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Sample records for chemical genetic screen

  1. Chemical genetics and drug screening in Drosophila cancer models

    Institute of Scientific and Technical Information of China (English)

    Mara Gladstone; Tin Tin Su

    2011-01-01

    Drug candidates often fail in preclinical and clinical testing because of reasons of efficacy and/or safety.It would be time- and cost-efficient to have screening models that reduce the rate of such false positive candidates that appear promising at first but fail later.In this regard,it would be particularly useful to have a rapid and inexpensive whole animal model that can pre-select hits from high-throughput screens but before testing in costly rodent assays.Drosophila melanogaster has emerged as a potential whole animal model for drug screening.Of particular interest have been drugs that must act in the context of multi-cellularity such as those for neurological disorders and cancer.A recent review provides a comprehensive summary of drug screening in Drosophila,but with an emphasis on neurodegenerative disorders.Here,we review Drosophila screens in the literature aimed at cancer therapeutics.

  2. Chemical genetic screening of KRAS-based synthetic lethal inhibitors for pancreatic cancer

    OpenAIRE

    Ji, Zhenyu; Mei, Fang C; Lory, Pedro L.; Gilbertson, Scott R.; Chen, Yijun; Cheng, Xiaodong

    2009-01-01

    Pancreatic cancer is one of the deadliest diseases largely due to difficulty in early diagnosis and the lack of effective treatments. KRAS is mutated in more than 90% of pancreatic cancer patients, and oncogenic KRAS contributes to pancreatic cancer tumorigenesis and progression. In this report, using an oncogenic KRASV12-based pancreatic cancer cell model, we developed a chemical genetic screen to identify small chemical inhibitors that selectively target pancreatic cancer cells with gain-of...

  3. Screening of herbal extracts influencing hematopoiesis and their chemical genetic effects in embryonic zebrafish

    Institute of Scientific and Technical Information of China (English)

    Rajaretinam Rajesh Kannan; Samuel Gnana Prakash Vincent

    2012-01-01

    Objective: To screen the herbal extracts influencing the hematopoietic stem cells (HSC) in zebrafish embryos and their chemical genetic effects. Methods: The herbals used in this study had been widely applicable in Siddha medicines in South India. Herbal extracts were treated in zebrafish embryos at 4 d post fertilization and the extracts inducing the HSC were enumerated in hemocytometer. The biocompatibility and the organogenesis of the screened extracts were assessed in the zebrafish embryos for their chemical genetic effects. The LC50 values were calculated with their parallel control. The blood cells were enumerated. Results: The level of RBC was found increased in the Bergera koenigii (B. koenigii) at 15 μg/mL (P<0.05), Mimosa pudica (M. pudica) at 20 μg/mL (P<0.05) and Solanum trilobatum (S. trilobatum) at 25 μg/mL (P<0.05) and decreased RBC level was found in Phyllanthus niruri (P. niruri) at 30 μg/mL (P<0.05). The WBC count was found increased in S. trilobatum at 20 μg/mL (P<0.05) and Annona muricata (Annona muricata) at 15 μg/mL (P<0.05) and the Vitis quadrangularis (V. quadrangularis) at 20 μg/mL (P<0.05) decreased the WBC level. There were no notable effects in heart beats and the chemical genetic effects were observed at higher concentration of the extract resulting in Pericardial bulging, trunk tail flexure with heart edema, fin fold deformities etc. Conclusions: This in vivo based screening of Hematopoiesis is an inexpensive assay to screen herbal compounds and found that S. trilobatum extract influenced embryonic HSC in zebrafish, which could be a therapeutic for blood related disorders.

  4. Prenatal screening and genetics

    DEFF Research Database (Denmark)

    Alderson, P; Aro, A R; Dragonas, T;

    2001-01-01

    Although the term 'genetic screening' has been used for decades, this paper discusses how, in its most precise meaning, genetic screening has not yet been widely introduced. 'Prenatal screening' is often confused with 'genetic screening'. As we show, these terms have different meanings, and we...... examine definitions of the relevant concepts in order to illustrate this point. The concepts are i) prenatal, ii) genetic screening, iii) screening, scanning and testing, iv) maternal and foetal tests, v) test techniques and vi) genetic conditions. So far, prenatal screening has little connection...... with precisely defined genetics. There are benefits but also disadvantages in overstating current links between them in the term genetic screening. Policy making and professional and public understandings about screening could be clarified if the distinct meanings of prenatal screening and genetic screening were...

  5. Prenatal screening and genetics

    NARCIS (Netherlands)

    Alderson, P.; Aro, A.R.; Dragonas, T.; Ettorre, E.; Hemminki, E.; Jalinoja, P.; Santalahti, P.; Tijmstra, T.

    2001-01-01

    Although the term 'genetic screening' has been used for decades, this paper discusses how, in its most precise meaning, genetic screening has not yet been widely introduced. 'Prenatal screening' is often confused with 'genetic screening'. As we show, these terms have different meanings, and we exami

  6. Chemical Screening in Zebrafish.

    Science.gov (United States)

    Brady, Colleen A; Rennekamp, Andrew J; Peterson, Randall T

    2016-01-01

    Phenotypic small molecule screens in zebrafish have gained popularity as an unbiased approach to probe biological processes. In this chapter we outline basic methods for performing chemical screens with larval zebrafish including breeding large numbers of embryos, plating larval fish into multi-well dishes, and adding small molecules to these wells. We also highlight important considerations when designing and interpreting the results of a phenotypic screen and possible follow-up approaches, including popular methods used to identify the mechanism of action of a chemical compound. PMID:27464797

  7. The genetic origins of biosynthesis and light-responsive control of the chemical UV screen of land plants

    International Nuclear Information System (INIS)

    Most land plants possess the capacity to protect themselves from UV light, and do so by producing pigments that absorb efficiently in the UV-A and UV-B regions of the spectrum while allowing transmission of nearly all photosynthetically useful wavelengths. These UV-absorbing pigments are mainly phenylpropanoids and flavonoids. This chapter summarizes current understanding of the mechanism of UV protection in higher land plants, evaluates the information available from lower land plants and their green-algal relatives, and then considers the possible evolutionary origins of this use of chemical filters for selectively screening UV light from solar radiation. It is proposed that photo control over the biosynthesis of UV-absorbing phenylpropanoids and flavonoids may have evolved in concert with the evolution of the high biosynthetic activity necessary for UV protection. The toxicity of phenylpropanoids and flavonoids has been postulated to have been a barrier to the evolution of an effective chemical UV screen, and that some means for sequestering these compounds and/or for controlling their synthesis probably evolved prior to, or in concert with, the evolution of high rates of biosynthesis. The original photoreceptor and signal transduction system is speculated to have been based on photo isomerization of a phenylpropanoid ester and a pre-existing product feedback mechanism for controlling phenylpropanoid biosynthesis. Understanding the original mechanism for photo control of the chemical UV screen of land plants could be valuable for understanding the adaptability of extant land plants to rising levels of solar UV-B radiation and may suggest genetic strategies for engineering improved UV tolerance in crop plants. (author)

  8. NeuroChip: a microfluidic electrophysiological device for genetic and chemical biology screening of Caenorhabditis elegans adult and larvae.

    Directory of Open Access Journals (Sweden)

    Chunxiao Hu

    Full Text Available Genetic and chemical biology screens of C. elegans have been of enormous benefit in providing fundamental insight into neural function and neuroactive drugs. Recently the exploitation of microfluidic devices has added greater power to this experimental approach providing more discrete and higher throughput phenotypic analysis of neural systems. Here we make a significant addition to this repertoire through the design of a semi-automated microfluidic device, NeuroChip, which has been optimised for selecting worms based on the electrophysiological features of the pharyngeal neural network. We demonstrate this device has the capability to sort mutant from wild-type worms based on high definition extracellular electrophysiological recordings. NeuroChip resolves discrete differences in excitatory, inhibitory and neuromodulatory components of the neural network from individual animals. Worms may be fed into the device consecutively from a reservoir and recovered unharmed. It combines microfluidics with integrated electrode recording for sequential trapping, restraining, recording, releasing and recovering of C. elegans. Thus mutant worms may be selected, recovered and propagated enabling mutagenesis screens based on an electrophysiological phenotype. Drugs may be rapidly applied during the recording thus permitting compound screening. For toxicology, this analysis can provide a precise description of sub-lethal effects on neural function. The chamber has been modified to accommodate L2 larval stages showing applicability for small size nematodes including parasitic species which otherwise are not tractable to this experimental approach. We also combine NeuroChip with optogenetics for targeted interrogation of the function of the neural circuit. NeuroChip thus adds a new tool for exploitation of C. elegans and has applications in neurogenetics, drug discovery and neurotoxicology.

  9. Characterization of pellicle inhibition in Gluconacetobacter xylinus 53582 by a small molecule, pellicin, identified by a chemical genetics screen.

    Directory of Open Access Journals (Sweden)

    Janice L Strap

    Full Text Available Pellicin ([2E]-3-phenyl-1-[2,3,4,5-tetrahydro-1,6-benzodioxocin-8-yl]prop-2-en-1-one was identified in a chemical genetics screen of 10,000 small molecules for its ability to completely abolish pellicle production in Gluconacetobacter xylinus. Cells grown in the presence of pellicin grew 1.5 times faster than untreated cells. Interestingly, growth in pellicin also caused G. xylinus cells to elongate. Measurement of cellulose synthesis in vitro showed that cellulose synthase activity was not directly inhibited by pellicin. Rather, when cellulose synthase activity was measured in cells that were pre-treated with the compound, the rate of cellulose synthesis increased eight-fold over that observed for untreated cells. This phenomenon was also apparent in the rapid production of cellulose when cells grown in the presence of pellicin were washed and transferred to media lacking the inhibitor. The rate at which cellulose was produced could not be accounted for by growth of the organism. Pellicin was not detected when intracellular contents were analyzed. Furthermore, it was found that pellicin exerts its effect extracellularly by interfering with the crystallization of pre-cellulosic tactoidal aggregates. This interference of the crystallization process resulted in enhanced production of cellulose II as evidenced by the ratio of acid insoluble to acid soluble product in in vitro assays and confirmed in vivo by scanning electron microscopy and powder X-ray diffraction. The relative crystallinity index, RCI, of pellicle produced by untreated G. xylinus cultures was 70% while pellicin-grown cultures had RCI of 38%. Mercerized pellicle of untreated cells had RCI of 42%, which further confirms the mechanism of action of pellicin as an inhibitor of the cellulose I crystallization process. Pellicin is a useful tool for the study of cellulose biosynthesis in G. xylinus.

  10. Optimal screening for genetic diseases.

    Science.gov (United States)

    Nævdal, Eric

    2014-12-01

    Screening for genetic diseases is performed in many regions and/or ethnic groups where there is a high prevalence of possibly malign genes. The propagation of such genes can be considered a dynamic externality. Given that many of these diseases are untreatable and give rise to truly tragic outcomes, they are a source of societal concern, and the screening process should perhaps be regulated. This paper incorporates a standard model of genetic propagation into an economic model of dynamic management to derive cost benefit rules for optimal screening. The highly non-linear nature of genetic dynamics gives rise to perhaps surprising results that include discontinuous controls and threshold effects. One insight is that any screening program that is in place for any amount of time should screen all individuals in a target population. The incorporation of genetic models may prove to be useful to several emerging fields in economics such as genoeconomics, neuroeconomics and paleoeconomics.

  11. Optimal screening for genetic diseases.

    Science.gov (United States)

    Nævdal, Eric

    2014-12-01

    Screening for genetic diseases is performed in many regions and/or ethnic groups where there is a high prevalence of possibly malign genes. The propagation of such genes can be considered a dynamic externality. Given that many of these diseases are untreatable and give rise to truly tragic outcomes, they are a source of societal concern, and the screening process should perhaps be regulated. This paper incorporates a standard model of genetic propagation into an economic model of dynamic management to derive cost benefit rules for optimal screening. The highly non-linear nature of genetic dynamics gives rise to perhaps surprising results that include discontinuous controls and threshold effects. One insight is that any screening program that is in place for any amount of time should screen all individuals in a target population. The incorporation of genetic models may prove to be useful to several emerging fields in economics such as genoeconomics, neuroeconomics and paleoeconomics. PMID:25203815

  12. Chemical genetics and regeneration.

    Science.gov (United States)

    Sengupta, Sumitra; Zhang, Liyun; Mumm, Jeff S

    2015-01-01

    Regeneration involves interactions between multiple signaling pathways acting in a spatially and temporally complex manner. As signaling pathways are highly conserved, understanding how regeneration is controlled in animal models exhibiting robust regenerative capacities should aid efforts to stimulate repair in humans. One way to discover molecular regulators of regeneration is to alter gene/protein function and quantify effect(s) on the regenerative process: dedifferentiation/reprograming, stem/progenitor proliferation, migration/remodeling, progenitor cell differentiation and resolution. A powerful approach for applying this strategy to regenerative biology is chemical genetics, the use of small-molecule modulators of specific targets or signaling pathways. Here, we review advances that have been made using chemical genetics for hypothesis-focused and discovery-driven studies aimed at furthering understanding of how regeneration is controlled.

  13. Chemical compatibility screening test results

    Energy Technology Data Exchange (ETDEWEB)

    Nigrey, P.J.; Dickens, T.G.

    1997-12-01

    A program for evaluating packaging components that may be used in transporting mixed-waste forms has been developed and the first phase has been completed. This effort involved the screening of ten plastic materials in four simulant mixed-waste types. These plastics were butadiene-acrylonitrile copolymer rubber, cross-linked polyethylene (XLPE), epichlorohydrin rubber, ethylene-propylene rubber (EPDM), fluorocarbon (Viton or Kel-F), polytetrafluoroethylene, high-density polyethylene (HDPE), isobutylene-isoprene copolymer rubber (butyl), polypropylene, and styrene-butadiene rubber (SBR). The selected simulant mixed wastes were (1) an aqueous alkaline mixture of sodium nitrate and sodium nitrite; (2) a chlorinated hydrocarbon mixture; (3) a simulant liquid scintillation fluid; and (4) a mixture of ketones. The testing protocol involved exposing the respective materials to 286,000 rads of gamma radiation followed by 14-day exposures to the waste types at 60{degrees}C. The seal materials were tested using vapor transport rate (VTR) measurements while the liner materials were tested using specific gravity as a metric. For these tests, a screening criterion of 0.9 g/hr/m{sup 2} for VTR and a specific gravity change of 10% was used. Based on this work, it was concluded that while all seal materials passed exposure to the aqueous simulant mixed waste, EPDM and SBR had the lowest VTRs. In the chlorinated hydrocarbon simulant mixed waste, only Viton passed the screening tests. In both the simulant scintillation fluid mixed waste and the ketone mixture simulant mixed waste, none of the seal materials met the screening criteria. For specific gravity testing of liner materials, the data showed that while all materials with the exception of polypropylene passed the screening criteria, Kel-F, HDPE, and XLPE offered the greatest resistance to the combination of radiation and chemicals.

  14. Chemical compatibility screening test results

    International Nuclear Information System (INIS)

    A program for evaluating packaging components that may be used in transporting mixed-waste forms has been developed and the first phase has been completed. This effort involved the screening of ten plastic materials in four simulant mixed-waste types. These plastics were butadiene-acrylonitrile copolymer rubber, cross-linked polyethylene (XLPE), epichlorohydrin rubber, ethylene-propylene rubber (EPDM), fluorocarbon (Viton or Kel-F), polytetrafluoroethylene, high-density polyethylene (HDPE), isobutylene-isoprene copolymer rubber (butyl), polypropylene, and styrene-butadiene rubber (SBR). The selected simulant mixed wastes were (1) an aqueous alkaline mixture of sodium nitrate and sodium nitrite; (2) a chlorinated hydrocarbon mixture; (3) a simulant liquid scintillation fluid; and (4) a mixture of ketones. The testing protocol involved exposing the respective materials to 286,000 rads of gamma radiation followed by 14-day exposures to the waste types at 60 degrees C. The seal materials were tested using vapor transport rate (VTR) measurements while the liner materials were tested using specific gravity as a metric. For these tests, a screening criterion of 0.9 g/hr/m2 for VTR and a specific gravity change of 10% was used. Based on this work, it was concluded that while all seal materials passed exposure to the aqueous simulant mixed waste, EPDM and SBR had the lowest VTRs. In the chlorinated hydrocarbon simulant mixed waste, only Viton passed the screening tests. In both the simulant scintillation fluid mixed waste and the ketone mixture simulant mixed waste, none of the seal materials met the screening criteria. For specific gravity testing of liner materials, the data showed that while all materials with the exception of polypropylene passed the screening criteria, Kel-F, HDPE, and XLPE offered the greatest resistance to the combination of radiation and chemicals

  15. Facile high-throughput forward chemical genetic screening by in situ monitoring of glucuronidase-based reporter gene expression in Arabidopsis thaliana

    Directory of Open Access Journals (Sweden)

    Vivek eHalder

    2015-01-01

    Full Text Available The use of biologically active small molecules to perturb biological functions holds enormous potential for investigating complex signaling networks. However, in contrast to animal systems, the search for and application of chemical tools for basic discovery in the plant sciences, generally referred to as ‘chemical genetics’, has only recently gained momentum. In addition to cultured cells, the well-characterized, small-sized model plant Arabidopsis thaliana is suitable for cultivation in microplates, which allows employing diverse cell- or phenotype-based chemical screens. In such screens, a chemical’s bioactivity is typically assessed either through scoring its impact on morphological traits or quantifying molecular attributes such as enzyme or reporter activities. Here, we describe a facile forward chemical screening methodology for intact Arabidopsis seedlings harboring the β-glucuronidase (GUS reporter by directly quantifying GUS activity in situ with 4-methylumbelliferyl-β-D-glucuronide (4-MUG as substrate. The quantitative nature of this screening assay has an obvious advantage over the also convenient histochemical GUS staining method, as it allows application of statistical procedures and unbiased hit selection based on threshold values as well as distinction between compounds with strong or weak bioactivity. At the same time, the in situ bioassay is very convenient requiring less effort and time for sample handling in comparison to the conventional quantitative in vitro GUS assay using 4-MUG, as validated with several Arabidopsis lines harboring different GUS reporter constructs. To demonstrate that the developed assays is particularly suitable for large-scale screening projects, we performed a pilot screen for chemical activators or inhibitors of salicylic acid-mediated defense signaling using the Arabidopsis PR1p::GUS line. Importantly, the screening methodology provided here can be adopted for any inducible GUS reporter line.

  16. Chemical Genetics: Budding Yeast as a Platform for Drug Discovery and Mapping of Genetic Pathways

    Directory of Open Access Journals (Sweden)

    Jorrit M. Enserink

    2012-08-01

    Full Text Available The budding yeast Saccharomyces cerevisiae is a widely used model organism, and yeast genetic methods are powerful tools for discovery of novel functions of genes. Recent advancements in chemical-genetics and chemical-genomics have opened new avenues for development of clinically relevant drug treatments. Systematic mapping of genetic networks by high-throughput chemical-genetic screens have given extensive insight in connections between genetic pathways. Here, I review some of the recent developments in chemical-genetic techniques in budding yeast.

  17. Population screening for genetic susceptibility to disease.

    OpenAIRE

    Clarke, A.

    1995-01-01

    Genetic screening for susceptibility to common diseases, such as the common cancers, cardiovascular disease, and diabetes, may soon be technically feasible. Commercial interests should not be allowed to introduce such screening before proper evaluation or without adequate counselling and support. The evaluation of such testing should include psychosocial and medical outcomes and outcomes for those given low risks as well as high risks. These tests may distract attention away from environmenta...

  18. Screening for fetal and genetic abnormalities.

    Science.gov (United States)

    Simpson, J L

    1991-09-01

    Screening for genetic abnormalities is an integral part of obstetrics. Prior to initiating screening, however, several prerequisites must be met: (i) capacity to alter clinical management, (ii) cost effectiveness, (iii) reliable means (usually assays) of assessment, and (iv) capacity to handle problems. In all pregnancies one should determine in systematic fashion whether family history places a pregnant woman at increased risk over the background risk of 2-3% congenital anomalies. All women over age 35 years at delivery should be offered prenatal cytogenetic testing, and women of all ages should be offered maternal serum alpha-fetoprotein screening for neural tube defects. Screening ostensibly normal populations is appropriate in certain ethnic groups to determine heterozygosity for selected disorders: Blacks for sickle-cell anaemia, Mediterranean people for beta-thalassaemia, Southeast Asians and Filipinos for alpha-thalassaemia, Ashkenazi Jews and perhaps French-Canadians for Tay-Sachs disease. Cystic fibrosis screening (delta F508 mutations) is not currently recommended for the general populations, but should be offered to relatives of an individual having delta F508 cystic fibrosis. Irrespective of the extent of screening programmes for Mendelian traits, the mutant allele will remain in the general population because by far the greatest genetic load lies in clinically normal heterozygotes, affected contributing far less to the load despite the obvious clinical effect. PMID:1720071

  19. Genetic screens in Caenorhabditis elegans models for neurodegenerative diseases

    NARCIS (Netherlands)

    Alvarenga Fernandes Sin, Olga; Michels, Helen; Nollen, Ellen A. A.

    2014-01-01

    Caenorhabditis elegans comprises unique features that make it an attractive model organism in diverse fields of biology. Genetic screens are powerful to identify genes and C. elegans can be customized to forward or reverse genetic screens and to establish gene function. These genetic screens can be

  20. Sources of Error in Mammalian Genetic Screens.

    Science.gov (United States)

    Sack, Laura Magill; Davoli, Teresa; Xu, Qikai; Li, Mamie Z; Elledge, Stephen J

    2016-01-01

    Genetic screens are invaluable tools for dissection of biological phenomena. Optimization of such screens to enhance discovery of candidate genes and minimize false positives is thus a critical aim. Here, we report several sources of error common to pooled genetic screening techniques used in mammalian cell culture systems, and demonstrate methods to eliminate these errors. We find that reverse transcriptase-mediated recombination during retroviral replication can lead to uncoupling of molecular tags, such as DNA barcodes (BCs), from their associated library elements, leading to chimeric proviral genomes in which BCs are paired to incorrect ORFs, shRNAs, etc This effect depends on the length of homologous sequence between unique elements, and can be minimized with careful vector design. Furthermore, we report that residual plasmid DNA from viral packaging procedures can contaminate transduced cells. These plasmids serve as additional copies of the PCR template during library amplification, resulting in substantial inaccuracies in measurement of initial reference populations for screen normalization. The overabundance of template in some samples causes an imbalance between PCR cycles of contaminated and uncontaminated samples, which results in a systematic artifactual depletion of GC-rich library elements. Elimination of contaminating plasmid DNA using the bacterial endonuclease Benzonase can restore faithful measurements of template abundance and minimize GC bias. PMID:27402361

  1. Genetic Screening for Familial Gastric Cancer

    Directory of Open Access Journals (Sweden)

    Oliveira Carla

    2004-05-01

    Full Text Available Abstract Approximately 10% of gastric cancer cases show familial clustering but only 1-3% of gastric carcinomas arise as a result of inherited gastric cancer predisposition syndromes. Direct proof that Hereditary Gastric Cancer a genetic disease with a germline gene defect has come from the demonstration of co-segregation of germline E-cadherin (CDH1 mutations with early onset diffuse gastric cancer in families with an autosomal dominant pattern of inheritance (HDGC. E-cadherin is a transmembrane calcium-dependent cell-adhesion molecule involved in cell-junction formation and the maintenance of epithelial integrity. In this review, we describe frequency and type of CDH1 mutations in sporadic and familial gastric cancer. Further we demonstrate the functional significance of some CDH1 germline missense mutations found in HDGC. We also discuss the CDH1 polymorphisms that have been associated to gastric cancer. We report other types of malignancies associated to HDGC, besides diffuse gastric cancer. Moreover, we review the data available on putative alternative candidate genes screened in familial gastric cancer. Finally, we briefly discuss the role of low-penetrance genes and Helicobacter pylori in gastric cancer. This knowledge is a fundamental step towards accurate genetic counselling, in which a highly specialised pre-symptomatic therapeutic intervention should be offered.

  2. The art and design of genetic screens: maize

    Science.gov (United States)

    Maize (Zea mays) is an excellent model for basic research. Genetic screens have informed our understanding of developmental processes, meiosis, epigenetics and biochemical pathways--not only in maize but also in other cereal crops. We discuss the forward and reverse genetic screens that are possible...

  3. Antenatal screening and the gendering of genetic responsibility

    OpenAIRE

    Reed Kate

    2007-01-01

    Abstract Background The objective of this study is to explore men's and women's perceptions of antenatal blood screening. The study will assess the impact of these perceptions on decision-making regarding diagnostic testing and selective abortion, and on parental feelings of genetic responsibility. By exploring gender and antenatal screening in this way, the research aims to contribute to our understanding of lay perceptions of genetic screening and increase our knowledge of the decision-maki...

  4. Comparative chemical screening and genetic analysis reveal tentoxin as a new virulence factor in Cochliobolus miyabeanus, the causal agent of brown spot disease on rice.

    Science.gov (United States)

    De Bruyne, Lieselotte; Van Poucke, Christof; Di Mavungu, Diana Jose; Zainudin, Nur Ain Izzati Mohd; Vanhaecke, Lynn; De Vleesschauwer, David; Turgeon, B Gillian; De Saeger, Sarah; Höfte, Monica

    2016-08-01

    Brown spot disease, caused by Cochliobolus miyabeanus, is currently considered to be one of the most important yield reducers of rice (Oryza sativa L.). Despite its agricultural importance, little is known about the virulence mechanisms deployed by the fungus. Therefore, we set out to identify novel virulence factors with a role in disease development. This article reports, for the first time, the production of tentoxin by C. miyabeanus as a virulence factor during brown spot disease and the identification of the non-ribosomal protein synthetase (NRPS) CmNps3, responsible for tentoxin biosynthesis. We compared the chemical compounds produced by C. miyabeanus strains differing in virulence ability using ultra-high-performance liquid chromatography (UHPLC) coupled to high-resolution Orbitrap mass spectrometry (HRMS). The production of tentoxin by a highly virulent strain was revealed by principal component analysis of the detected ions and confirmed by UHPLC coupled to tandem-quadrupole mass spectrometry (MS/MS). The corresponding NRPS was identified by in silico genome analysis and confirmed by gene deletion. Infection tests with wild-type and Cmnps3 mutants showed that tentoxin acts as a virulence factor and is correlated with chlorosis development during the second phase of infection. Although rice has previously been classified as a tentoxin-insensitive plant species, our data demonstrate that tentoxin production by C. miyabeanus affects symptom development. PMID:26456797

  5. Genetic Recombination as a Chemical Reaction Network

    OpenAIRE

    Müller, Stefan; Hofbauer, Josef

    2015-01-01

    The process of genetic recombination can be seen as a chemical reaction network with mass-action kinetics. We review the known results on existence, uniqueness, and global stability of an equilibrium in every compatibility class and for all rate constants, from both the population genetics and the reaction networks point of view.

  6. Antenatal screening and the gendering of genetic responsibility

    Directory of Open Access Journals (Sweden)

    Reed Kate

    2007-09-01

    Full Text Available Abstract Background The objective of this study is to explore men's and women's perceptions of antenatal blood screening. The study will assess the impact of these perceptions on decision-making regarding diagnostic testing and selective abortion, and on parental feelings of genetic responsibility. By exploring gender and antenatal screening in this way, the research aims to contribute to our understanding of lay perceptions of genetic screening and increase our knowledge of the decision-making process in screening. Research design This qualitative study will be based on semi-structured interviews with twenty pregnant women and twenty male partners in the post-industrial city of Sheffield, UK. All interviews will be taped, transcribed and analysed thematically using NVIVO, a qualitative software package. Discussion The findings of this study have relevance to existing debates on the social and ethical implications of reproductive genetics. A better understanding of male and female perceptions of the screening process could improve guidance and practice in antenatal screening and genetic counselling. It will also inform and contribute to the development of theory on gender and genetic screening.

  7. Emergency preparedness for newborn screening and genetic services.

    Science.gov (United States)

    Pass, Kenneth A; Thoene, Jess; Watson, Michael S

    2009-06-01

    Patients identified in newborn screening programs can be among the most vulnerable during a disaster due to their need to have prompt diagnosis and medical management. Recent disasters have challenged the ability of newborn screening programs to maintain the needed continuity during emergency situations. This has significant implications for the newborn screening laboratories, the diagnostic confirmation providers, and the patients who either require diagnosis or maintenance of their therapeutic interventions. In 2007, the National Coordinating Center (NCC) for the Regional Genetics and Newborn Screening Collaboratives (RCs) sponsored a meeting involving representatives of the Regional Genetics and Newborn Screening Collaborative Groups, state newborn screening programs, providers of diagnosis and confirmation services, manufacturers of equipment, medical foods, and other treatments used in patients identified in newborn screening programs, and individuals from agencies involved in disaster response including the National Disaster Medical Service, the Centers for Disease Control and Prevention, the Emergency Management Assistance Compact, the Federal Emergency Management Agency, and others. In addition to developing contingency plans for newborn screening, we have considered other uses of genetics as it is used in DNA-based kinship identification of mass casualties. The meeting resulted in the description of a wide range of issues facing newborn screening programs, provider groups, and patients for which emergency preparedness development is needed in order that appropriate response is enabled. PMID:19444127

  8. Private and public eugenics: genetic and screening in India

    NARCIS (Netherlands)

    Gupta, J.A.

