed for spermatogenesis in XX self-fertilizing hermaphrodites and for male development in both soma and germ line of XO animals. Loss-of-function alleles of fem-3 transform XX and XO animals into females (spermless hermaphrodites). In contrast, fem-3 (gf) alleles masculinize only one tissue, the hermaphrodite germ line. Thus, XX fem-3 (gf) mutant animals have a normal hermaphrodite soma, but the germ line produces a vast excess of sperm and no oocytes. All nine fem-3 (gf) alleles are temperature sensitive. The temperature-sensitive period is from late L4 to early adult, a period just preceding the first signs of oogenesis. The finding of gain-of-function alleles which confer a phenotype opposite to that of loss-of-function alleles supports the idea that fem-3 plays a critical role in germ-line sex determination. Furthermore, the germ-line specificity of the fem-3 (gf) mutant phenotype and the late temperature-sensitive period suggest that, in the wild-type XX hermaphrodite, fem-3 is negatively regulated so that the hermaphrodite stops making sperm and starts making oocytes. Temperature shift experiments also show that, in the germ line, sexual commitment appears to be a continuing process. Spermatogenesis can resume even after oogenesis has begun, and oogenesis can be initiated much later than normal.

  6. Molecular determinants of dysregulated GABAergic gene expression in the prefrontal cortex of subjects with schizophrenia.

    Science.gov (United States)

    Mellios, Nikolaos; Huang, Hsien-Sung; Baker, Stephen P; Galdzicka, Marzena; Ginns, Edward; Akbarian, Schahram

    2009-06-15

    Prefrontal deficits in gamma-aminobutyric acid (GABA)ergic gene expression, including neuropeptide Y (NPY), somatostatin (SST), and parvalbumin (PV) messenger RNAs (mRNAs), have been reported for multiple schizophrenia cohorts. Preclinical models suggest that a subset of these GABAergic markers (NPY/SST) is regulated by brain-derived neurotrophic factor (BDNF), which in turn is under the inhibitory influence of small noncoding RNAs. However, it remains unclear if these mechanisms are important determinants for dysregulated NPY and SST expression in prefrontal cortex (PFC) of subjects with schizophrenia. Using a postmortem case-control design, the association between BDNF protein, NPY/SST/PV mRNAs, and two BDNF-regulating microRNAs (miR-195 and miR-30a-5p) was determined in samples from the PFC of 20 schizophrenia and 20 control subjects. Complementary studies were conducted in cerebral cortex of mice subjected to antipsychotic treatment or a brain-specific ablation of the Bdnf gene. Subjects with schizophrenia showed deficits in NPY and PV mRNAs. Within-pair differences in BDNF protein levels showed strong positive correlations with NPY and SST and a robust inverse association with miR-195 levels, which in turn were not affected by antipsychotic treatment or genetic ablation of Bdnf. Taken together, these results suggest that prefrontal deficits in a subset of GABAergic mRNAs, including NPY, are dependent on the regional supply of BDNF, which in turn is fine-tuned through a microRNA (miRNA)-mediated mechanism.

  7. Gene Regulatory Network Analysis Reveals Differences in Site-specific Cell Fate Determination in Mammalian Brain

    Directory of Open Access Journals (Sweden)

    Gokhan eErtaylan

    2014-12-01

    Full Text Available Neurogenesis - the generation of new neurons - is an ongoing process that persists in the adult mammalian brain of several species, including humans. In this work we analyze two discrete brain regions: the subventricular zone (SVZ lining the walls of the lateral ventricles; and the subgranular zone (SGZ of the dentate gyrus of the hippocampus in mice and shed light on the SVZ and SGZ specific neurogenesis. We propose a computational model that relies on the construction and analysis of region specific gene regulatory networks from the publicly available data on these two regions. Using this model a number of putative factors involved in neuronal stem cell (NSC identity and maintenance were identified. We also demonstrate potential gender and niche-derived differences based on cell surface and nuclear receptors via Ar, Hif1a and Nr3c1.We have also conducted cell fate determinant analysis for SVZ NSC populations to Olfactory Bulb interneurons and SGZ NSC populations to the granule cells of the Granular Cell Layer. We report thirty-one candidate cell fate determinant gene pairs, ready to be validated. We focus on Ar - Pax6 in SVZ and Sox2 - Ncor1 in SGZ. Both pairs are expressed and localized in the suggested anatomical structures as shown by in situ hybridization and found to physically interact.Finally, we conclude that there are fundamental differences between SGZ and SVZ neurogenesis. We argue that these regulatory mechanisms are linked to the observed differential neurogenic potential of these regions. The presence of nuclear and cell surface receptors in the region specific regulatory circuits indicate the significance of niche derived extracellular factors, hormones and region specific factors such as the oxygen sensitivity, dictating SGZ and SVZ specific neurogenesis.

  8. A Novel Method to Determine the Local Stability of the n-Species Lotka-Volterra System with Multiple Delays

    Directory of Open Access Journals (Sweden)

    Xiao-Ping Chen

    2016-01-01

    Full Text Available The n-species Lotka-Volterra system with discrete delays is considered. The local asymptotic stability of positive equilibrium is investigated based on a contour integral method. The main purpose of this paper is to propose a new and general algorithm to study the local asymptotic stability of the positive equilibrium for the n-dimensional Lotka-Volterra system. Some numerical experiments are carried out to show the effectiveness of the proposed method.

  9. Pathway-based analysis of GWAs data identifies association of sex determination genes with susceptibility to testicular germ cell tumors.

    Science.gov (United States)

    Koster, Roelof; Mitra, Nandita; D'Andrea, Kurt; Vardhanabhuti, Saran; Chung, Charles C; Wang, Zhaoming; Loren Erickson, R; Vaughn, David J; Litchfield, Kevin; Rahman, Nazneen; Greene, Mark H; McGlynn, Katherine A; Turnbull, Clare; Chanock, Stephen J; Nathanson, Katherine L; Kanetsky, Peter A

    2014-11-15

    Genome-wide association (GWA) studies of testicular germ cell tumor (TGCT) have identified 18 susceptibility loci, some containing genes encoding proteins important in male germ cell development. Deletions of one of these genes, DMRT1, lead to male-to-female sex reversal and are associated with development of gonadoblastoma. To further explore genetic association with TGCT, we undertook a pathway-based analysis of SNP marker associations in the Penn GWAs (349 TGCT cases and 919 controls). We analyzed a custom-built sex determination gene set consisting of 32 genes using three different methods of pathway-based analysis. The sex determination gene set ranked highly compared with canonical gene sets, and it was associated with TGCT (FDRG = 2.28 × 10(-5), FDRM = 0.014 and FDRI = 0.008 for Gene Set Analysis-SNP (GSA-SNP), Meta-Analysis Gene Set Enrichment of Variant Associations (MAGENTA) and Improved Gene Set Enrichment Analysis for Genome-wide Association Study (i-GSEA4GWAS) analysis, respectively). The association remained after removal of DMRT1 from the gene set (FDRG = 0.0002, FDRM = 0.055 and FDRI = 0.009). Using data from the NCI GWA scan (582 TGCT cases and 1056 controls) and UK scan (986 TGCT cases and 4946 controls), we replicated these findings (NCI: FDRG = 0.006, FDRM = 0.014, FDRI = 0.033, and UK: FDRG = 1.04 × 10(-6), FDRM = 0.016, FDRI = 0.025). After removal of DMRT1 from the gene set, the sex determination gene set remains associated with TGCT in the NCI (FDRG = 0.039, FDRM = 0.050 and FDRI = 0.055) and UK scans (FDRG = 3.00 × 10(-5), FDRM = 0.056 and FDRI = 0.044). With the exception of DMRT1, genes in the sex determination gene set have not previously been identified as TGCT susceptibility loci in these GWA scans, demonstrating the complementary nature of a pathway-based approach for genome-wide analysis of TGCT.

  10. Sexual epigenetics: gender-specific methylation of a gene in the sex determining region of Populus balsamifera

    Science.gov (United States)

    Bräutigam, Katharina; Soolanayakanahally, Raju; Champigny, Marc; Mansfield, Shawn; Douglas, Carl; Campbell, Malcolm M.; Cronk, Quentin

    2017-01-01

    Methylation has frequently been implicated in gender determination in plants. The recent discovery of the sex determining region (SDR) of balsam poplar, Populus balsamifera, pinpointed 13 genes with differentiated X and Y copies. We tested these genes for differential methylation using whole methylome sequencing of xylem tissue of multiple individuals grown under field conditions in two common gardens. The only SDR gene to show a marked pattern of gender-specific methylation is PbRR9, a member of the two component response regulator (type-A) gene family, involved in cytokinin signalling. It is an ortholog of Arabidopsis genes ARR16 and ARR17. The strongest patterns of differential methylation (mostly male-biased) are found in the putative promoter and the first intron. The 4th intron is strongly methylated in both sexes and the 5th intron is unmethylated in both sexes. Using a statistical learning algorithm we find that it is possible accurately to assign trees to gender using genome-wide methylation patterns alone. The strongest predictor is the region coincident with PbRR9, showing that this gene stands out against all genes in the genome in having the strongest sex-specific methylation pattern. We propose the hypothesis that PbRR9 has a direct, epigenetically mediated, role in poplar sex determination. PMID:28345647

  11. Minor Components and Oxidative Stability as Determined by DSC of Fractionated and Lipase-catalyzed Structured Rapeseed Oil

    Directory of Open Access Journals (Sweden)

    Alim, M. A.

    2013-06-01

    Full Text Available Natural fats and oils can be modified by various methods to prepare products with desired physical, chemical and nutritional properties. The enrichment and retention of the minor lipid components, the incorporation of capric acid and oxidative stability in low temperature fractionated rapeseed oil (RSO in acetone were assessed in this study. The fractionated liquid part (L-RSO, the solid part (S-RSO and the RSO were transesterified with capric acid (CA at different mole ratios using lipase. The yields of L-RSO and S-RSO were 30 and 70 g per 100 g, respectively. The L-RSO contained higher levels of linoleic acid and linolenic acid, and a lower level of oleic acid compared to the S-RSO. The S-RSO contained a higher amount of total sterols than the RSO and the L-RSO. In contrast, the L-RSO contained a higher amount of total tocopherols than the RSO and the S-RSO. The incorporation of CA was ideal at a mole ratio of 1:3. The content of sterols and tocopherols gradually decreased with an increased mole ratio for the CA incorporation. The oxidative stability shown as onset temperature, determined by DSC, of the S-RSO was higher compared to those of the L-RSO and RSO.Las grasas y aceites naturales pueden ser modificados mediante diversos métodos para preparar productos con propiedades físicas, químicas y nutricionales deseadas. El enriquecimiento y la retención de componentes lipídicos menores, la incorporación de ácido cáprico y estabilidad a la oxidación a baja temperatura de aceites de colza (RSO fraccionado en acetona, se evaluaron en este estudio. La fracción líquida (L-RSO, la fracción sólida (S-RSO y el RSO son transesterificados con ácido cáprico (CA en diversas relaciones molares utilizando lipasa. Los rendimientos de las fracciones L-RSO y RSO-S fueron de 30 y 70 g por 100 g, respectivamente. La fracción líquida L-RSO contenía un mayor nivel de los ácidos linoleico y linolénico, y un menor nivel de ácido oleico en

  12. Benchmarking the GENE and GYRO codes through the relative roles of electromagnetic and E  ×  B stabilization in JET high-performance discharges

    Science.gov (United States)

    Bravenec, R.; Citrin, J.; Candy, J.; Mantica, P.; Görler, T.; contributors, JET

    2016-12-01

    Nonlinear gyrokinetic simulations using the GENE code have previously predicted a significant nonlinear enhanced electromagnetic stabilization in certain JET discharges with high neutral-beam power and low core magnetic shear (Citrin et al 2013 Phys. Rev. Lett. 111 155001, 2015 Plasma Phys. Control. Fusion 57 014032). This dominates over the impact of E  ×  B flow shear in these discharges. Furthermore, fast ions were shown to be a major contributor to the electromagnetic stabilization. These conclusions were based on results from the GENE gyrokinetic turbulence code. In this work we verify these results using the GYRO code. Comparing results (linear frequencies, eigenfunctions, and nonlinear fluxes) from different gyrokinetic codes as a means of verification (benchmarking) is only convincing if the codes agree for more than one discharge. Otherwise, agreement may simply be fortuitous. Therefore, we analyze three discharges, all with a carbon wall: a simplified, two-species, circular geometry case based on an actual JET discharge; an L-mode discharge with a significant fast-ion pressure fraction; and a low-triangularity high-β hybrid discharge. All discharges were analyzed at normalized toroidal flux coordinate ρ  =  0.33 where significant ion temperature peaking is observed. The GYRO simulations support the conclusion that electromagnetic stabilization is strong, and dominates E  ×  B shear stabilization.

  13. Increased expression of X-linked genes in mammals is associated with a higher stability of transcripts and an increased ribosome density.

    Science.gov (United States)

    Faucillion, Marie-Line; Larsson, Jan

    2015-03-18

    Mammalian sex chromosomes evolved from the degeneration of one homolog of a pair of ancestral autosomes, the proto-Y. This resulted in a gene dose imbalance that is believed to be restored (partially or fully) through upregulation of gene expression from the single active X-chromosome in both sexes by a dosage compensatory mechanism. We analyzed multiple genome-wide RNA stability data sets and found significantly longer average half-lives for X-chromosome transcripts than for autosomal transcripts in various human cell lines, both male and female, and in mice. Analysis of ribosome profiling data shows that ribosome density is higher on X-chromosome transcripts than on autosomal transcripts in both humans and mice, suggesting that the higher stability is causally linked to a higher translation rate. Our results and observations are in accordance with a dosage compensatory upregulation of expressed X-linked genes. We therefore propose that differential mRNA stability and translation rates of the autosomes and sex chromosomes contribute to an evolutionarily conserved dosage compensation mechanism in mammals.

  14. Determination of amlodipine in human plasma by electrospray ionization LC-MS/MS method: validation and its stability studies

    Directory of Open Access Journals (Sweden)

    Anusak Sirikatitham

    2008-07-01

    Full Text Available A sensitive and specific high-performance liquid chromatography combined with electrospray ionization (ESI tandemmass spectrometry (LC-MS/MS method, operating in the positive ionization mode, for quantifying of amlodipine in humanplasma using tizanidine as internal standard (I.S. was developed and validated. The analyte and I.S. were extracted bysimple one step liquid/liquid extraction with a mixture of diethylether/dichloromethane (70/30, v/v. The chromatographicseparation was performed on a C18 analytical column under isocratic conditions using a mixture of 10mM ammoniumformate/methanol/acetonitrile (30/50/20, v/v/v as mobile phase at a flow rate of 1.0 mL/min. Total chromatographic runtime was 5.0 min. Detection was performed on a API 2000 QTRAP quadrupole linear ion trap mass spectrometer via turboion spray ionization. Quantitation was performed using multiple reaction monitoring (MRM mode to study parent ®product ion transitions of m/z 409.4 ® 238.1 for amlodipine and m/z 254.2 ® 44.1 for I.S., respectively. The validation andstability studies were performed according to the Thai FDA guidance for assessment of bioequivalence study in Thailand.The results were within the accepted criteria as stated in the aforementioned guidance. Linearity in plasma was obtained overthe concentration range 0.3-15.0 ng/mL, with a coefficient of determination (r2 of 0.9993. Lower limit of quantification(LLOQ was identifiable and reproducible at 0.3 ng/mL. The within- and between-run precision values were below 10%and the accuracy was ranged from 94.87 to 102.44% at all three quality controls samples levels. The analyte was found tobe stable in plasma samples under three freeze-thaw cycles, long-term storage (3 months at -20oC, short-term storage (4hours at room temperature, post-preparative and stock-solution stability. The robust and rapid LC-MS/MS method has beensuccessfully applied for routine assay to support bioequivalence or pharmacokinetics studies

  15. Development and Validation of a Stability Indicating LC Method for the Assay and Related Substances Determination of a Proteasome Inhibitor Bortezomib

    Directory of Open Access Journals (Sweden)

    Kasa Srinivasulu

    2012-01-01

    Full Text Available A novel, simple, sensitive, stability indicating HPLC method was developed and validated for quantification of impurities (process related and degradants and assay determination of bortezomib. Stability indicating power of the method was established by forced degradation experiments and mass balance study. The chromatographic separation was achieved with Waters SymmetryShield RP18 column using gradient elution using the mobile phase-A consists of a mixture of water-acetonitrile-formic acid (715 : 285 : 1, v/v/v and the mobile phase-B consists a mixture of methanol-water-formic acid (800 : 200 : 1, v/v/v, respectively. The developed method is validated for parameters like precision, accuracy, linearity, LOD, LOQ, and ruggedness. Central composite experimental design (CCD was applied to check the robustness of the method. The stability tests were also performed on drug substances as per ICH norms.

  16. Common genetic variations in the CYP2R1 and GC genes are determinants of vitamin D status in Danes

    DEFF Research Database (Denmark)

    Nissen, Ioanna

    ), after 6 months intake of vitamin D3-fortified bread and milk (paper II) and in 92 participants in the VitDgen study after artificial UVB irradiation during winter (paper III). Common genetic variations in the CYP2R1 and GC genes were found to be important determinants of vitamin D status in three out...... by genetic variation in vitamin D modulating genes. Twin and family-based studies indicate that genetic variation may have an appreciable influence on vitamin D status. Moreover, several candidate gene studies including two genome-wide association studies (GWAS) have found single nucleotide polymorphisms...... (SNPs) in CYP2R1, CYP24A1, CYP27B1, C10orf88, DHCR7/NADSYN1, GC and VDR genes to be associated with vitamin D status. The main hypothesis of this work was that genetically determined variation in vitamin D metabolism would influence the effect of vitamin D sources (vitamin D...

  17. Identification of learning and memory genes in canine; promoter investigation and determining the selective pressure.

    Science.gov (United States)

    Seifi Moroudi, Reihane; Masoudi, Ali Akbar; Vaez Torshizi, Rasoul; Zandi, Mohammad

    2014-12-01

    One of the important behaviors of dogs is trainability which is affected by learning and memory genes. These kinds of the genes have not yet been identified in dogs. In the current research, these genes were found in animal models by mining the biological data and scientific literatures. The proteins of these genes were obtained from the UniProt database in dogs and humans. Not all homologous proteins perform similar functions, thus comparison of these proteins was studied in terms of protein families, domains, biological processes, molecular functions, and cellular location of metabolic pathways in Interpro, KEGG, Quick Go and Psort databases. The results showed that some of these proteins have the same performance in the rat or mouse, dog, and human. It is anticipated that the protein of these genes may be effective in learning and memory in dogs. Then, the expression pattern of the recognized genes was investigated in the dog hippocampus using the existing information in the GEO profile. The results showed that BDNF, TAC1 and CCK genes are expressed in the dog hippocampus, therefore, these genes could be strong candidates associated with learning and memory in dogs. Subsequently, due to the importance of the promoter regions in gene function, this region was investigated in the above genes. Analysis of the promoter indicated that the HNF-4 site of BDNF gene and the transcription start site of CCK gene is exposed to methylation. Phylogenetic analysis of protein sequences of these genes showed high similarity in each of these three genes among the studied species. The dN/dS ratio for BDNF, TAC1 and CCK genes indicates a purifying selection during the evolution of the genes.

  18. Determination of ploidy level and isolation of genes encoding acetyl-CoA carboxylase in Japanese Foxtail (Alopecurus japonicus.

    Directory of Open Access Journals (Sweden)

    Hongle Xu

    Full Text Available Ploidy level is important in biodiversity studies and in developing strategies for isolating important plant genes. Many herbicide-resistant weed species are polyploids, but our understanding of these polyploid weeds is limited. Japanese foxtail, a noxious agricultural grass weed, has evolved herbicide resistance. However, most studies on this weed have ignored the fact that there are multiple copies of target genes. This may complicate the study of resistance mechanisms. Japanese foxtail was found to be a tetraploid by flow cytometer and chromosome counting, two commonly used methods in the determination of ploidy levels. We found that there are two copies of the gene encoding plastidic acetyl-CoA carboxylase (ACCase in Japanese foxtail and all the homologous genes are expressed. Additionally, no difference in ploidy levels or ACCase gene copy numbers was observed between an ACCase-inhibiting herbicide-resistant and a herbicide-sensitive population in this study.

  19. Stabilization of Arabidopsis DREB2A is required but not sufficient for the induction of target genes under conditions of stress.

    Directory of Open Access Journals (Sweden)

    Kyoko Morimoto

    Full Text Available The Arabidopsis thaliana transcription factor DEHYDRATION-RESPONSIVE ELEMENT-BINDING PROTEIN2A (DREB2A controls the expression of many genes involved in the plant's response to dehydration and heat stress. Despite the significance of post-translational regulation in DREB2A activation, the mechanism underlying this activation remains unclear. Here, with the aid of a newly produced antibody against DREB2A, we characterized the regulation of DREB2A stability in plants exposed to stress stimuli. Endogenous DREB2A accumulated in wild-type Arabidopsis plants subjected to dehydration and heat stress. A degradation assay using Arabidopsis T87 suspension-cultured cells revealed that DREB2A protein degradation was inhibited at high temperatures. The proteasome-dependent degradation of DREB2A required the import of this protein into the nucleus. The E3 ligases DRIP1 and DRIP2 were involved in this process under both normal and stressful conditions; however, other E3 ligases may have also been involved, at least during the late stages of the heat stress response. Although the constitutive expression of DREB2A resulted in an overproduction of DREB2A and enhanced target gene induction during stress in transgenic plants, the accumulation of DREB2A caused by proteasome inhibitors did not induce target gene expression. Thus, the stabilization of DREB2A is important but not sufficient to induce target gene expression; further activation processes are required.

  20. Genetics and Molecular Biology of Epstein-Barr Virus-Encoded BART MicroRNA: A Paradigm for Viral Modulation of Host Immune Response Genes and Genome Stability

    Directory of Open Access Journals (Sweden)

    David H. Dreyfus

    2017-01-01

    Full Text Available Epstein-Barr virus, a ubiquitous human herpesvirus, is associated through epidemiologic evidence with common autoimmune syndromes and cancers. However, specific genetic mechanisms of pathogenesis have been difficult to identify. In this review, the author summarizes evidence that recently discovered noncoding RNAs termed microRNA encoded by Epstein-Barr virus BARF (BamHI A right frame termed BART (BamHI A right transcripts are modulators of human immune response genes and genome stability in infected and bystander cells. BART expression is apparently regulated by complex feedback loops with the host immune response regulatory NF-κB transcription factors. EBV-encoded BZLF-1 (ZEBRA protein could also regulate BART since ZEBRA contains a terminal region similar to ankyrin proteins such as IκBα that regulate host NF-κB. BALF-2 (BamHI A left frame transcript, a viral homologue of the immunoglobulin and T cell receptor gene recombinase RAG-1 (recombination-activating gene-1, may also be coregulated with BART since BALF-2 regulatory sequences are located near the BART locus. Viral-encoded microRNA and viral mRNA transferred to bystander cells through vesicles, defective viral particles, or other mechanisms suggest a new paradigm in which bystander or hit-and-run mechanisms enable the virus to transiently or chronically alter human immune response genes as well as the stability of the human genome.

  1. Integrative framework for identification of key cell identity genes uncovers determinants of ES cell identity and homeostasis.