    2007-01-01

    Epidemiologists and geneticists claim that genetics has an increasing role to play in public health policies and programs in the future. Within this perspective, genetic testing and screening are instrumental in avoiding the birth of children with serious, costly or untreatable disorders. This paper

  9. Genetic dissection of mammalian ERAD through comparative haploid and CRISPR forward genetic screens

    DEFF Research Database (Denmark)

    Timms, Richard T.; Menzies, Sam A.; Tchasovnikarova, Iva A.;

    2016-01-01

    The application of forward genetic screens to cultured human cells represents a powerful method to study gene function. The repurposing of the bacterial CRISPR/Cas9 system provides an effective method to disrupt gene function in mammalian cells, and has been applied to genome-wide screens. Here, we...... compare the efficacy of genome-wide CRISPR/Cas9-mediated forward genetic screens versus gene-trap mutagenesis screens in haploid human cells, which represent the existing ‘gold standard’ method. This head-to-head comparison aimed to identify genes required for the endoplasmic reticulum....../3-associated disulphide reductase. Genome-wide CRISPR/Cas9-mediated screens together with haploid genetic screens provide a powerful addition to the forward genetic toolbox....

  10. 75 FR 70248 - Endocrine Disruptor Screening Program; Second List of Chemicals for Tier 1 Screening

    Science.gov (United States)

    2010-11-17

    ... more specific chemical effects on the endocrine system, and are currently in the process of being... substance interferes with the endocrine systems of humans or other species simply because it has been listed... AGENCY Endocrine Disruptor Screening Program; Second List of Chemicals for Tier 1 Screening...

  11. Screening Jews and genes: a consideration of the ethics of genetic screening within the Jewish community: challenges and responses.

    Science.gov (United States)

    Levin, M

    1999-01-01

    Screening for genetic disorders, particularly Tay-Sachs Disease, has been traditionally welcome by the Jewish community. I review the history of genetic screening among Jews and the views from the Jewish tradition on the subject, and then discuss ethical challenges of screening and the impact of historical memories upon future acceptance of screening programs. Some rational principles to guide future design of genetic screening programs among Jews are proposed. PMID:10464669

  12. Big screens with small RNAs : loss of function genetic screens to identify novel cancer genes

    NARCIS (Netherlands)

    Mullenders, J.

    2009-01-01

    This thesis described the construction and screening of one of the first large scale RNAi libraries for use in human cells. Functional genetic screens with this library have led to the identification of novel cancer genes. These cancer genes function in several pathways including the p53 tumor suppr

  13. Recent advances in prenatal screening and diagnosis of genetic disorders.

    Science.gov (United States)

    Bozzette, Maryann

    2002-11-01

    In any pregnancy, there is an approximate 3% to 5% chance that a fetal complication will occur. The most familiar prenatal diagnostics cannot be performed until the fetus is well into gestation, and most involve invasive procedures along with their inherent risks. In light of these facts, many noninvasive prenatal screening and diagnostic tests have been developed, the newest using recombinant deoxyribonucleic acid (DNA) technology in the examination of fetal cells. Through these procedures, genetic coding errors and chromosomal disruptions may be detected. This article discusses the currently available prenatal and screening diagnostic tests for genetic disorders with a focus on the latest technology. PMID:12473913

  14. Systematic Mapping of Chemical-Genetic Interactions in Saccharomyces cerevisiae.

    Science.gov (United States)

    Suresh, Sundari; Schlecht, Ulrich; Xu, Weihong; Bray, Walter; Miranda, Molly; Davis, Ronald W; Nislow, Corey; Giaever, Guri; Lokey, R Scott; St Onge, Robert P

    2016-01-01

    Chemical-genetic interactions (CGIs) describe a phenomenon where the effects of a chemical compound (i.e., a small molecule) on cell growth are dependent on a particular gene. CGIs can reveal important functional information about genes and can also be powerful indicators of a compound's mechanism of action. Mapping CGIs can lead to the discovery of new chemical probes, which, in contrast to genetic perturbations, operate at the level of the gene product (or pathway) and can be fast-acting, tunable, and reversible. The simple culture conditions required for yeast and its rapid growth, as well as the availability of a complete set of barcoded gene deletion strains, facilitate systematic mapping of CGIs in this organism. This process involves two basic steps: first, screening chemical libraries to identify bioactive compounds affecting growth and, second, measuring the effects of these compounds on genome-wide collections of mutant strains. Here, we introduce protocols for both steps that have great potential for the discovery and development of new small-molecule tools and medicines. PMID:27587783

  15. Genetic dissection of mammalian ERAD through comparative haploid and CRISPR forward genetic screens

    Science.gov (United States)

    Timms, Richard T.; Menzies, Sam A.; Tchasovnikarova, Iva A.; Christensen, Lea C.; Williamson, James C.; Antrobus, Robin; Dougan, Gordon; Ellgaard, Lars; Lehner, Paul J.

    2016-01-01

    The application of forward genetic screens to cultured human cells represents a powerful method to study gene function. The repurposing of the bacterial CRISPR/Cas9 system provides an effective method to disrupt gene function in mammalian cells, and has been applied to genome-wide screens. Here, we compare the efficacy of genome-wide CRISPR/Cas9-mediated forward genetic screens versus gene-trap mutagenesis screens in haploid human cells, which represent the existing ‘gold standard' method. This head-to-head comparison aimed to identify genes required for the endoplasmic reticulum-associated degradation (ERAD) of MHC class I molecules. The two approaches show high concordance (>70%), successfully identifying the majority of the known components of the canonical glycoprotein ERAD pathway. Both screens also identify a role for the uncharacterized gene TXNDC11, which we show encodes an EDEM2/3-associated disulphide reductase. Genome-wide CRISPR/Cas9-mediated screens together with haploid genetic screens provide a powerful addition to the forward genetic toolbox. PMID:27283361

  16. Combination of hearing screening and genetic screening for deafness-susceptibility genes in newborns.

    Science.gov (United States)

    Yao, Gen-Dong; Li, Shou-Xia; Chen, Ding-Li; Feng, Hai-Qin; Zhao, Su-Bin; Liu, Yong-Jie; Guo, Li-Li; Yang, Zhi-Ming; Zhang, Xiao-Fang; Sun, Cai-Xia; Wang, Ze-Hui; Zhang, Wei-Yong

    2014-01-01

    The aim of this study was to determine the clinical significance of the results of screening of newborn hearing and the incidence of deafness-susceptibility genes. One thousand newborn babies in the Handan Center Hospital (Handan, China) underwent screening of hearing and deafness-susceptibility genes. The first screening test was carried out using otoacoustic emissions (OAEs). Babies with hearing loss who failed to pass the initial screening were scheduled for rescreening at 42 days after birth. Cord blood was used for the screening of deafness-susceptibility genes, namely the GJB2, SLC26A4 and mitochondrial 12S rRNA (MTRNR1) genes. Among the 1,000 neonates that underwent the first hearing screening, 25 exhibited left-sided hearing loss, 21 exhibited right-sided hearing loss and 15 cases had binaural hearing loss. After rescreening 42 days later, only one of the initial 61 cases exhibited hearing loss under OAE testing. The neonatal deafness gene tests showed two cases with 1555A>G mutation and two cases with 1494C>T mutation of the MTRNR1 gene. In the SLC26A4 gene screening, four cases exhibited the heterozygous IVS7-2A>G mutation and one case exhibited heterozygous 1226G>A mutation. In the GJB2 gene screening, two cases exhibited the homozygous 427C>T mutation and 10 exhibited the heterozygous 235delC mutation. The genetic screening revealed 21 newborns with mutations in the three deafness-susceptibility genes. The overall carrier rate was 2.1% (21/1,000). The association of hearing and gene screening may be the promising screening strategy for the diagnosis of hearing loss.

  17. A comprehensive platform for highly multiplexed mammalian functional genetic screens

    Directory of Open Access Journals (Sweden)

    Cheung-Ong Kahlin

    2011-05-01

    Full Text Available Abstract Background Genome-wide screening in human and mouse cells using RNA interference and open reading frame over-expression libraries is rapidly becoming a viable experimental approach for many research labs. There are a variety of gene expression modulation libraries commercially available, however, detailed and validated protocols as well as the reagents necessary for deconvolving genome-scale gene screens using these libraries are lacking. As a solution, we designed a comprehensive platform for highly multiplexed functional genetic screens in human, mouse and yeast cells using popular, commercially available gene modulation libraries. The Gene Modulation Array Platform (GMAP is a single microarray-based detection solution for deconvolution of loss and gain-of-function pooled screens. Results Experiments with specially constructed lentiviral-based plasmid pools containing ~78,000 shRNAs demonstrated that the GMAP is capable of deconvolving genome-wide shRNA "dropout" screens. Further experiments with a larger, ~90,000 shRNA pool demonstrate that equivalent results are obtained from plasmid pools and from genomic DNA derived from lentivirus infected cells. Parallel testing of large shRNA pools using GMAP and next-generation sequencing methods revealed that the two methods provide valid and complementary approaches to deconvolution of genome-wide shRNA screens. Additional experiments demonstrated that GMAP is equivalent to similar microarray-based products when used for deconvolution of open reading frame over-expression screens. Conclusion Herein, we demonstrate four major applications for the GMAP resource, including deconvolution of pooled RNAi screens in cells with at least 90,000 distinct shRNAs. We also provide detailed methodologies for pooled shRNA screen readout using GMAP and compare next-generation sequencing to GMAP (i.e. microarray based deconvolution methods.

  18. Innovation in academic chemical screening: Filling the gaps in chemical biology

    OpenAIRE

    Hasson, Samuel A.; Inglese, James

    2013-01-01

    Academic screening centers across the world have endeavored to discover small molecules that can modulate biological systems. To increase the reach of functional-genomic and chemical screening programs, universities, research institutes, and governments have followed their industrial counterparts in adopting high-throughput paradigms. As academic screening efforts have steadily grown in scope and complexity, so have the ideas of what is possible with the union of technology and biology. This ...

  19. Application of Plagiarism Screening Software in the Chemical Engineering Curriculum

    Science.gov (United States)

    Cooper, Matthew E.; Bullard, Lisa G.

    2014-01-01

    Plagiarism is an area of increasing concern for written ChE assignments, such as laboratory and design reports, due to ease of access to text and other materials via the internet. This study examines the application of plagiarism screening software to four courses in a university chemical engineering curriculum. The effectiveness of plagiarism…

  20. A Chemical Genetic Approach To The Study Of Cellular Transport

    NARCIS (Netherlands)

    Nieland, T.J.F.

    2005-01-01

    The focus of this thesis is the use of chemical genetics to study two different aspects of membrane biology, (a) the mechanisms underlying cellular lipid transport and (b) the intersection between endocytic and exocytic traffic. The broad goals of chemical genetics are to find novel chemical tool

  1. Innovation in academic chemical screening: filling the gaps in chemical biology.

    Science.gov (United States)

    Hasson, Samuel A; Inglese, James

    2013-06-01

    Academic screening centers across the world have endeavored to discover small molecules that can modulate biological systems. To increase the reach of functional-genomic and chemical screening programs, universities, research institutes, and governments have followed their industrial counterparts in adopting high-throughput paradigms. As academic screening efforts have steadily grown in scope and complexity, so have the ideas of what is possible with the union of technology and biology. This review addresses the recent conceptual and technological innovation that has been propelling academic screening into its own unique niche. In particular, high-content and whole-organism screening are changing how academics search for novel bioactive compounds. Importantly, we recognize examples of successful chemical probe development that have punctuated the changing technology landscape. PMID:23683346

  2. A Chemical-Genomic Screen of Neglected Antibiotics Reveals Illicit Transport of Kasugamycin and Blasticidin S.

    Science.gov (United States)

    Shiver, Anthony L; Osadnik, Hendrik; Kritikos, George; Li, Bo; Krogan, Nevan; Typas, Athanasios; Gross, Carol A

    2016-06-01

    Fighting antibiotic resistance requires a deeper understanding of the genetic factors that determine the antibiotic susceptibility of bacteria. Here we describe a chemical-genomic screen in Escherichia coli K-12 that was designed to discover new aspects of antibiotic resistance by focusing on a set of 26 antibiotics and other stresses with poorly characterized mode-of-action and determinants of resistance. We show that the screen identifies new resistance determinants for these antibiotics including a common signature from two antimicrobials, kasugamycin and blasticidin S, used to treat crop diseases like rice blast and fire blight. Following this signature, we further investigated the mechanistic basis for susceptibility to kasugamycin and blasticidin S in E. coli using both genetic and biochemical approaches. We provide evidence that these compounds hijack an overlapping set of peptide ABC-importers to enter the bacterial cell. Loss of uptake may be an underappreciated mechanism for the development of kasugamycin resistance in bacterial plant pathogens. PMID:27355376

  3. A Chemical-Genomic Screen of Neglected Antibiotics Reveals Illicit Transport of Kasugamycin and Blasticidin S.

    Directory of Open Access Journals (Sweden)

    Anthony L Shiver

    2016-06-01

    Full Text Available Fighting antibiotic resistance requires a deeper understanding of the genetic factors that determine the antibiotic susceptibility of bacteria. Here we describe a chemical-genomic screen in Escherichia coli K-12 that was designed to discover new aspects of antibiotic resistance by focusing on a set of 26 antibiotics and other stresses with poorly characterized mode-of-action and determinants of resistance. We show that the screen identifies new resistance determinants for these antibiotics including a common signature from two antimicrobials, kasugamycin and blasticidin S, used to treat crop diseases like rice blast and fire blight. Following this signature, we further investigated the mechanistic basis for susceptibility to kasugamycin and blasticidin S in E. coli using both genetic and biochemical approaches. We provide evidence that these compounds hijack an overlapping set of peptide ABC-importers to enter the bacterial cell. Loss of uptake may be an underappreciated mechanism for the development of kasugamycin resistance in bacterial plant pathogens.

  4. Global burden of genetic disease and the role of genetic screening.

    Science.gov (United States)

    Verma, I C; Puri, R D

    2015-10-01

    It is estimated that 5.3% of newborns will suffer from a genetic disorder, when followed up until the age of 25 years. In developing, as compared to western countries, hemoglobinopathies and glucose-6-phosphate dehydrogenase deficiency have a higher incidence due to severe falciparum malaria in the distant past, and autosomal recessive disorders have a higher frequency due to greater proportion of consanguineous marriages. Chromosomal disorders have a combined frequency of 1 in 153 births, therefore screening for chromosomal disorders is essential, using biochemical markers, ultrasonography, and recently by non-invasive prenatal diagnosis based on cell-free fetal DNA in maternal plasma. Preconceptional counseling should be encouraged. For genetic disorders screening should be carried out, ideally after marriage, but before pregnancy. The disorders to be screened depend upon ethnicity. Metabolic disorders have a high incidence in developing countries due to greater rate of consanguineous marriages. Newborn screening is recommended to reduce the burden of these disorders, as many metabolic disorders can be treated. Hearing and critical congenital heart disease should both be screened in the newborn period.

  5. Pooled-matrix protein interaction screens using Barcode Fusion Genetics.

    Science.gov (United States)

    Yachie, Nozomu; Petsalaki, Evangelia; Mellor, Joseph C; Weile, Jochen; Jacob, Yves; Verby, Marta; Ozturk, Sedide B; Li, Siyang; Cote, Atina G; Mosca, Roberto; Knapp, Jennifer J; Ko, Minjeong; Yu, Analyn; Gebbia, Marinella; Sahni, Nidhi; Yi, Song; Tyagi, Tanya; Sheykhkarimli, Dayag; Roth, Jonathan F; Wong, Cassandra; Musa, Louai; Snider, Jamie; Liu, Yi-Chun; Yu, Haiyuan; Braun, Pascal; Stagljar, Igor; Hao, Tong; Calderwood, Michael A; Pelletier, Laurence; Aloy, Patrick; Hill, David E; Vidal, Marc; Roth, Frederick P

    2016-04-22

    High-throughput binary protein interaction mapping is continuing to extend our understanding of cellular function and disease mechanisms. However, we remain one or two orders of magnitude away from a complete interaction map for humans and other major model organisms. Completion will require screening at substantially larger scales with many complementary assays, requiring further efficiency gains in proteome-scale interaction mapping. Here, we report Barcode Fusion Genetics-Yeast Two-Hybrid (BFG-Y2H), by which a full matrix of protein pairs can be screened in a single multiplexed strain pool. BFG-Y2H uses Cre recombination to fuse DNA barcodes from distinct plasmids, generating chimeric protein-pair barcodes that can be quantified via next-generation sequencing. We applied BFG-Y2H to four different matrices ranging in scale from ~25 K to 2.5 M protein pairs. The results show that BFG-Y2H increases the efficiency of protein matrix screening, with quality that is on par with state-of-the-art Y2H methods.

  6. Pooled-matrix protein interaction screens using Barcode Fusion Genetics.

    Science.gov (United States)

    Yachie, Nozomu; Petsalaki, Evangelia; Mellor, Joseph C; Weile, Jochen; Jacob, Yves; Verby, Marta; Ozturk, Sedide B; Li, Siyang; Cote, Atina G; Mosca, Roberto; Knapp, Jennifer J; Ko, Minjeong; Yu, Analyn; Gebbia, Marinella; Sahni, Nidhi; Yi, Song; Tyagi, Tanya; Sheykhkarimli, Dayag; Roth, Jonathan F; Wong, Cassandra; Musa, Louai; Snider, Jamie; Liu, Yi-Chun; Yu, Haiyuan; Braun, Pascal; Stagljar, Igor; Hao, Tong; Calderwood, Michael A; Pelletier, Laurence; Aloy, Patrick; Hill, David E; Vidal, Marc; Roth, Frederick P

    2016-01-01

    High-throughput binary protein interaction mapping is continuing to extend our understanding of cellular function and disease mechanisms. However, we remain one or two orders of magnitude away from a complete interaction map for humans and other major model organisms. Completion will require screening at substantially larger scales with many complementary assays, requiring further efficiency gains in proteome-scale interaction mapping. Here, we report Barcode Fusion Genetics-Yeast Two-Hybrid (BFG-Y2H), by which a full matrix of protein pairs can be screened in a single multiplexed strain pool. BFG-Y2H uses Cre recombination to fuse DNA barcodes from distinct plasmids, generating chimeric protein-pair barcodes that can be quantified via next-generation sequencing. We applied BFG-Y2H to four different matrices ranging in scale from ~25 K to 2.5 M protein pairs. The results show that BFG-Y2H increases the efficiency of protein matrix screening, with quality that is on par with state-of-the-art Y2H methods. PMID:27107012

  7. Chemical Genetics — A Versatile Method to Combine Science and Higher Level Teaching in Molecular Genetics

    Directory of Open Access Journals (Sweden)

    Björn Sandrock

    2012-10-01

    Full Text Available Phosphorylation is a key event in many cellular processes like cell cycle, transformation of environmental signals to transcriptional activation or polar growth. The chemical genetics approach can be used to analyse the effect of highly specific inhibition in vivo and is a promising method to screen for kinase targets. We have used this approach to study the role of the germinal centre kinase Don3 during the cell division in the phytopathogenic fungus Ustilago maydis. Due to the easy determination of the don3 phenotype we have chosen this approach for a genetic course for M.Sc. students and for IMPRS (International Max-Planck research school students. According to the principle of “problem-based learning” the aim of this two-week course is to transfer knowledge about the broad spectrum of kinases to the students and that the students acquire the ability to design their own analog-sensitive kinase of interest. In addition to these training goals, we benefit from these annual courses the synthesis of basic constructs for genetic modification of several kinases in our model system U. maydis.

  8. Applying theological developments to bioethical issues such as genetic screening.

    Science.gov (United States)

    Mallia, Pierre; ten Have, Henk

    2005-01-01

    Catholic movements within the centre of Roman Catholic doctrine recently have discussed Trinitarian theology as applied to sciences, arts, economics, health and other social areas. We explore the possibilities Trinitarian theology offers to bioethical debate, concentrating particularly on genetic screening and testing. It is important therefore to analyse the philosophical implications of this approach onto the bioethical world, where much disagreement occurs on fundamental issues. It is Catholic basic teaching to recognize and see God's hand in plurality, not merely as a cliche and then doing what we feel is right, but to recognize how to live in a pluralistic world. We recognize, in agreement with these theologians, that in order for a Trinitarian mode of understanding to be used by those doing bioethical debate, there is a need to depart from fundamentalism.

  9. A microfluidic-based genetic screen to identify microbial virulence factors that inhibit dendritic cell migration.

    Science.gov (United States)

    McLaughlin, Laura M; Xu, Hui; Carden, Sarah E; Fisher, Samantha; Reyes, Monique; Heilshorn, Sarah C; Monack, Denise M

    2014-04-01

    Microbial pathogens are able to modulate host cells and evade the immune system by multiple mechanisms. For example, Salmonella injects effector proteins into host cells and evades the host immune system in part by inhibiting dendritic cell (DC) migration. The identification of microbial factors that modulate normal host functions should lead to the development of new classes of therapeutics that target these pathways. Current screening methods to identify either host or pathogen genes involved in modulating migration towards a chemical signal are limited because they do not employ stable, precisely controlled chemical gradients. Here, we develop a positive selection microfluidic-based genetic screen that allows us to identify Salmonella virulence factors that manipulate DC migration within stable, linear chemokine gradients. Our screen identified 7 Salmonella effectors (SseF, SifA, SspH2, SlrP, PipB2, SpiC and SseI) that inhibit DC chemotaxis toward CCL19. This method is widely applicable for identifying novel microbial factors that influence normal host cell chemotaxis as well as revealing new mammalian genes involved in directed cell migration. PMID:24599496

  10. Quantitative Chemical-Genetic Interaction Map Connects Gene Alterations to Drug Responses | Office of Cancer Genomics

    Science.gov (United States)

    In a recent Cancer Discovery report, CTD2 researchers at the University of California in San Francisco developed a new quantitative chemical-genetic interaction mapping approach to evaluate drug sensitivity or resistance in isogenic cell lines. Performing a high-throughput screen with isogenic cell lines allowed the researchers to explore the impact of a panel of emerging and established drugs on cells overexpressing a single cancer-associated gene in isolation.

  11. Mining Chemical Activity Status from High-Throughput Screening Assays

    KAUST Repository

    Soufan, Othman

    2015-12-14

    High-throughput screening (HTS) experiments provide a valuable resource that reports biological activity of numerous chemical compounds relative to their molecular targets. Building computational models that accurately predict such activity status (active vs. inactive) in specific assays is a challenging task given the large volume of data and frequently small proportion of active compounds relative to the inactive ones. We developed a method, DRAMOTE, to predict activity status of chemical compounds in HTP activity assays. For a class of HTP assays, our method achieves considerably better results than the current state-of-the-art-solutions. We achieved this by modification of a minority oversampling technique. To demonstrate that DRAMOTE is performing better than the other methods, we performed a comprehensive comparison analysis with several other methods and evaluated them on data from 11 PubChem assays through 1,350 experiments that involved approximately 500,000 interactions between chemicals and their target proteins. As an example of potential use, we applied DRAMOTE to develop robust models for predicting FDA approved drugs that have high probability to interact with the thyroid stimulating hormone receptor (TSHR) in humans. Our findings are further partially and indirectly supported by 3D docking results and literature information. The results based on approximately 500,000 interactions suggest that DRAMOTE has performed the best and that it can be used for developing robust virtual screening models. The datasets and implementation of all solutions are available as a MATLAB toolbox online at www.cbrc.kaust.edu.sa/dramote and can be found on Figshare.

  12. Chemical genetics approaches for selective intervention in epigenetics.

    Science.gov (United States)

    Runcie, Andrew C; Chan, Kwok-Ho; Zengerle, Michael; Ciulli, Alessio

    2016-08-01

    Chemical genetics is the use of biologically active small molecules (chemical probes) to investigate the functions of gene products, through the modulation of protein activity. Recent years have seen significant progress in the application of chemical genetics to study epigenetics, following the development of new chemical probes, a growing appreciation of the role of epigenetics in disease and a recognition of the need and utility of high-quality, cell-active chemical probes. In this review, we single out the bromodomain reader domains as a prime example of both the success, and challenges facing chemical genetics. The difficulty in generating single-target selectivity has long been a thorn in the side of chemical genetics, however, recent developments in advanced forms of chemical genetics promise to bypass this, and other, limitations. The 'bump-and-hole' approach has now been used to probe - for the first time - the BET bromodomain subfamily with single-target selectivity and may be applicable to other epigenetic domains. Meanwhile, PROTAC compounds have been shown to be significantly more efficacious than standard domain inhibitors, and have the potential to enhance target selectivity.

  13. Genetic Counseling and Screening of Consanguineous Couples and Their Offspring: Recommendations of the National Society of Genetic Counselors.

    Science.gov (United States)

    Bennett, Robin L; Motulsky, Arno G; Bittles, Alan; Hudgins, Louanne; Uhrich, Stefanie; Doyle, Debra Lochner; Silvey, Kerry; Scott, C Ronald; Cheng, Edith; McGillivray, Barbara; Steiner, Robert D; Olson, Debra

    2002-04-01

    The objective of this document is to provide recommendations for genetic counseling and screening for consanguineous couples (related as second cousins or closer) and their offspring with the goals of1. providing preconception reproductive options2. improving pregnancy outcome and identifying reproductive choices3. reducing morbidity and mortality in the 1st years of life, and4. respecting psychosocial and multicultural issues.The recommendations are the opinions of a multicenter working group (the Consanguinity Working Group (CWG)) with expertise in genetic counseling, medical genetics, biochemical genetics, genetic epidemiology, pediatrics, perinatology, and public health genetics, which was convened by the National Society of Genetic Counselors (NSGC). The consensus of the CWG and NSGC reviewers is that beyond a thorough medical family history with follow-up of significant findings, no additional preconception screening is recommended for consanguineous couples. Consanguineous couples should be offered similar genetic screening as suggested for any couple of their ethnic group. During pregnancy, consanguineous couples should be offered maternal-fetal serum marker screening and high-resolution fetal ultrasonography. Newborns should be screened for impaired hearing and detection of treatable inborn errors of metabolism. These recommendations should not be construed as dictating an exclusive course of management, nor does use of such recommendations guarantee a particular outcome. The professional judgment of a health care provider, familiar with the facts and circumstances of a specific case, will always supersede these recommendations.

  14. Wildlife ecological screening levels for inhalation of volatile organic chemicals.

    Science.gov (United States)

    Gallegos, Patricia; Lutz, Jill; Markwiese, James; Ryti, Randall; Mirenda, Rich

    2007-06-01

    For most chemicals, evaluation of ecological risk typically does not address inhalation because ingestion dominates exposure. However, burrowing ecological receptors have an increased exposure potential from inhalation at sites contaminated with volatile chemicals in the subsurface. Evaluation of ecological risk from contaminants like volatile organic chemicals (VOCs) is constrained by a lack of relevant ecological screening levels (ESLs). To address this need, inhalation ESLs were developed for 16 VOCs: Acetone, benzene, carbon tetrachloride, chloroform, chloromethane, dichlorodifluoromethane, 1,1-dichloroethane, 1,2-dichloroethane, 1,1-dichloroethene, methylene chloride, tetrachloroethene, toluene, 1,1,1-trichloroethane, trichloroethene, trichlorofluoromethane, and total xylene. These ESLs are based on Botta's pocket gopher (Thomomys bottae) as a representative fossorial receptor. The ESLs are presented with an emphasis on the process for developing inhalation toxicity reference values to illustrate the selection of suitable toxicity data and effect levels from the literature. The resulting ESLs provide a quantitative method for evaluating ecological risk of VOCs through comparison to relevant exposure data such as direct burrow-air measurements. The toxicity reference value development and ESL calculation processes and assumptions detailed here are provided as bases from which risk assessors can use or refine to suit site-specific needs with respect to toxicity and exposure inputs.

  15. Mutagenicity of chemicals in genetically modified animals

    NARCIS (Netherlands)

    Willems MI; van Benthem J; LEO

    2001-01-01

    The strategy for assessing human health risks of chemicals consists of a large number of tests in different research disciplines. Tests include acute and chronic toxicity, genotoxicity, reproduction toxicity and carcinogenicity. Genotoxic properties of chemicals are assessed in short-term in vitro

  16. The TSCA interagency testing committee`s approaches to screening and scoring chemicals and chemical groups: 1977-1983

    Energy Technology Data Exchange (ETDEWEB)

    Walker, J.D. [Environmental Protection Agency, Washington, DC (United States)

    1990-12-31

    This paper describes the TSCA interagency testing committee`s (ITC) approaches to screening and scoring chemicals and chemical groups between 1977 and 1983. During this time the ITC conducted five scoring exercises to select chemicals and chemical groups for detailed review and to determine which of these chemicals and chemical groups should be added to the TSCA Section 4(e) Priority Testing List. 29 refs., 1 fig., 2 tabs.

  17. Film loss-free cleaning chemicals for EUV mask lifetime elongation developed through combinatorial chemical screening

    Science.gov (United States)

    Choi, Jaehyuck; Kim, Jinsu; Lowe, Jeff; Dattilo, Davide; Koh, Soowan; Choi, Jun Yeol; Dietze, Uwe; Shoki, Tsutomu; Kim, Byung Gook; Jeon, Chan-Uk

    2015-10-01

    EUV masks include many different layers of various materials rarely used in optical masks, and each layer of material has a particular role in enhancing the performance of EUV lithography. Therefore, it is crucial to understand how the mask quality and patterning performance can change during mask fabrication, EUV exposure, maintenance cleaning, shipping, or storage. SPM (Sulfuric acid peroxide mixture) which has been extensively used for acid cleaning of photomask and wafer has serious drawback for EUV mask cleaning. It shows severe film loss of tantalum-based absorber layers and limited removal efficiency of EUV-generated carbon contaminants on EUV mask surface. Here, we introduce such novel cleaning chemicals developed for EUV mask as almost film loss free for various layers of the mask and superior carbon removal performance. Combinatorial chemical screening methods allowed us to screen several hundred combinations of various chemistries and additives under several different process conditions of temperature and time, eventually leading to development of the best chemistry selections for EUV mask cleaning. Recently, there have been many activities for the development of EUV pellicle, driven by ASML and core EUV scanner customer companies. It is still important to obtain film-loss free cleaning chemicals because cleaning cycle of EUV mask should be much faster than that of optic mask mainly due to EUV pellicle lifetime. More frequent cleaning, combined with the adoption of new materials for EUV masks, necessitates that mask manufacturers closely examine the performance change of EUV masks during cleaning process. We have investigated EUV mask quality changes and film losses during 50 cleaning cycles using new chemicals as well as particle and carbon contaminant removal characteristics. We have observed that the performance of new chemicals developed is superior to current SPM or relevant cleaning chemicals for EUV mask cleaning and EUV mask lifetime elongation.