    Science.gov (United States)

    Cinghu, Senthilkumar; Yellaboina, Sailu; Freudenberg, Johannes M; Ghosh, Swati; Zheng, Xiaofeng; Oldfield, Andrew J; Lackford, Brad L; Zaykin, Dmitri V; Hu, Guang; Jothi, Raja

    2014-04-22

    Identification of genes associated with specific biological phenotypes is a fundamental step toward understanding the molecular basis underlying development and pathogenesis. Although RNAi-based high-throughput screens are routinely used for this task, false discovery and sensitivity remain a challenge. Here we describe a computational framework for systematic integration of published gene expression data to identify genes defining a phenotype of interest. We applied our approach to rank-order all genes based on their likelihood of determining ES cell (ESC) identity. RNAi-mediated loss-of-function experiments on top-ranked genes unearthed many novel determinants of ESC identity, thus validating the derived gene ranks to serve as a rich and valuable resource for those working to uncover novel ESC regulators. Underscoring the value of our gene ranks, functional studies of our top-hit Nucleolin (Ncl), abundant in stem and cancer cells, revealed Ncl's essential role in the maintenance of ESC homeostasis by shielding against differentiation-inducing redox imbalance-induced oxidative stress. Notably, we report a conceptually novel mechanism involving a Nucleolin-dependent Nanog-p53 bistable switch regulating the homeostatic balance between self-renewal and differentiation in ESCs. Our findings connect the dots on a previously unknown regulatory circuitry involving genes associated with traits in both ESCs and cancer and might have profound implications for understanding cell fate decisions in cancer stem cells. The proposed computational framework, by helping to prioritize and preselect candidate genes for tests using complex and expensive genetic screens, provides a powerful yet inexpensive means for identification of key cell identity genes.

  2. Sexual Dimorphism of Body Size Is Controlled by Dosage of the X-Chromosomal Gene Myc and by the Sex-Determining Gene tra in Drosophila.

    Science.gov (United States)

    Wehr Mathews, Kristina; Cavegn, Margrith; Zwicky, Monica

    2017-01-06

    Drosophila females are larger than males. In this paper, we describe how X chromosome dosage drives sexual dimorphism of body size through two means: first, through unbalanced expression of a key X-linked growth regulating gene and second, through female-specific activation of the sex-determination pathway. X-chromosome dosage determines phenotypic sex by regulating the genes of the sex-determining pathway. In the presence of two sets of X-chromosome signal elements (XSEs), Sex-lethal (Sxl) is activated in female (XX) but not male (XY) animals. Sxl activates transformer (tra), a gene that encodes a splicing factor essential for female-specific development. It has previously been shown that null mutations in the tra gene result in only a partial reduction of body size of XX animals, which shows that other factors must contribute to size determination. We tested whether X dosage directly affects animal size by analyzing males with duplications of X chromosomal segments. Upon tiling across the X chromosome, we found four duplications that increase male size by over 9%. Within these, we identified several genes that promote growth as a result of duplication. Only one of these, Myc, was found not to be dosage compensated. Together, our results indicate that both Myc dosage and tra expression play crucial roles in determining sex-specific size in Drosophila larvae and adult tissue. Since Myc also acts as an XSE that contributes to tra activation in early, development, a double dose of Myc in females serves at least twice in development to promote sexual size dimorphism.

  3. The Am-tra2 gene is an essential regulator of female splice regulation at two levels of the sex determination hierarchy of the honeybee.

    Science.gov (United States)

    Nissen, Inga; Müller, Miriam; Beye, Martin

    2012-11-01

    Heteroallelic and homo- or hemiallelic Complementary sex determiner (Csd) proteins determine sexual fate in the honeybee (Apis mellifera) by controlling the alternative splicing of the downstream gene fem (feminizer). Thus far, we have little understanding of how heteroallelic Csd proteins mediate the splicing of female fem messenger RNAs (mRNAs) or how Fem proteins direct the splicing of honeybee dsx (Am-dsx) pre-mRNAs. Here, we report that Am-tra2, which is an ortholog of Drosophila melanogaster tra2, is an essential component of female splicing of the fem and Am-dsx transcripts in the honeybee. The Am-tra2 transcripts are alternatively (but non-sex-specifically) spliced, and they are translated into six protein isoforms that all share the basic RNA-binding domain/RS (arginine/serine) domain structure. Knockdown studies showed that the Am-tra2 gene is required to splice fem mRNAs into the productive female and nonproductive male forms. We suggest that the Am-Tra2 proteins are essential regulators of fem pre-mRNA splicing that, together with heteroallelic Csd proteins and/or Fem proteins, implement the female pathway. In males, the Am-Tra2 proteins may enhance the switch of fem transcripts into the nonproductive male form when heteroallelic Csd proteins are absent. This dual function of Am-Tra2 proteins possibly enhances and stabilizes the binary decision process of male/female splicing. Our knockdown studies also imply that the Am-Tra2 protein is an essential regulator for Am-dsx female splice regulation, suggesting an ancestral role in holometabolous insects. We also provide evidence that the Am-tra2 gene has an essential function in honeybee embryogenesis that is unrelated to sex determination.

  4. Sox9 gene regulation and the loss of the XY/XX sex-determining mechanism in the mole vole Ellobius lutescens.

    Science.gov (United States)

    Bagheri-Fam, Stefan; Sreenivasan, Rajini; Bernard, Pascal; Knower, Kevin C; Sekido, Ryohei; Lovell-Badge, Robin; Just, Walter; Harley, Vincent R

    2012-01-01

    In most mammals, the Y chromosomal Sry gene initiates testis formation within the bipotential gonad, resulting in male development. SRY is a transcription factor and together with SF1 it directly up-regulates the expression of the pivotal sex-determining gene Sox9 via a 1.3-kb cis-regulatory element (TESCO) which contains an evolutionarily conserved region (ECR) of 180 bp. Remarkably, several rodent species appear to determine sex in the absence of Sry and a Y chromosome, including the mole voles Ellobius lutescens and Ellobius tancrei, whereas Ellobius fuscocapillus of the same genus retained Sry. The sex-determining mechanisms in the Sry-negative species remain elusive. We have cloned and sequenced 1.1 kb of E. lutescens TESCO which shares 75% sequence identity with mouse TESCO indicating that testicular Sox9 expression in E. lutescens might still be regulated via TESCO. We have also cloned and sequenced the ECRs of E. tancrei and E. fuscocapillus. While the three Ellobius ECRs are highly similar (94-97% sequence identity), they all display a 14-bp deletion (Δ14) removing a highly conserved SOX/TCF site. Introducing Δ14 into mouse TESCO increased both basal activity and SF1-mediated activation of TESCO in HEK293T cells. We propose a model whereby Δ14 may have triggered up-regulation of Sox9 in XX gonads leading to destabilization of the XY/XX sex-determining mechanism in Ellobius. E. lutescens/E. tancrei and E. fuscocapillus could have independently stabilized their sex determination mechanisms by Sry-independent and Sry-dependent approaches, respectively.

  5. Optimization and validation of a stability-indicating RP-HPLC method for determination of azithromycin and its related compounds.

    Science.gov (United States)

    El-Gindy, Alaa; Attia, Khalid A; Nassar, Mohammad Wafaa; Al Abasawi, Nasr M; Al-Shabrawi, Maisra

    2011-01-01

    A validated stability-indicating HPLC method was developed for the analysis of azithromycin (AZ) and its related compounds in raw materials, capsule, and suspension using an Xterra RP C18 column at 50 degrees C with UV detection at 215 nm. Isocratic elution was employed using the mobile phase 14 mM disodium hydrogen phosphate (pH 10.5, adjusted by 1 M NaOH)-methanol-acetonitrile-tetrahydrofuran (40.0 + 30.0 + 30.0 + 0.1, v/v/v/v). AZ and 14 of its related compounds were separated and quantified. The described method was linear over the range of 2-1800 microg/mL AZ with (r = 0.9999). The stability of AZ was studied under accelerated acidic, alkaline, and oxidative conditions. The proposed method was used to investigate the kinetics of acidic and alkaline hydrolysis process of AZ at different temperatures, and the apparent pseudo first-order rate constant, half-life, and activation energy were calculated. The major peak detected from the degradation of AZ in alkaline and acidic conditions was decladinosylazithromycine, while azithromycin N-oxide was detected from the oxidative degradation. Long-term stability studies for capsule and oral suspension were carried out. The proposed stability-indicating method was completely validated according to the U.S. Food and Drug Administration requirements.

  6. Women's opinions about attending for breast cancer screening: Stability of cognitive determinants during three rounds of screening.

    NARCIS (Netherlands)

    Drossaert, Constance H.C.; Boer, Hendrik; Seydel, E.R.

    2005-01-01

    Examines women's opinions about attending breast cancer screening. Stability of beliefs and intentions towards repeat attendance at breast cancer screening; Assessment of whether cognitions changed in the course of the programme; Increase of attendance in subsequent rounds of breast cancer screening

  7. Determination of the stability constant of Np(V) fluoride complex using a fluoride ion selective electrode

    Energy Technology Data Exchange (ETDEWEB)

    Sawant, R.M.; Rizvi, G.H.; Chaudhuri, N.K.; Patil, S.K. (Bhabha Atomic Research Centre, Bombay (India). Radiochemistry Div.)

    1985-04-01

    Fluoride complexing of Np(V) was studied using fluoride ion selective electrode (F-ISE). Free fluoride ion concentrations in the presence of Np(V) were measured at 0.1 and 0.01M ionic strength. The data were used to calculate the stability constant of the fluoride complex of Np(V) and the values obtained are reported.

  8. Factors That Determine the Career Stability of Assistant Principals in a Large Urban School District in the Southeast

    Science.gov (United States)

    Edwards, Danielle Felder

    2009-01-01

    This study examined the career stability (career choices assistant principals intend to make over the next five to ten years) in a large, urban school district in the southeastern region of the United States in order to identify factors significantly related to their career aspirations. The study invited a purposive sample (n = 177) of assistant…

  9. Women's opinions about attending for breast cancer screening: Stability of cognitive determinants during three rounds of screening.

    NARCIS (Netherlands)

    Drossaert, C.H.C.; Boer, H.; Seydel, E.R.

    2005-01-01

    Examines women's opinions about attending breast cancer screening. Stability of beliefs and intentions towards repeat attendance at breast cancer screening; Assessment of whether cognitions changed in the course of the programme; Increase of attendance in subsequent rounds of breast cancer screening

  10. Cloning, Expression, and Chromosomal Stabilization of the Propionibacterium shermanii Proline Iminopeptidase Gene (pip) for Food-Grade Application in Lactococcus lactis

    Science.gov (United States)

    Leenhouts, Kees; Bolhuis, Albert; Boot, Johan; Deutz, Inge; Toonen, Marjolein; Venema, Gerard; Kok, Jan; Ledeboer, Aat

    1998-01-01

    Proline iminopeptidase produced by Propionibacterium shermanii plays an essential role in the flavor development of Swiss-type cheeses. The enzyme (Pip) was purified and characterized, and the gene (pip) was cloned and expressed in Escherichia coli and Lactococcus lactis, the latter species being an extensively studied, primary cheese starter culture that is less fastidious in its growth condition requirements than P. shermanii. The levels of expression of the pip gene could be enhanced with a factor 3 to 5 by using a strong constitutive promoter in L. lactis or the inducible tac promoter in E. coli. Stable replication of the rolling-circle replicating (rcr) plasmid, used to express pip in L. lactis, could only be obtained by providing the repA gene in trans. Upon the integration of pip, clear gene dosage effects were observed and stable multicopy integrants could be maintained upon growth under the selective pressure of sucrose. The multicopy integrants demonstrated a high degree of stability in the presence of glucose. This study examines the possibilities to overexpress genes that play an important role in food fermentation processes and shows a variety of options to obtain stable food-grade expression of such genes in L. lactis. PMID:9835556

  11. How ion properties determine the stability of a lipase enzyme in ionic liquids: a molecular dynamics study.

    Science.gov (United States)

    Klähn, Marco; Lim, Geraldine S; Wu, Ping

    2011-11-07

    The influence of eight different ionic liquid (IL) solvents on the stability of the lipase Candida antarctica lipase B (CAL-B) is investigated with molecular dynamics (MD) simulations. Considered ILs contain cations that are based either on imidazolium or guanidinium as well as nitrate, tetrafluoroborate or hexafluorophosphate anions. Partial unfolding of CAL-B is observed during high-temperature MD simulations and related changes of CAL-B regarding its radius of gyration, surface area, secondary structure, amount of solvent close to the backbone and interaction strength with the ILs are evaluated. CAL-B stability is influenced primarily by anions in the order NO(3)(-)≪ BF(4)(-) helices and an increase of surface area, radius of gyration and protein-IL total interaction strength of CAL-B, all of which describe a destabilization of the folded protein state. On the other hand, a destabilization of the protein core is facilitated when direct core-IL interactions are feasible. This is the case when long alkyl chains are involved or when particularly hydrophobic ILs induce major conformational changes that enable ILs direct access to the protein core. This core instability is characterized by a disintegration of β-sheets, diffusion of ions into CAL-B and increasing protein-IL van der Waals interactions. This process describes a stabilization of the unfolded protein state. Both of these processes reduce the folding free energy and thus destabilize CAL-B. The results of this work clarify the impact of ions on CAL-B stabilization. An extrapolation of the observed trends enables proposing novel ILs in which protein stability could be enhanced further.

  12. Differences in gene expression of human xylosyltransferases and determination of acceptor specificities for various proteoglycans

    Energy Technology Data Exchange (ETDEWEB)

    Roch, Christina; Kuhn, Joachim; Kleesiek, Knut [Institut fuer Laboratoriums- und Transfusionsmedizin, Herz- und Diabeteszentrum NRW, Universitaetsklinik der Ruhr-Universitaet Bochum, 32545 Bad Oeynhausen (Germany); Goetting, Christian, E-mail: cgoetting@hdz-nrw.de [Institut fuer Laboratoriums- und Transfusionsmedizin, Herz- und Diabeteszentrum NRW, Universitaetsklinik der Ruhr-Universitaet Bochum, 32545 Bad Oeynhausen (Germany)

    2010-01-01

    The xylosyltransferase (XT) isoforms XT-I and XT-II initiate the posttranslational glycosaminoglycan (GAG) synthesis. Here, we determined the relative expression of both isoforms in 33 human cell lines. The majority of tested cell lines showed dominant XYLT2 gene expression, while only in 23132/87, JAR, NCI-H510A and THP-1 was the XT-I mRNA expression higher. Nearly equal expression levels were detected in six cell lines. Additionally, to shed light on putative differences in acceptor specificities the acceptor properties of potential acceptor sequences were determined. Peptides were expressed as glutathione-S-transferase fusion proteins containing putative or known GAG attachment sites of in vivo proteoglycans. Kinetic analysis showed that K{sub m} and V{sub max} values for XT-I mediated xylosylation were slightly higher than those for XT-II, and that XT-I showed a lesser stringency concerning the acceptor sequence. Mutagenesis of the bikunin peptide sequence in the G-S-G attachment site and flanking regions generated potential acceptor molecules. Here, mutations on the N-terminal side and the attachment site were found to be more susceptible to a loss of acceptor function than mutations in the C-terminus. Altogether the known consensus sequence a-a-a-a-G-S-G-a-a/G-a ('a' representing Asp or Glu) for XT-I mediated xylosylation could be approved and additionally extended to apply to XT-II as well.

  13. Expression profile of the sex determination gene doublesex in a gynandromorph of bumblebee, Bombus ignitus

    Science.gov (United States)

    Ugajin, Atsushi; Matsuo, Koshiro; Kubo, Ryohei; Sasaki, Tetsuhiko; Ono, Masato

    2016-04-01

    Gynandromorphy that has both male and female features is known in many insect orders, including Hymenoptera. In most cases, however, only external morphology and behavioral aspects have been studied. We found a gynandromorph of bumblebee, Bombus ignitus, that showed almost bilateral distribution of external sexual traits, with male characters observed on the left side and female characters on the right side. This individual never exhibited sexual behavior toward new queens. The dissection of the head part showed that it had bilaterally dimorphic labial glands, only the left of which was well developed and synthesized male-specific pheromone components. In contrast, the gynandromorph possessed an ovipositor and a pair of ovaries in the abdominal part, suggesting that it had a uniformly female reproductive system. Furthermore, we characterized several internal organs of the gynandromorph by a molecular biological approach. The expression analyses of a sex determination gene, doublesex, in the brain, the fat bodies, the hindgut, and the ovaries of the gynandromorph revealed a male-type expression pattern exclusively in the left brain hemisphere and consistent female-type expression in other tissues. These findings clearly indicate the sexual discordance between external traits and internal organs in the gynandromorph. The results of genetic analyses using microsatellite markers suggested that this individual consisted of both genetically male- and female-type tissues.

  14. Determination of the differentially expressed genes in microarray experiments using local FDR

    Directory of Open Access Journals (Sweden)

    Daudin J-J

    2004-09-01

    Full Text Available Abstract Background Thousands of genes in a genomewide data set are tested against some null hypothesis, for detecting differentially expressed genes in microarray experiments. The expected proportion of false positive genes in a set of genes, called the False Discovery Rate (FDR, has been proposed to measure the statistical significance of this set. Various procedures exist for controlling the FDR. However the threshold (generally 5% is arbitrary and a specific measure associated with each gene would be worthwhile. Results Using process intensity estimation methods, we define and give estimates of the local FDR, which may be considered as the probability for a gene to be a false positive. After a global assessment rule controlling the false positive error, the local FDR is a valuable guideline for deciding wether a gene is differentially expressed. The interest of the method is illustrated on three well known data sets. A R routine for computing local FDR estimates from p-values is available at http://www.inapg.fr/ens_rech/mathinfo/recherche/mathematique/outil.html. Conclusions The local FDR associated with each gene measures the probability that it is a false positive. It gives the opportunity to compute the FDR of any given group of clones (of the same gene or genes pertaining to the same regulation network or the same chromosomic region.

  15. Determination of the composition of the organic matter chemically stabilized by agricultural soil clay minerals: Spectroscopy and Density Fractionation

    Science.gov (United States)

    Oufqir, Sofia; Bloom, Paul; Toner, Brandy; Hatcher, Patrick

    2014-05-01

    The interactions between soil organic matter and clay minerals are considered important processes because of their ability to sequester C in soil for long periods of time, and hence control C in the global C cycle when present. However, differing results have been reported regarding the composition of the soil organic matter - aromatic fractions versus aliphatic fractions - associated with clay minerals. To clarify this critical issue and better understand the C sequestration process in soils, we aimed to determine the nature of the chemically bound natural organic matter on clay surfaces, and to probe the speciation and spatial distribution of C in the soil clay nanoparticles using direct spectroscopic measurements namely solid-state CP-MAS and DP-MAS 13C NMR spectroscopy, x-ray diffraction spectroscopy (XRD), and scanning transmission x-ray microscopy (STXM). We tested the hypotheses that peptides and polysaccharides are stabilized by the smectite-illite clay while the lipids and black carbon are a separate phase; and that they are evenly distributed on clay surfaces. A soil clay fraction (5.5% organic C) was isolated from the surface of a prairie soil (Mollisol) in southwestern Minnesota, characterized by a pH 6.0, 32.5% clay content, and 3.7% organic carbon, using a sonication-sedimentation-siphoning process in distilled water. Then was subjected to density separation combined with low energy ultrasonic dispersion to separate the free organic and black C (light fraction) from the chemically bound C (heavy fraction). The XRD results indicated a dominance of interstratified smectite-illite clays in soil. The 13C-NMR spectra of the soil clay fraction suggested that polysaccharides and polypeptides are the prevailing components of the organic matter associated with the mineral clay, with only a minor component of aromatic C. The light fraction has strong alkyl C-H bands characteristic of fatty acids plus strong C-O bands characteristic of polysaccharides, including

  16. The role of the transformer gene in sex determination and reproduction in the tephritid fruit fly, Bactrocera dorsalis (Hendel).

    Science.gov (United States)

    Peng, Wei; Zheng, Wenping; Handler, Alfred M; Zhang, Hongyu

    2015-12-01

    Transformer (tra) is a switch gene in the somatic sex-determination hierarchy that regulates sexual dimorphism based on RNA splicing in many insects. In tephritids, a Y-linked male determining gene (M) controls sex in the sex-determination pathway. Here, homologues of Drosophila tra and transformer-2 (tra-2) genes were isolated and characterized in Bactrocera dorsalis (Hendel), one of the most destructive agricultural insect pests in many Asian countries. Two male-specific and one female-specific isoforms of B. dorsalis transformer (Bdtra) were identified. The presence of multiple TRA/TRA-2 binding sites in Bdtra suggests that the TRA/TRA-2 proteins are splicing regulators promoting and maintaining, epigenetically, female sex determination by a tra positive feedback loop in XX individuals during development. The expression patterns of female-specific Bdtra transcripts during early embryogenesis shows that a peak appears at 15 h after egg laying. Using dsRNA to knock-down Bdtra expression in the embryo and adult stages, we showed that sexual formation is determined early in the embryo stage and that parental RNAi does not lead to the production of all male progeny as in Tribolium castaneum. RNAi results from adult abdominal dsRNA injections show that Bdtra has a positive influence on female yolk protein gene (Bdyp1) expression and fecundity.

  17. A Stability-Indicating HPLC-DAD Method for Determination of Stiripentol: Development, Validation, Kinetics, Structure Elucidation and Application to Commercial Dosage Form

    OpenAIRE

    Hany W. Darwish; Abdelhameed, Ali S; Mohamed I. Attia; Bakheit, Ahmed H.; Khalil, Nasr Y; Al-Majed, Abdulrahman A.

    2014-01-01

    A rapid, simple, sensitive, and accurate isocratic reversed-phase stability-indicating high performance liquid chromatography method has been developed and validated for the determination of stiripentol and its degradation product in its bulk form and pharmaceutical dosage form. Chromatographic separation was achieved on a Symmetry C18 column and quantification was achieved using photodiode array detector (DAD). The method was validated in accordance with the ICH requirements showing specific...