  18. Quantitative genetic activity graphical profiles for use in chemical evaluation

    Energy Technology Data Exchange (ETDEWEB)

    Waters, M.D. [Environmental Protection Agency, Washington, DC (United States); Stack, H.F.; Garrett, N.E.; Jackson, M.A. [Environmental Health Research and Testing, Inc., Research Triangle Park, NC (United States)

    1990-12-31

    A graphic approach, terms a Genetic Activity Profile (GAP), was developed to display a matrix of data on the genetic and related effects of selected chemical agents. The profiles provide a visual overview of the quantitative (doses) and qualitative (test results) data for each chemical. Either the lowest effective dose or highest ineffective dose is recorded for each agent and bioassay. Up to 200 different test systems are represented across the GAP. Bioassay systems are organized according to the phylogeny of the test organisms and the end points of genetic activity. The methodology for producing and evaluating genetic activity profile was developed in collaboration with the International Agency for Research on Cancer (IARC). Data on individual chemicals were compiles by IARC and by the US Environmental Protection Agency (EPA). Data are available on 343 compounds selected from volumes 1-53 of the IARC Monographs and on 115 compounds identified as Superfund Priority Substances. Software to display the GAPs on an IBM-compatible personal computer is available from the authors. Structurally similar compounds frequently display qualitatively and quantitatively similar profiles of genetic activity. Through examination of the patterns of GAPs of pairs and groups of chemicals, it is possible to make more informed decisions regarding the selection of test batteries to be used in evaluation of chemical analogs. GAPs provided useful data for development of weight-of-evidence hazard ranking schemes. Also, some knowledge of the potential genetic activity of complex environmental mixtures may be gained from an assessment of the genetic activity profiles of component chemicals. The fundamental techniques and computer programs devised for the GAP database may be used to develop similar databases in other disciplines. 36 refs., 2 figs.

  19. Flow method and apparatus for screening chemicals using micro x-ray fluorescence

    Science.gov (United States)

    Warner, Benjamin P.; Havrilla, George J.; Miller, Thomasin C.; Lewis, Cris; Mahan, Cynthia A.; Wells, Cyndi A.

    2009-04-14

    Method and apparatus for screening chemicals using micro x-ray fluorescence. A method for screening a mixture of potential pharmaceutical chemicals for binding to at least one target binder involves flow-separating a solution of chemicals and target binders into separated components, exposing them to an x-ray excitation beam, detecting x-ray fluorescence signals from the components, and determining from the signals whether or not a binding event between a chemical and target binder has occurred.

  20. Medical and lay attitudes towards genetic screening and testing in Finland

    DEFF Research Database (Denmark)

    Toiviainen, Hanna; Jallinoja, Piia; Aro, Arja R;

    2003-01-01

    peoples' expectations as regards to genetic testing are too high. Having more medical education was related to having less 'cannot say' and missing responses. Our results do not suggest that major conflicts about the direction of genetic testing and screening would arise in near future. However, different......The purpose of this study was to compare physicians', midwives' and lay people's attitudes towards genetic screening and testing to find out whether medical education and experience influence attitudes of genetic screening and testing. The study was based on comparison of answers to joint questions...... referred to as midwives in the following; n=800, response rate 79%), and lay people (n=2000, response rate 62%). Midwives were more worried about the consequences of genetic testing and stressed the autonomy of the customer more strongly than lay people did. Furthermore, professionals considered that lay...

  1. Chemical characteristics and volatile profile of genetically modified peanut cultivars.

    Science.gov (United States)

    Ng, Ee Chin; Dunford, Nurhan T; Chenault, Kelly

    2008-10-01

    Genetic engineering has been used to modify peanut cultivars for improving agronomic performance and pest resistance. Food products developed through genetic engineering have to be assessed for their safety before approval for human consumption. Preservation of desirable chemical, flavor and aroma attributes of the peanut cultivars during the genetic modifications is critical for acceptance of genetically modified peanuts (GMP) by the food industry. Hence, the main objective of this study is to examine chemical characteristics and volatile profile of GMP. The genetically modified peanut cultivars, 188, 540 and 654 were obtained from the USDA-ARS in Stillwater, Oklahoma. The peanut variety Okrun was examined as a control. The volatile analysis was performed using a gas chromatograph/mass spectrometer (GC/MS) equipped with an olfactory detector. The peanut samples were also analyzed for their moisture, ash, protein, sugar and oil compositions. Experimental results showed that the variations in nutritional composition of peanut lines examined in this study were within the values reported for existing cultivars. There were minor differences in volatile profile among the samples. The implication of this study is significant, since it shows that peanut cultivars with greater pest and fungal resistance were successfully developed without major changes in their chemical characteristics.

  2. Chemical characteristics and volatile profile of genetically modified peanut cultivars.

    Science.gov (United States)

    Ng, Ee Chin; Dunford, Nurhan T; Chenault, Kelly

    2008-10-01

    Genetic engineering has been used to modify peanut cultivars for improving agronomic performance and pest resistance. Food products developed through genetic engineering have to be assessed for their safety before approval for human consumption. Preservation of desirable chemical, flavor and aroma attributes of the peanut cultivars during the genetic modifications is critical for acceptance of genetically modified peanuts (GMP) by the food industry. Hence, the main objective of this study is to examine chemical characteristics and volatile profile of GMP. The genetically modified peanut cultivars, 188, 540 and 654 were obtained from the USDA-ARS in Stillwater, Oklahoma. The peanut variety Okrun was examined as a control. The volatile analysis was performed using a gas chromatograph/mass spectrometer (GC/MS) equipped with an olfactory detector. The peanut samples were also analyzed for their moisture, ash, protein, sugar and oil compositions. Experimental results showed that the variations in nutritional composition of peanut lines examined in this study were within the values reported for existing cultivars. There were minor differences in volatile profile among the samples. The implication of this study is significant, since it shows that peanut cultivars with greater pest and fungal resistance were successfully developed without major changes in their chemical characteristics. PMID:19000610

  3. Identification of genetic and chemical modulators of zebrafish mechanosensory hair cell death.

    Directory of Open Access Journals (Sweden)

    Kelly N Owens

    2008-02-01

    Full Text Available Inner ear sensory hair cell death is observed in the majority of hearing and balance disorders, affecting the health of more than 600 million people worldwide. While normal aging is the single greatest contributor, exposure to environmental toxins and therapeutic drugs such as aminoglycoside antibiotics and antineoplastic agents are significant contributors. Genetic variation contributes markedly to differences in normal disease progression during aging and in susceptibility to ototoxic agents. Using the lateral line system of larval zebrafish, we developed an in vivo drug toxicity interaction screen to uncover genetic modulators of antibiotic-induced hair cell death and to identify compounds that confer protection. We have identified 5 mutations that modulate aminoglycoside susceptibility. Further characterization and identification of one protective mutant, sentinel (snl, revealed a novel conserved vertebrate gene. A similar screen identified a new class of drug-like small molecules, benzothiophene carboxamides, that prevent aminoglycoside-induced hair cell death in zebrafish and in mammals. Testing for interaction with the sentinel mutation suggests that the gene and compounds may operate in different pathways. The combination of chemical screening with traditional genetic approaches is a new strategy for identifying drugs and drug targets to attenuate hearing and balance disorders.

  4. The application and use of chemical space mapping to interpret crystallization screening results

    OpenAIRE

    Snell, Edward H.; Nagel, Ray M.; Wojtaszcyk, Ann; O’Neill, Hugh; Wolfley, Jennifer L.; Joseph R Luft

    2008-01-01

    Macromolecular crystallization screening is an empirical process. It often begins by setting up experiments with a number of chemically diverse cocktails designed to sample chemical space known to promote crystallization. Where a potential crystal is seen a refined screen is set up, optimizing around that condition. By using an incomplete factorial sampling of chemical space to formulate the cocktails and presenting the results graphically, it is possible to readily identify trends relevant t...

  5. Coeliac disease and autoimmune disease-genetic overlap and screening

    NARCIS (Netherlands)

    Lundin, Knut E. A.; Wijmenga, Cisca

    2015-01-01

    Coeliac disease is a treatable, gluten-induced disease that often occurs concurrently with other autoimmune diseases. In genetic studies since 2007, a partial genetic overlap between these diseases has been revealed and further insights into the pathophysiology of coeliac disease and autoimmunity ha

  6. Colorectal Cancer Survivors' Interest in Genetic Testing for Hereditary Cancer: Implications for Universal Tumor Screening

    OpenAIRE

    Cragun, Deborah; Malo, Teri L.; Pal, Tuya; Shibata, David; Vadaparampil, Susan T

    2012-01-01

    Aims: Benefits of universal tumor screening for Lynch syndrome (LS), the most common form of hereditary colorectal cancer (CRC), will be realized only if patients are interested in genetic counseling and testing. This study explores interest in genetic testing for hereditary CRC among CRC patients who have never received genetic counseling or testing. Methods Using results from a cross-sectional survey of CRC patients (n=91) at varying categories of risk for hereditary CRC, bivariate and mult...

  7. Genetic susceptibility factors for multiple chemical sensitivity revisited

    DEFF Research Database (Denmark)

    Rasmussen, Henrik Berg; Linneberg, Allan; Andersen, Charlotte Brasch;

    2010-01-01

    Multiple chemical sensitivity (MCS) is characterised by adverse effects due to exposure to low levels of chemical substances. Various genes, especially genes of importance to the metabolism of xenobiotic compounds, have been associated with MCS, but findings are inconsistent. The purpose of this ...... examined are of less importance to MCS than previously reported or that gene-environment interactions or significant degrees of genetic heterogeneity in MCS underlie inconsistent findings in the literature....... of this study was to investigate genetic susceptibility factors for MCS and self-reported chemical sensitivity in a population sample. Ninety six MCS patients and 1,207 controls from a general population divided into four severity groups of chemical sensitivity were genotyped for variants in the genes encoding...... compared in post hoc analyses with all individuals from the population sample (p=0.02). Genetic variants in paraoxonase 1 and methylene tetrahydrofolate reductase were not associated with MSC or with self-reported chemical sensitivity in the population sample. Our results suggest that variants in the genes...

  8. A chemical genetics approach for specific differentiation of stem cells to somatic cells: a new promising therapeutical approach.

    Science.gov (United States)

    Sachinidis, Agapios; Sotiriadou, Isaia; Seelig, Bianca; Berkessel, Albrecht; Hescheler, Jürgen

    2008-01-01

    Cell replacement therapy of severe degenerative diseases such as diabetes, myocardial infarction and Parkinson's disease through transplantation of somatic cells generated from embryonic stem (ES) cells is currently receiving considerable attention for the therapeutic applications. ES cells harvested from the inner cell mass (ICM) of the early embryo, can proliferate indefinitely in vitro while retaining the ability to differentiate into all somatic cells thereby providing an unlimited renewable source of somatic cells. In this context, identifying soluble factors, in particular chemically synthesized small molecules, and signal cascades involved in specific differentiation processes toward a defined tissue specific cell type are crucial for optimizing the generation of somatic cells in vitro for therapeutic approaches. However, experimental models are required allowing rapid and "easy-to-handle" parallel screening of chemical libraries to achieve this goal. Recently, the forward chemical genetic screening strategy has been postulated to screen small molecules in cellular systems for a specific desired phenotypic effect. The current review is focused on the progress of ES cell research in the context of the chemical genetics to identify small molecules promoting specific differentiation of ES cells to desired cell phenotype. Chemical genetics in the context of the cell ES-based cell replacement therapy remains a challenge for the near future for several scientific fields including chemistry, molecular biology, medicinal physics and robotic technologies.

  9. Applying theological developments to bioethical issues such as genetic screening.

    NARCIS (Netherlands)

    Mallia, P.; Have, H.A.M.J. ten

    2005-01-01

    Catholic movements within the centre of Roman Catholic doctrine recently have discussed Trinitarian theology as applied to sciences, arts, economics, health and other social areas. We explore the possibilities Trinitarian theology offers to bioethical debate, concentrating particularly on genetic sc

  10. Pathways and barriers to genetic testing and screening: Molecular genetics meets the high-risk family. Final report

    Energy Technology Data Exchange (ETDEWEB)

    Duster, T.

    1998-11-01

    The proliferation of genetic screening and testing is requiring increasing numbers of Americans to integrate genetic knowledge and interventions into their family life and personal experience. This study examines the social processes that occur as families at risk for two of the most common autosomal recessive diseases, sickle cell disease (SC) and cystic fibrosis (CF), encounter genetic testing. Each of these diseases is found primarily in a different ethnic/racial group (CF in Americans of North European descent and SC in Americans of West African descent). This has permitted them to have a certain additional lens on the role of culture in integrating genetic testing into family life and reproductive planning. A third type of genetic disorder, the thalassemias was added to the sample in order to extent the comparative frame and to include other ethnic and racial groups.

  11. Genetic screening in couples experiencing recurrent assisted procreation failure.

    Science.gov (United States)

    Dada, Rima; Kumar, R; Shamsi, M B; Tanwar, M; Pathak, D; Venkatesh, S; Kumar, M; Singh, H; Singh, K; Aron, M; Kumar, R; Singh, G; Sharma, R K; Gupta, N P

    2008-04-01

    Infertility is a major health problem affecting about 10-20% of couples in the reproductive age group. Male factor is assumed to be responsible in about 50% cases of infertility. The origin of reduced testicular sperm function is unknown in about 50-70% of cases and for such couples assisted reproduction techniques (ART) are a boon. Male infertility is often due to poor semen quality and may be associated with genetic defects. ART has revolutionized management of infertility and intracytoplasmic sperm injection (ICSI) is the ART procedure of choice in 60-80% cases. Despite major technological advancements and professional expertise in ART, the success rate and carry-home live birth rate of ICSI is low (18-25%). This study was aimed to understand the genetic etiopathology of recurrent ART failure. For this, 110 couples with 3 or more failed ART cycles were recruited. A detailed history was taken and only idiopathic ART failure cases were enrolled for this study. They were subjected to cytogenetic and Yq microdeletion analysis. Genetic abnormalities were detected in 19 couples. Since a large number (18.2%) cases harboured genetic abnormalities, it is important for all couples opting for ART to undergo a thorough genetic analysis to prevent recurrent emotional, physical and financial stress. PMID:21086725

  12. 76 FR 49473 - Petition to Maximize Practical Utility of List 1 Chemicals Screened Through EPA's Endocrine...

    Science.gov (United States)

    2011-08-10

    ... AGENCY Petition to Maximize Practical Utility of List 1 Chemicals Screened Through EPA's Endocrine... decisions on data received in response to the test orders issued under the Endocrine Disruptor Screening...'' system, which means EPA will not know your identity or contact information unless you provide it in...

  13. Pooled‐matrix protein interaction screens using Barcode Fusion Genetics

    OpenAIRE

    Yachie, Nozomu; Petsalaki, Evangelia; Mellor, Joseph C.; Weile, Jochen; Jacob, Yves; Verby, Marta; Ozturk, Sedide B.; Li, Siyang; Cote, Atina G; Mosca, Roberto; Knapp, Jennifer J; Ko, Minjeong; Yu, Analyn; Gebbia, Marinella; Sahni, Nidhi

    2016-01-01

    Abstract High‐throughput binary protein interaction mapping is continuing to extend our understanding of cellular function and disease mechanisms. However, we remain one or two orders of magnitude away from a complete interaction map for humans and other major model organisms. Completion will require screening at substantially larger scales with many complementary assays, requiring further efficiency gains in proteome‐scale interaction mapping. Here, we report Barcode Fusion Genetics‐Yeast Tw...

  14. Chemical Genetics Uncovers Novel Inhibitors of Lignification, Including p-Iodobenzoic Acid Targeting CINNAMATE-4-HYDROXYLASE.

    Science.gov (United States)

    Van de Wouwer, Dorien; Vanholme, Ruben; Decou, Raphaël; Goeminne, Geert; Audenaert, Dominique; Nguyen, Long; Höfer, René; Pesquet, Edouard; Vanholme, Bartel; Boerjan, Wout

    2016-09-01

    Plant secondary-thickened cell walls are characterized by the presence of lignin, a recalcitrant and hydrophobic polymer that provides mechanical strength and ensures long-distance water transport. Exactly the recalcitrance and hydrophobicity of lignin put a burden on the industrial processing efficiency of lignocellulosic biomass. Both forward and reverse genetic strategies have been used intensively to unravel the molecular mechanism of lignin deposition. As an alternative strategy, we introduce here a forward chemical genetic approach to find candidate inhibitors of lignification. A high-throughput assay to assess lignification in Arabidopsis (Arabidopsis thaliana) seedlings was developed and used to screen a 10-k library of structurally diverse, synthetic molecules. Of the 73 compounds that reduced lignin deposition, 39 that had a major impact were retained and classified into five clusters based on the shift they induced in the phenolic profile of Arabidopsis seedlings. One representative compound of each cluster was selected for further lignin-specific assays, leading to the identification of an aromatic compound that is processed in the plant into two fragments, both having inhibitory activity against lignification. One fragment, p-iodobenzoic acid, was further characterized as a new inhibitor of CINNAMATE 4-HYDROXYLASE, a key enzyme of the phenylpropanoid pathway synthesizing the building blocks of the lignin polymer. As such, we provide proof of concept of this chemical biology approach to screen for inhibitors of lignification and present a broad array of putative inhibitors of lignin deposition for further characterization. PMID:27485881

  15. SCREENING CHEMICALS FOR ESTROGEN RECEPTOR BIOACTIVITY USING A COMPUTATIONAL MODEL

    Science.gov (United States)

    The U.S. Environmental Protection Agency (EPA) is considering the use high-throughput and computational methods for regulatory applications in the Endocrine Disruptor Screening Program (EDSP). To use these new tools for regulatory decision making, computational methods must be a...

  16. Integrated screening concept in women with genetic predisposition for breast cancer

    International Nuclear Information System (INIS)

    Breast cancer is in 5% of cases due to a genetic disposition. BRCA1 and BRCA2 are by far the most common breast cancer susceptibility genes. For a woman with a genetic predisposition, the individual risk of developing breast cancer sometime in her life is between 70 and 90%. Compared to the spontaneous forms of breast cancer, woman with a genetic predisposition often develop breast cancer at a much younger age. This is why conventional screening programs on the basis of mammography alone cannot be applied without modification to this high-risk group. In this article, an integrated screening concept for women with genetic prodisposition for breast cancer using breast self-examination, clinical examination, ultrasound, mammography and magnetic resonance imaging is introduced. (orig.)

  17. Evaluation of food-relevant chemicals in the ToxCast high-throughput screening program

    Science.gov (United States)

    There are thousands of chemicals that are directly added to or come in contact with food, many of which have undergone little to no toxicological evaluation. The ToxCast high-throughput screening (HTS) program has evaluated over 1,800 chemicals in concentration-response across ~8...

  18. Effect of screening for cystic-fibrosis on the influence of genetic-counseling

    NARCIS (Netherlands)

    Dankert-Roelse, J E; te Meerman, G J; Knol, K; ten Kate, L P

    1987-01-01

    We studied the influence of genetic counseling for cystic fibrosis on family planning, using neonatal screening, family size at time of diagnosis, and maternal age as possible determinants for reproductive behaviour. The expected number of children born to mothers of equal age and parity in the same

  19. Digital microfluidic three-dimensional cell culture and chemical screening platform using alginate hydrogels.

    Science.gov (United States)

    George, Subin M; Moon, Hyejin

    2015-03-01

    Electro wetting-on-dielectric (EWOD) digital microfluidics (DMF) can be used to develop improved chemical screening platforms using 3-dimensional (3D) cell culture. Alginate hydrogels are one common method by which a 3D cell culture environment is created. This paper presents a study of alginate gelation on EWOD DMF and investigates designs to obtain uniform alginate hydrogels that can be repeatedly addressed by any desired liquids. A design which allows for gels to be retained in place during liquid delivery and removal without using any physical barriers or hydrophilic patterning of substrates is presented. A proof of concept screening platform is demonstrated by examining the effects of different concentrations of a test chemical on 3D cells in alginate hydrogels. In addition, the temporal effects of the various chemical concentrations on different hydrogel posts are demonstrated, thereby establishing the benefits of an EWOD DMF 3D cell culture and chemical screening platform using alginate hydrogels. PMID:25945142

  20. CRISPR-Cas9 for medical genetic screens: applications and future perspectives.

    Science.gov (United States)

    Xue, Hui-Ying; Ji, Li-Juan; Gao, Ai-Mei; Liu, Ping; He, Jing-Dong; Lu, Xiao-Jie

    2016-02-01

    CRISPR-Cas9 (clustered regularly interspaced short palindromic repeats-CRISPR associated nuclease 9) systems have emerged as versatile and convenient (epi)genome editing tools and have become an important player in medical genetic research. CRISPR-Cas9 and its variants such as catalytically inactivated Cas9 (dead Cas9, dCas9) and scaffold-incorporating single guide sgRNA (scRNA) have been applied in various genomic screen studies. CRISPR screens enable high-throughput interrogation of gene functions in health and diseases. Compared with conventional RNAi screens, CRISPR screens incur less off-target effects and are more versatile in that they can be used in multiple formats such as knockout, knockdown and activation screens, and can target coding and non-coding regions throughout the genome. This powerful screen platform holds the potential of revolutionising functional genomic studies in the near future. Herein, we introduce the mechanisms of (epi)genome editing mediated by CRISPR-Cas9 and its variants, introduce the procedures and applications of CRISPR screen in functional genomics, compare it with conventional screen tools and at last discuss current challenges and opportunities and propose future directions.

  1. Largest Common Chemical Feature Subtree as a Virtual Screening Method

    DEFF Research Database (Denmark)

    Kristensen, Thomas Greve; Pedersen, Christian Storm; Thomsen, Rene;

    subtree. Our experiments show that this results in a speed up of the calculation and an improvement in the quality of the result. We test our method on the DUD (Directory of Useful Decoys) test set and compare it to other ligand based screening methods (topological descriptors and CDK fingerprints), using...... the enrichment and AUC (Area Under Curve) as a performance measure. We also compare the performance with the docking program DOCK. The results of our experiments indicate that our method performs better than the topological descriptor and that it is comparable to the CDK fingerprint. The docking based method...

  2. An integrated hybrid microfluidic device for oviposition-based chemical screening of adult Drosophila melanogaster.

    Science.gov (United States)

    Leung, Jacob C K; Hilliker, Arthur J; Rezai, Pouya

    2016-02-21

    Chemical screening using Drosophila melanogaster (the fruit fly) is vital in drug discovery, agricultural, and toxicological applications. Oviposition (egg laying) on chemically-doped agar plates is an important read-out metric used to quantitatively assess the biological fitness and behavioral responses of Drosophila. Current oviposition-based chemical screening studies are inaccurate, labor-intensive, time-consuming, and inflexible due to the manual chemical doping of agar. In this paper, we have developed a novel hybrid agar-polydimethylsiloxane (PDMS) microfluidic device for single- and multi-concentration chemical dosing and on-chip oviposition screening of free-flying adult stage Drosophila. To achieve this, we have devised a novel technique to integrate agar with PDMS channels using ice as a sacrificial layer. Subsequently, we have conducted single-chemical toxicity and multiple choice chemical preference assays on adult Drosophila melanogaster using zinc and acetic acid at various concentrations. Our device has enabled us to 1) demonstrate that Drosophila is capable of sensing the concentration of different chemicals on a PDMS-agar microfluidic device, which plays significant roles in determining oviposition site selection and 2) investigate whether oviposition preference differs between single- and multi-concentration chemical environments. This device may be used to study fundamental and applied biological questions in Drosophila and other egg laying insects. It can also be extended in design to develop sophisticated and dynamic chemical dosing and high-throughput screening platforms in the future that are not easily achievable with the existing oviposition screening techniques. PMID:26768402

  3. Genetic alterations in sporadic and hereditary colorectal cancer: implementations for screening and follow-up.

    Science.gov (United States)

    Souglakos, John

    2007-01-01

    The genetics underlying an inherited predisposition to cancer are rapidly being uncovered. This fact may ultimately lead to the routine use of molecular tools to diagnose these disorders, and establish interventions to prevent the development of cancer. Among the multiple cancer family syndromes, several are known to be associated with the development of colon cancer. These disorders may be diagnosed during evaluation of the index patient or during screening of family members who are at risk. Although the effectiveness of screening and surveillance strategies is unproven in controlled clinical trials for any of these syndromes, the high cancer risk warrants screening, and reasonable recommendations can be made. Several other genetic syndromes are associated with gastrointestinal polyposis. The risk of colon cancer in these diseases is uncertain, and may not be increased and they are not mentioned in this review. Examples include Cowden disease, intestinal ganglioneuromatosis, Ruvalcaba-Myhre-Smith syndrome, Devon family syndrome, and Cronkite-Canada syndrome. PMID:17384504

  4. Incidental findings, genetic screening and the challenge of personalisation

    Directory of Open Access Journals (Sweden)

    Carlo Petrini

    2014-12-01

    Full Text Available Genetic tests frequently produce more information than is initially expected. Several documents have addressed this issue and offer suggestions regarding how this information should be managed and, in particular, concerning the expedience of revealing (or not revealing it to the persons concerned. While the approaches to the management of these incidental findings (IFs vary, it is usually recommended that the information be disclosed if there is confirmed clinical utility and the possibility of treatment or prevention. However, this leaves unsolved some fundamental issues such as the different ways of interpreting "clinical utility", countless sources of uncertainty and varying ways of defining the notion of "incidental". Guidelines and other reference documents can offer indications to those responsible for managing IFs but should not be allowed to relieve researchers and healthcare professionals of their responsibilities.

  5. Statistically designed experiments to screen chemical mixtures for possible interactions

    NARCIS (Netherlands)

    Groten, J.P.; Tajima, O.; Feron, V.J.; Schoen, E.D.

    1998-01-01

    For the accurate analysis of possible interactive effects of chemicals in a defined mixture, statistical designs are necessary to develop clear and manageable experiments. For instance, factorial designs have been successfully used to detect two-factor interactions. Particularly useful for this purp

  6. Chemical genetics reveals an RGS/G-protein role in the action of a compound.

    Directory of Open Access Journals (Sweden)

    Kevin Fitzgerald

    2006-04-01

    Full Text Available We report here on a chemical genetic screen designed to address the mechanism of action of a small molecule. Small molecules that were active in models of urinary incontinence were tested on the nematode Caenorhabditis elegans, and the resulting phenotypes were used as readouts in a genetic screen to identify possible molecular targets. The mutations giving resistance to compound were found to affect members of the RGS protein/G-protein complex. Studies in mammalian systems confirmed that the small molecules inhibit muscarinic G-protein coupled receptor (GPCR signaling involving G-alphaq (G-protein alpha subunit. Our studies suggest that the small molecules act at the level of the RGS/G-alphaq signaling complex, and define new mutations in both RGS and G-alphaq, including a unique hypo-adapation allele of G-alphaq. These findings suggest that therapeutics targeted to downstream components of GPCR signaling may be effective for treatment of diseases involving inappropriate receptor activation.

  7. [Screening for hereditary neuromuscular disorders with molecular genetic methods in the Roma population of Hungary].

    Science.gov (United States)

    Herczegfalvi, Agnes; Pikó, Henriett; Karcagi, Veronika

    2008-11-30

    Recent medical genetic research has identified a number of novel, or previously known, but rare conditions, caused by private founder mutations. The Finnish and Ashkenazi Jew populations provide the best examples for identifying genes in unique genetic disorders. In these populations, research efforts and high-level medical services resulted in intense improvements of medical care and in organization of population-based screening programs. Hereditary disorders of the Roma populations are known for a long time. The genetic background of these diseases has been established by extensive molecular genetic studies. The Romas represent 6% of the Hungarian population and live under extremely bad health conditions. Therefore, our aim was to map the incidence of the hereditary neuromuscular disorders among the Hungarian Roma population. Moreover, we intended to provide proper information, genetic counseling and possible prevention strategies for the families at risk, which should represent a primer task in public health. Because of our experience in neuromuscular disorders, we choose six, frequent, autosomal recessive disorders for these clinical and genetic studies: hereditary motor and sensory neuropathy type Lom (HMSNL), hereditary motor and sensory neuropathy type Russe (HMSNR), congenital cataracts facial dysmorphism syndrome (CCFDN), limb-girdle muscular dystrophy 2C (LGMD2C), congenital myasthenic syndrome (CMS) and spinal muscular atrophy (SMA). Following identification of the founder mutations, the possibility of prenatal diagnosis and carrier screening for family members will contribute to the decrease of the recurrence risk for these severe, mostly untreatable disorders. PMID:19070320

  8. Genome-Wide Synthetic Genetic Screening by Transposon Mutagenesis in Candida albicans

    Science.gov (United States)

    Horton, Brooke N.; Kumar, Anuj

    2016-01-01

    Transposon-based mutagenesis is an effective method for genetic screening on a genome-wide scale, with particular applicability in organisms possessing compact genomes where transforming DNA tends to integrate by homologous recombination. Methods for transposon mutagenesis have been applied with great success in the budding yeast Saccharomyces cerevisiae and in the related pathogenic yeast Candida albicans. In C. albicans, we have implemented transposon mutagenesis to generate heterozygous mutations for the analysis of complex haploinsufficiency, a type of synthetic genetic interaction wherein a pair of non-complementing heterozygous mutations results in a stronger phenotype then either individual mutation in isolation. Genes exhibiting complex haploinsufficiency typically function within a regulatory pathway, in parallel pathways, or in parallel branches within a single pathway. Here, we present protocols to implement transposon mutagenesis for complex haploinsufficiency screening in C. albicans, indicating methods for transposon construction, mutagenesis, phenotypic screening, and identification of insertion sites in strains of interest. In total, the approach is a useful means to implement large-scale synthetic genetic screening in the diploid C. albicans. PMID:25636616

  9. Expanding the fragrance chemical space for virtual screening

    OpenAIRE

    Ruddigkeit, Lars; Awale, Mahendra; Reymond, Jean-Louis

    2014-01-01

    The properties of fragrance molecules in the public databases SuperScent and Flavornet were analyzed to define a “fragrance-like” (FL) property range (Heavy Atom Count ≤ 21, only C, H, O, S, (O + S) ≤ 3, Hydrogen Bond Donor ≤ 1) and the corresponding chemical space including FL molecules from PubChem (NIH repository of molecules), ChEMBL (bioactive molecules), ZINC (drug-like molecules), and GDB-13 (all possible organic molecules up to 13 atoms of C, N, O, S, Cl). The FL subsets of these data...