  18. A new method for determination of most likely landslide initiation points and the evaluation of digital terrain model scale in terrain stability mapping

    Directory of Open Access Journals (Sweden)

    P. Tarolli

    2006-01-01

    Full Text Available This paper introduces a new approach for determining the most likely initiation points for landslides from potential instability mapped using a terrain stability model. This approach identifies the location with critical stability index from a terrain stability model on each downslope path from ridge to valley. Any measure of terrain stability may be used with this approach, which here is illustrated using results from SINMAP, and from simply taking slope as an index of potential instability. The relative density of most likely landslide initiation points within and outside mapped landslide scars provides a way to evaluate the effectiveness of a terrain stability measure, even when mapped landslide scars include run out zones, rather than just initiation locations. This relative density was used to evaluate the utility of high resolution terrain data derived from airborne laser altimetry (LIDAR for a small basin located in the Northeastern Region of Italy. Digital Terrain Models were derived from the LIDAR data for a range of grid cell sizes (from 2 to 50 m. We found appreciable differences between the density of most likely landslide initiation points within and outside mapped landslides with ratios as large as three or more with the highest ratios for a digital terrain model grid cell size of 10 m. This leads to two conclusions: (1 The relative density from a most likely landslide initiation point approach is useful for quantifying the effectiveness of a terrain stability map when mapped landslides do not or can not differentiate between initiation, runout, and depositional areas; and (2 in this study area, where landslides occurred in complexes that were sometimes more than 100 m wide, a digital terrain model scale of 10 m is optimal. Digital terrain model scales larger than 10 m result in loss of resolution that degrades the results, while for digital terrain model scales smaller than 10 m the physical processes responsible for triggering

  19. Analiza rezultata određivanja sadržaja stabilizatora u prirodno starenim barutima / Content of stabilizer determination results analysis of naturally aged gun propellants

    Directory of Open Access Journals (Sweden)

    Luka Grbović

    2006-04-01

    Full Text Available Praćenje sadržaja stabilizatora jedna je od suvremenih i pouzdanih metoda koja se u svetu, a i kod nas, primenjuje za kontrolu kemijske stabilnosti i prognoziranje veka upotrebljivosti baruta. Radi ocene kemijske stabilnosti baruta domaće proizvodnje u radu su analizirani rezultati određivanja sadržaja stabilizatora u prirodno starenim jednobaznim barutima tipa NC i NCD i dvobaznim barutima tipa NGB i NGH. Pri tome su uporedno analizirani rezultati uzoraka iz kolekcija baruta iz Kragujevca (KB-1 i Zelenike (KB-2. Dobijeni rezultati ukazuju na znatno brži pad stabilnosti uzoraka baruta iz kolekcije KB-2, što se može objasniti značajnim uticajem klimatskih uslova. Osim toga, dobijeni rezultati potvrđuju uticaj mnogih faktora na kemijsku stabilnost baruta, kao što su: sastav, kvalitet sirovina, tehnološki postupak, oblik i dimenzije barutnog zrna i sl. / Monitoring of the stabilizer content is one of the modern and reliable methods in the world, and here also, which is used to control chemical stability and to predict the usage time of gun propellants. For the purpose of evaluation of gun propellants' chemical stability manufactured in our country, in this paper -we analyzed results from the content of stabilizer determination in naturally aged single base, NC and NCD type, and double base, NGB and NGH type of gun propellants. At the same time -we analyzed results from both collections of gun propellant samples, one from Kragujevac factory (KB-1 and the other from Zelenika factory (KB-2. Obtained results point that the stability decays faster for KB-2 collection samples, -which can be explained due to climate factors. Beside this influence, obtained results prove the influence of many factors on chemical stability of gun propellants: used components, quality of the ingredients, procedures of manufacture, shape and dimensions of gun propellant grains, etc.

  20. Effect of various salts on the stability of lansoprazole, omeprazole, and pantoprazole as determined by high-performance liquid chromatography.

    Science.gov (United States)

    Ekpe, A; Jacobsen, T

    1999-09-01

    A fast and reproducible reverse-phase high-performance liquid chromatography (HPLC) assay method has been developed for the simultaneous quantitation of omeprazole, lansoprazole, and pantoprazole. The three compounds were monitored at 280 nm using Zorbax Eclipse XDB C8 (5 microns, 150 cm x 4.6 mm i.d.) and a mobile phase consisting of 700:300 phosphate buffer:acetonitrile with the pH adjusted to 7.0 with phosphoric acid. The method was used to study the effect of pH and various salts on the stability of the three compounds. The pH rate profile curve showed that pantoprazole was the most stable compound and lansoprazole the least stable. The stabilities of the compounds in salt solutions were found to be in the following order: phosphate buffer bicarbonate < sodium chloride < water. The rate of degradation had a direct relationship with the H+ and salt concentration.

  1. DETERMINATION OF THE STABILITY AND SHELF LIFE OF OINTMENT WITH ZINC SALT OF HYALURONIC ACID AND THIOTRIAZOLINE

    Directory of Open Access Journals (Sweden)

    Bezrukaviy Y.A.

    2013-10-01

    Full Text Available Based on the results of physical-chemical andmicrobiological research investigated the stability andshelf life of ointment with zinc salt of hyaluronic acid andthiotriazoline. Ointment which stored in aluminum tubes,during all term of researches met all the indicators ofdrugs QC. Ointment which kept in jars from orange glassdid not meet the indicator "acidic number" in defining ofindicators per 24 months of storage. Was established shelflife of medication with zinc salt of hyaluronic acid andthiotriazoline - 2 years.

  2. Characterization of molecular determinants of the conformational stability of macrophage migration inhibitory factor: leucine 46 hydrophobic pocket.

    Directory of Open Access Journals (Sweden)

    Farah El-Turk

    Full Text Available Macrophage Migration Inhibitory Factor (MIF is a key mediator of inflammatory responses and innate immunity and has been implicated in the pathogenesis of several inflammatory and autoimmune diseases. The oligomerization of MIF, more specifically trimer formation, is essential for its keto-enol tautomerase activity and probably mediates several of its interactions and biological activities, including its binding to its receptor CD74 and activation of certain signaling pathways. Therefore, understanding the molecular factors governing the oligomerization of MIF and the role of quaternary structure in modulating its structural stability and multifunctional properties is crucial for understanding the function of MIF in health and disease. Herein, we describe highly conserved intersubunit interactions involving the hydrophobic packing of the side chain of Leu46 onto the β-strand β3 of one monomer within a hydrophobic pocket from the adjacent monomer constituted by residues Arg11, Val14, Phe18, Leu19, Val39, His40, Val41, Val42, and Pro43. To elucidate the structural significance of these intersubunit interactions and their relative contribution to MIF's trimerization, structural stability and catalytic activity, we generated three point mutations where Leu46 was replaced by glycine (L46G, alanine (L46A and phenylalanine (L46F, and their structural properties, stability, oligomerization state, and catalytic activity were characterized using a battery of biophysical methods and X-ray crystallography. Our findings provide new insights into the role of the Leu46 hydrophobic pocket in stabilizing the conformational state of MIF in solution. Disrupting the Leu46 hydrophobic interaction perturbs the secondary and tertiary structure of the protein but has no effect on its oligomerization state.

  3. Environmental Sex Determination in the Branchiopod Crustacean Daphnia magna: Deep Conservation of a Doublesex Gene in the Sex-Determining Pathway

    Science.gov (United States)

    Kato, Yasuhiko; Kobayashi, Kaoru; Watanabe, Hajime; Iguchi, Taisen

    2011-01-01

    Sex-determining mechanisms are diverse among animal lineages and can be broadly divided into two major categories: genetic and environmental. In contrast to genetic sex determination (GSD), little is known about the molecular mechanisms underlying environmental sex determination (ESD). The Doublesex (Dsx) genes play an important role in controlling sexual dimorphism in genetic sex-determining organisms such as nematodes, insects, and vertebrates. Here we report the identification of two Dsx genes from Daphnia magna, a freshwater branchiopod crustacean that parthenogenetically produces males in response to environmental cues. One of these genes, designated DapmaDsx1, is responsible for the male trait development when expressed during environmental sex determination. The domain organization of DapmaDsx1 was similar to that of Dsx from insects, which are thought to be the sister group of branchiopod crustaceans. Intriguingly, the molecular basis for sexually dimorphic expression of DapmaDsx1 is different from that of insects. Rather than being regulated sex-specifically at the level of pre–mRNA splicing in the coding region, DapmaDsx1 exhibits sexually dimorphic differences in the abundance of its transcripts. During embryogenesis, expression of DapmaDsx1 was increased only in males and its transcripts were primarily detected in male-specific structures. Knock-down of DapmaDsx1 in male embryos resulted in the production of female traits including ovarian maturation, whereas ectopic expression of DapmaDsx1 in female embryos resulted in the development of male-like phenotypes. Expression patterns of another D. magna Dsx gene, DapmaDsx2, were similar to those of DapmaDsx1, but silencing and overexpression of this gene did not induce any clear phenotypic changes. These results establish DapmaDsx1 as a key regulator of the male phenotype. Our findings reveal how ESD is implemented by selective expression of a fundamental genetic component that is functionally conserved

  4. Techniques to determine ignition, flame stability and burnout of blended coals in p.f. power station boilers

    Energy Technology Data Exchange (ETDEWEB)

    Su, S.; Pohl, J.H.; Holcombe, D.; Hart, J.A. [University of Queensland, Brisbane, Qld. (Australia). Dept. of Chemical Engineering

    2001-07-01

    The blending of coals has become popular to improve the performance of coals, to meet specifications of power plants and to reduce the cost of coals. This article reviews the results and provides new information on ignition, flame stability, and carbon burnout studies of blended coals. The reviewed studies were conducted in laboratory-, pilot-, and full-scale facilities. The new information was taken in pilot-scale studies. The results generally show that blending a high-volatile coal with a low-volatile coal or anthracite can improve the ignition, flame stability and burnout of the blends. This paper discusses two general methods to predict the performance of blended coals: (1) experiment; and (2) indices. Laboratory- and pilot-scale tests, at least, provide a relative ranking of the combustion performance of coal/blends in power station boilers. Several indices, volatile matter content, heating value and a maceral index, can be used to predict the relative ranking of ignitability and flame stability of coals and blends. The maceral index, fuel ratio, and vitrinite reflectance can also be used to predict the absolute carbon burnout of coal and blends within limits. 59 refs., 20 figs., 4 tabs.

  5. Determining the Composition and Stability of Protein Complexes Using an Integrated Label-Free and Stable Isotope Labeling Strategy

    Science.gov (United States)

    Greco, Todd M.; Guise, Amanda J.; Cristea, Ileana M.

    2016-01-01

    In biological systems, proteins catalyze the fundamental reactions that underlie all cellular functions, including metabolic processes and cell survival and death pathways. These biochemical reactions are rarely accomplished alone. Rather, they involve a concerted effect from many proteins that may operate in a directed signaling pathway and/or may physically associate in a complex to achieve a specific enzymatic activity. Therefore, defining the composition and regulation of protein complexes is critical for understanding cellular functions. In this chapter, we describe an approach that uses quantitative mass spectrometry (MS) to assess the specificity and the relative stability of protein interactions. Isolation of protein complexes from mammalian cells is performed by rapid immunoaffinity purification, and followed by in-solution digestion and high-resolution mass spectrometry analysis. We employ complementary quantitative MS workflows to assess the specificity of protein interactions using label-free MS and statistical analysis, and the relative stability of the interactions using a metabolic labeling technique. For each candidate protein interaction, scores from the two workflows can be correlated to minimize nonspecific background and profile protein complex composition and relative stability. PMID:26867737

  6. Genomic organisation and characterisation of the neural sex-determination gene fruitless (fru) in the Hawaiian species Drosophila heteroneura.

    Science.gov (United States)

    Davis, T; Kurihara, J; Yamamoto, D

    2000-04-04

    There are several mechanisms for the determination of sex. Sexual behaviour is part of the sex-determination cascade, and in Drosophila melanogaster male courtship is controlled in part by the fruitless gene. As part of a study of sexual behaviour in Hawaiian Drosophila, we have cloned the neural sex-determination gene fru from the Hawaiian picture-wing species Drosophila heteroneura. The fru gene has at least seven exons covering a region of 18kb and encodes three transcripts, fruA, fruB and fruC. Each transcript encodes a single ORF of 841, 678 and 691aa, respectively. The FRUA and FRUB proteins have a BTB protein-protein-binding domain and two zinc finger-like domains and are well conserved with the D. melanogaster proteins. The FRUC protein has a BTB domain but no zinc finger-like domains. The fru gene is expressed in 1-7 day old adult males as a 5.1kb transcript. This transcript is not seen in adult females, so the fru gene has a different pattern of sex-differential expression in the Hawaiian Drosophila compared with D. melanogaster.