  10. A post-developmental genetic screen for zebrafish models of inherited liver disease.

    Science.gov (United States)

    Kim, Seok-Hyung; Wu, Shu-Yu; Baek, Jeong-In; Choi, Soo Young; Su, Yanhui; Flynn, Charles R; Gamse, Joshua T; Ess, Kevin C; Hardiman, Gary; Lipschutz, Joshua H; Abumrad, Naji N; Rockey, Don C

    2015-01-01

    Nonalcoholic fatty liver disease (NAFLD) is one of the most common causes of chronic liver disease such as simple steatosis, nonalcoholic steatohepatitis (NASH), cirrhosis and fibrosis. However, the molecular pathogenesis and genetic variations causing NAFLD are poorly understood. The high prevalence and incidence of NAFLD suggests that genetic variations on a large number of genes might be involved in NAFLD. To identify genetic variants causing inherited liver disease, we used zebrafish as a model system for a large-scale mutant screen, and adopted a whole genome sequencing approach for rapid identification of mutated genes found in our screen. Here, we report on a forward genetic screen of ENU mutagenized zebrafish. From 250 F2 lines of ENU mutagenized zebrafish during post-developmental stages (5 to 8 days post fertilization), we identified 19 unique mutant zebrafish lines displaying visual evidence of hepatomegaly and/or steatosis with no developmental defects. Histological analysis of mutants revealed several specific phenotypes, including common steatosis, micro/macrovesicular steatosis, hepatomegaly, ballooning, and acute hepatocellular necrosis. This work has identified multiple post-developmental mutants and establishes zebrafish as a novel animal model for post-developmental inherited liver disease.

  11. A post-developmental genetic screen for zebrafish models of inherited liver disease.

    Directory of Open Access Journals (Sweden)

    Seok-Hyung Kim

    Full Text Available Nonalcoholic fatty liver disease (NAFLD is one of the most common causes of chronic liver disease such as simple steatosis, nonalcoholic steatohepatitis (NASH, cirrhosis and fibrosis. However, the molecular pathogenesis and genetic variations causing NAFLD are poorly understood. The high prevalence and incidence of NAFLD suggests that genetic variations on a large number of genes might be involved in NAFLD. To identify genetic variants causing inherited liver disease, we used zebrafish as a model system for a large-scale mutant screen, and adopted a whole genome sequencing approach for rapid identification of mutated genes found in our screen. Here, we report on a forward genetic screen of ENU mutagenized zebrafish. From 250 F2 lines of ENU mutagenized zebrafish during post-developmental stages (5 to 8 days post fertilization, we identified 19 unique mutant zebrafish lines displaying visual evidence of hepatomegaly and/or steatosis with no developmental defects. Histological analysis of mutants revealed several specific phenotypes, including common steatosis, micro/macrovesicular steatosis, hepatomegaly, ballooning, and acute hepatocellular necrosis. This work has identified multiple post-developmental mutants and establishes zebrafish as a novel animal model for post-developmental inherited liver disease.

  12. Screening chemicals for thyroid-disrupting activity: A critical comparison of mammalian and amphibian models.

    Science.gov (United States)

    Pickford, Daniel B

    2010-11-01

    In order to minimize risks to human and environmental health, chemical safety assessment programs are being reinforced with toxicity tests more specifically designed for detecting endocrine disrupters. This includes the necessity to detect thyroid-disrupting chemicals, which may operate through a variety of modes of action, and have potential to impair neurological development in humans, with resulting deficits of individual and social potential. Mindful of these concerns, the consensus favors in vivo models for both hazard characterization (testing) and hazard identification (screening) steps, in order to minimize false negatives. Owing to its obligate dependence on thyroid hormones, it has been proposed that amphibian metamorphosis be used as a generalized vertebrate model for thyroid function in screening batteries for detection of thyroid disrupters. However, it seems unlikely that such an assay would ever fully replace in vivo mammalian assays currently being validated for human health risk assessment: in its current form the amphibian metamorphosis screening assay would not provide capacity for reliably detecting other modes of endocrine-disrupting activity. Conversely, several candidate mammalian screening assays appear to offer robust capacity to detect a variety of modes of endocrine-disrupting activity, including thyroid activity. To assess whether omission of an amphibian metamorphosis assay from an in vivo screening battery would generate false negatives, the response of amphibian and mammalian assays to a variety known thyroid disrupters, as reported in peer-reviewed literature or government agency reports, was critically reviewed. Of the chemicals investigated from the literature selected (41), more had been tested in mammalian studies with thyroid-relevant endpoints (32) than in amphibian assays with appropriate windows of exposure and developmental endpoints (27). One chemical (methoxychlor) was reported to exhibit thyroid activity in an appropriate

  13. Human genetics in rheumatoid arthritis guides a high-throughput drug screen of the CD40 signaling pathway.

    Directory of Open Access Journals (Sweden)

    Gang Li

    2013-05-01

    Full Text Available Although genetic and non-genetic studies in mouse and human implicate the CD40 pathway in rheumatoid arthritis (RA, there are no approved drugs that inhibit CD40 signaling for clinical care in RA or any other disease. Here, we sought to understand the biological consequences of a CD40 risk variant in RA discovered by a previous genome-wide association study (GWAS and to perform a high-throughput drug screen for modulators of CD40 signaling based on human genetic findings. First, we fine-map the CD40 risk locus in 7,222 seropositive RA patients and 15,870 controls, together with deep sequencing of CD40 coding exons in 500 RA cases and 650 controls, to identify a single SNP that explains the entire signal of association (rs4810485, P = 1.4×10(-9. Second, we demonstrate that subjects homozygous for the RA risk allele have ∼33% more CD40 on the surface of primary human CD19+ B lymphocytes than subjects homozygous for the non-risk allele (P = 10(-9, a finding corroborated by expression quantitative trait loci (eQTL analysis in peripheral blood mononuclear cells from 1,469 healthy control individuals. Third, we use retroviral shRNA infection to perturb the amount of CD40 on the surface of a human B lymphocyte cell line (BL2 and observe a direct correlation between amount of CD40 protein and phosphorylation of RelA (p65, a subunit of the NF-κB transcription factor. Finally, we develop a high-throughput NF-κB luciferase reporter assay in BL2 cells activated with trimerized CD40 ligand (tCD40L and conduct an HTS of 1,982 chemical compounds and FDA-approved drugs. After a series of counter-screens and testing in primary human CD19+ B cells, we identify 2 novel chemical inhibitors not previously implicated in inflammation or CD40-mediated NF-κB signaling. Our study demonstrates proof-of-concept that human genetics can be used to guide the development of phenotype-based, high-throughput small-molecule screens to identify potential novel

  14. Human Genetics in Rheumatoid Arthritis Guides a High-Throughput Drug Screen of the CD40 Signaling Pathway

    Science.gov (United States)

    Li, Gang; Diogo, Dorothée; Wu, Di; Spoonamore, Jim; Dancik, Vlado; Franke, Lude; Kurreeman, Fina; Rossin, Elizabeth J.; Duclos, Grant; Hartland, Cathy; Zhou, Xuezhong; Li, Kejie; Liu, Jun; De Jager, Philip L.; Siminovitch, Katherine A.; Zhernakova, Alexandra; Raychaudhuri, Soumya; Bowes, John; Eyre, Steve; Padyukov, Leonid; Gregersen, Peter K.; Worthington, Jane; Gupta, Namrata; Clemons, Paul A.; Stahl, Eli; Tolliday, Nicola; Plenge, Robert M.

    2013-01-01

    Although genetic and non-genetic studies in mouse and human implicate the CD40 pathway in rheumatoid arthritis (RA), there are no approved drugs that inhibit CD40 signaling for clinical care in RA or any other disease. Here, we sought to understand the biological consequences of a CD40 risk variant in RA discovered by a previous genome-wide association study (GWAS) and to perform a high-throughput drug screen for modulators of CD40 signaling based on human genetic findings. First, we fine-map the CD40 risk locus in 7,222 seropositive RA patients and 15,870 controls, together with deep sequencing of CD40 coding exons in 500 RA cases and 650 controls, to identify a single SNP that explains the entire signal of association (rs4810485, P = 1.4×10−9). Second, we demonstrate that subjects homozygous for the RA risk allele have ∼33% more CD40 on the surface of primary human CD19+ B lymphocytes than subjects homozygous for the non-risk allele (P = 10−9), a finding corroborated by expression quantitative trait loci (eQTL) analysis in peripheral blood mononuclear cells from 1,469 healthy control individuals. Third, we use retroviral shRNA infection to perturb the amount of CD40 on the surface of a human B lymphocyte cell line (BL2) and observe a direct correlation between amount of CD40 protein and phosphorylation of RelA (p65), a subunit of the NF-κB transcription factor. Finally, we develop a high-throughput NF-κB luciferase reporter assay in BL2 cells activated with trimerized CD40 ligand (tCD40L) and conduct an HTS of 1,982 chemical compounds and FDA–approved drugs. After a series of counter-screens and testing in primary human CD19+ B cells, we identify 2 novel chemical inhibitors not previously implicated in inflammation or CD40-mediated NF-κB signaling. Our study demonstrates proof-of-concept that human genetics can be used to guide the development of phenotype-based, high-throughput small-molecule screens to identify potential novel therapies in

  15. Genetic screening technology: ethical issues in access to tests by employers and health insurance committees.

    Science.gov (United States)

    Faden, R R; Kass, N E

    1993-01-01

    Whereas the introduction of new technologies previously has raised the ethical question of who ought to have access to a new procedure or device, genetic testing technology raises the new ethical question of to whom access to a new technology ought to be limited. In this article we discuss the implications of employers and private health insurance companies having access to genetic testing technology. Although there may be legitimate business interests in allowing employers and insurers to conduct genetic screening, there are other valid societal interests in regulating or limiting the use of this technology by third parties. Public policy developed in the area of new genetic technology must reflect such interests. PMID:17165239

  16. Chinese geneticists' views of ethical issues in genetic testing and screening: evidence for eugenics in China.

    Science.gov (United States)

    Mao, X

    1998-09-01

    To identify Chinese geneticists' views of ethical issues in genetic testing and screening, a national survey was conducted. Of 402 Chinese geneticists asked to participate, 255 (63%) returned by mail anonymous questionnaires. The majority of respondents thought that genetic testing should be offered in the workplace for alpha-antitrypsin deficiency (95%) and the predisposition of executives to heart disease, cancer, and diabetes (94%); that genetic testing should be included in preemployment physical examinations (86%); that governments should require premarital carrier tests (86%), newborn screening for sickle cell (77%), and Duchenne muscular dystrophy (71%); and that children should be tested for genes for late-onset disorders such as Huntington disease (85%), susceptibility to cancers (85%), familial hypercholesterolemia (84%), alcoholism (69%), and Alzheimer disease (61%). Most believed that partners should know each other's genetic status before marriage (92%), that carriers of the same defective gene should not mate with each other (91%), and that women should have a prenatal diagnosis if medically indicated (91%). The majority said that in China decisions about family planning were shared by the couple (82%). More than half had views that, in China, there were no laws to prohibit disability discrimination (64%), particularly to protect people with adult polycystic kidney disease (57%), cystic fibrosis (56%), or genetic predisposition to other diseases (50%). To some extent, these results might provide a basis for a discussion of eugenics in China, particularly about China's Maternal and Infant Health Care Law (1994). PMID:9718350

  17. Screening of perfluorinated chemicals (PFCs) in various aquatic organisms

    Energy Technology Data Exchange (ETDEWEB)

    Fernandez-Sanjuan, Maria; Meyer, Johan; Damasio, Joana; Faria, Melissa; Barata, Carlos; Lacorte, Silvia [IDAEA-CSIC, Department of Environmental Chemistry, Barcelona (Spain)

    2010-10-15

    The aim of this study was to evaluate the occurrence of five perfluorinated chemicals (perfluorooctane sulfonic acid (PFOS), perfluorooctanoic acid (PFOA), perfluorononanoic acid (PFNA), perfluorohexane sulfonic acid (PFHxS), and perfluorobutane sulfonic acid) in aquatic organisms dwelling in either freshwater or marine ecosystems. Organisms selected were insect larvae, oysters, zebra mussels, sardines, and crabs, which are widespread in the environment and may represent potential bioindicators of exposure to PFCs. The study comprises the optimization of a solid-liquid extraction method and determination by high-performance liquid chromatography coupled to tandem mass spectrometry. Using spiked zebra mussels at 10 and 100 ng/g level, the method developed provided recoveries of 96% and 122%, and 82% to 116%, respectively, and a limit of detection between 0.07 and 0.22 ng/g ww. The method was highly sensitivity and robust to determine PFC compounds in a wide array of biological matrices, and no matrix interferents nor blank contamination was observed. Among organisms studied, none of the bivalves accumulated PFCs, and contrarily, insect larvae, followed by fish and crabs contained levels ranging from 0.23 to 144 ng/g ww of PFOS, from 0.14 to 4.3 ng/g ww of PFOA, and traces of PFNA and PFHxS. Assessment of the potential use of aquatic organisms for biomonitoring studies is further discussed. (orig.)

  18. Cyanobactins from Cyanobacteria: Current Genetic and Chemical State of Knowledge.

    Science.gov (United States)

    Martins, Joana; Vasconcelos, Vitor

    2015-11-01

    Cyanobacteria are considered to be one of the most promising sources of new, natural products. Apart from non-ribosomal peptides and polyketides, ribosomally synthesized and post-translationally modified peptides (RiPPs) are one of the leading groups of bioactive compounds produced by cyanobacteria. Among these, cyanobactins have sparked attention due to their interesting bioactivities and for their potential to be prospective candidates in the development of drugs. It is assumed that the primary source of cyanobactins is cyanobacteria, although these compounds have also been isolated from marine animals such as ascidians, sponges and mollusks. The aim of this review is to update the current knowledge of cyanobactins, recognized as being produced by cyanobacteria, and to emphasize their genetic clusters and chemical structures as well as their bioactivities, ecological roles and biotechnological potential.

  19. Genome-wide Screening Reveals the Genetic Determinants of an Antibiotic Insecticide in Bacillus thuringiensis*

    OpenAIRE

    Liu, Xiao-Yan; Ruan, Li-Fang; Hu, Zhen-Fei; Peng, Dong-hai; Cao, Shi-Yun; Yu, Zi-Niu; Liu, Yao; Zheng, Jin-Shui; Sun, Ming

    2010-01-01

    Thuringiensin is a thermostable secondary metabolite in Bacillus thuringiensis and has insecticidal activity against a wide range of insects. Until now, the regulatory mechanisms and genetic determinants involved in thuringiensin production have remained unclear. Here, we successfully used heterologous expression-guided screening in an Escherichia coli–Bacillus thuringiensis shuttle bacterial artificial chromosome library, to clone the intact thuringiensin synthesis (thu) cluster. Then the th...

  20. A Zebrafish Genetic Screen Identifies Neuromedin U as a Regulator of Sleep/Wake States

    OpenAIRE

    Chiu, Cindy N.; Rihel, Jason; Lee, Daniel A.; Singh, Chanpreet; Mosser, Eric A.; Chen, Shijia; Sapin, Viveca; Pham, Uyen; Engle, Jae; Niles, Brett J.; Montz, Christin J.; Chakravarthy, Sridhara; Zimmerman, Steven; Salehi-Ashtiani, Kourosh; Vidal, Marc

    2016-01-01

    Neuromodulation of arousal states ensures that an animal appropriately responds to its environment and engages in behaviors necessary for survival. However, the molecular and circuit properties underlying neuromodulation of arousal states such as sleep and wakefulness remain unclear. To tackle this challenge in a systematic and unbiased manner, we performed a genetic overexpression screen to identify genes that affect larval zebrafish arousal. We found that the neuropeptide neuromedin U (Nmu)...

  1. Statement of The American Society of Human Genetics on cystic fibrosis carrier screening

    Energy Technology Data Exchange (ETDEWEB)

    1992-12-01

    The identification in 1989 of the cystic fibrosis (CF) gene and its most common mutation immediately raised the possibility of CF carrier detection by DNA analysis. The American Society of Human Genetics (ASHG) issued a statement recommending that CF carrier testing should be made available to individuals with a family history of CF. It was also stated that screening of individuals or couples in the general population should not be offered until the rate of CF carrier detection improves. An additional prerequisite emphasized the need for the establishment of effective educational and counseling programs consistent with previous widely accepted principles. An NIH workshop reached similar conclusions. ASHG recommendations are that screening be limited to individuals with a family history of CF, testing should be accompanied by education and counseling, screening should be voluntary and confidential with appropriate laboratory quality controls, and efforts should be expanded to educate health care providers and the public.

  2. Preimplantation Genetic Screening: An Effective Testing for Infertile and Repeated Miscarriage Patients?

    Directory of Open Access Journals (Sweden)

    Ning Wang

    2010-01-01

    Full Text Available Aneuploidy in pregnancy is known to increase with advanced maternal age (AMA and associate with repeated implantation failure (RIF, and repeated miscarriage (RM. Preimplantation genetic screening (PGS has been introduced into clinical practice, screening, and eliminating aneuploidy embryos, which can improve the chance of conceptions for infertility cases with poor prognosis. These patients are a good target group to assess the possible benefit of aneuploidy screening. Although practiced widely throughout the world, there still exist some doubts about the efficacy of this technique. Recent randomized trials were not as desirable as we expected, suggesting that PGS needs to be reconsidered. The aim of this review is to discuss the efficacy of PGS.

  3. "It gives them more options": preferences for preconception genetic carrier screening for fragile X syndrome in primary healthcare.

    Science.gov (United States)

    Archibald, Alison D; Hickerton, Chriselle L; Wake, Samantha A; Jaques, Alice M; Cohen, Jonathan; Metcalfe, Sylvia A

    2016-04-01

    This study aims to explore stakeholder views about offering population-based genetic carrier screening for fragile X syndrome. A qualitative study using interviews and focus groups with stakeholders was undertaken to allow for an in-depth exploration of views and perceptions about practicalities of, and strategies for, offering carrier screening for fragile X syndrome to the general population in healthcare settings. A total of 188 stakeholders took part including healthcare providers (n = 81), relatives of people with fragile X syndrome (n = 29), and members of the general community (n = 78). The importance of raising community awareness about screening and providing appropriate support for carriers was emphasized. There was a preference for preconception carrier screening and for providing people with the opportunity to make an informed decision about screening. Primary care was highlighted as a setting which would ensure screening is accessible; however, challenges of offering screening in primary care were identified including time to discuss screening, knowledge about the test and possible outcomes, and the health professionals' approach to offering screening. With the increasing availability of genetic carrier tests, it is essential that research now focuses on evaluating approaches for the delivery of carrier screening programs. Primary healthcare is perceived as an appropriate setting through which to access the target population, and raising awareness is essential to making genetic screening more accessible to the general community. PMID:26842720

  4. Data-driven prioritization of chemicals for various water types using suspect screening LC-HRMS

    NARCIS (Netherlands)

    Sjerps, Rosa M.A.; Vughs, Dennis; van Leerdam, Jan A.; ter Laak, Thomas L.; van Wezel, Annemarie P.

    2016-01-01

    For the prioritization of more than 5200 anthropogenic chemicals authorized on the European market, we use a large scale liquid chromatography-high resolution mass spectrometry (LC-HRMS) suspect screening study. The prioritization is based on occurrence in 151 water samples including effluent, surfa

  5. Recommendations for Developing Alternative Test Methods for Screening and Prioritization of Chemicals for Developmental Neurotoxicity

    Science.gov (United States)

    Developmental neurotoxicity testing (DNT) is perceived by many stakeholders to be an area in critical need of alternative methods to current animal testing protocols and gUidelines. An immediate goal is to develop test methods that are capable of screening large numbers of chemic...

  6. Postmortem genetic screening for the identification, verification, and reporting of genetic variants contributing to the sudden death of the young.

    Science.gov (United States)

    Methner, D Nicole R; Scherer, Steven E; Welch, Katherine; Walkiewicz, Magdalena; Eng, Christine M; Belmont, John W; Powell, Mark C; Korchina, Viktoriya; Doddapaneni, Harsha Vardhan; Muzny, Donna M; Gibbs, Richard A; Wolf, Dwayne A; Sanchez, Luis A; Kahn, Roger

    2016-09-01

    Each year in the United States, thousands of cases of sudden and unexpected deaths of infants, children, and young adults are assigned an undetermined cause of death after postmortem investigation and autopsy. Heritable genetic variants have been suggested as the cause of up to a third of sudden death (SD) cases. Elucidation of the genetic variants involved in SD cases is important to not only help establish cause and manner of death of these individuals, but to also aid in determining whether familial genetic testing should be considered. Previously, these types of postmortem screenings have not been a feasible option for most county medical examiners' and coroners' offices. We sequenced full exons of 64 genes associated with SD in the largest known cohort (351) of infant and young SD decedents using massively parallel sequencing at 1 yr of age), were found to have a reportable genetic variant contributing to SD. These percentages represent an estimate lower than those previously reported. Overall yields and results likely vary between studies due to differences in evaluation techniques and reporting. Additionally, we recommend ongoing assessment of data, including nonreported novel variants, as technology and literature continually advance. This study demonstrates a strategy to implement molecular autopsies in medicolegal investigations of young SD decedents. PMID:27435932

  7. Understanding undergraduate nursing students’ knowledge of, and attitudes to, prenatal genetic screening in Malaysia: A preliminary study

    Directory of Open Access Journals (Sweden)

    Lean Keng Soon

    2013-03-01

    Full Text Available Background: Nursing students must be able to ensure that patients understand important health screening information. However, studies have reported a deficit in nursing students’ knowledge of genetics and genetic disorders. Aim: The purpose of this study was to investigate nursing students’ knowledge of, and attitudes to prenatal genetic screening. Methods: A descriptive, cross-sectional survey was conducted with final year undergraduate nursing students recruited from a tertiary teaching hospital in Kelantan, Malaysia. Thirty-four of the 37 students returned a self-administered questionnaire (response rate=97% that assessed knowledge of, and attitudes to, prenatal genetic screening. Findings: The majority of the respondents demonstrated an adequate level of knowledge regarding prenatal genetic screening (65%, and 62% were more supportive than others of screening. Approximately one third (35% had limited knowledge. Simple linear regression analysis uncovered a significant linear relationship between the attitude score and the knowledge score (R2=0.48, P<0.05, 95% confidence interval. Conclusions: These findings suggest the need to improve and upgrade the nursing education strategy concerning genetics. The deficit in nursing students’ knowledge of prenatal genetic screening may result in inappropriate counselling, health teaching and referrals, which can have serious repercussions. 

  8. Chemical genetics suggests a critical role for lysyl oxidase in zebrafish notochord morphogenesis.

    Science.gov (United States)

    Anderson, Carrie; Bartlett, Stephen J; Gansner, John M; Wilson, Duncan; He, Ling; Gitlin, Jonathan D; Kelsh, Robert N; Dowden, James

    2007-01-01

    As a result of a chemical genetic screen for modulators of metalloprotease activity, we report that 2-mercaptopyridine-N-oxide induces a conspicuous undulating notochord defect in zebrafish embryos, a phenocopy of the leviathan mutant. The location of the chemically-induced wavy notochord correlated with the timing of application, thus defining a narrow chemical sensitivity window during segmentation stages. Microscopic observations revealed that notochord undulations appeared during the phase of notochord cell vacuolation and notochord elongation. Notochord cells become swollen as well as disorganized, while electron microscopy revealed disrupted organization of collagen fibrils in the surrounding sheath. We demonstrate by assay in zebrafish extracts that 2-mercaptopyridine-N-oxide inhibits lysyl oxidase. Thus, we provide insight into notochord morphogenesis and reveal novel compounds for lysyl oxidase inhibition. Taken together, these data underline the utility of small molecules for elucidating the dynamic mechanisms of early morphogenesis and provide a potential explanation for the recently established role of copper in zebrafish notochord formation.

  9. A Fluorescence-Based Genetic Screen to Study Retinal Degeneration in Drosophila.

    Directory of Open Access Journals (Sweden)

    Yu Huang

    Full Text Available The Drosophila visual system has been proved to be a powerful genetic model to study eye disease such as retinal degeneration. Here, we describe a genetic method termed "Rh1::GFP ey-flp/hid" that is based on the fluorescence of GFP-tagged major rhodopsin Rh1 in the eyes of living flies and can be used to monitor the integrity of photoreceptor cells. Through combination of this method and ERG recording, we examined a collection of 667 mutants and identified 18 genes that are required for photoreceptor cell maintenance, photoresponse, and rhodopsin synthesis. Our findings demonstrate that this "Rh1::GFP ey-flp/hid" method enables high-throughput F1 genetic screens to rapidly and precisely identify mutations of retinal degeneration.

  10. Chemical compatibility screening results of plastic packaging to mixed waste simulants

    Energy Technology Data Exchange (ETDEWEB)

    Nigrey, P.J.; Dickens, T.G.

    1995-12-01

    We have developed a chemical compatibility program for evaluating transportation packaging components for transporting mixed waste forms. We have performed the first phase of this experimental program to determine the effects of simulant mixed wastes on packaging materials. This effort involved the screening of 10 plastic materials in four liquid mixed waste simulants. The testing protocol involved exposing the respective materials to {approximately}3 kGy of gamma radiation followed by 14 day exposures to the waste simulants of 60 C. The seal materials or rubbers were tested using VTR (vapor transport rate) measurements while the liner materials were tested using specific gravity as a metric. For these tests, a screening criteria of {approximately}1 g/m{sup 2}/hr for VTR and a specific gravity change of 10% was used. It was concluded that while all seal materials passed exposure to the aqueous simulant mixed waste, EPDM and SBR had the lowest VTRs. In the chlorinated hydrocarbon simulant mixed waste, only VITON passed the screening tests. In both the simulant scintillation fluid mixed waste and the ketone mixture simulant mixed waste, none of the seal materials met the screening criteria. It is anticipated that those materials with the lowest VTRs will be evaluated in the comprehensive phase of the program. For specific gravity testing of liner materials the data showed that while all materials with the exception of polypropylene passed the screening criteria, Kel-F, HDPE, and XLPE were found to offer the greatest resistance to the combination of radiation and chemicals.

  11. Screening of heavy quark free energies at finite temperature and non-zero baryon chemical potential

    CERN Document Server

    Döring, M; Kaczmarek, O; Karsch, Frithjof; Laermann, E

    2006-01-01

    We analyze the dependence of heavy quark free energies on the baryon chemical potential (mu_b) in 2-flavour QCD using improved (p4) staggered fermions with a bare quark mass of m/T = 0.4. By performing a 6th order Taylor expansion in the chemical potential which circumvents the sign problem. The Taylor expansion coefficients of colour singlet and colour averaged free energies are calculated and from this the expansion coefficients for the corresponding screening masses are determined. We find that for small mu_b the free energies of a static quark anti-quark pair decrease in a medium with a net excess of quarks and that screening is well described by a screening mass which increases with increasing mu_b. The mu_b-dependent corrections to the screening masses are well described by perturbation theory for T > 2 T_c. In particular, we find for all temperatures above T_c that the expansion coefficients for singlet and colour averaged screening masses differ by a factor 2.

  12. Molecular, chemical and biological screening of soil actinomycete isolates in seeking bioactive peptide metabolites

    Directory of Open Access Journals (Sweden)

    Javad Hamedi

    2015-10-01

    Full Text Available Background and Objective: Due to the evolution of multidrug-resistant strains, screening of natural resources, especially actinomycetes, for new therapeutic agents discovery has become the interests of researchers. In this study, molecular, chemical and biological screening of soil actinomycetes was carried out in order to search for peptide-producing actinomycetes.Materials and Methods: 60 actinomycetes were isolated from soils of Iran. The isolates were subjected to molecular screening for detection NRPS (non-ribosomal peptide synthetases gene. Phylogenic identification of NRPS containing isolates was performed. Chemical screening of the crude extracts was performed using chlorine o-dianisidine as peptide detector reagent and bioactivity of peptide producing strains was determined by antimicrobial bioassay. High pressure liquid chromatography- mass spectrometry (HPLC-MS with UV-visible spectroscopy was performed for detection of the metabolite diversity in selected strain.Results: Amplified NRPS adenylation gene (700 bp was detected among 30 strains. Phylogenic identification of these isolates showed presence of rare actinomycetes genera among the isolates and 10 out of 30 strains were subjected to chemical screening. Nocardia sp. UTMC 751 showed antimicrobial activity against bacterial and fungal test pathogens. HPLC-MSand UV-visible spectroscopy results from the crude extract showed that this strain has probably the ability to produce new metabolites.Conclusion: By application of a combined approach, including molecular, chemical and bioactivity analysis, a promising strain of Nocardia sp. UTMC 751 was obtained. This strain had significant activity against Staphylococcus aureus and Pseudomonas aeruginosa. Strain Nocardia sp. UTMC 751 produce five unknown and most probably new metabolites with molecular weights of 274.2, 390.3, 415.3, 598.4 and 772.5. This strain had showed 99% similarity to Nocardia ignorata DSM 44496 T.

  13. Maximising the efficiency of clinical screening programmes: balancing predictive genetic testing with a right not to know.

    Science.gov (United States)

    Schuurman, Agnes G; van der Kolk, Dorina M; Verkerk, Marian A; Birnie, Erwin; Ranchor, Adelita V; Plantinga, Mirjam; van Langen, Irene M

    2015-09-01

    We explored the dilemma between patients' right not to know their genetic status and the efficient use of health-care resources in the form of clinical cancer screening programmes. Currently, in the Netherlands, 50% risk carriers of heritable cancer syndromes who choose not to know their genetic status have access to the same screening programmes as proven mutation carriers. This implies an inefficient use of health-care resources, because half of this group will not carry the familial mutation. At the moment, only a small number of patients are involved; however, the expanding possibilities for genetic risk profiling means this issue must be addressed because of potentially adverse societal and financial impact. The trade-off between patients' right not to know their genetic status and efficient use of health-care resources was discussed in six focus groups with health-care professionals and patients from three Dutch university hospitals. Professionals prefer patients to undergo a predictive DNA test as a prerequisite for entering cancer screening programmes. Professionals prioritise treating sick patients or proven mutation carriers over screening untested individuals. Participation in cancer screening programmes without prior DNA testing is, however, supported by most professionals, as testing is usually delayed and relatively few patients are involved at present. Reducing the number of 50% risk carriers undergoing screening is expected to be achieved by: offering more psychosocial support, explaining the iatrogenic risks of cancer screening, increasing out-of-pocket costs, and offering a less stringent screening programme for 50% risk carriers. PMID:25564039

  14. Maximising the efficiency of clinical screening programmes: balancing predictive genetic testing with a right not to know

    Science.gov (United States)

    Schuurman, Agnes G; van der Kolk, Dorina M; Verkerk, Marian A; Birnie, Erwin; Ranchor, Adelita V; Plantinga, Mirjam; van Langen, Irene M

    2015-01-01

    We explored the dilemma between patients' right not to know their genetic status and the efficient use of health-care resources in the form of clinical cancer screening programmes. Currently, in the Netherlands, 50% risk carriers of heritable cancer syndromes who choose not to know their genetic status have access to the same screening programmes as proven mutation carriers. This implies an inefficient use of health-care resources, because half of this group will not carry the familial mutation. At the moment, only a small number of patients are involved; however, the expanding possibilities for genetic risk profiling means this issue must be addressed because of potentially adverse societal and financial impact. The trade-off between patients' right not to know their genetic status and efficient use of health-care resources was discussed in six focus groups with health-care professionals and patients from three Dutch university hospitals. Professionals prefer patients to undergo a predictive DNA test as a prerequisite for entering cancer screening programmes. Professionals prioritise treating sick patients or proven mutation carriers over screening untested individuals. Participation in cancer screening programmes without prior DNA testing is, however, supported by most professionals, as testing is usually delayed and relatively few patients are involved at present. Reducing the number of 50% risk carriers undergoing screening is expected to be achieved by: offering more psychosocial support, explaining the iatrogenic risks of cancer screening, increasing out-of-pocket costs, and offering a less stringent screening programme for 50% risk carriers. PMID:25564039

  15. Identification of common genetic modifiers of neurodegenerative diseases from an integrative analysis of diverse genetic screens in model organisms

    Directory of Open Access Journals (Sweden)

    Chen Xi

    2012-02-01

    Full Text Available Abstract Background An array of experimental models have been developed in the small model organisms C. elegans, S. cerevisiae and D. melanogaster for the study of various neurodegenerative diseases including Alzheimer's disease, Parkinson's disease, and expanded polyglutamine diseases as exemplified by Huntington's disease (HD and related ataxias. Genetic approaches to determine the nature of regulators of the disease phenotypes have ranged from small scale to essentially whole genome screens. The published data covers distinct models in all three organisms and one important question is the extent to which shared genetic factors can be uncovered that affect several or all disease models. Surprisingly it has appeared that there may be relatively little overlap and that many of the regulators may be organism or disease-specific. There is, however, a need for a fully integrated analysis of the available genetic data based on careful comparison of orthologues across the species to determine the real extent of overlap. Results We carried out an integrated analysis using C. elegans as the baseline model organism since this is the most widely studied in this context. Combination of data from 28 published studies using small to large scale screens in all three small model organisms gave a total of 950 identifications of genetic regulators. Of these 624 were separate genes with orthologues in C. elegans. In addition, 34 of these genes, which all had human orthologues, were found to overlap across studies. Of the common genetic regulators some such as chaperones, ubiquitin-related enzymes (including the E3 ligase CHIP which directly links the two pathways and histone deacetylases were involved in expected pathways whereas others such as the peroxisomal acyl CoA-oxidase suggest novel targets for neurodegenerative disease therapy Conclusions We identified a significant number of overlapping regulators of neurodegenerative disease models. Since the diseases

  16. Reevaluation of 1999 Health-Based Environmental Screening Levels (HBESLs) for Chemical Warfare Agents

    Energy Technology Data Exchange (ETDEWEB)

    Watson, Annetta Paule [ORNL; Dolislager, Fredrick G [ORNL

    2007-05-01

    This report evaluates whether new information and updated scientific models require that changes be made to previously published health-based environmental soil screening levels (HBESLs) and associated environmental fate/breakdown information for chemical warfare agents (USACHPPM 1999). Specifically, the present evaluation describes and compares changes that have been made since 1999 to U.S. Environmental Protection Agency (EPA) risk assessment models, EPA exposure assumptions, as well as to specific chemical warfare agent parameters (e.g., toxicity values). Comparison was made between screening value estimates recalculated with current assumptions and earlier health-based environmental screening levels presented in 1999. The chemical warfare agents evaluated include the G-series and VX nerve agents and the vesicants sulfur mustard (agent HD) and Lewisite (agent L). In addition, key degradation products of these agents were also evaluated. Study findings indicate that the combined effect of updates and/or changes to EPA risk models, EPA default exposure parameters, and certain chemical warfare agent toxicity criteria does not result in significant alteration to the USACHPPM (1999) health-based environmental screening level estimates for the G-series and VX nerve agents or the vesicant agents HD and L. Given that EPA's final position on separate Tier 1 screening levels for indoor and outdoor worker screening assessments has not yet been released as of May 2007, the study authors find that the 1999 screening level estimates (see Table ES.1) are still appropriate and protective for screening residential as well as nonresidential sites. As such, risk management decisions made on the basis of USACHPPM (1999) recommendations do not require reconsideration. While the 1999 HBESL values are appropriate for continued use as general screening criteria, the updated '2007' estimates (presented below) that follow the new EPA protocols currently under development

  17. Y-STR genetic screening by high-resolution melting analysis.

    Science.gov (United States)

    Deng, J Q; Liu, B Q; Wang, Y; Liu, W; Cai, J F; Long, R; Li, W H

    2016-01-01

    Currently, the widely used automated capillary electrophoresis-based short tandem repeat (STR) genotyping method for genetic screening in forensic practice is laborious, time-consuming, expensive, and technically challenging in some cases. Thus, new molecular-based strategies for conclusively identifying forensically relevant biological evidence are required. Here, we used high-resolution melting analysis (HRM) for Y-chromosome STR genotyping for forensic genetic screening. The reproducibility of the melting profile over dilution, sensitivity, discrimination power, and other factors was preliminarily studied in 10 Y-STR loci. The results showed that HRM-based approaches revealed more genotypes (compared to capillary electrophoresis), showed higher uniformity in replicate tests and diluted samples, and enabled successful detection of DNA at concentrations as low as 0.25 ng. For mixed samples, the melting curve profiles discriminated between mixed samples based on reference samples with high efficiency. The triplex Y-chromosome STR HRM assay was performed and provided a foundation for further studies such as a multiplex HRM assay. The HRM approach is a one-step application and the entire procedure can be completed within 2 h at a low cost. In conclusion, our findings demonstrate that the HRM-based Y-STR assay is a useful screening tool that can be used in forensic practice. PMID:26909950

  18. Mining Natural-Products Screening Data for Target-Class Chemical Motifs.

    Science.gov (United States)

    Coma, Isabel; Bandyopadhyay, Deepak; Diez, Emilio; Ruiz, Emilio Alvarez; de los Frailes, Maria Teresa; Colmenarejo, Gonzalo

    2014-06-01

    In this article, we describe two complementary data-mining approaches used to characterize the GlaxoSmithKline (GSK) natural-products set (NPS) based on information from the high-throughput screening (HTS) databases. Both methods rely on the aggregation and analysis of a large set of single-shot screening data for a number of biological assays, with the goal to reveal natural-product chemical motifs. One of them is an established method based on the data-driven clustering of compounds using a wide range of descriptors,(1)whereas the other method partitions and hierarchically clusters the data to identify chemical cores.(2,3)Both methods successfully find structural scaffolds that significantly hit different groups of discrete drug targets, compared with their relative frequency of demonstrating inhibitory activity in a large number of screens. We describe how these methods can be applied to unveil hidden information in large single-shot HTS data sets. Applied prospectively, this type of information could contribute to the design of new chemical templates for drug-target classes and guide synthetic efforts for lead optimization of tractable hits that are based on natural-product chemical motifs. Relevant findings for 7TM receptors (7TMRs), ion channels, class-7 transferases (protein kinases), hydrolases, and oxidoreductases will be discussed. PMID:24518065

  19. A neuron-based screening platform for optimizing genetically-encoded calcium indicators.

    Directory of Open Access Journals (Sweden)

    Trevor J Wardill

    Full Text Available Fluorescent protein-based sensors for detecting neuronal activity have been developed largely based on non-neuronal screening systems. However, the dynamics of neuronal state variables (e.g., voltage, calcium, etc. are typically very rapid compared to those of non-excitable cells. We developed an electrical stimulation and fluorescence imaging platform based on dissociated rat primary neuronal cultures. We describe its use in testing genetically-encoded calcium indicators (GECIs. Efficient neuronal GECI expression was achieved using lentiviruses containing a neuronal-selective gene promoter. Action potentials (APs and thus neuronal calcium levels were quantitatively controlled by electrical field stimulation, and fluorescence images were recorded. Images were segmented to extract fluorescence signals corresponding to individual GECI-expressing neurons, which improved sensitivity over full-field measurements. We demonstrate the superiority of screening GECIs in neurons compared with solution measurements. Neuronal screening was useful for efficient identification of variants with both improved response kinetics and high signal amplitudes. This platform can be used to screen many types of sensors with cellular resolution under realistic conditions where neuronal state variables are in relevant ranges with respect to timing and amplitude.

  20. Testing for direct genetic effects using a screening step in family-based association studies

    Directory of Open Access Journals (Sweden)

    Sharon M Lutz

    2013-11-01

    Full Text Available In genome wide association studies (GWAS, families based studies tend to have less power to detect genetic associations than population based studies, such as case-control studies. This can be an issue when testing if genes in a family based GWAS have a direct effect on the phenotype of interest or if the genes act indirectly through a secondary phenotype. When multiple SNPs are tested for a direct effect in the family based study, a screening step can be used to minimize the burden of multiple comparisons in the causal analysis. We propose a 2-stage screening step that can be incorporated into the family based association test (FBAT approach similar to the conditional mean model approach in the VanSteen-algorithm [1]. Simulations demonstrate that the type 1 error is preserved and this method is advantageous when multiple markers are tested. This method is illustrated by an application to the Framingham Heart Study.

  1. Genetically modified organisms in food-screening and specific detection by polymerase chain reaction.

    Science.gov (United States)

    Vollenhofer, S; Burg, K; Schmidt, J; Kroath, H

    1999-12-01

    PCR methods for the detection of genetically modified organisms (GMOs) were developed that can be used for screening purposes and for specific detection of glyphosate-tolerant soybean and insect-resistant maize in food. Primers were designed to amplify parts of the 35S promoter derived from Cauliflower Mosaic Virus, the NOS terminator derived from Agrobacterium tumefaciens and the antibiotic marker gene NPTII (neomycin-phosphotransferase II), to allow for general screening of foods. PCR/hybridization protocols were established for the detection of glyphosate-tolerant RoundUp Ready soybean and insect-resistant Bt-maize. Besides hybridization, confirmation of the results using restriction analysis was also possible. The described methods enabled a highly sensitive and specific detection of GMOs and thus provide a useful tool for routine analysis of raw and processed food products.

  2. Large-Scale Forward Genetic Screening Analysis of Development of Hematopoiesis in Zebrafish

    Institute of Scientific and Technical Information of China (English)

    Kun Wang; Ning Ma; Yiyue Zhang; Wenqing Zhang; Zhibin Huang; Lingfeng Zhao; Wei Liu; Xiaohui Chen; Ping Meng; Qing Lin; Yali Chi; Mengchang Xu

    2012-01-01

    Zebrafish is a powerful model for the investigation of hematopoiesis.In order to isolate novel mutants with hematopoietic defects,large-scale mutagenesis screening of zebrafish was performed.By scoring specific hematopoietic markers,52 mutants were identified and then classified into four types based on specific phenotypic traits.Each mutant represented a putative mutation of a gene regulating the relevant aspect of hematopoiesis,including early macrophage development,early granulopoiesis,embryonic myelopoiesis,and definitive erythropoiesis/lymphopoiesis.Our method should be applicable for other types of genetic screening in zebrafish.In addition,further study of the mutants we identified may help to unveil the molecular basis of hematopoiesis.

  3. Genetic screens and functional genomics using CRISPR/Cas9 technology.

    Science.gov (United States)

    Hartenian, Ella; Doench, John G

    2015-04-01

    Functional genomics attempts to understand the genome by perturbing the flow of information from DNA to RNA to protein, in order to learn how gene dysfunction leads to disease. CRISPR/Cas9 technology is the newest tool in the geneticist's toolbox, allowing researchers to edit DNA with unprecedented ease, speed and accuracy, and representing a novel means to perform genome-wide genetic screens to discover gene function. In this review, we first summarize the discovery and characterization of CRISPR/Cas9, and then compare it to other genome engineering technologies. We discuss its initial use in screening applications, with a focus on optimizing on-target activity and minimizing off-target effects. Finally, we comment on future challenges and opportunities afforded by this technology.

  4. Screening for genetic haemochromatosis in blood samples with raised alanine aminotransferase

    OpenAIRE

    Bhavnani, M; Lloyd, D; Bhattacharyya, A.; Marples, J; Elton, P; Worwood, M.

    2000-01-01

    BACKGROUND—In the UK approximately 1 in 140 people are homozygous for the C282Y mutation of the HFE gene and are at risk from iron overload caused by genetic haemochromatosis (GH). Early detection can prevent organ damage secondary to iron deposition and increase life expectancy.
AIM—To screen for GH in all blood samples sent to the laboratory for routine liver function tests in which raised serum alanine aminotransferase (ALT) activity was detected.
METHODS—ALT was measured in sera sent to t...

  5. Screening of genetic parameters for soluble protein expression in Escherichia coli

    DEFF Research Database (Denmark)

    Vernet, Erik; Kotzsch, Alexander; Voldborg, Bjørn;

    2011-01-01

    . Here we present a screening strategy for expression of biomedically relevant proteins in Escherichia coli using a panel of six different genetic variations. These include engineered strains for rare codon supplementation, increased disulfide bond formation in the cytoplasm and novel vectors...... for secretion to the periplasm or culture medium. Combining these variants with expression construct truncations design, we report on parallel cloning and expression of more than 300 constructs representing 24 selected proteins; including full-length variants of human growth factors, interleukins and growth...

  6. A chemical screen identifies class A G-protein coupled receptors as regulators of cilia

    OpenAIRE

    Avasthi, Prachee; Marley, Aaron; Lin, Henry; Gregori-Puigjane, Elisabet; Shoichet, Brian K.; von Zastrow, Mark; Marshall, Wallace F.

    2012-01-01

    Normal cilia length and motility are critical for proper cellular function. Prior studies of the regulation of ciliary structure and length have primarily focused on the intraflagellar transport machinery and motor proteins required for ciliary assembly and disassembly. However, several mutants with abnormal length flagella highlight the importance of signaling proteins as well. In this study, an unbiased chemical screen was performed to uncover signaling pathways that are critical for ciliog...

  7. Risk-based high-throughput chemical screening and prioritization using exposure models and in vitro bioactivity assays

    DEFF Research Database (Denmark)

    Shin, Hyeong-Moo; Ernstoff, Alexi; Arnot, Jon;

    2015-01-01

    We present a risk-based high-throughput screening (HTS) method to identify chemicals for potential health concerns or for which additional information is needed. The method is applied to 180 organic chemicals as a case study. We first obtain information on how the chemical is used and identify...

  8. Screen for genetic modifiers of stbm reveals that photoreceptor fate and rotation can be genetically uncoupled in the Drosophila eye.

    Directory of Open Access Journals (Sweden)

    Tanya Wolff

    Full Text Available BACKGROUND: Polarity of the Drosophila compound eye arises primarily as a consequence of two events that are tightly linked in time and space: fate specification of two photoreceptor cells, R3 and R4, and the subsequent directional movement of the unit eyes of the compound eye, or ommatidia. While it is thought that these fates dictate the direction of ommatidial rotation, the phenotype of mutants in the genes that set up this polarity led to the hypothesis that these two events could be uncoupled. METHODOLOGY/PRINCIPAL FINDINGS: To definitively demonstrate these events are genetically separable, we conducted a dominant modifier screen to determine if genes, when misexpressed, could selectively enhance subclasses of mutant ommatidia in which the direction of rotation does not follow the R3/R4 cell fates, yet not affect the number of ommatidia in which rotation follows the R3/R4 cell fates. We identified a subset of P element lines that exhibit this selective enhancement. We also identified lines that behave in the opposite manner: They enhance the number of ommatidia that rotate in the right direction, but do not alter the number of ommatidia that rotate incorrectly with respect to the R3/R4 fates. CONCLUSIONS/SIGNIFICANCE: These results indicate that fate and direction of rotation can be genetically separated, and that there are genes that act between R3/R4 fate specification and direction of ommatidial rotation. These data affirm what has been a long-standing assumption about the genetic control of ommatidial polarity.

  9. A forward genetic screen reveals that calcium-dependent protein kinase 3 regulates egress in Toxoplasma.

    Directory of Open Access Journals (Sweden)

    Erin Garrison

    Full Text Available Egress from the host cell is a crucial and highly regulated step in the biology of the obligate intracellular parasite, Toxoplasma gondii. Active egress depends on calcium fluxes and appears to be a crucial step in escaping the attack from the immune system and, potentially, in enabling the parasites to shuttle into appropriate cells for entry into the brain of the host. Previous genetic screens have yielded mutants defective in both ionophore-induced egress and ionophore-induced death. Using whole genome sequencing of one mutant and subsequent analysis of all mutants from these screens, we find that, remarkably, four independent mutants harbor a mis-sense mutation in the same gene, TgCDPK3, encoding a calcium-dependent protein kinase. All four mutations are predicted to alter key regions of TgCDPK3 and this is confirmed by biochemical studies of recombinant forms of each. By complementation we confirm a crucial role for TgCDPK3 in the rapid induction of parasite egress and we establish that TgCDPK3 is critical for formation of latent stages in the brains of mice. Genetic knockout of TgCDPK3 confirms a crucial role for this kinase in parasite egress and a non-essential role for it in the lytic cycle.

  10. A haploid genetic screen identifies the G1/S regulatory machinery as a determinant of Wee1 inhibitor sensitivity.

    Science.gov (United States)

    Heijink, Anne Margriet; Blomen, Vincent A; Bisteau, Xavier; Degener, Fabian; Matsushita, Felipe Yu; Kaldis, Philipp; Foijer, Floris; van Vugt, Marcel A T M

    2015-12-01

    The Wee1 cell cycle checkpoint kinase prevents premature mitotic entry by inhibiting cyclin-dependent kinases. Chemical inhibitors of Wee1 are currently being tested clinically as targeted anticancer drugs. Wee1 inhibition is thought to be preferentially cytotoxic in p53-defective cancer cells. However, TP53 mutant cancers do not respond consistently to Wee1 inhibitor treatment, indicating the existence of genetic determinants of Wee1 inhibitor sensitivity other than TP53 status. To optimally facilitate patient selection for Wee1 inhibition and uncover potential resistance mechanisms, identification of these currently unknown genes is necessary. The aim of this study was therefore to identify gene mutations that determine Wee1 inhibitor sensitivity. We performed a genome-wide unbiased functional genetic screen in TP53 mutant near-haploid KBM-7 cells using gene-trap insertional mutagenesis. Insertion site mapping of cells that survived long-term Wee1 inhibition revealed enrichment of G1/S regulatory genes, including SKP2, CUL1, and CDK2. Stable depletion of SKP2, CUL1, or CDK2 or chemical Cdk2 inhibition rescued the γ-H2AX induction and abrogation of G2 phase as induced by Wee1 inhibition in breast and ovarian cancer cell lines. Remarkably, live cell imaging showed that depletion of SKP2, CUL1, or CDK2 did not rescue the Wee1 inhibition-induced karyokinesis and cytokinesis defects. These data indicate that the activity of the DNA replication machinery, beyond TP53 mutation status, determines Wee1 inhibitor sensitivity, and could serve as a selection criterion for Wee1-inhibitor eligible patients. Conversely, loss of the identified S-phase genes could serve as a mechanism of acquired resistance, which goes along with development of severe genomic instability. PMID:26598692

  11. New multiplex PCR methods for rapid screening of genetically modified organisms in foods.

    Science.gov (United States)

    Datukishvili, Nelly; Kutateladze, Tamara; Gabriadze, Inga; Bitskinashvili, Kakha; Vishnepolsky, Boris

    2015-01-01

    We present novel multiplex PCR methods for rapid and reliable screening of genetically modified organisms (GMOs). New designed PCR primers targeting four frequently used GMO specific sequences permitted identification of new DNA markers, in particular 141 bp fragment of cauliflower mosaic virus (CaMV) 35S promoter, 224 bp fragment of Agrobacterium tumefaciens nopaline synthase (NOS) terminator, 256 bp fragment of 5-enolppyruvylshikimate-phosphate synthase (epsps) gene and 258 bp fragment of Cry1Ab delta-endotoxin (cry1Ab) gene for GMO screening. The certified reference materials containing Roundup Ready soybean (RRS) and maize MON 810 were applied for the development and optimization of uniplex and multiplex PCR systems. Evaluation of amplification products by agarose gel electrophoresis using negative and positive controls confirmed high specificity and sensitivity at 0.1% GMO for both RRS and MON 810. The fourplex PCR was developed and optimized that allows simultaneous detection of three common transgenic elements, such as: CaMV 35S promoter, NOS terminator, epsps gene together with soybean-specific lectin gene. The triplex PCR developed enables simultaneous identification of transgenic elements, such as: 35S promoter and cry1Ab gene together with maize zein gene. The analysis of different processed foods demonstrated that multiplex PCR methods developed in this study are useful for accurate and fast screening of GM food products.

  12. New multiplex PCR methods for rapid screening of genetically modified organisms in foods

    Directory of Open Access Journals (Sweden)

    Nelly eDatukishvili

    2015-07-01

    Full Text Available We present novel multiplex PCR methods for rapid and reliable screening of genetically modified organisms (GMOs. New designed PCR primers targeting four frequently used GMO specific sequences permitted identification of new DNA markers, in particular 141 bp fragment of cauliflower mosaic virus (CaMV 35S promoter, 224 bp fragment of Agrobacterium tumefaciens nopaline synthase (NOS terminator, 256 bp fragment of 5-enolppyruvylshikimate-phosphate synthase (epsps gene and 258 bp fragment of Cry1Ab delta-endotoxin (cry1Ab gene for GMO screening. The certified reference materials containing Roundup Ready soybean (RRS and maize MON 810 were applied for the development and optimization of uniplex and multiplex PCR systems. Evaluation of amplification products by agarose gel electrophoresis using negative and positive controls confirmed high specificity and sensitivity at 0.1% GMO for both RRS and MON 810. The fourplex PCR was developed and optimized that allows simultaneous detection of three common transgenic elements, such as: CaMV 35S promoter, NOS terminator, epsps gene together with soybean-specific lectin gene. The triplex PCR developed enables simultaneous identification of transgenic elements, such as: 35S promoter and cry1Ab gene together with maize zein gene. The analysis of different processed foods demonstrated that multiplex PCR methods developed in this study are useful for accurate and fast screening of GM food products.

  13. Flow Cytometry Enables Multiplexed Measurements of Genetically Encoded Intramolecular FRET Sensors Suitable for Screening.

    Science.gov (United States)

    Doucette, Jaimee; Zhao, Ziyan; Geyer, Rory J; Barra, Melanie M; Balunas, Marcy J; Zweifach, Adam

    2016-07-01

    Genetically encoded sensors based on intramolecular FRET between CFP and YFP are used extensively in cell biology research. Flow cytometry has been shown to offer a means to measure CFP-YFP FRET; we suspected it would provide a unique way to conduct multiplexed measurements from cells expressing different FRET sensors, which is difficult to do with microscopy, and that this could be used for screening. We confirmed that flow cytometry accurately measures FRET signals using cells transiently transfected with an ERK activity reporter, comparing responses measured with imaging and cytometry. We created polyclonal long-term transfectant lines, each expressing a different intramolecular FRET sensor, and devised a way to bar-code four distinct populations of cells. We demonstrated the feasibility of multiplexed measurements and determined that robust multiplexed measurements can be conducted in plate format. To validate the suitability of the method for screening, we measured responses from a plate of bacterial extracts that in unrelated experiments we had determined contained the protein kinase C (PKC)-activating compound teleocidin A-1. The multiplexed assay correctly identifying the teleocidin A-1-containing well. We propose that multiplexed cytometric FRET measurements will be useful for analyzing cellular function and for screening compound collections. PMID:26908592

  14. Genetic analysis and SOD1 mutation screening in Iranian amyotrophic lateral sclerosis patients.

    Science.gov (United States)

    Alavi, Afagh; Nafissi, Shahriar; Rohani, Mohammad; Zamani, Babak; Sedighi, Behnaz; Shamshiri, Hosein; Fan, Jian-Bing; Ronaghi, Mostafa; Elahi, Elahe

    2013-05-01

    Amyotrophic lateral sclerosis (ALS) is a fatal motor neuron disease, and the most common in European populations. Results of genetic analysis and mutation screening of SOD1 in a cohort of 60 Iranian ALS patients are here reported. Initially, linkage analysis in 4 families identified a disease-linked locus that included the known ALS gene, SOD1. Screening of SOD1 identified homozygous p.Asp90Ala causing mutations in all the linked families. Haplotype analysis suggests that the p.Asp90Ala alleles in the Iranian patients might share a common founder with the renowned Scandinavian recessive p.Asp90Ala allele. Subsequent screening in all the patients resulted in identification of 3 other mutations in SOD1, including p.Leu84Phe in the homozygous state. Phenotypic features of the mutation-bearing patients are presented. SOD1 mutations were found in 11.7% of the cohort, 38.5% of the familial ALS probands, and 4.25% of the sporadic ALS cases. SOD1 mutations contribute significantly to ALS among Iranians.

  15. Flow Cytometry Enables Multiplexed Measurements of Genetically Encoded Intramolecular FRET Sensors Suitable for Screening.

    Science.gov (United States)

    Doucette, Jaimee; Zhao, Ziyan; Geyer, Rory J; Barra, Melanie M; Balunas, Marcy J; Zweifach, Adam

    2016-07-01

    Genetically encoded sensors based on intramolecular FRET between CFP and YFP are used extensively in cell biology research. Flow cytometry has been shown to offer a means to measure CFP-YFP FRET; we suspected it would provide a unique way to conduct multiplexed measurements from cells expressing different FRET sensors, which is difficult to do with microscopy, and that this could be used for screening. We confirmed that flow cytometry accurately measures FRET signals using cells transiently transfected with an ERK activity reporter, comparing responses measured with imaging and cytometry. We created polyclonal long-term transfectant lines, each expressing a different intramolecular FRET sensor, and devised a way to bar-code four distinct populations of cells. We demonstrated the feasibility of multiplexed measurements and determined that robust multiplexed measurements can be conducted in plate format. To validate the suitability of the method for screening, we measured responses from a plate of bacterial extracts that in unrelated experiments we had determined contained the protein kinase C (PKC)-activating compound teleocidin A-1. The multiplexed assay correctly identifying the teleocidin A-1-containing well. We propose that multiplexed cytometric FRET measurements will be useful for analyzing cellular function and for screening compound collections.

  16. [Embryonic stem cell test in screening of medicine and other chemicals].