  7. Genome editing reveals dmrt1 as an essential male sex-determining gene in Chinese tongue sole (Cynoglossus semilaevis)

    Science.gov (United States)

    Cui, Zhongkai; Liu, Yun; Wang, Wenwen; Wang, Qian; Zhang, Ning; Lin, Fan; Wang, Na; Shao, Changwei; Dong, Zhongdian; Li, Yangzhen; Yang, Yingming; Hu, Mengzhu; Li, Hailong; Gao, Fengtao; Wei, Zhanfei; Meng, Liang; Liu, Yang; Wei, Min; Zhu, Ying; Guo, Hua; Cheng, Christopher H. K.; Schartl, Manfred; Chen, Songlin

    2017-01-01

    Chinese tongue sole is a marine fish with ZW sex determination. Genome sequencing suggested that the Z-linked dmrt1 is a putative male determination gene, but direct genetic evidence is still lacking. Here we show that TALEN of dmrt1 efficiently induced mutations of this gene. The ZZ dmrt1 mutant fish developed ovary-like testis, and the spermatogenesis was disrupted. The female-related genes foxl2 and cyp19a1a were significantly increased in the gonad of the ZZ dmrt1 mutant. Conversely, the male-related genes Sox9a and Amh were significantly decreased. The dmrt1 deficient ZZ fish grew much faster than ZZ male control. Notably, we obtained an intersex ZW fish with a testis on one side and an ovary on the other side. This fish was chimeric for a dmrt1 mutation in the ovary, and wild-type dmrt1 in the testis. Our data provide the first functional evidence that dmrt1 is a male determining gene in tongue sole. PMID:28205594

  8. HIV-1 frameshift efficiency is primarily determined by the stability of base pairs positioned at the mRNA entrance channel of the ribosome.

    Science.gov (United States)

    Mouzakis, Kathryn D; Lang, Andrew L; Vander Meulen, Kirk A; Easterday, Preston D; Butcher, Samuel E

    2013-02-01

    The human immunodeficiency virus (HIV) requires a programmed -1 ribosomal frameshift for Pol gene expression. The HIV frameshift site consists of a heptanucleotide slippery sequence (UUUUUUA) followed by a spacer region and a downstream RNA stem-loop structure. Here we investigate the role of the RNA structure in promoting the -1 frameshift. The stem-loop was systematically altered to decouple the contributions of local and overall thermodynamic stability towards frameshift efficiency. No correlation between overall stability and frameshift efficiency is observed. In contrast, there is a strong correlation between frameshift efficiency and the local thermodynamic stability of the first 3-4 bp in the stem-loop, which are predicted to reside at the opening of the mRNA entrance channel when the ribosome is paused at the slippery site. Insertion or deletions in the spacer region appear to correspondingly change the identity of the base pairs encountered 8 nt downstream of the slippery site. Finally, the role of the surrounding genomic secondary structure was investigated and found to have a modest impact on frameshift efficiency, consistent with the hypothesis that the genomic secondary structure attenuates frameshifting by affecting the overall rate of translation.

  9. Determining the effect of DNA methylation on gene expression in cancer cells.

    Science.gov (United States)

    Lee, Chai-Jin; Evans, Jared; Kim, Kwangsoo; Chae, Heejoon; Kim, Sun

    2014-01-01

    DNA methylation, a DNA modification by adding methyl group to cytosine, has an important role in the regulation of gene expression. DNA methylation is known to be associated with gene transcription by interfering with DNA-binding proteins, such as transcription factors. DNA methylation is closely related to tumorigenesis, and the methylation state of some genes can be used as a biomarker for tumorigenesis. Aberrant DNA methylation of genomic regions, including CpG islands, CpG shores, and first exons, is related to the altered gene expression pattern characteristics of all human cancers. Subheading 1 surveys recent developments on DNA methylation and gene expressions in cancer. Then we provide analysis of DNA methylation and gene expression in 30 breast cancer cell lines representing different tumor phenotypes. This study conducted an integrated analysis to identify the relationship between DNA methylation in various genomic regions and expression levels of downstream genes, using MethylCapseq data (affinity purification followed by next-generation sequencing of eluted DNA) and Affymetrix gene expression microarray data. The goal of this study was to assess genome-wide methylation profiles associated with different molecular subtypes of human breast cancer (luminal, basal A, and basal B) and to comprehensively investigate the effect of DNA methylation on gene expression in breast cancer phenotypes. This showed that methylation of genomic regions near transcription start sites, CpG island, CpG shore, and first exon was strongly associated with gene repression, and the effects of the regions on gene expression patterns were different for different molecular subtypes of breast cancer. The results further indicated that aberrant methylation of specific genomic regions was significantly associated with different breast cancer subtypes.

  10. Matrix attachment regions included in a bicistronic vector enhances and stabilizes follistatin gene expressions in both transgenic cells and transgenic mice

    Directory of Open Access Journals (Sweden)

    Xiaoming HU,Jing GUO,Chunling BAI,Zhuying WEI,Li GAO,Tingmao HU,Shorgan BOU,Guangpeng LI

    2016-03-01

    Full Text Available In the present study, follistatin (FST gene expression vectors with either a bicistronic gene transfer cassette alone, or a bicistron gene cassette carrying a matrix attachment region (MAR were constructed and transfected to bovine fetal fibroblasts. Evaluations of both the integration and expression of exogenous FST indicated that the pMAR-CAG-FST-IRES-AcGFP1-polyA-MAR (pMAR-FST vector had higher capacity to form monoclonal transgenic cells than the vector without MAR, though transient transfection and integration efficiency were similar with either construct. Remarkably, protein expression in transgenic cells with the pMAR-FST vector was significantly higher than that from the bicistronic vector. Exogenous FST was expressed in all of the pMAR-FST transgenic mice at F0, F1 and F2. Total muscle growth in F0 mice was significantly greater than in wild-type mice, with larger muscles in fore and hind limbs of transgenic mice. pMAR-FST transgenic mice were also found with more evenly distributed muscle bundles and thinner spaces between sarcolemma, which suggests a correlation between transgene expression-associated muscle development and the trend of muscle growth. In conclusion, a pMAR-FST vector, which excluded the resistant genes and frame structure, enhances and stabilizes FST gene expressions in both transfected cells and transgenic mice.

  11. Determination of variants in the 3'-region of the Tyrosinase gene requires locus specific amplification.

    NARCIS (Netherlands)

    Chaki, M.; Mukhopadhyay, A.; Ray, K.

    2005-01-01

    Mutations in the Tyrosinase gene (TYR, 11q14-q21) cause oculocutaneous albinism type 1 (OCA1). The 3'-region of the TYR shows 98.55% sequence identity with a pseudogene, known as Tyrosinase-Like Gene (TYRL, 11p11.2-cen). A large number of publicly available nucleotide variants of TYR in this region

  12. The Bsister MADS gene FST determines ovule patterning and development of the zygotic embryo and endosperm.

    Directory of Open Access Journals (Sweden)

    Dong Sun Lee

    Full Text Available Many homeotic MADS-box genes have been identified as controllers of the floral transition and floral development. However, information regarding Bsister (Bs-function genes in monocots is still limited. Here, we describe the functional characterization of a Bs-group MADS-box gene FEMALE-STERILE (FST, whose frame-shift mutation (fst results in abnormal ovules and the complete abortion of zygotic embryos and endosperms in rice. Anatomical analysis showed that the defective development in the fst mutant exclusively occurred in sporophytic tissues including integuments, fertilized proembryos and endosperms. Analyses of the spatio-temporal expression pattern revealed that the prominent FST gene products accumulated in the inner integument, nucellar cell of the micropylar side, apical and base of the proembryos and free endosperm nuclei. Microarray and gene ontology analysis unraveled substantial changes in the expression level of many genes in the fst mutant ovules and seeds, with a subset of genes involved in several developmental and hormonal pathways appearing to be down-regulated. Using both forward and reverse genetics approaches, we demonstrated that rice FST plays indispensable roles and multiple functions during ovule and early seed development. These findings support a novel function for the Bs-group MADS-box genes in plants.

  13. A Preliminary List of Horizontally Transferred Genes in Prokaryotes Determined by Tree Reconstruction and Reconciliation

    Directory of Open Access Journals (Sweden)

    Hyeonsoo Jeong

    2017-08-01

    Full Text Available Genome-wide global detection of genes involved in horizontal gene transfer (HGT remains an active area of research in medical microbiology and evolutionary genomics. Utilizing the explicit evolutionary method of comparing topologies of a total of 154,805 orthologous gene trees against corresponding 16S rRNA “reference” trees, we previously detected a total of 660,894 candidate HGT events in 2,472 completely-sequenced prokaryotic genomes. Here, we report an HGT-index for each individual gene-reference tree pair reconciliation, representing the total number of detected HGT events on the gene tree divided by the total number of genomes (taxa member of that tree. HGT-index is thus a simple measure indicating the sensitivity of prokaryotic genes to participate (or not participate in HGT. Our preliminary list provides HGT-indices for a total of 69,365 genes (detected in >10 and <50% available prokaryotic genomes that are involved in a wide range of biological processes such as metabolism, information, and bacterial response to environment. Identification of horizontally-derived genes is important to combat antibiotic resistance and is a step forward toward reconstructions of improved phylogenies describing the history of life. Our effort is thus expected to benefit ongoing research in the fields of clinical microbiology and evolutionary biology.

  14. Conserved regulation of the soybean early nodulin ENOD2 gene promoter in determinate and indeterminate transgenic root nodules.

    NARCIS (Netherlands)

    Lauridsen, P.; Franssen, H.; Stougaard, J.; Bisseling, T.; Marcker, K.A.

    1993-01-01

    The beta-glucuronidase (GUS) activity expressed from the soybean early nodulin ENOD2(B) gene promoter was localized histochemically in nodules of Lotus corniculatus and Trifolium repens. In both the determinate Lotus nodules and the indeterminate Trifolium nodules, activity was found in the parenchy

  15. Simultaneous determination of gene expression and bacterial identity in single cells in defined mixtures of pure cultures

    DEFF Research Database (Denmark)

    Poulsen, Lars K.; Dalton, Helen M.; Angels, Mark;

    1997-01-01

    A protocol was developed to achieve the simultaneous determination of gene expression and bacterial identity at the level of single cells: a chromogenic beta-galactosidase activity assay was combined with in situ hybridization of Fluorescently labelled oligonucleotide probes to rRNA. The method a...

  16. Determining lower limits of detection of digital PCR assays for cancer-related gene mutations

    Directory of Open Access Journals (Sweden)

    Coren A. Milbury

    2014-09-01

    Full Text Available Digital PCR offers very high sensitivity compared to many other technologies for processing molecular detection assays. Herein, a process is outlined for determining the lower limit of detection (LoD of two droplet-based digital PCR assays for point mutations of the epidermal growth factor receptor (EGFR gene. Hydrolysis probe mutation-detection assays for EGFR p.L858R and p.T790M mutations were characterized in detail. Furthermore, sixteen additional cancer-related mutation assays were explored by the same approach. For the EGFR L8585R assay, the assay sensitivity is extremely good, and thus, the LoD is limited by the amount of amplifiable DNA that is analyzed. With 95% confidence limits, the LoD is one mutant in 180,000 wild-type molecules for the evaluation of 3.3 μg of genomic DNA, and detection of one mutant molecule in over 4 million wild-type molecules was achieved when 70 million copies of DNA were processed. The measured false-positive rate for the EGFR L8585R assay is one in 14 million, which indicates the theoretical LoD if an unlimited amount of DNA is evaluated. For the EFGR T790M assay, the LoD is one mutant in 13,000 for analysis of a 3.3 μg sample of genomic DNA, and the dPCR assay limit sensitivity approaches one mutant in 22,000 wild-type molecules.

  17. Genome-wide experimental determination of barriers to horizontal gene transfer

    Energy Technology Data Exchange (ETDEWEB)

    Rubin, Edward; Sorek, Rotem; Zhu, Yiwen; Creevey, Christopher J.; Francino, M. Pilar; Bork, Peer; Rubin, Edward M.