    Science.gov (United States)

    Zhao, Qing; Xu, Jinsen

    2005-02-01

    The technique of embryonic stem cell test (EST) has been developed and used in vitro to screen new medicines and other chemicals. According to toxicity, such medicines and chemicals can be classified as: non-toxic, weak toxic and/or strong toxic. EST shows merits such as no requirement of the sacrifice of pregnant animals, no side-effects on human or animals for candidate medicines and chemicals, higher sensitivity of embryonic stem cells when compared with the sensitivity of ordinary tissues of adult samples in toxicologic researches, higher accuracy when combined with computing techniques, and possible quantitation based on techniques of molecular biology. Advances in utility of EST technique were reviewed and the prospect of technique was also discussed in this paper. PMID:15762145

  17. Chemical Genetics Uncovers Novel Inhibitors of Lignification, Including p-Iodobenzoic Acid Targeting CINNAMATE-4-HYDROXYLASE1[OPEN

    Science.gov (United States)

    Van de Wouwer, Dorien; Decou, Raphaël; Audenaert, Dominique; Nguyen, Long

    2016-01-01

    Plant secondary-thickened cell walls are characterized by the presence of lignin, a recalcitrant and hydrophobic polymer that provides mechanical strength and ensures long-distance water transport. Exactly the recalcitrance and hydrophobicity of lignin put a burden on the industrial processing efficiency of lignocellulosic biomass. Both forward and reverse genetic strategies have been used intensively to unravel the molecular mechanism of lignin deposition. As an alternative strategy, we introduce here a forward chemical genetic approach to find candidate inhibitors of lignification. A high-throughput assay to assess lignification in Arabidopsis (Arabidopsis thaliana) seedlings was developed and used to screen a 10-k library of structurally diverse, synthetic molecules. Of the 73 compounds that reduced lignin deposition, 39 that had a major impact were retained and classified into five clusters based on the shift they induced in the phenolic profile of Arabidopsis seedlings. One representative compound of each cluster was selected for further lignin-specific assays, leading to the identification of an aromatic compound that is processed in the plant into two fragments, both having inhibitory activity against lignification. One fragment, p-iodobenzoic acid, was further characterized as a new inhibitor of CINNAMATE 4-HYDROXYLASE, a key enzyme of the phenylpropanoid pathway synthesizing the building blocks of the lignin polymer. As such, we provide proof of concept of this chemical biology approach to screen for inhibitors of lignification and present a broad array of putative inhibitors of lignin deposition for further characterization. PMID:27485881

  18. Screening values for Non-Carcinogenic Hanford Waste Tank Vapor Chemicals that Lack Established Occupational Exposure Limits

    Energy Technology Data Exchange (ETDEWEB)

    Poet, Torka S.; Mast, Terryl J.; Huckaby, James L.

    2006-02-06

    Over 1,500 different volatile chemicals have been reported in the headspaces of tanks used to store high-level radioactive waste at the U.S. Department of Energy's Hanford Site. Concern about potential exposure of tank farm workers to these chemicals has prompted efforts to evaluate their toxicity, identify chemicals that pose the greatest risk, and incorporate that information into the tank farms industrial hygiene worker protection program. Established occupation exposure limits for individual chemicals and petroleum hydrocarbon mixtures have been used elsewhere to evaluate about 900 of the chemicals. In this report headspace concentration screening values were established for the remaining 600 chemicals using available industrial hygiene and toxicological data. Screening values were intended to be more than an order of magnitude below concentrations that may cause adverse health effects in workers, assuming a 40-hour/week occupational exposure. Screening values were compared to the maximum reported headspace concentrations.

  19. Functional Genetic Screen to Identify Interneurons Governing Behaviorally Distinct Aspects of Drosophila Larval Motor Programs.

    Science.gov (United States)

    Clark, Matt Q; McCumsey, Stephanie J; Lopez-Darwin, Sereno; Heckscher, Ellie S; Doe, Chris Q

    2016-01-01

    Drosophila larval crawling is an attractive system to study rhythmic motor output at the level of animal behavior. Larval crawling consists of waves of muscle contractions generating forward or reverse locomotion. In addition, larvae undergo additional behaviors, including head casts, turning, and feeding. It is likely that some neurons (e.g., motor neurons) are used in all these behaviors, but the identity (or even existence) of neurons dedicated to specific aspects of behavior is unclear. To identify neurons that regulate specific aspects of larval locomotion, we performed a genetic screen to identify neurons that, when activated, could elicit distinct motor programs. We used 165 Janelia CRM-Gal4 lines-chosen for sparse neuronal expression-to ectopically express the warmth-inducible neuronal activator TrpA1, and screened for locomotor defects. The primary screen measured forward locomotion velocity, and we identified 63 lines that had locomotion velocities significantly slower than controls following TrpA1 activation (28°). A secondary screen was performed on these lines, revealing multiple discrete behavioral phenotypes, including slow forward locomotion, excessive reverse locomotion, excessive turning, excessive feeding, immobile, rigid paralysis, and delayed paralysis. While many of the Gal4 lines had motor, sensory, or muscle expression that may account for some or all of the phenotype, some lines showed specific expression in a sparse pattern of interneurons. Our results show that distinct motor programs utilize distinct subsets of interneurons, and provide an entry point for characterizing interneurons governing different elements of the larval motor program. PMID:27172197

  20. Evaluating the Impact of Uncertainties in Clearance and Exposure When Prioritizing Chemicals Screened in High-Throughput Assays

    Science.gov (United States)

    The toxicity-testing paradigm has evolved to include high-throughput (HT) methods for addressing the increasing need to screen hundreds to thousands of chemicals rapidly. Approaches that involve in vitro screening assays, in silico predictions of exposure concentrations, and phar...

  1. An in vitro screening method to evaluate chemicals as potential chemotherapeutants to control Aeromonas hydrophila infection in channel catfish

    Science.gov (United States)

    Using catfish gill cells G1B and four chemicals (hydrogen peroxide, sodium chloride, potassium permanganate, and D-mannose), the feasibility of using an in vitro screening method to identify potential effective chemotherapeutants was evaluated in this study. In vitro screening results revealed that,...

  2. A Chemical Screening Approach to Identify Novel Key Mediators of Erythroid Enucleation

    Science.gov (United States)

    Wölwer, Christina B.; Pase, Luke B.; Pearson, Helen B.; Gödde, Nathan J.; Lackovic, Kurt; Huang, David C. S.; Russell, Sarah M.; Humbert, Patrick O.

    2015-01-01

    Erythroid enucleation is critical for terminal differentiation of red blood cells, and involves extrusion of the nucleus by orthochromatic erythroblasts to produce reticulocytes. Due to the difficulty of synchronizing erythroblasts, the molecular mechanisms underlying the enucleation process remain poorly understood. To elucidate the cellular program governing enucleation, we utilized a novel chemical screening approach whereby orthochromatic cells primed for enucleation were enriched ex vivo and subjected to a functional drug screen using a 324 compound library consisting of structurally diverse, medicinally active and cell permeable drugs. Using this approach, we have confirmed the role of HDACs, proteasomal regulators and MAPK in erythroid enucleation and introduce a new role for Cyclin-dependent kinases, in particular CDK9, in this process. Importantly, we demonstrate that when coupled with imaging analysis, this approach provides a powerful means to identify and characterize rate limiting steps involved in the erythroid enucleation process. PMID:26569102

  3. Randomized comparison of next-generation sequencing and array comparative genomic hybridization for preimplantation genetic screening: a pilot study

    OpenAIRE

    Yang, Zhihong; Lin, James; Zhang, John; Fong, Wai Ieng; Li, Pei; Zhao, Rong; Liu, Xiaohong; Podevin, William; Kuang, Yanping; Liu, Jiaen

    2015-01-01

    Background Recent advances in next-generation sequencing (NGS) have provided new methods for preimplantation genetic screening (PGS) of human embryos from in vitro fertilization (IVF) cycles. However, there is still limited information about clinical applications of NGS in IVF and PGS (IVF-PGS) treatments. The present study aimed to investigate the effects of NGS screening on clinical pregnancy and implantation outcomes for PGS patients in comparison to array comparative genomic hybridization...

  4. Genetically Engineered Protein Modules: Development and Applications in Anti-Viral Agent Screening and Cancer Marker Detection

    OpenAIRE

    Biswas, Payal

    2010-01-01

    ABSTRACT OF THE DISSERTATION Genetically Engineered Protein Modules: Development and Applications in Anti-Viral Agent Screening and Cancer Marker Detection byPayal BiswasDoctor of Philosophy Cell Molecular and Developmental Biology Graduate ProgramUniversity of California, Riverside, August 2010Dr. Wilfred Chen, ChairpersonOne of the most critical aspects in drug discovery is the bioactivity screening assay, by which compounds that most effectively inhibit the target are identified. During t...

  5. Chemical genetic effects of Sargassum wightii during embryonic development in zebrafish

    Directory of Open Access Journals (Sweden)

    Rajaretinam Rajesh Kannan

    2015-01-01

    Conclusions: This study demonstrated that the phytomolecules from S. wightii exhibited potential molecular switches on the developmental process, which might have significant role in understanding the development based chemical genetic studies in zebrafish.

  6. Identification of autophagosome-associated proteins and regulators by quantitative proteomic analysis and genetic screens

    DEFF Research Database (Denmark)

    Dengjel, Jörn; Høyer-Hansen, Maria; Nielsen, Maria O;

    2012-01-01

    Autophagy is one of the major intracellular catabolic pathways, but little is known about the composition of autophagosomes. To study the associated proteins, we isolated autophagosomes from human breast cancer cells using two different biochemical methods and three stimulus types: amino acid......, and the activation of autophagy significantly decreased the cellular proteasome level and activity supporting interplay between the two degradation pathways. A screen of yeast strains defective in the orthologs of the human genes encoding for a common set of autophagosome-associated proteins revealed several...... regulators of autophagy, including subunits of the retromer complex. The combined spatiotemporal proteomic and genetic data sets presented here provide a basis for further characterization of autophagosome biogenesis and cargo selection....

  7. A Zebrafish Genetic Screen Identifies Neuromedin U as a Regulator of Sleep/Wake States.

    Science.gov (United States)

    Chiu, Cindy N; Rihel, Jason; Lee, Daniel A; Singh, Chanpreet; Mosser, Eric A; Chen, Shijia; Sapin, Viveca; Pham, Uyen; Engle, Jae; Niles, Brett J; Montz, Christin J; Chakravarthy, Sridhara; Zimmerman, Steven; Salehi-Ashtiani, Kourosh; Vidal, Marc; Schier, Alexander F; Prober, David A

    2016-02-17

    Neuromodulation of arousal states ensures that an animal appropriately responds to its environment and engages in behaviors necessary for survival. However, the molecular and circuit properties underlying neuromodulation of arousal states such as sleep and wakefulness remain unclear. To tackle this challenge in a systematic and unbiased manner, we performed a genetic overexpression screen to identify genes that affect larval zebrafish arousal. We found that the neuropeptide neuromedin U (Nmu) promotes hyperactivity and inhibits sleep in zebrafish larvae, whereas nmu mutant animals are hypoactive. We show that Nmu-induced arousal requires Nmu receptor 2 and signaling via corticotropin releasing hormone (Crh) receptor 1. In contrast to previously proposed models, we find that Nmu does not promote arousal via the hypothalamic-pituitary-adrenal axis, but rather probably acts via brainstem crh-expressing neurons. These results reveal an unexpected functional and anatomical interface between the Nmu system and brainstem arousal systems that represents a novel wake-promoting pathway. PMID:26889812

  8. ACOG Committee Opinion No. 442: Preconception and prenatal carrier screening for genetic diseases in individuals of Eastern European Jewish descent.

    Science.gov (United States)

    2009-10-01

    Certain autosomal recessive disease conditions are more prevalent in individuals of Eastern European Jewish (Ashkenazi) descent. Previously, the American College of Obstetricians and Gynecologists recommended that individuals of Eastern European Jewish ancestry be offered carrier screening for Tay-Sachs disease, Canavan disease, and cystic fibrosis as part of routine obstetric care. Based on the criteria used to justify offering carrier screening for Tay-Sachs disease, Canavan disease, and cystic fibrosis, the American College of Obstetricians and Gynecologists' Committee on Genetics recommends that couples of Ashkenazi Jewish ancestry also should be offered carrier screening for familial dysautonomia. Individuals of Ashkenazi Jewish descent may inquire about the availability of carrier screening for other disorders. Carrier screening is available for mucolipidosis IV, Niemann-Pick disease type A, Fanconi anemia group C, Bloom syndrome, and Gaucher disease. PMID:19888064

  9. The minisequencing method: a simple strategy for genetic screening of MEN 2 families

    Directory of Open Access Journals (Sweden)

    Domingues Rita

    2002-05-01

    Full Text Available Abstract Background Multiple endocrine neoplasia type 2 is an autosomal dominant disorder. MEN 2A is characterized by medullary thyroid carcinoma, pheochromocytoma and hyperparathyroidism; MEN 2B by medullary thyroid carcinoma, pheochromocytoma and characteristic stigmata. Activating germline mutations of the RET proto oncogene are responsible for this hereditary syndrome. Codon 634 mutations are the most common mutations occurring in MEN 2A families whereas a specific mutation at codon 918 is observed in the great majority of MEN 2B families. Analysis of these codons will provide a final diagnosis in the great majority of affected families making unnecessary further studies. To specifically study the codons 634 and 918 we used a minisequencing method as an alternative method to complete sequencing. Results Using this mutation detection method we were able to reproduce in all cases, representative of 7 families, the information previously obtained by direct sequencing of PCR products. Depending on the number of primers used in the minisequencing reaction, we were able to interrogate either only one nucleotide of the target codon or the three nucleotides simultaneously. Conclusions This technique appears as a simple, rapid and efficient method for genetic screening of MEN 2 families. It can be utilized to seek for unknown mutations at specific codons or to screen for previously identified mutations and is therefore of interest to study index cases or individuals at risk. Results suggest that complete sequencing is unnecessary.

  10. Genetic modifier screens reveal new components that interact with the Drosophila dystroglycan-dystrophin complex.

    Directory of Open Access Journals (Sweden)

    Mariya M Kucherenko

    Full Text Available The Dystroglycan-Dystrophin (Dg-Dys complex has a capacity to transmit information from the extracellular matrix to the cytoskeleton inside the cell. It is proposed that this interaction is under tight regulation; however the signaling/regulatory components of Dg-Dys complex remain elusive. Understanding the regulation of the complex is critical since defects in this complex cause muscular dystrophy in humans. To reveal new regulators of the Dg-Dys complex, we used a model organism Drosophila melanogaster and performed genetic interaction screens to identify modifiers of Dg and Dys mutants in Drosophila wing veins. These mutant screens revealed that the Dg-Dys complex interacts with genes involved in muscle function and components of Notch, TGF-beta and EGFR signaling pathways. In addition, components of pathways that are required for cellular and/or axonal migration through cytoskeletal regulation, such as Semaphorin-Plexin, Frazzled-Netrin and Slit-Robo pathways show interactions with Dys and/or Dg. These data suggest that the Dg-Dys complex and the other pathways regulating extracellular information transfer to the cytoskeletal dynamics are more intercalated than previously thought.

  11. Volatile organic chemical emissions from carpet cushions: Screening measurements. Final report

    Energy Technology Data Exchange (ETDEWEB)

    Hodgson, A.T.; Phan, T.A.

    1994-05-01

    The US Consumer Product Safety Commission (CPSC) has received complaints from consumers regarding the occurrence of adverse health effects following the installation of new carpeting (Schachter, 1990). Carpet systems are suspected of emitting chemicals which may be the cause of these complaints, as well as objectionable odors. Carpets themselves have been shown to emit a variety of volatile organic compounds (VOCs). The objective of this study was to screen the representative samples of carpet cushions for emissions of individual VOCS, total VOCs (TVOC), formaldehyde, and, for the two types of polyurethane cushions, isomers of toluene diisocyanate (TDI). The measurements of VOCS, TVOC and formaldehyde were made over six-hour periods using small-volume (4-L) dynamic chambers. Sensitive gas chromatography-mass spectrometry (GC-MS) techniques were used to identify many of the VOCs emitted by the cushion samples and to obtain quantitative estimates of the emission rates of selected compounds. Separate screening measurements were conducted for TDI. The data from the screening measurements were used by the CPSC`s Health Sciences Laboratory to help design and conduct week-long measurements of emission rates of selected compounds.

  12. GAPscreener: An automatic tool for screening human genetic association literature in PubMed using the support vector machine technique

    Directory of Open Access Journals (Sweden)

    Khoury Muin J

    2008-04-01

    Full Text Available Abstract Background Synthesis of data from published human genetic association studies is a critical step in the translation of human genome discoveries into health applications. Although genetic association studies account for a substantial proportion of the abstracts in PubMed, identifying them with standard queries is not always accurate or efficient. Further automating the literature-screening process can reduce the burden of a labor-intensive and time-consuming traditional literature search. The Support Vector Machine (SVM, a well-established machine learning technique, has been successful in classifying text, including biomedical literature. The GAPscreener, a free SVM-based software tool, can be used to assist in screening PubMed abstracts for human genetic association studies. Results The data source for this research was the HuGE Navigator, formerly known as the HuGE Pub Lit database. Weighted SVM feature selection based on a keyword list obtained by the two-way z score method demonstrated the best screening performance, achieving 97.5% recall, 98.3% specificity and 31.9% precision in performance testing. Compared with the traditional screening process based on a complex PubMed query, the SVM tool reduced by about 90% the number of abstracts requiring individual review by the database curator. The tool also ascertained 47 articles that were missed by the traditional literature screening process during the 4-week test period. We examined the literature on genetic associations with preterm birth as an example. Compared with the traditional, manual process, the GAPscreener both reduced effort and improved accuracy. Conclusion GAPscreener is the first free SVM-based application available for screening the human genetic association literature in PubMed with high recall and specificity. The user-friendly graphical user interface makes this a practical, stand-alone application. The software can be downloaded at no charge.

  13. Genetic and chemical diversity of high mucilaginous plants of Sida complex by ISSR markers and chemical fingerprinting.

    Science.gov (United States)

    Thul, Sanjog T; Srivastava, Ankit K; Singh, Subhash C; Shanker, Karuna

    2011-09-01

    A method was developed based on multiple approaches wherein DNA and chemical analysis was carried out toward differentiation of important species of Sida complex that is being used for commercial preparation. Isolated DNA samples were successfully performed through PCR amplification using ISSR markers and degree of genetic diversity among the different species of Sida is compared with that of chemical diversity. For genetic fingerprint investigation, selected 10 ISSR primers generating reproducible banding patterns were used. Among the total of 63 amplicons, 62 were recorded as polymorphic, genetic similarity index deduced from ISSR profiles ranged from 12 to 51%. Based on similarity index, S. acuta and S. rhombifolia found to be most similar (51%). High number of species-specific bands played pivotal role to delineate species at genetic level. Investigation based on HPTLC fingerprints analysis revealed 23 bands representing to characteristic chemicals and similarity index ranged from 73 to 91%. Prominent distinguishable bands were observed only in S. acuta, while S. cordifolia and S. rhombifolia shared most bands making them difficult to identify on chemical fingerprint basis. This report summarizes the genotypic and chemotypic diversity and the use of profiles for authentication of species of Sida complex.

  14. Data-driven prioritization of chemicals for various water types using suspect screening LC-HRMS.

    Science.gov (United States)

    Sjerps, Rosa M A; Vughs, Dennis; van Leerdam, Jan A; ter Laak, Thomas L; van Wezel, Annemarie P

    2016-04-15

    For the prioritization of more than 5200 anthropogenic chemicals authorized on the European market, we use a large scale liquid chromatography-high resolution mass spectrometry (LC-HRMS) suspect screening study. The prioritization is based on occurrence in 151 water samples including effluent, surface water, ground water and drinking water. The suspect screening linked over 700 detected compounds with known accurate masses to one or multiple suspects. Using a prioritization threshold and removing false positives reduced this to 113 detected compounds linked to 174 suspects, 24 compounds reflect a confirmed structure by comparison with the pure reference standard. The prioritized compounds and suspects are relevant for detailed risk assessments after confirmation of their identity. Only one of the 174 prioritized compounds and suspects is mentioned in water quality regulations, and only 20% is mentioned on existing lists of potentially relevant chemicals. This shows the complementarity to commonly used target-based methods. The semi-quantitative total concentration, expressed as internal standard equivalents of detected compounds linked to suspects, in effluents is approximately 10 times higher than in surface waters, while ground waters and drinking waters show the lowest response. The average retention time, a measure for hydrophobicity, of the detected compounds per sample decreased from effluent to surface- and groundwater to drinking water, confirming the occurrence of more polar compounds in drinking water. The semi-quantitative total concentrations exceed the conservative and precautionary threshold of toxicological concern. Therefore, adverse effects of mixtures cannot be neglected without a more thorough risk assessment. PMID:26921851

  15. Data-driven prioritization of chemicals for various water types using suspect screening LC-HRMS.

    Science.gov (United States)

    Sjerps, Rosa M A; Vughs, Dennis; van Leerdam, Jan A; ter Laak, Thomas L; van Wezel, Annemarie P

    2016-04-15

    For the prioritization of more than 5200 anthropogenic chemicals authorized on the European market, we use a large scale liquid chromatography-high resolution mass spectrometry (LC-HRMS) suspect screening study. The prioritization is based on occurrence in 151 water samples including effluent, surface water, ground water and drinking water. The suspect screening linked over 700 detected compounds with known accurate masses to one or multiple suspects. Using a prioritization threshold and removing false positives reduced this to 113 detected compounds linked to 174 suspects, 24 compounds reflect a confirmed structure by comparison with the pure reference standard. The prioritized compounds and suspects are relevant for detailed risk assessments after confirmation of their identity. Only one of the 174 prioritized compounds and suspects is mentioned in water quality regulations, and only 20% is mentioned on existing lists of potentially relevant chemicals. This shows the complementarity to commonly used target-based methods. The semi-quantitative total concentration, expressed as internal standard equivalents of detected compounds linked to suspects, in effluents is approximately 10 times higher than in surface waters, while ground waters and drinking waters show the lowest response. The average retention time, a measure for hydrophobicity, of the detected compounds per sample decreased from effluent to surface- and groundwater to drinking water, confirming the occurrence of more polar compounds in drinking water. The semi-quantitative total concentrations exceed the conservative and precautionary threshold of toxicological concern. Therefore, adverse effects of mixtures cannot be neglected without a more thorough risk assessment.

  16. Use of biosensors to screen urine samples for potentially toxic chemicals.

    Science.gov (United States)

    Horswell, Jacqui; Dickson, Stuart

    2003-09-01

    Forensic toxicology laboratories are often required to implicate or exclude poisoning as a factor in a death or unexplained illness. An analytical tool which enables toxicologists to screen a wide variety of common poisons would be extremely useful. In this paper, we describe the use of a bacterial biosensor for detecting the presence of commonly encountered potentially toxic chemicals in urine. The biosensor responds to any chemical that causes metabolic stress to the bacterial cell and the response is in direct proportion to the concentration of the stressor. This allows a measure of the concentration of a toxicant in urine, without knowing exactly what the toxic compound(s) may be. This affords a distinct advantage over conventional analytical techniques, which require an extensive screening program before it is even known that a toxic compound is present. This preliminary investigation has shown that this biosensor can indicate the presence, in urine, of herbicides such as glyphosate, 2,4-dichlorophenoxyacetic acid, and 2,4,5-trichlorophenoxyacetic acid; the biocide pentachlorophenol; or inorganic poisons such as arsenic, mercury, and cyanide. The biosensor was also shown to be sensitive to a concentration range of these toxicants likely to be found in samples submitted for toxicological analysis.

  17. Chemical and Genetic Wrappers for Improved Phage and RNA Display

    OpenAIRE

    Lamboy, Jorge A.; Tam, Phillip Y.; Lee, Lucie S.; Jackson, Pilgrim J.; Avrantinis, Sara K; Lee, Hye Jin; Corn, Robert M.; Weiss, Gregory A.

    2008-01-01

    An Achilles heel inherent to all molecular display formats, background binding between target and display system introduces false positives into screens and selections. For example, the negatively charged surfaces of phage, mRNA, and ribosome display systems bind with unacceptably high non-specificity to positively charged target molecules, which represent an estimated 35% of proteins in the human proteome. We report the first systematic attempt to understand why a broad class of molecular di...

  18. A Genetic Screen for Mutants with Supersized Lipid Droplets in Caenorhabditis elegans

    Science.gov (United States)

    Li, Shiwei; Xu, Shibin; Ma, Yanli; Wu, Shuang; Feng, Yu; Cui, Qingpo; Chen, Lifeng; Zhou, Shuang; Kong, Yuanyuan; Zhang, Xiaoyu; Yu, Jialei; Wu, Mengdi; Zhang, Shaobing O.

    2016-01-01

    To identify genes that regulate the dynamics of lipid droplet (LD) size, we have used the genetically tractable model organism Caenorhabditis elegans, whose wild-type LD population displays a steady state of size with an upper limit of 3 μm in diameter. From a saturated forward genetic screen of 6.7 × 105 mutagenized haploid genomes, we isolated 118 mutants with supersized intestinal LDs often reaching 10 μm. These mutants define nine novel complementation groups, in addition to four known genes (maoc-1, dhs-28, daf-22, and prx-10). The nine groups are named drop (lipid droplet abnormal) and categorized into four classes. Class I mutants drop-5 and drop-9, similar to prx-10, are up-regulated in ACS-22-DGAT-2-dependent LD growth, resistant to LD hydrolysis, and defective in peroxisome import. Class II mutants drop-2, drop-3, drop-6, and drop-7 are up-regulated in LD growth, are resistant to LD hydrolysis, but are not defective in peroxisome import. Class III mutants drop-1 and drop-8 are neither up-regulated in LD growth nor resistant to LD hydrolysis, but seemingly up-regulated in LD fusion. Class IV mutant drop-4 is cloned as sams-1 and, different to the other three classes, is ACS-22-independent and hydrolysis-resistant. These four classes of supersized LD mutants should be valuable for mechanistic studies of LD cellular processes including growth, hydrolysis, and fusion. PMID:27261001

  19. Phyto chemical and antioxidant screening of extracts of Aquilaria malaccensis leaves

    International Nuclear Information System (INIS)

    Aquilaria malaccensis is an endangered economic plant used for production of agar wood worldwide. The sequential maceration extraction methods utilizing solvents with different polarities namely hexane, ethyl acetate and methanol yielded the corresponding crude extract. The aqueous and methanol extracts along with dry powder of leaf of the plant was screened for the presence of phytochemicals. They were also tested for antioxidant activities. The result indicates the presence of alkaloids, flavanoids, triterpenoids, steroids and tannins. The phyto chemical screening suggests that flavanoids present in this species might provide a great value of antioxidant activity. Preliminary screenings of the free radical scavenging activity on the extracts of the plants with 2, 2-Diphenyl-1-picrylhydrazyl (DPPH) were tested and showed positive result. Quarcetine was used as reference standard. The extracts exhibited strong antioxidant activity radical scavenging activity with IC50 value of 8.0 x 102 μg/ ml, 1.6 x 102 μg/ ml, 1.4 x 102 μg/ ml, 30.0 μg/ ml and 3.33 μg/ ml for hexane, DCM, ethyl acetate, methanol and quercetine respectively. Determination on antioxidant activity of each crude extract showed that methanol crude extract had the highest IC50 value than ethyl acetate, dichloromethane and hexane crude extract. This means that methanol possess the highest inhibition of DPPH radical scavenging activity compared to the other crudes but still lower than Quercetin (standard). Phyto chemical analysis on the hexane extract of Aquilaria malaccensis has been conducted. Several chromatographic methods have been employed to the hexane of the leaves which led to the isolation of three compounds namely Stigmasterol, β-sitosterol and 3-fridelanol. The present study has proved the usefulness of agar wood tree for medicinal purposes and its potential as a source of useful drugs. (author)

  20. Electrochemically induced chemical sensor properties in graphite screen-printed electrodes: The case of a chemical sensor for uranium

    Energy Technology Data Exchange (ETDEWEB)

    Kostaki, Vasiliki T.; Florou, Ageliki B. [Laboratory of Analytical Chemistry, Department of Chemistry, University of Ioannina, 451 10 Ioannina (Greece); Prodromidis, Mamas I., E-mail: mprodrom@cc.uoi.gr [Laboratory of Analytical Chemistry, Department of Chemistry, University of Ioannina, 451 10 Ioannina (Greece)

    2011-10-01

    Highlights: > Electrochemical treatment endows analytical characteristics to SPEs. > A sensitive chemical sensor for uranium is described. > Performance is due to a synergy between electrochemical treatment and ink's solvents. > The amount of the solvent controls the achievable sensitivity. - Abstract: We report for the first time on the possibility to develop chemical sensors based on electrochemically treated, non-modified, graphite screen-printed electrodes (SPEs). The applied galvanostatic treatment (5 {mu}A for 6 min in 0.1 M H{sub 2}SO{sub 4}) is demonstrated to be effective for the development of chemical sensors for the determination of uranium in aqueous solutions. A detailed study of the effect of various parameters related to the fabrication of SPEs on the performance of the resulting sensors along with some diagnostic experiments on conventional graphite electrodes showed that the inducible analytical characteristics are due to a synergy between electrochemical treatment and ink's solvents. Indeed, the amount of the latter onto the printed working layer controls the achievable sensitivity. The preconcentration of the analyte was performed in an electroless mode in an aqueous solutions of U(VI), pH 4.6, and then, the accumulated species was reduced by means of a differential pulse voltammetry scan in 0.1 M H{sub 3}BO{sub 3}, pH 3. Under selected experimental conditions, a linear calibration curve over the range 5 x 10{sup -9} to 10{sup -7} M U(VI) was constructed. The 3{sigma} limit of detection at a preconcentration time of 30 min, and the relative standard deviation of the method were 4.5 x 10{sup -9} M U(VI) and >12% (n = 5, 5 x 10{sup -8} M U(VI)), respectively. The effect of potential interferences was also examined.

  1. Patients' ratings of genetic conditions validate a taxonomy to simplify decisions about preconception carrier screening via genome sequencing.

    Science.gov (United States)

    Leo, Michael C; McMullen, Carmit; Wilfond, Benjamin S; Lynch, Frances L; Reiss, Jacob A; Gilmore, Marian J; Himes, Patricia; Kauffman, Tia L; Davis, James V; Jarvik, Gail P; Berg, Jonathan S; Harding, Cary; Kennedy, Kathleen A; Simpson, Dana Kostiner; Quigley, Denise I; Richards, C Sue; Rope, Alan F; Goddard, Katrina A B

    2016-03-01

    Advances in genome sequencing and gene discovery have created opportunities to efficiently assess more genetic conditions than ever before. Given the large number of conditions that can be screened, the implementation of expanded carrier screening using genome sequencing will require practical methods of simplifying decisions about the conditions for which patients want to be screened. One method to simplify decision making is to generate a taxonomy based on expert judgment. However, expert perceptions of condition attributes used to classify these conditions may differ from those used by patients. To understand whether expert and patient perceptions differ, we asked women who had received preconception genetic carrier screening in the last 3 years to fill out a survey to rate the attributes (predictability, controllability, visibility, and severity) of several autosomal recessive or X-linked genetic conditions. These conditions were classified into one of five taxonomy categories developed by subject experts (significantly shortened lifespan, serious medical problems, mild medical problems, unpredictable medical outcomes, and adult-onset conditions). A total of 193 women provided 739 usable ratings across 20 conditions. The mean ratings and correlations demonstrated that participants made distinctions across both attributes and categories. Aggregated mean attribute ratings across categories demonstrated logical consistency between the key features of each attribute and category, although participants perceived little difference between the mild and serious categories. This study provides empirical evidence for the validity of our proposed taxonomy, which will simplify patient decisions for results they would like to receive from preconception carrier screening via genome sequencing.

  2. Human Genetic Marker for Resistance to Radiation and Chemicals

    Energy Technology Data Exchange (ETDEWEB)

    DR. Howard B. Lieberman

    2001-05-11

    TO characterize the human HRDAD9 gene and evaluate its potential as a biomarker to predict susceptibility to the deleterious health effects potentially caused by exposure to radiations or chemicals present at DOE hazardous waste cleanup sites. HRAD9 is a human gene that is highly conserved throughout evolution. Related genes have been isolated from yeasts and mice, underscoring its biological significance. Most of our previous work involved characterization of the yeast gene cognate, wherein it was determined that the corresponding protein plays a significant role in promoting resistance of cells to radiations and chemicals, and in particular, controlling cell growth in response to DNA damage.

  3. Human Genetic Marker for Resistance to Radiation and Chemicals

    International Nuclear Information System (INIS)

    TO characterize the human HRDAD9 gene and evaluate its potential as a biomarker to predict susceptibility to the deleterious health effects potentially caused by exposure to radiations or chemicals present at DOE hazardous waste cleanup sites. HRAD9 is a human gene that is highly conserved throughout evolution. Related genes have been isolated from yeasts and mice, underscoring its biological significance. Most of our previous work involved characterization of the yeast gene cognate, wherein it was determined that the corresponding protein plays a significant role in promoting resistance of cells to radiations and chemicals, and in particular, controlling cell growth in response to DNA damage

  4. Genetic effects of combined chemical-X-ray treatments in male mouse germ cells

    International Nuclear Information System (INIS)

    Several studies have shown that the yield of genetic damage induced by radiation in male mouse germ cells can be modified by chemical treatments. Pre-treatments with radio-protecting agents have given contradictory results but this appears to be largely attributable to the different germ cell stages tested and dependent upon the level of radiation damage induced. Pre-treatments which enhance the yield of genetic damage have been reported although, as yet, no tests have been conducted with radio-sensitizers. Another form of interaction between chemicals and radiation is specifically found with spermatogonial stem cells. Chemicals that kill cells can, by population depletion, substantially and predictably modify the genetic response to subsequent radiation exposure over a period of several days, or even weeks. Enhancement and reduction in the genetic yield can be attained, dependent upon the interval between treatments, with the modification also varying with the type of genetic damage scored. Post-treatment with one chemical (TEM) has been shown to reduce the genetic response to radiation exposure. (author)

  5. Genetic dissection of Flaviviridae host factors through genome-scale CRISPR screens.

    Science.gov (United States)

    Marceau, Caleb D; Puschnik, Andreas S; Majzoub, Karim; Ooi, Yaw Shin; Brewer, Susan M; Fuchs, Gabriele; Swaminathan, Kavya; Mata, Miguel A; Elias, Joshua E; Sarnow, Peter; Carette, Jan E

    2016-07-01

    The Flaviviridae are a family of viruses that cause severe human diseases. For example, dengue virus (DENV) is a rapidly emerging pathogen causing an estimated 100 million symptomatic infections annually worldwide. No approved antivirals are available to date and clinical trials with a tetravalent dengue vaccine showed disappointingly low protection rates. Hepatitis C virus (HCV) also remains a major medical problem, with 160 million chronically infected patients worldwide and only expensive treatments available. Despite distinct differences in their pathogenesis and modes of transmission, the two viruses share common replication strategies. A detailed understanding of the host functions that determine viral infection is lacking. Here we use a pooled CRISPR genetic screening strategy to comprehensively dissect host factors required for these two highly important Flaviviridae members. For DENV, we identified endoplasmic-reticulum (ER)-associated multi-protein complexes involved in signal sequence recognition, N-linked glycosylation and ER-associated degradation. DENV replication was nearly completely abrogated in cells deficient in the oligosaccharyltransferase (OST) complex. Mechanistic studies pinpointed viral RNA replication and not entry or translation as the crucial step requiring the OST complex. Moreover, we show that viral non-structural proteins bind to the OST complex. The identified ER-associated protein complexes were also important for infection by other mosquito-borne flaviviruses including Zika virus, an emerging pathogen causing severe birth defects. By contrast, the most significant genes identified in the HCV screen were distinct and included viral receptors, RNA-binding proteins and enzymes involved in metabolism. We found an unexpected link between intracellular flavin adenine dinucleotide (FAD) levels and HCV replication. This study shows notable divergence in host-depenency factors between DENV and HCV, and illuminates new host targets for

  6. Genetic dissection of Flaviviridae host factors through genome-scale CRISPR screens.

    Science.gov (United States)

    Marceau, Caleb D; Puschnik, Andreas S; Majzoub, Karim; Ooi, Yaw Shin; Brewer, Susan M; Fuchs, Gabriele; Swaminathan, Kavya; Mata, Miguel A; Elias, Joshua E; Sarnow, Peter; Carette, Jan E

    2016-07-01

    The Flaviviridae are a family of viruses that cause severe human diseases. For example, dengue virus (DENV) is a rapidly emerging pathogen causing an estimated 100 million symptomatic infections annually worldwide. No approved antivirals are available to date and clinical trials with a tetravalent dengue vaccine showed disappointingly low protection rates. Hepatitis C virus (HCV) also remains a major medical problem, with 160 million chronically infected patients worldwide and only expensive treatments available. Despite distinct differences in their pathogenesis and modes of transmission, the two viruses share common replication strategies. A detailed understanding of the host functions that determine viral infection is lacking. Here we use a pooled CRISPR genetic screening strategy to comprehensively dissect host factors required for these two highly important Flaviviridae members. For DENV, we identified endoplasmic-reticulum (ER)-associated multi-protein complexes involved in signal sequence recognition, N-linked glycosylation and ER-associated degradation. DENV replication was nearly completely abrogated in cells deficient in the oligosaccharyltransferase (OST) complex. Mechanistic studies pinpointed viral RNA replication and not entry or translation as the crucial step requiring the OST complex. Moreover, we show that viral non-structural proteins bind to the OST complex. The identified ER-associated protein complexes were also important for infection by other mosquito-borne flaviviruses including Zika virus, an emerging pathogen causing severe birth defects. By contrast, the most significant genes identified in the HCV screen were distinct and included viral receptors, RNA-binding proteins and enzymes involved in metabolism. We found an unexpected link between intracellular flavin adenine dinucleotide (FAD) levels and HCV replication. This study shows notable divergence in host-depenency factors between DENV and HCV, and illuminates new host targets for

  7. Automated screening for small organic ligands using DNA-encoded chemical libraries.

    Science.gov (United States)

    Decurtins, Willy; Wichert, Moreno; Franzini, Raphael M; Buller, Fabian; Stravs, Michael A; Zhang, Yixin; Neri, Dario; Scheuermann, Jörg

    2016-04-01

    DNA-encoded chemical libraries (DECLs) are collections of organic compounds that are individually linked to different oligonucleotides, serving as amplifiable identification barcodes. As all compounds in the library can be identified by their DNA tags, they can be mixed and used in affinity-capture experiments on target proteins of interest. In this protocol, we describe the screening process that allows the identification of the few binding molecules within the multiplicity of library members. First, the automated affinity selection process physically isolates binding library members. Second, the DNA codes of the isolated binders are PCR-amplified and subjected to high-throughput DNA sequencing. Third, the obtained sequencing data are evaluated using a C++ program and the results are displayed using MATLAB software. The resulting selection fingerprints facilitate the discrimination of binding from nonbinding library members. The described procedures allow the identification of small organic ligands to biological targets from a DECL within 10 d. PMID:26985574

  8. A Chemical Screen Identifies Novel Compounds That Overcome Glial-Mediated Inhibition Of Neuronal Regeneration

    Science.gov (United States)

    Usher, Lynn C.; Johnstone, Andrea; Ertürk, Ali; Hu, Ying; Strikis, Dinara; Wanner, Ina B.; Moorman, Sanne; Lee, Jae-Wook; Min, Jaeki; Ha, Hyung-Ho; Duan, Yuanli; Hoffman, Stanley; Goldberg, Jeffrey L.; Bradke, Frank; Chang, Young-Tae; Lemmon, Vance P.; Bixby, John L.

    2010-01-01

    A major barrier to regeneration of central nervous system (CNS) axons is the presence of growth-inhibitory proteins associated with myelin and the glial scar. To identify chemical compounds with the ability to overcome the inhibition of regeneration, we screened a novel triazine library, based on the ability of compounds to increase neurite outgrowth from cerebellar neurons on inhibitory myelin substrates. The screen produced 4 “hit compounds”, which act with nM potency on several different neuronal types, and on several distinct substrates relevant to glial inhibition. Moreover, the compounds selectively overcome inhibition rather than promote growth in general. The compounds do not affect neuronal cAMP levels, PKC activity, or EGFR activation. Interestingly, one of the compounds alters microtubule dynamics and increases microtubule density in both fibroblasts and neurons. This same compound promotes regeneration of dorsal column axons after acute lesions, and potentiates regeneration of optic nerve axons after nerve crush in vivo. These compounds should provide insight into the mechanisms through which glial-derived inhibitors of regeneration act, and could lead to the development of novel therapies for CNS injury. PMID:20357120

  9. Evaluation of a novel electronic genetic screening and clinical decision support tool in prenatal clinical settings.

    Science.gov (United States)

    Edelman, Emily A; Lin, Bruce K; Doksum, Teresa; Drohan, Brian; Edelson, Vaughn; Dolan, Siobhan M; Hughes, Kevin; O'Leary, James; Vasquez, Lisa; Copeland, Sara; Galvin, Shelley L; DeGroat, Nicole; Pardanani, Setul; Gregory Feero, W; Adams, Claire; Jones, Renee; Scott, Joan

    2014-07-01

    "The Pregnancy and Health Profile" (PHP) is a free prenatal genetic screening and clinical decision support (CDS) software tool for prenatal providers. PHP collects family health history (FHH) during intake and provides point-of-care risk assessment for providers and education for patients. This pilot study evaluated patient and provider responses to PHP and effects of using PHP in practice. PHP was implemented in four clinics. Surveys assessed provider confidence and knowledge and patient and provider satisfaction with PHP. Data on the implementation process were obtained through semi-structured interviews with administrators. Quantitative survey data were analyzed using Chi square test, Fisher's exact test, paired t tests, and multivariate logistic regression. Open-ended survey questions and interviews were analyzed using qualitative thematic analysis. Of the 83% (513/618) of patients that provided feedback, 97% felt PHP was easy to use and 98% easy to understand. Thirty percent (21/71) of participating physicians completed both pre- and post-implementation feedback surveys [13 obstetricians (OBs) and 8 family medicine physicians (FPs)]. Confidence in managing genetic risks significantly improved for OBs on 2/6 measures (p values ≤0.001) but not for FPs. Physician knowledge did not significantly change. Providers reported value in added patient engagement and reported mixed feedback about the CDS report. We identified key steps, resources, and staff support required to implement PHP in a clinical setting. To our knowledge, this study is the first to report on the integration of patient-completed, electronically captured and CDS-enabled FHH software into primary prenatal practice. PHP is acceptable to patients and providers. Key to successful implementation in the future will be customization options and interoperability with electronic health records.

  10. Identification of cell surface targets for HIV-1 therapeutics using genetic screens

    International Nuclear Information System (INIS)

    Human immunodeficiency virus (HIV) drugs designed to interfere with obligatory utilization of certain host cell factors by virus are less likely to encounter development of resistant strains than drugs directed against viral components. Several cellular genes required for productive infection by HIV were identified by the use of genetic suppressor element (GSE) technology as potential targets for anti-HIV drug development. Fragmented cDNA libraries from various pools of human peripheral blood mononuclear cells (PBMC) were expressed in vitro in human immunodeficiency virus type 1 (HIV-1)-susceptible cell lines and subjected to genetic screens to identify GSEs that interfered with viral replication. After three rounds of selection, more than 15 000 GSEs were sequenced, and the cognate genes were identified. The GSEs that inhibited the virus were derived from a diverse set of genes including cell surface receptors, cytokines, signaling proteins, transcription factors, as well as genes with unknown function. Approximately 2.5% of the identified genes were previously shown to play a role in the HIV-1 life cycle; this finding supports the biological relevance of the assay. GSEs were derived from the following 12 cell surface proteins: CXCR4, CCR4, CCR7, CD11C, CD44, CD47, CD68, CD69, CD74, CSF3R, GABBR1, and TNFR2. Requirement of some of these genes for viral infection was also investigated by using RNA interference (RNAi) technology; accordingly, 10 genes were implicated in early events of the viral life cycle, before viral DNA synthesis. Thus, these cell surface proteins represent novel targets for the development of therapeutics against HIV-1 infection and AIDS

  11. High-Throughput Screening of Chemical Effects on Steroidogenesis Using H295R Human Adrenocortical Carcinoma Cells.

    Science.gov (United States)

    Karmaus, Agnes L; Toole, Colleen M; Filer, Dayne L; Lewis, Kenneth C; Martin, Matthew T

    2016-04-01

    Disruption of steroidogenesis by environmental chemicals can result in altered hormone levels causing adverse reproductive and developmental effects. A high-throughput assay using H295R human adrenocortical carcinoma cells was used to evaluate the effect of 2060 chemical samples on steroidogenesis via high-performance liquid chromatography followed by tandem mass spectrometry quantification of 10 steroid hormones, including progestagens, glucocorticoids, androgens, and estrogens. The study employed a 3 stage screening strategy. The first stage established the maximum tolerated concentration (MTC; ≥ 70% viability) per sample. The second stage quantified changes in hormone levels at the MTC whereas the third stage performed concentration-response (CR) on a subset of samples. At all stages, cells were prestimulated with 10 µM forskolin for 48 h to induce steroidogenesis followed by chemical treatment for 48 h. Of the 2060 chemical samples evaluated, 524 samples were selected for 6-point CR screening, based in part on significantly altering at least 4 hormones at the MTC. CR screening identified 232 chemical samples with concentration-dependent effects on 17β-estradiol and/or testosterone, with 411 chemical samples showing an effect on at least one hormone across the steroidogenesis pathway. Clustering of the concentration-dependent chemical-mediated steroid hormone effects grouped chemical samples into 5 distinct profiles generally representing putative mechanisms of action, including CYP17A1 and HSD3B inhibition. A distinct pattern was observed between imidazole and triazole fungicides suggesting potentially distinct mechanisms of action. From a chemical testing and prioritization perspective, this assay platform provides a robust model for high-throughput screening of chemicals for effects on steroidogenesis.

  12. Comparison of three marine screening tests and four Oslo and Paris Commission procedures to evaluate toxicity of offshore chemicals

    Energy Technology Data Exchange (ETDEWEB)

    Weideborg, M.; Vik, E.A.; Oefjord, G.D.; Kjoennoe, O. [Aquateam-Norwegian Water Technology Centre A/S, Oslo (Norway)

    1997-02-01

    The results from the screening toxicity tests Artemia salina, Microtox{reg_sign}, and Mitochondria RET test were compared with those obtained from OSPAR (Oslo and Paris Commissions)-authorized procedures for testing of offshore chemicals (Skeletonema costatum, Acartia tonsa, Abra alba, and Corophium volutator). In this study 82 test substances (26 non-water soluble) were included. The Microtox test was found to be the most sensitive of the three screening tests. Microtox and Mitochondria RET test results showed good correlation with results from Acartia and Skeletonema testing, and it was concluded that the Microtox test was a suitable screening test as a base for assessment of further testing, especially regarding water-soluble chemicals. Sensitivity of Artemia salina to the tested chemicals was too low for it to be an appropriate bioassay organism for screening testing. A very good correlation was found between the results obtained with the Skeletonema and Acartia tests. The results indicated no need for more than one of the Skeletonema or Acartia tests if the Skeletonema median effective concentration or Acartia median lethal concentration was greater than 200 mg/L. The sediment-reworker tests (A. Alba or C. volutator) for chemicals that are likely to end up in the sediments (non-water soluble or surfactants) should be performed, independent of results from screening tests and other OSPAR species.

  13. Data on screening and identification of genetically modified papaya in food supplements.

    Science.gov (United States)

    Prins, Theo W; Scholtens, Ingrid M J; Bak, Arno W; van Dijk, Jeroen P; Voorhuijzen, Marleen M; Laurensse, Emile J; Kok, Esther J

    2016-12-01

    This article contains data related to the research article entitled "A case study to determine the geographical origin of unknown GM papaya in routine food sample analysis, followed by identification of papaya events 16-0-1 and 18-2-4" (Prins et al., 2016) [1]. Quantitative real-time PCR (qPCR) with targets that are putatively present in genetically modified (GM) papaya was used as a first screening to narrow down the vast array of candidates. The combination of elements P-nos and nptII was further confirmed by amplification and subsequent sequencing of the P-nos/nptII construct. Next, presence of the candidate GM papayas 16-0-1 and 18-2-4 were investigated by amplification and sequencing of event-spanning regions on the left and right border. This data article reports the Cq values for GM elements, the nucleotide sequence of the P-nos/nptII construct and the presence of GM papaya events 18-2-4 and/or 16-0-1 in five samples that were randomly sampled to be analysed in the framework of the official Dutch GMO monitoring program for food.

  14. A Transposon Screen Identifies Genetic Determinants of Vibrio cholerae Resistance to High-Molecular-Weight Antibiotics.

    Science.gov (United States)

    Dörr, Tobias; Delgado, Fernanda; Umans, Benjamin D; Gerding, Matthew A; Davis, Brigid M; Waldor, Matthew K

    2016-08-01

    Gram-negative bacteria are notoriously resistant to a variety of high-molecular-weight antibiotics due to the limited permeability of their outer membrane (OM). The basis of OM barrier function and the genetic factors required for its maintenance remain incompletely understood. Here, we employed transposon insertion sequencing to identify genes required for Vibrio cholerae resistance to vancomycin and bacitracin, antibiotics that are thought to be too large to efficiently penetrate the OM. The screen yielded several genes whose protein products are predicted to participate in processes important for OM barrier functions and for biofilm formation. In addition, we identified a novel factor, designated vigA (for vancomycin inhibits growth), that has not previously been characterized or linked to outer membrane function. The vigA open reading frame (ORF) codes for an inner membrane protein, and in its absence, cells became highly sensitive to glycopeptide antibiotics (vancomycin and ramoplanin) and bacitracin but not to other large antibiotics or detergents. In contrast to wild-type (WT) cells, the vigA mutant was stained with fluorescent vancomycin. These observations suggest that VigA specifically prevents the periplasmic accumulation of certain large antibiotics without exerting a general role in the maintenance of OM integrity. We also observed marked interspecies variability in the susceptibilities of Gram-negative pathogens to glycopeptides and bacitracin. Collectively, our findings suggest that the OM barrier is not absolute but rather depends on specific OM-antibiotic interactions. PMID:27216069

  15. Screening genetically modified organisms using multiplex-PCR coupled with oligonucleotide microarray.

    Science.gov (United States)

    Xu, Jia; Miao, Haizhen; Wu, Houfei; Huang, Wensheng; Tang, Rong; Qiu, Minyan; Wen, Jianguo; Zhu, Shuifang; Li, Yao

    2006-07-15

    In this research, we developed a multiplex polymerase chain reaction (multiplex-PCR) coupled with a DNA microarray system simultaneously aiming at many targets in a consecutive reaction to detect a genetically modified organism (GMO). There are a total of 20 probes for detecting a GMO in a DNA microarray which can be classified into three categories according to their purpose: the first for screening GMO from un-transgenic plants based on the common elements such as promoter, reporter and terminator genes; the second for specific gene confirmation based on the target gene sequences such as herbicide-resistance or insect-resistance genes; the third for species-specific genes which the sequences are unique for different plant species. To ensure the reliability of this method, different kinds of positive and negative controls were used in DNA microarray. Commercial GM soybean, maize, rapeseed and cotton were identified by means of this method and further confirmed by PCR analysis and sequencing. The results indicate that this method discriminates between the GMOs very quickly and in a cost-saving and more time efficient way. It can detect more than 95% of currently commercial GMO plants and the limits of detection are 0.5% for soybean and 1% for maize. This method is proved to be a new method for routine analysis of GMOs.

  16. A Sleeping Beauty forward genetic screen identifies new genes and pathways driving osteosarcoma development and metastasis

    Science.gov (United States)

    Moriarity, Branden S; Otto, George M; Rahrmann, Eric P; Rathe, Susan K; Wolf, Natalie K; Weg, Madison T; Manlove, Luke A; LaRue, Rebecca S; Temiz, Nuri A; Molyneux, Sam D; Choi, Kwangmin; Holly, Kevin J; Sarver, Aaron L; Scott, Milcah C; Forster, Colleen L; Modiano, Jaime F; Khanna, Chand; Hewitt, Stephen M; Khokha, Rama; Yang, Yi; Gorlick, Richard; Dyer, Michael A; Largaespada, David A

    2016-01-01

    Osteosarcomas are sarcomas of the bone, derived from osteoblasts or their precursors, with a high propensity to metastasize. Osteosarcoma is associated with massive genomic instability, making it problematic to identify driver genes using human tumors or prototypical mouse models, many of which involve loss of Trp53 function. To identify the genes driving osteosarcoma development and metastasis, we performed a Sleeping Beauty (SB) transposon-based forward genetic screen in mice with and without somatic loss of Trp53. Common insertion site (CIS) analysis of 119 primary tumors and 134 metastatic nodules identified 232 sites associated with osteosarcoma development and 43 sites associated with metastasis, respectively. Analysis of CIS-associated genes identified numerous known and new osteosarcoma-associated genes enriched in the ErbB, PI3K-AKT-mTOR and MAPK signaling pathways. Lastly, we identified several oncogenes involved in axon guidance, including Sema4d and Sema6d, which we functionally validated as oncogenes in human osteosarcoma. PMID:25961939

  17. A Forward Genetic Screen for Molecules Involved in Pheromone-Induced Dauer Formation in Caenorhabditis elegans

    Science.gov (United States)

    Neal, Scott J.; Park, JiSoo; DiTirro, Danielle; Yoon, Jason; Shibuya, Mayumi; Choi, Woochan; Schroeder, Frank C.; Butcher, Rebecca A.; Kim, Kyuhyung; Sengupta, Piali

    2016-01-01

    Animals must constantly assess their surroundings and integrate sensory cues to make appropriate behavioral and developmental decisions. Pheromones produced by conspecific individuals provide critical information regarding environmental conditions. Ascaroside pheromone concentration and composition are instructive in the decision of Caenorhabditis elegans to either develop into a reproductive adult or enter into the stress-resistant alternate dauer developmental stage. Pheromones are sensed by a small set of sensory neurons, and integrated with additional environmental cues, to regulate neuroendocrine signaling and dauer formation. To identify molecules required for pheromone-induced dauer formation, we performed an unbiased forward genetic screen and identified phd (pheromone response-defective dauer) mutants. Here, we describe new roles in dauer formation for previously identified neuronal molecules such as the WD40 domain protein QUI-1 and MACO-1 Macoilin, report new roles for nociceptive neurons in modulating pheromone-induced dauer formation, and identify tau tubulin kinases as new genes involved in dauer formation. Thus, phd mutants define loci required for the detection, transmission, or integration of pheromone signals in the regulation of dauer formation. PMID:26976437

  18. Data on screening and identification of genetically modified papaya in food supplements.

    Science.gov (United States)

    Prins, Theo W; Scholtens, Ingrid M J; Bak, Arno W; van Dijk, Jeroen P; Voorhuijzen, Marleen M; Laurensse, Emile J; Kok, Esther J

    2016-12-01

    This article contains data related to the research article entitled "A case study to determine the geographical origin of unknown GM papaya in routine food sample analysis, followed by identification of papaya events 16-0-1 and 18-2-4" (Prins et al., 2016) [1]. Quantitative real-time PCR (qPCR) with targets that are putatively present in genetically modified (GM) papaya was used as a first screening to narrow down the vast array of candidates. The combination of elements P-nos and nptII was further confirmed by amplification and subsequent sequencing of the P-nos/nptII construct. Next, presence of the candidate GM papayas 16-0-1 and 18-2-4 were investigated by amplification and sequencing of event-spanning regions on the left and right border. This data article reports the Cq values for GM elements, the nucleotide sequence of the P-nos/nptII construct and the presence of GM papaya events 18-2-4 and/or 16-0-1 in five samples that were randomly sampled to be analysed in the framework of the official Dutch GMO monitoring program for food. PMID:27626052

  19. Screening and genetic manipulation of green organisms for establishment of biological life support systems in space.

    Science.gov (United States)

    Saei, Amir Ata; Omidi, Amir Ali; Barzegari, Abolfazl

    2013-01-01

    Curiosity has driven humankind to explore and conquer space. However, today, space research is not a means to relieve this curiosity anymore, but instead has turned into a need. To support the crew in distant expeditions, supplies should either be delivered from the Earth, or prepared for short durations through physiochemical methods aboard the space station. Thus, research continues to devise reliable regenerative systems. Biological life support systems may be the only answer to human autonomy in outposts beyond Earth. For construction of an artificial extraterrestrial ecosystem, it is necessary to search for highly adaptable super-organisms capable of growth in harsh space environments. Indeed, a number of organisms have been proposed for cultivation in space. Meanwhile, some manipulations can be done to increase their photosynthetic potential and stress tolerance. Genetic manipulation and screening of plants, microalgae and cyanobacteria is currently a fascinating topic in space bioengineering. In this commentary, we will provide a viewpoint on the realities, limitations and promises in designing biological life support system based on engineered and/or selected green organism. Special focus will be devoted to the engineering of key photosynthetic enzymes in pioneer green organisms and their potential use in establishment of transgenic photobioreactors in space. PMID:22992434

  20. Chemical-genetic profile analysis of five inhibitory compounds in yeast

    Directory of Open Access Journals (Sweden)

    Alamgir Md

    2010-08-01

    Full Text Available Abstract Background Chemical-genetic profiling of inhibitory compounds can lead to identification of their modes of action. These profiles can help elucidate the complex interactions between small bioactive compounds and the cell machinery, and explain putative gene function(s. Results Colony size reduction was used to investigate the chemical-genetic profile of cycloheximide, 3-amino-1,2,4-triazole, paromomycin, streptomycin and neomycin in the yeast Saccharomyces cerevisiae. These compounds target the process of protein biosynthesis. More than 70,000 strains were analyzed from the array of gene deletion mutant yeast strains. As expected, the overall profiles of the tested compounds were similar, with deletions for genes involved in protein biosynthesis being the major category followed by metabolism. This implies that novel genes involved in protein biosynthesis could be identified from these profiles. Further investigations were carried out to assess the activity of three profiled genes in the process of protein biosynthesis using relative fitness of double mutants and other genetic assays. Conclusion Chemical-genetic profiles provide insight into the molecular mechanism(s of the examined compounds by elucidating their potential primary and secondary cellular target sites. Our follow-up investigations into the activity of three profiled genes in the process of protein biosynthesis provided further evidence concerning the usefulness of chemical-genetic analyses for annotating gene functions. We termed these genes TAE2, TAE3 and TAE4 for translation associated elements 2-4.