    2007-09-24

    Horizontal gene transfer, in which genetic material is transferred from the genome of one organism to another, has been investigated in microbial species mainly through computational sequence analyses. To address the lack of experimental data, we studied the attempted movement of 246,045 genes from 79 prokaryotic genomes into E. coli and identified genes that consistently fail to transfer. We studied the mechanisms underlying transfer inhibition by placing coding regions from different species under the control of inducible promoters. Their toxicity to the host inhibited transfer regardless of the species of origin and our data suggest that increased gene dosage and associated increased expression is a predominant cause for transfer failure. While these experimental studies examined transfer solely into E. coli, a computational analysis of gene transfer rates across available bacterial and archaeal genomes indicates that the barriers observed in our study are general across the tree of life.

  18. The two-component system CpxR/A represses the expression of Salmonella virulence genes by affecting the stability of the transcriptional regulator HilD

    Science.gov (United States)

    De la Cruz, Miguel A.; Pérez-Morales, Deyanira; Palacios, Irene J.; Fernández-Mora, Marcos; Calva, Edmundo; Bustamante, Víctor H.

    2015-01-01

    Salmonella enterica can cause intestinal or systemic infections in humans and animals mainly by the presence of pathogenicity islands SPI-1 and SPI-2, containing 39 and 44 genes, respectively. The AraC-like regulator HilD positively controls the expression of the SPI-1 genes, as well as many other Salmonella virulence genes including those located in SPI-2. A previous report indicates that the two-component system CpxR/A regulates the SPI-1 genes: the absence of the sensor kinase CpxA, but not the absence of its cognate response regulator CpxR, reduces their expression. The presence and absence of cell envelope stress activates kinase and phosphatase activities of CpxA, respectively, which in turn controls the level of phosphorylated CpxR (CpxR-P). In this work, we further define the mechanism for the CpxR/A-mediated regulation of SPI-1 genes. The negative effect exerted by the absence of CpxA on the expression of SPI-1 genes was counteracted by the absence of CpxR or by the absence of the two enzymes, AckA and Pta, which render acetyl-phosphate that phosphorylates CpxR. Furthermore, overexpression of the lipoprotein NlpE, which activates CpxA kinase activity on CpxR, or overexpression of CpxR, repressed the expression of SPI-1 genes. Thus, our results provide several lines of evidence strongly supporting that the absence of CpxA leads to the phosphorylation of CpxR via the AckA/Pta enzymes, which represses both the SPI-1 and SPI-2 genes. Additionally, we show that in the absence of the Lon protease, which degrades HilD, the CpxR-P-mediated repression of the SPI-1 genes is mostly lost; moreover, we demonstrate that CpxR-P negatively affects the stability of HilD and thus decreases the expression of HilD-target genes, such as hilD itself and hilA, located in SPI-1. Our data further expand the insight on the different regulatory pathways for gene expression involving CpxR/A and on the complex regulatory network governing virulence in Salmonella. PMID:26300871

  19. Determinants of human adipose tissue gene expression: impact of diet, sex, metabolic status, and cis genetic regulation.

    Directory of Open Access Journals (Sweden)

    Nathalie Viguerie

    2012-09-01

    Full Text Available Weight control diets favorably affect parameters of the metabolic syndrome and delay the onset of diabetic complications. The adaptations occurring in adipose tissue (AT are likely to have a profound impact on the whole body response as AT is a key target of dietary intervention. Identification of environmental and individual factors controlling AT adaptation is therefore essential. Here, expression of 271 transcripts, selected for regulation according to obesity and weight changes, was determined in 515 individuals before, after 8-week low-calorie diet-induced weight loss, and after 26-week ad libitum weight maintenance diets. For 175 genes, opposite regulation was observed during calorie restriction and weight maintenance phases, independently of variations in body weight. Metabolism and immunity genes showed inverse profiles. During the dietary intervention, network-based analyses revealed strong interconnection between expression of genes involved in de novo lipogenesis and components of the metabolic syndrome. Sex had a marked influence on AT expression of 88 transcripts, which persisted during the entire dietary intervention and after control for fat mass. In women, the influence of body mass index on expression of a subset of genes persisted during the dietary intervention. Twenty-two genes revealed a metabolic syndrome signature common to men and women. Genetic control of AT gene expression by cis signals was observed for 46 genes. Dietary intervention, sex, and cis genetic variants independently controlled AT gene expression. These analyses help understanding the relative importance of environmental and individual factors that control the expression of human AT genes and therefore may foster strategies aimed at improving AT function in metabolic diseases.

  20. Stability-indicating method for the determination of clorazepate dipotassium-II. Via N-desmethyldiazepam and determination of its degradation products.

    Science.gov (United States)

    El-Bardicy, M G; Bebawy, L I; Amer, M M

    1992-10-01

    A spectrophotometric method for the determination of the intact clorazepate dipotassium in the presence of its degradation products is developed. It depends upon preliminary hydrolysis of clorazepate dipotassium-thus liberating its equivalent of N-desmethyldiazepam which is extracted, with benzene-methylene chloride (9:1). The extract is evaporated, the residue dissolved in methanol and its absorbance measured at about 315 nm. The procedure determines 0.4-1.6 mg of clorazepate dipotassium with an accuracy of 100.2+/-0.7%. The procedure is applied successfully for the determination of clorazepate dipotassium in bulk powder and in capsules; retaining its accuracy in the presence of up to 80% degradation. Determination of the different degradation products is also possible. Thus, N-desmethyl diazepam is determined after preliminary extraction with benzene-methylene chloride mixture, followed by TLC separation, 2-amino-5-chlorobenzophenone by directly applying the first derivative spectrophotometric technique, and glycine in the aqueous layer determined colorimetrically with ninhydrin reagent in the presence of pyridine.

  1. Comprehensive analysis of slope stability and determination of stable slopes in the Chador-Malu iron ore mine using numerical and limit equilibrium methods

    Institute of Scientific and Technical Information of China (English)

    ATAEIM; BODAGHABADIS

    2008-01-01

    One of the critical aspects in mine design is slope stability analysis and the determination of stable slopes. In the Chador Malu iron ore mine, one of the most important iron ore mines in central Iran, it was considered vital to perform a comprehensive slope stability analysis. At first, we divided the existing rock hosting pit into six zones and a geotechnical map was prepared. Then,the value of MRMR (Mining Rock Mass Rating) was determined for each zone. Owing to the fact that the Chador-Malu iron ore mine is located in a highly tectonic area and the rock mass completely crushed, the Hock-Brown failure criterion was found suitable to estimate geo-mechanical parameters. After that, the value of cohesion (c) and friction angle (e) were calculated for different geotechnical zones and relative graphs and equations were derived as a function of slope height. The stability analyses using numerical and limit equilibrium methods showed that some instability problems might occur by increasing the slope height.Therefore, stable slopes for each geotechnical zone and prepared sections were calculated and presented as a function of slope height.

  2. Gene expression profiling of genetically determined growth variation in bivalve larvae (Crassostrea gigas).

    Science.gov (United States)

    Meyer, E; Manahan, D T

    2010-03-01

    Growth rates in animals are governed by a wide range of biological factors, many of which remain poorly understood. To identify the genes that establish growth differences in bivalve larvae, we compared expression patterns in contrasting phenotypes (slow- and fast-growth) that were experimentally produced by genetic crosses of the Pacific oyster Crassostrea gigas. Based on transcriptomic profiling of 4.5 million cDNA sequence tags, we sequenced and annotated 181 cDNA clones identified by statistical analysis as candidates for differential growth. Significant matches were found in GenBank for 43% of clones (N=78), including 34 known genes. These sequences included genes involved in protein metabolism, energy metabolism and regulation of feeding activity. Ribosomal protein genes were predominant, comprising half of the 34 genes identified. Expression of ribosomal protein genes showed non-additive inheritance - i.e. expression in fast-growing hybrid larvae was different from average levels in inbred larvae from these parental families. The expression profiles of four ribosomal protein genes (RPL18, RPL31, RPL352 and RPS3) were validated by RNA blots using additional, independent crosses from the same families. Expression of RPL35 was monitored throughout early larval development, revealing that these expression patterns were established early in development (in 2-day-old larvae). Our findings (i) provide new insights into the mechanistic bases of growth and highlight genes not previously considered in growth regulation, (ii) support the general conclusion that genes involved in protein metabolism and feeding regulation are key regulators of growth, and (iii) provide a set of candidate biomarkers for predicting differential growth rates during animal development.

  3. Effective Surfactants Blend Concentration Determination for O/W Emulsion Stabilization by Two Nonionic Surfactants by Simple Linear Regression

    National Research Council Canada - National Science Library

    Hassan, A K

    2015-01-01

    ...°. Applying the simple linear regression least squares method statistical analysis to the temperature-conductivity obtained data determines the effective surfactants blend concentration required...

  4. Expectation-maximization algorithm for determining natural selection of Y-linked genes through two-sex branching processes.

    Science.gov (United States)

    González, M; Gutiérrez, C; Martínez, R

    2012-09-01

    A two-dimensional bisexual branching process has recently been presented for the analysis of the generation-to-generation evolution of the number of carriers of a Y-linked gene. In this model, preference of females for males with a specific genetic characteristic is assumed to be determined by an allele of the gene. It has been shown that the behavior of this kind of Y-linked gene is strongly related to the reproduction law of each genotype. In practice, the corresponding offspring distributions are usually unknown, and it is necessary to develop their estimation theory in order to determine the natural selection of the gene. Here we deal with the estimation problem for the offspring distribution of each genotype of a Y-linked gene when the only observable data are each generation's total numbers of males of each genotype and of females. We set out the problem in a non parametric framework and obtain the maximum likelihood estimators of the offspring distributions using an expectation-maximization algorithm. From these estimators, we also derive the estimators for the reproduction mean of each genotype and forecast the distribution of the future population sizes. Finally, we check the accuracy of the algorithm by means of a simulation study.

  5. Fluoroquinolone resistance in atypical pneumococci and oral streptococci: evidence of horizontal gene transfer of fluoroquinolone resistance determinants from Streptococcus pneumoniae.

    Science.gov (United States)

    Ip, Margaret; Chau, Shirley S L; Chi, Fang; Tang, Julian; Chan, Paul K

    2007-08-01

    Atypical strains, presumed to be pneumococcus, with ciprofloxacin MICs of > or =4.0 microg/ml and unique sequence variations within the quinolone resistance-determining regions (QRDRs) of the gyrase and topoisomerase genes in comparison with the Streptococcus pneumoniae R6 strain, were examined. These strains were reidentified using phenotypic methods, including detection of optochin susceptibility, bile solubility, and agglutination by serotype-specific antisera, and genotypic methods, including detection of pneumolysin and autolysin genes by PCR, 16S rRNA sequencing, and multilocus sequence typing (MLST). The analysis based on concatenated sequences of the six MLST loci distinguished the "atypical" strains from pneumococci, and these strains clustered closely with S. mitis. However, all these strains and five of nine strains from the viridans streptococcal group possessed one to three gyrA, gyrB, parC, and parE genes whose QRDR sequences clustered with those of S. pneumoniae, providing evidence of horizontal transfer of the QRDRs of the gyrase and topoisomerase genes from pneumococci into viridans streptococci. These genes also conferred fluoroquinolone resistance to viridans streptococci. In addition, the fluoroquinolone resistance determinants of 32 well-characterized Streptococcus mitis and Streptococcus oralis strains from bacteremic patients were also compared. These strains have unique amino acid substitutions in GyrA and ParC that were distinguishable from those in fluoroquinolone-resistant pneumococci and the "atypical" isolates. Both recombinational events and de novo mutations play an important role in the development of fluoroquinolone resistance.

  6. Sex determination in 58 bird species and evaluation of CHD gene as a universal molecular marker in bird sexing.

    Science.gov (United States)

    Vucicevic, Milos; Stevanov-Pavlovic, Marija; Stevanovic, Jevrosima; Bosnjak, Jasna; Gajic, Bojan; Aleksic, Nevenka; Stanimirovic, Zoran

    2013-01-01

    The aim of this research was to test the CHD gene (Chromo Helicase DNA-binding gene) as a universal molecular marker for sexing birds of relatively distant species. The CHD gene corresponds to the aim because of its high degree of conservation and different lengths in Z and W chromosomes due to different intron sizes. DNA was isolated from feathers and the amplification of the CHD gene was performed with the following sets of polymerase chain reaction (PCR) primers: 2550F/2718R and P2/P8. Sex determination was attempted in 284 samples of 58 bird species. It was successful in 50 bird species; in 16 of those (Alopochen aegyptiacus, Ara severus, Aratinga acuticaudata, Bucorvus leadbeateri, Cereopsis novaehollandiae, Columba arquatrix, Corvus corax, C. frugilegus, Cyanoliseus patagonus, Guttera plumifera, Lamprotornis superbus, Milvus milvus, Neophron percnopterus, Ocyphaps lophotes, Podiceps cristatus, and Poicephalus senegalus), it was carried out for the first time using molecular markers and PCR. It is reasonable to assume that extensive research is necessary to define the CHD gene as a universal molecular marker for successful sex determination in all bird species (with exception of ratites). The results of this study may largely contribute to the aim.

  7. Stability of Retroviral Vectors Against Ultracentrifugation Is Determined by the Viral Internal Core and Envelope Proteins Used for Pseudotyping.

    Science.gov (United States)

    Kim, Soo-Hyun; Lim, Kwang-Il

    2017-05-31

    Retroviral and lentiviral vectors are mostly pseudotyped and often purified and concentrated via ultracentrifugation. In this study, we quantified and compared the stabilities of retroviral [murine leukemia virus (MLV)-based] and lentiviral [human immunodeficiency virus (HIV)-1-based] vectors pseudotyped with relatively mechanically stable envelope proteins, vesicular stomatitis virus glycoproteins (VSVGs), and the influenza virus WSN strain envelope proteins against ultracentrifugation. Lentiviral genomic and functional particles were more stable than the corresponding retroviral particles against ultracentrifugation when pseudotyped with VSVGs. However, both retroviral and lentiviral particles were unstable when pseudotyped with the influenza virus WSN strain envelope proteins. Therefore, the stabilities of pseudotyped retroviral and lentiviral vectors against ultracentrifugation process are a function of not only the type of envelope proteins, but also the type of viral internal core (MLV or HIV-1 core). In addition, the fraction of functional viral particles among genomic viral particles greatly varied at times during packaging, depending on the type of envelope proteins used for pseudotyping and the viral internal core.

  8. A Stability Indicating UPLC Method for the Determination of Levofloxacin Hemihydrate in Pharmaceutical Dosage Form: Application to Pharmaceutical Analysis

    Directory of Open Access Journals (Sweden)

    Batuk Dabhi

    2013-01-01

    Full Text Available A reliable and sensitive isocratic stability indicating RP-UPLC method has been developed and validated for quantitative analysis and content uniformity study of levofloxacin hemihydrate in tablets. An isocratic method for analysis of levofloxacin hemihydrate was archived on ACQUITY UPLC BEH C18 (100*2.1 mm particle size 1.7 μ columns within shorter runtime of 4 min with a flow rate of 0.400 mL/min and using a photodiode array detector to monitor the eluate at 294 nm. The mobile phase consisted of acetonitrile-buffer (23 : 77 v/v, (buffer: 20 mM K2HPO4 + 1 mL triethylamine in 1 L water, pH=2.50 by orthophosphoric acid. Response was a liner function of drug concentration in the range of 0.5–80 μg/mL (r2=0.999 with a limit of detection and quantification of 0.1 and 0.5 μg/mL, respectively. Accuracy (recovery was between 99.77% and 101.55%. The drug was subjected to oxidation, hydrolysis, photolysis, and thermal degradation. Degradation products resulting from the stress studies did not interfere with the detection of levofloxacin hemihydrate, and the assay is stability indicating.

  9. Sex determination of ovine embryos by SRY and amelogenin (AMEL) genes using maternal circulating cell free DNA.

    Science.gov (United States)

    Saberivand, Adel; Ahsan, Sima

    2016-01-01

    Simple and precise methods for sex determination in animals are a pre-requisite for a number of applications in animal production and forensics. Some of the existing methods depend only on the detection of Y-chromosome specific sequences. However, the detection of Y and X-chromosome specific sequences is advantageous. In the present study the accuracy of sex determination by SRY (sex-determining region Y) and AMEL (Amelogenin) gene detection was assessed using a polymerase chain reaction (PCR) of DNA extracted from free fetal cells in maternal blood, which is noninvasive for fetus and easier to collect. The PCR amplification of SRY primers produced a single band of 171bp from ewes bearing a male fetus, whereas no band was amplified from the DNA extracted from ewes pregnant to a female fetus. Moreover, two bands of 182 and 242bp in male and a single band of 242 in female fetuses were produced by AMEL gene primers in the PCR reaction. Using this technique 100% of samples were successfully sexed, excluding twins. In conclusion, we demonstrated that sex determination using DNA of free fetal cells in maternal plasma is efficient using both SRY and AMEL gene sequences. It also is evident that this method is not suitable for sex determination of twin pregnancies.

  10. Determination on thermal stability of milk powder by differential scanning calorimetry%差示扫描量热法测定奶粉的热稳定性

    Institute of Scientific and Technical Information of China (English)

    陈文亮; 于鹏; 刘翠平; 高长永

    2012-01-01

    Thermal stabilities of four kinds of milk powder were determined by DSC, including whole milk powder, skim milk powder, sweeten milk powder and milk powder with vegetable oil for elder. Their activation energies were calculated by the Ozawa and Kissinger methods respectively and their thermal stabilities were compared. Moreover, the influences of components in milk powder on thermal stability were also analyzed. The results indicated that the order of thermal stability from high to low was skim milk powder, whole milk powder, milk powder with vegetable oil for elder and sweeten milk powder. And thermal stability could be reduced by milk fat, vegetable oil and sugar.%以差示扫描量热法对全脂奶粉、脱脂奶粉、全脂甜奶粉及中老年奶粉的热稳定性进行了研究,运用Ozawa和Kissinger方程求得各种奶粉的活化能,比较了这几种不同成分的奶粉的热稳定性,分析了奶粉成分对奶粉热稳定性的影响.结果表明,热稳定从高到低的顺序依次为脱脂奶粉、全脂奶粉、中老年奶粉、全脂甜奶粉,并且牛乳脂肪、植物油、蔗糖会降低奶粉的热稳定性.

  11. Analytic theory for the determination of velocity and stability of bubbles in a Hele-Shaw cell. Part 1: Velocity selection

    Science.gov (United States)

    Tanveer, Saleh

    1989-01-01

    An asymptotic theory is presented for the determination of velocity and linear stability of a steady symmetric bubble in a Hele-Shaw cell for small surface tension. In the first part, the bubble velocity U relative to the fluid velocity at infinity is determined for small surface tension T by determining transcendentally small correction to the asymptotic series solution. It is found that for any relative bubble velocity U in the interval (U(c),2), solutions exist at a countably infinite set of values of T (which has zero as its limit point) corresponding to the different branches of bubble solutions. U(c) decreases monotonically from 2 to 1 as the bubble area increases from 0 to infinity. However, for a bubble of arbitrarily given size, as T approaches 0, solution exists on any given branch with relative bubble velocity U satisfying the relation 2-U = cT to the 2/3 power, where c depends on the branch but is independent of the bubble area. The analytical evidence further suggests that there are no solutions for U greater than 2. These results are in agreement with earlier analytical results for a finger. In Part 2, an analytic theory is presented for the determination of the linear stability of the bubble in the limit of zero surface tension. Only the solution branch corresponding to the largest possible U for given surface tension is found to be stable, while all the others are unstable, in accordance with earlier numerical results.

  12. Analysis of the stability of housekeeping gene expression in the left cardiac ventricle of rats submitted to chronic intermittent hypoxia

    Directory of Open Access Journals (Sweden)

    Guilherme Silva Julian

    Full Text Available ABSTRACT Obstructive sleep apnea (OSA has been associated with oxidative stress and various cardiovascular consequences, such as increased cardiovascular disease risk. Quantitative real-time PCR is frequently employed to assess changes in gene expression in experimental models. In this study, we analyzed the effects of chronic intermittent hypoxia (an experimental model of OSA on housekeeping gene expression in the left cardiac ventricle of rats. Analyses via four different approaches-use of the geNorm, BestKeeper, and NormFinder algorithms; and 2−ΔCt (threshold cycle data analysis-produced similar results: all genes were found to be suitable for use, glyceraldehyde-3-phosphate dehydrogenase and 18S being classified as the most and the least stable, respectively. The use of more than one housekeeping gene is strongly advised.

  13. The PH gene determines fruit acidity and contributes to the evolution of sweet melons.

    Science.gov (United States)

    Cohen, Shahar; Itkin, Maxim; Yeselson, Yelena; Tzuri, Galil; Portnoy, Vitaly; Harel-Baja, Rotem; Lev, Shery; Sa'ar, Uzi; Davidovitz-Rikanati, Rachel; Baranes, Nadine; Bar, Einat; Wolf, Dalia; Petreikov, Marina; Shen, Shmuel; Ben-Dor, Shifra; Rogachev, Ilana; Aharoni, Asaph; Ast, Tslil; Schuldiner, Maya; Belausov, Eduard; Eshed, Ravit; Ophir, Ron; Sherman, Amir; Frei, Benedikt; Neuhaus, H Ekkehard; Xu, Yimin; Fei, Zhangjun; Giovannoni, Jim; Lewinsohn, Efraim; Tadmor, Yaakov; Paris, Harry S; Katzir, Nurit; Burger, Yosef; Schaffer, Arthur A

    2014-06-05

    Taste has been the subject of human selection in the evolution of agricultural crops, and acidity is one of the three major components of fleshy fruit taste, together with sugars and volatile flavour compounds. We identify a family of plant-specific genes with a major effect on fruit acidity by map-based cloning of C. melo PH gene (CmPH) from melon, Cucumis melo taking advantage of the novel natural genetic variation for both high and low fruit acidity in this species. Functional silencing of orthologous PH genes in two distantly related plant families, cucumber and tomato, produced low-acid, bland tasting fruit, showing that PH genes control fruit acidity across plant families. A four amino-acid duplication in CmPH distinguishes between primitive acidic varieties and modern dessert melons. This fortuitous mutation served as a preadaptive antecedent to the development of sweet melon cultigens in Central Asia over 1,000 years ago.

  14. Determinants of Power in Gene-Based Burden Testing for Monogenic Disorders.

    Science.gov (United States)

    Guo, Michael H; Dauber, Andrew; Lippincott, Margaret F; Chan, Yee-Ming; Salem, Rany M; Hirschhorn, Joel N

    2016-09-01

    Whole-exome sequencing has enabled new approaches for discovering genes associated with monogenic disorders. One such approach is gene-based burden testing, in which the aggregate frequency of "qualifying variants" is compared between case and control subjects for each gene. Despite substantial successes of this approach, the genetic causes for many monogenic disorders remain unknown or only partially known. It is possible that particular genetic architectures lower rates of discovery, but the influence of these factors on power has not been rigorously evaluated. Here, we leverage large-scale exome-sequencing data to create an empirically based simulation framework to evaluate the impact of key parameters (background variation rates, locus heterogeneity, mode of inheritance, penetrance) on power in gene-based burden tests in the contex