  1. Prediction of Synergism from Chemical-Genetic Interactions by Machine Learning.

    Science.gov (United States)

    Wildenhain, Jan; Spitzer, Michaela; Dolma, Sonam; Jarvik, Nick; White, Rachel; Roy, Marcia; Griffiths, Emma; Bellows, David S; Wright, Gerard D; Tyers, Mike

    2015-12-23

    The structure of genetic interaction networks predicts that, analogous to synthetic lethal interactions between non-essential genes, combinations of compounds with latent activities may exhibit potent synergism. To test this hypothesis, we generated a chemical-genetic matrix of 195 diverse yeast deletion strains treated with 4,915 compounds. This approach uncovered 1,221 genotype-specific inhibitors, which we termed cryptagens. Synergism between 8,128 structurally disparate cryptagen pairs was assessed experimentally and used to benchmark predictive algorithms. A model based on the chemical-genetic matrix and the genetic interaction network failed to accurately predict synergism. However, a combined random forest and Naive Bayesian learner that associated chemical structural features with genotype-specific growth inhibition had strong predictive power. This approach identified previously unknown compound combinations that exhibited species-selective toxicity toward human fungal pathogens. This work demonstrates that machine learning methods trained on unbiased chemical-genetic interaction data may be widely applicable for the discovery of synergistic combinations in different species. PMID:27136353

  2. Design and optimization of pulsed Chemical Exchange Saturation Transfer MRI using a multiobjective genetic algorithm.

    Science.gov (United States)

    Yoshimaru, Eriko S; Randtke, Edward A; Pagel, Mark D; Cárdenas-Rodríguez, Julio

    2016-02-01

    Pulsed Chemical Exchange Saturation Transfer (CEST) MRI experimental parameters and RF saturation pulse shapes were optimized using a multiobjective genetic algorithm. The optimization was carried out for RF saturation duty cycles of 50% and 90%, and results were compared to continuous wave saturation and Gaussian waveform. In both simulation and phantom experiments, continuous wave saturation performed the best, followed by parameters and shapes optimized by the genetic algorithm and then followed by Gaussian waveform. We have successfully demonstrated that the genetic algorithm is able to optimize pulse CEST parameters and that the results are translatable to clinical scanners. PMID:26778301

  3. Screening of 397 chemicals and development of a quantitative structure-activity relationship model for androgen receptor antagonism

    DEFF Research Database (Denmark)

    Vinggaard, Annemarie; Niemelä, Jay Russell; Wedebye, Eva Bay;

    2008-01-01

    We have screened 397 chemicals for human androgen receptor (AR) antagonism by a sensitive reporter gene assay to generate data for the development of a quantitative structure-activity relationship (QSAR) model. A total of 523 chemicals comprising data on 292 chemicals from our laboratory and data...... on 231 chemicals from the literature constituted the training set for the model. The chemicals were selected with the purpose of representing a wide range of chemical structures (e.g., organochlorines and polycyclic aromatic hydrocarbons) and various functions (e.g., natural hormones, pesticides......, plastizicers, plastic additives, brominated flame retardants, and roast mutagens). In addition, the intention was to obtain an equal number of positive and negative chemicals. Among our own data for the training set, 45.7% exhibited inhibitory activity against the transcriptional activity induced...

  4. A genetic screen for replication initiation defective (rid mutants in Schizosaccharomyces pombe

    Directory of Open Access Journals (Sweden)

    Locovei Alexandra M

    2010-08-01

    Full Text Available Abstract In fission yeast the intra-S phase and DNA damage checkpoints are activated in response to inhibition of DNA replication or DNA damage, respectively. The intra-S phase checkpoint responds to stalled replication forks leading to the activation of the Cds1 kinase that both delays cell cycle progression and stabilizes DNA replication forks. The DNA damage checkpoint, that operates during the G2 phase of the cell cycle delays mitotic progression through activation of the checkpoint kinase, Chk1. Delay of the cell cycle is believed to be essential to allow time for either replication restart (in S phase or DNA damage repair (in G2. Previously, our laboratory showed that fission yeast cells deleted for the N-terminal half of DNA polymerase ε (Cdc20 are delayed in S phase, but surprisingly require Chk1 rather than Cds1 to maintain cell viability. Several additional DNA replication mutants were then tested for their dependency on Chk1 or Cds1 when grown under semi-permissive temperatures. We discovered that mutants defective in DNA replication initiation are sensitive only to loss of Chk1, whilst mutations that inhibit DNA replication elongation are sensitive to loss of both Cds1 and Chk1. To confirm that the Chk1-sensitive, Cds1-insensitive phenotype (rid phenotype is specific to mutants defective in DNA replication initiation, we completed a genetic screen for cell cycle mutants that require Chk1, but not Cds1 to maintain cell viability when grown at semi-permissive temperatures. Our screen identified two mutants, rid1-1 and rid2-1, that are defective in Orc1 and Mcm4, respectively. Both mutants show defects in DNA replication initiation consistent with our hypothesis that the rid phenotype is replication initiation specific. In the case of Mcm4, the mutation has been mapped to a highly conserved region of the protein that appears to be required for DNA replication initiation, but not elongation. Therefore, we conclude that the cellular

  5. piggyBac transposon somatic mutagenesis with an activated reporter and tracker (PB-SMART for genetic screens in mice.

    Directory of Open Access Journals (Sweden)

    Sean F Landrette

    Full Text Available Somatic forward genetic screens have the power to interrogate thousands of genes in a single animal. Retroviral and transposon mutagenesis systems in mice have been designed and deployed in somatic tissues for surveying hematopoietic and solid tumor formation. In the context of cancer, the ability to visually mark mutant cells would present tremendous advantages for identifying tumor formation, monitoring tumor growth over time, and tracking tumor infiltrations and metastases into wild-type tissues. Furthermore, locating mutant clones is a prerequisite for screening and analyzing most other somatic phenotypes. For this purpose, we developed a system using the piggyBac (PB transposon for somatic mutagenesis with an activated reporter and tracker, called PB-SMART. The PB-SMART mouse genetic screening system can simultaneously induce somatic mutations and mark mutated cells using bioluminescence or fluorescence. The marking of mutant cells enable analyses that are not possible with current somatic mutagenesis systems, such as tracking cell proliferation and tumor growth, detecting tumor cell infiltrations, and reporting tissue mutagenesis levels by a simple ex vivo visual readout. We demonstrate that PB-SMART is highly mutagenic, capable of tumor induction with low copy transposons, which facilitates the mapping and identification of causative insertions. We further integrated a conditional transposase with the PB-SMART system, permitting tissue-specific mutagenesis with a single cross to any available Cre line. Targeting the germline, the system could also be used to conduct F1 screens. With these features, PB-SMART provides an integrated platform for individual investigators to harness the power of somatic mutagenesis and phenotypic screens to decipher the genetic basis of mammalian biology and disease.

  6. Increasing Live Birth Rate by Preimplantation Genetic Screening of Pooled Polar Bodies Using Array Comparative Genomic Hybridization

    OpenAIRE

    Michael Feichtinger; Tina Stopp; Christian Göbl; Elisabeth Feichtinger; Enrico Vaccari; Ulrike Mädel; Franco Laccone; Monika Stroh-Weigert; Markus Hengstschläger; Wilfried Feichtinger; Jürgen Neesen

    2015-01-01

    Meiotic errors during oocyte maturation are considered the major contributors to embryonic aneuploidy and failures in human IVF treatment. Various technologies have been developed to screen polar bodies, blastomeres and trophectoderm cells for chromosomal aberrations. Array-CGH analysis using bacterial artificial chromosome (BAC) arrays is widely applied for preimplantation genetic diagnosis (PGD) using single cells. Recently, an increase in the pregnancy rate has been demonstrated using arra...

  7. Economic Efficiency of Genetic Screening to Inform the Use of Abacavir Sulfate in the Treatment of HIV

    OpenAIRE

    Kauf, Teresa L.; Farkouh, Raymond A.; Earnshaw, Stephanie R; Maria E. Watson; Penny Maroudas; Mike G. Chambers

    2010-01-01

    Background: Abacavir sulfate (abacavir) is associated with a hypersensitivity reaction (HSR) that affects 5-8% of patients. While serious complications are rare, failure to identify it, or abacavir re-challenge following HSR, can be fatal. Genetic screening for HLA-B*5701 can identify patients who are likely to experience an HSR and reduces the incidence of the reaction. Objective: We assessed the intrinsic and practical value, from the US healthcare system perspective, of prospective HLA-B*5...

  8. Screening and genetic counseling for beta-thalassemia trait in a population unselected for interest: comparison of three counseling methods.

    OpenAIRE

    Rowley, P T; Lipkin, M; Fisher, L

    1984-01-01

    We have assessed the effects of screening and genetic counseling for beta-thalassemia trait on knowledge, attitudes, and behavior in a prospective, controlled study of randomly selected adult members of a health maintenance organization. We report here that knowledge of manifestations and of inheritance of thalassemia, previously reported to be high immediately after counseling, were well maintained at 2 and 10 months following counseling. There was no detectable impairment of self-concept. M...

  9. An evaluation of chemical screening test kits for lead in paint

    Energy Technology Data Exchange (ETDEWEB)

    Oglesby, L.S.

    1996-04-01

    The Residential Lead-Based Paint Hazard Reduction Act (Title X) requires abatement and management of lead-based paint. The purpose of this study was to evaluate three chemical screening test kits using materials and methods from one study and subjecting the results to the statistical analysis of another. The three kits were used to predict the presence of lead in paint at ten weight concentrations from 0.04 to 3.97%. Paint was applied to four wood boards yielding a sample size of 40. Four boards were painted with lead-free paint and used as blanks. All of the boards were tested with the three test kits by an untrained individual having no knowledge of the actual lead content. Sensitivity, specificity, and false positive and negative rates were calculated for the test kit results. The manufactures` detection limits, the observed sensitivity ranged from 1.00 to 0.80, specificity ranged from 1.00 to 0.42, false positive ranged from 0 to 58%, and false negatives ranged from 0 to 20%. At the 0.5% Federal threshold level, the observed sensitivity ranged from 1.00 to 0.94, specificity ranged from 1.00 to 0.5, false positives ranged from 0 to 11.1%, and false negatives ranged from 0 to 20%. The observed false positive and false negative rates for all three kits were found to be significantly lower than those reported in a previous study. These results indicate that the kits perform very well at the Federal threshold, with two of the kits having false negative rates below 12.5% and false positive rates of 3.13%. These results indicate that these two kits would probably be acceptable screening tests for lead in paint.

  10. [Genetic screening of patients with familial hypercholesterolemia and insurability for life insurance policies and disability cover policies].

    Science.gov (United States)

    Homsma, S J; Lansberg, P J; Kastelein, J J

    2004-03-01

    In the Netherlands, people with familial hypercholesterolaemia (FH) have been actively screened since 1994 by means of DNA analysis. Recently, the Stichting Opsporing Erfelijke Hypercholesterolemie (Foundation for the Detection of Familial Hypercholesterolaemia) initiated a large scale-screening programme aimed at finding all 40,000 people. The Dutch ministry of Health, Welfare and Sport is providing the financial support. Genetic screening has social implications and raises questions on insurability. The Dutch Medical Examination Act prohibits insurers from posing questions about untreatable, serious inheritable conditions for insured sums under a certain value: for life-insurance policies policies insurers can request information for the purpose of an accurate risk classification. Insurance contracts can be accepted at normal rates if the target value of LDL-cholesterol < 4 mmol/l and additional risk factors such as smoking and an abnormal BMI are absent; the risk is determined by the phenotype and clinical factors and not by the genotype.

  11. Electrochemical sensor for multiplex screening of genetically modified DNA: identification of biotech crops by logic-based biomolecular analysis.

    Science.gov (United States)

    Liao, Wei-Ching; Chuang, Min-Chieh; Ho, Ja-An Annie

    2013-12-15

    Genetically modified (GM) technique, one of the modern biomolecular engineering technologies, has been deemed as profitable strategy to fight against global starvation. Yet rapid and reliable analytical method is deficient to evaluate the quality and potential risk of such resulting GM products. We herein present a biomolecular analytical system constructed with distinct biochemical activities to expedite the computational detection of genetically modified organisms (GMOs). The computational mechanism provides an alternative to the complex procedures commonly involved in the screening of GMOs. Given that the bioanalytical system is capable of processing promoter, coding and species genes, affirmative interpretations succeed to identify specified GM event in terms of both electrochemical and optical fashions. The biomolecular computational assay exhibits detection capability of genetically modified DNA below sub-nanomolar level and is found interference-free by abundant coexistence of non-GM DNA. This bioanalytical system, furthermore, sophisticates in array fashion operating multiplex screening against variable GM events. Such a biomolecular computational assay and biosensor holds great promise for rapid, cost-effective, and high-fidelity screening of GMO.

  12. A conditional piggyBac transposition system for genetic screening in mice identifies oncogenic networks in pancreatic cancer.

    Science.gov (United States)

    Rad, Roland; Rad, Lena; Wang, Wei; Strong, Alexander; Ponstingl, Hannes; Bronner, Iraad F; Mayho, Matthew; Steiger, Katja; Weber, Julia; Hieber, Maren; Veltkamp, Christian; Eser, Stefan; Geumann, Ulf; Öllinger, Rupert; Zukowska, Magdalena; Barenboim, Maxim; Maresch, Roman; Cadiñanos, Juan; Friedrich, Mathias; Varela, Ignacio; Constantino-Casas, Fernando; Sarver, Aaron; Ten Hoeve, Jelle; Prosser, Haydn; Seidler, Barbara; Bauer, Judith; Heikenwälder, Mathias; Metzakopian, Emmanouil; Krug, Anne; Ehmer, Ursula; Schneider, Günter; Knösel, Thomas; Rümmele, Petra; Aust, Daniela; Grützmann, Robert; Pilarsky, Christian; Ning, Zemin; Wessels, Lodewyk; Schmid, Roland M; Quail, Michael A; Vassiliou, George; Esposito, Irene; Liu, Pentao; Saur, Dieter; Bradley, Allan

    2015-01-01

    Here we describe a conditional piggyBac transposition system in mice and report the discovery of large sets of new cancer genes through a pancreatic insertional mutagenesis screen. We identify Foxp1 as an oncogenic transcription factor that drives pancreatic cancer invasion and spread in a mouse model and correlates with lymph node metastasis in human patients with pancreatic cancer. The propensity of piggyBac for open chromatin also enabled genome-wide screening for cancer-relevant noncoding DNA, which pinpointed a Cdkn2a cis-regulatory region. Histologically, we observed different tumor subentities and discovered associated genetic events, including Fign insertions in hepatoid pancreatic cancer. Our studies demonstrate the power of genetic screening to discover cancer drivers that are difficult to identify by other approaches to cancer genome analysis, such as downstream targets of commonly mutated human cancer genes. These piggyBac resources are universally applicable in any tissue context and provide unique experimental access to the genetic complexity of cancer.

  13. Identification of genes important for cutaneous function revealed by a large scale reverse genetic screen in the mouse.

    Directory of Open Access Journals (Sweden)

    Tia DiTommaso

    2014-10-01

    Full Text Available The skin is a highly regenerative organ which plays critical roles in protecting the body and sensing its environment. Consequently, morbidity and mortality associated with skin defects represent a significant health issue. To identify genes important in skin development and homeostasis, we have applied a high throughput, multi-parameter phenotype screen to the conditional targeted mutant mice generated by the Wellcome Trust Sanger Institute's Mouse Genetics Project (Sanger-MGP. A total of 562 different mouse lines were subjected to a variety of tests assessing cutaneous expression, macroscopic clinical disease, histological change, hair follicle cycling, and aberrant marker expression. Cutaneous lesions were associated with mutations in 23 different genes. Many of these were not previously associated with skin disease in the organ (Mysm1, Vangl1, Trpc4ap, Nom1, Sparc, Farp2, and Prkab1, while others were ascribed new cutaneous functions on the basis of the screening approach (Krt76, Lrig1, Myo5a, Nsun2, and Nf1. The integration of these skin specific screening protocols into the Sanger-MGP primary phenotyping pipelines marks the largest reported reverse genetic screen undertaken in any organ and defines approaches to maximise the productivity of future projects of this nature, while flagging genes for further characterisation.

  14. An Evolution-Based Screen for Genetic Differentiation between Anopheles Sister Taxa Enriches for Detection of Functional Immune Factors.

    Directory of Open Access Journals (Sweden)

    Christian Mitri

    2015-12-01

    Full Text Available Nucleotide variation patterns across species are shaped by the processes of natural selection, including exposure to environmental pathogens. We examined patterns of genetic variation in two sister species, Anopheles gambiae and Anopheles coluzzii, both efficient natural vectors of human malaria in West Africa. We used the differentiation signature displayed by a known coordinate selective sweep of immune genes APL1 and TEP1 in A. coluzzii to design a population genetic screen trained on the sweep, classified a panel of 26 potential immune genes for concordance with the signature, and functionally tested their immune phenotypes. The screen results were strongly predictive for genes with protective immune phenotypes: genes meeting the screen criteria were significantly more likely to display a functional phenotype against malaria infection than genes not meeting the criteria (p = 0.0005. Thus, an evolution-based screen can efficiently prioritize candidate genes for labor-intensive downstream functional testing, and safely allow the elimination of genes not meeting the screen criteria. The suite of immune genes with characteristics similar to the APL1-TEP1 selective sweep appears to be more widespread in the A. coluzzii genome than previously recognized. The immune gene differentiation may be a consequence of adaptation of A. coluzzii to new pathogens encountered in its niche expansion during the separation from A. gambiae, although the role, if any of natural selection by Plasmodium is unknown. Application of the screen allowed identification of new functional immune factors, and assignment of new functions to known factors. We describe biochemical binding interactions between immune proteins that underlie functional activity for malaria infection, which highlights the interplay between pathogen specificity and the structure of immune complexes. We also find that most malaria-protective immune factors display phenotypes for either human or rodent

  15. An Evolution-Based Screen for Genetic Differentiation between Anopheles Sister Taxa Enriches for Detection of Functional Immune Factors.

    Science.gov (United States)

    Mitri, Christian; Bischoff, Emmanuel; Takashima, Eizo; Williams, Marni; Eiglmeier, Karin; Pain, Adrien; Guelbeogo, Wamdaogo M; Gneme, Awa; Brito-Fravallo, Emma; Holm, Inge; Lavazec, Catherine; Sagnon, N'Fale; Baxter, Richard H; Riehle, Michelle M; Vernick, Kenneth D

    2015-12-01

    Nucleotide variation patterns across species are shaped by the processes of natural selection, including exposure to environmental pathogens. We examined patterns of genetic variation in two sister species, Anopheles gambiae and Anopheles coluzzii, both efficient natural vectors of human malaria in West Africa. We used the differentiation signature displayed by a known coordinate selective sweep of immune genes APL1 and TEP1 in A. coluzzii to design a population genetic screen trained on the sweep, classified a panel of 26 potential immune genes for concordance with the signature, and functionally tested their immune phenotypes. The screen results were strongly predictive for genes with protective immune phenotypes: genes meeting the screen criteria were significantly more likely to display a functional phenotype against malaria infection than genes not meeting the criteria (p = 0.0005). Thus, an evolution-based screen can efficiently prioritize candidate genes for labor-intensive downstream functional testing, and safely allow the elimination of genes not meeting the screen criteria. The suite of immune genes with characteristics similar to the APL1-TEP1 selective sweep appears to be more widespread in the A. coluzzii genome than previously recognized. The immune gene differentiation may be a consequence of adaptation of A. coluzzii to new pathogens encountered in its niche expansion during the separation from A. gambiae, although the role, if any of natural selection by Plasmodium is unknown. Application of the screen allowed identification of new functional immune factors, and assignment of new functions to known factors. We describe biochemical binding interactions between immune proteins that underlie functional activity for malaria infection, which highlights the interplay between pathogen specificity and the structure of immune complexes. We also find that most malaria-protective immune factors display phenotypes for either human or rodent malaria, with

  16. A Genetic Mosaic Screen Reveals Ecdysone-Responsive Genes Regulating Drosophila Oogenesis.

    Science.gov (United States)

    Ables, Elizabeth T; Hwang, Grace H; Finger, Danielle S; Hinnant, Taylor D; Drummond-Barbosa, Daniela

    2016-01-01

    Multiple aspects of Drosophila oogenesis, including germline stem cell activity, germ cell differentiation, and follicle survival, are regulated by the steroid hormone ecdysone. While the transcriptional targets of ecdysone signaling during development have been studied extensively, targets in the ovary remain largely unknown. Early studies of salivary gland polytene chromosomes led to a model in which ecdysone stimulates a hierarchical transcriptional cascade, wherein a core group of ecdysone-sensitive transcription factors induce tissue-specific responses by activating secondary branches of transcriptional targets. More recently, genome-wide approaches have identified hundreds of putative ecdysone-responsive targets. Determining whether these putative targets represent bona fide targets in vivo, however, requires that they be tested via traditional mutant analysis in a cell-type specific fashion. To investigate the molecular mechanisms whereby ecdysone signaling regulates oogenesis, we used genetic mosaic analysis to screen putative ecdysone-responsive genes for novel roles in the control of the earliest steps of oogenesis. We identified a cohort of genes required for stem cell maintenance, stem and progenitor cell proliferation, and follicle encapsulation, growth, and survival. These genes encode transcription factors, chromatin modulators, and factors required for RNA transport, stability, and ribosome biogenesis, suggesting that ecdysone might control a wide range of molecular processes during oogenesis. Our results suggest that, although ecdysone target genes are known to have cell type-specific roles, many ecdysone response genes that control larval or pupal cell types at developmental transitions are used reiteratively in the adult ovary. These results provide novel insights into the molecular mechanisms by which ecdysone signaling controls oogenesis, laying new ground for future studies. PMID:27226164

  17. Automated, quantitative cognitive/behavioral screening of mice: for genetics, pharmacology, animal cognition and undergraduate instruction.

    Science.gov (United States)

    Gallistel, C R; Balci, Fuat; Freestone, David; Kheifets, Aaron; King, Adam

    2014-02-26

    We describe a high-throughput, high-volume, fully automated, live-in 24/7 behavioral testing system for assessing the effects of genetic and pharmacological manipulations on basic mechanisms of cognition and learning in mice. A standard polypropylene mouse housing tub is connected through an acrylic tube to a standard commercial mouse test box. The test box has 3 hoppers, 2 of which are connected to pellet feeders. All are internally illuminable with an LED and monitored for head entries by infrared (IR) beams. Mice live in the environment, which eliminates handling during screening. They obtain their food during two or more daily feeding periods by performing in operant (instrumental) and Pavlovian (classical) protocols, for which we have written protocol-control software and quasi-real-time data analysis and graphing software. The data analysis and graphing routines are written in a MATLAB-based language created to simplify greatly the analysis of large time-stamped behavioral and physiological event records and to preserve a full data trail from raw data through all intermediate analyses to the published graphs and statistics within a single data structure. The data-analysis code harvests the data several times a day and subjects it to statistical and graphical analyses, which are automatically stored in the "cloud" and on in-lab computers. Thus, the progress of individual mice is visualized and quantified daily. The data-analysis code talks to the protocol-control code, permitting the automated advance from protocol to protocol of individual subjects. The behavioral protocols implemented are matching, autoshaping, timed hopper-switching, risk assessment in timed hopper-switching, impulsivity measurement, and the circadian anticipation of food availability. Open-source protocol-control and data-analysis code makes the addition of new protocols simple. Eight test environments fit in a 48 in x 24 in x 78 in cabinet; two such cabinets (16 environments) may be

  18. Improving toxicity screening and drug development by using genetically defined strains.

    Science.gov (United States)

    Festing, Michael F W

    2010-01-01

    According to the US Food and Drugs Administration (Food and Drug Administration (2004) Challenge and opportunity on the critical path to new medical products.) "The inability to better assess and predict product safety leads to failures during clinical development and, occasionally, after marketing". This increases the cost of new drugs as clinical trials are even more expensive than pre-clinical testing.One relatively easy way of improving toxicity testing is to improve the design of animal experiments. A fundamental principle when designing an experiment is to control all variables except the one of interest: the treatment. Toxicologist and pharmacologists have widely ignored this principle by using genetically heterogeneous "outbred" rats and mice, increasing the chance of false-negative results. By using isogenic (inbred or F1 hybrid, see Note 1) rats and mice instead of outbred stocks the signal/noise ratio and the power of the experiments can be increased at little extra cost whilst using no more animals. Moreover, the power of the experiment can be further increased by using more than one strain, as this reduces the chance of selecting one which is resistant to the test chemical. This can also be done without increasing the total number of animals by using a factorial experimental design, e.g. if the ten outbred animals per treatment group in a 28-day toxicity test were replaced by two animals of each of five strains (still ten animals per treatment group) selected to be as genetically diverse as possible, this would increase the signal/noise ratio and power of the experiment. This would allow safety to be assessed using the most sensitive strain.Toxicologists should also consider making more use of the mouse instead of the rat. They are less costly to maintain, use less test substance, there are many inbred and genetically modified strains, and it is easier to identify gene loci controlling variation in response to xenobiotics in this species.We demonstrate

  19. Back-propagation network improved by conjugate gradient based on genetic algorithm in QSAR study on endocrine disrupting chemicals

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Since the complexity and structural diversity of man-made compounds are considered,quantitative structure-activity relationships(QSARs)-based fast screening approaches are urgently needed for the assessment of the potential risk of endocrine disrupting chemicals(EDCs).The artificial neural networks(ANN)are capable of recognizing highly nonlinear relationships,so it will have a bright application prospect in building high-quality QSAR models.As a popular supervised training algorithm in ANN,back-propagation(BP)converges slowly and immerses in vibration frequently.In this paper,a research strategy that BP neural network was improved by conjugate gradient(CG)algorithm with a variable selection method based on genetic algorithm was applied to investigate the QSAR of EDCs.This resulted in a robust and highly predictive ANN model with R2 of 0.845 for the training set,q2 pred of 0.81 and root-mean-square error(RMSE) of 0.688 for the test set.The result shows that our method can provide a feasible and practical tool for the rapid screening of the estrogen activity of organic compounds.

  20. Metabolic and Genetic Screening of Electromagnetic Hypersensitive Subjects as a Feasible Tool for Diagnostics and Intervention

    Directory of Open Access Journals (Sweden)

    Chiara De Luca

    2014-01-01

    Full Text Available Growing numbers of “electromagnetic hypersensitive” (EHS people worldwide self-report severely disabling, multiorgan, non-specific symptoms when exposed to low-dose electromagnetic radiations, often associated with symptoms of multiple chemical sensitivity (MCS and/or other environmental “sensitivity-related illnesses” (SRI. This cluster of chronic inflammatory disorders still lacks validated pathogenetic mechanism, diagnostic biomarkers, and management guidelines. We hypothesized that SRI, not being merely psychogenic, may share organic determinants of impaired detoxification of common physic-chemical stressors. Based on our previous MCS studies, we tested a panel of 12 metabolic blood redox-related parameters and of selected drug-metabolizing-enzyme gene polymorphisms, on 153 EHS, 147 MCS, and 132 control Italians, confirming MCS altered (P<0.05–0.0001 glutathione-(GSH, GSH-peroxidase/S-transferase, and catalase erythrocyte activities. We first described comparable—though milder—metabolic pro-oxidant/proinflammatory alterations in EHS with distinctively increased plasma coenzyme-Q10 oxidation ratio. Severe depletion of erythrocyte membrane polyunsaturated fatty acids with increased ω6/ω3 ratio was confirmed in MCS, but not in EHS. We also identified significantly (P=0.003 altered distribution-versus-control of the CYP2C19*1/*2 SNP variants in EHS, and a 9.7-fold increased risk (OR: 95% C.I.=1.3–74.5 of developing EHS for the haplotype (nullGSTT1 + (nullGSTM1 variants. Altogether, results on MCS and EHS strengthen our proposal to adopt this blood metabolic/genetic biomarkers’ panel as suitable diagnostic tool for SRI.

  1. Physico-chemical screening of accessions of Jatropha curcas for biodiesel production

    International Nuclear Information System (INIS)

    Biodiesel is an alternative environmentally friendly fuel made from renewable biological sources such as vegetable oils and animal fats. The present report deals with screening of 14 accessions of Jatropha curcas collected from all over India to find the most suitable ones for production of Biodiesel. From the 14 accessions of J. curcas located in the plantation at Osmania University, 4 accessions were initially selected on the basis of traits like general appearance, pest resistance, seed yield and seed-oil content. Further, the seed-oil of these 4 accessions was characterized by physico-chemical analysis to identify the elite accessions for production of biodiesel. Highest 1000-seed weight (640 g) and highest percentage seed-oil content (50.16) (extracted by Soxhlet method with hexane as the solvent) was recorded in the “KM” accession. The transesterification process is affected by the presence of high free fatty acids (recorded in “MB” accession) and high moisture content (recorded in “KM” accession) of the seed-oil which also interfere with the separation of fatty esters and glycerol during production of Biodiesel. Further, high phosphorus content and iodine number (recorded in “MB” accession) interfere with conversion of seed-oil to Biodiesel. In the above context, in spite of its yield being lower, the seed-oil of the “RSAD” accession was found to be most suitable for Biodiesel production followed by “KM”, “F.W.B” and “MB” accessions, since it contains lower free fatty acids, acid value, viscosity, diglycerides and iodine number. -- Highlights: ► We analyzed Indian Jatropha accessions for yield and quality. ► Elite accessions were selected for physico-chemical analysis of seed-oil. ► Four elite accessions identified as good candidates for Biodiesel production. ► The “RSAD” accession was found to be the best suited for biodiesel.

  2. Zebrafish chemical screening reveals the impairment of dopaminergic neuronal survival by cardiac glycosides.

    Directory of Open Access Journals (Sweden)

    Yaping Sun

    Full Text Available Parkinson's disease is a neurodegenerative disorder characterized by the prominent degeneration of dopaminergic (DA neurons among other cell types. Here we report a first chemical screen of over 5,000 compounds in zebrafish, aimed at identifying small molecule modulators of DA neuron development or survival. We find that Neriifolin, a member of the cardiac glycoside family of compounds, impairs survival but not differentiation of both zebrafish and mammalian DA neurons. Cardiac glycosides are inhibitors of Na(+/K(+ ATPase activity and widely used for treating heart disorders. Our data suggest that Neriifolin impairs DA neuronal survival by targeting the neuronal enriched Na(+/K(+ ATPase α3 subunit (ATP1A3. Modulation of ionic homeostasis, knockdown of p53, or treatment with antioxidants protects DA neurons from Neriifolin-induced death. These results reveal a previously unknown effect of cardiac glycosides on DA neuronal survival and suggest that it is mediated through ATP1A3 inhibition, oxidative stress, and p53. They also elucidate potential approaches for counteracting the neurotoxicity of this valuable class of medications.

  3. A chemical screen to identify inducers of the mitochondrial unfolded protein response in C. elegans.

    Science.gov (United States)

    Rauthan, Manish; Pilon, Marc

    2015-01-01

    We previously showed that inhibition of the mevalonate pathway in C. elegans causes inhibition of protein prenylation, developmental arrest and lethality. We also showed that constitutive activation of the mitochondrial unfolded protein response, UPR(mt), is an effective way for C. elegans to become resistant to the negative effects of mevalonate pathway inhibition. This was an important finding since statins, a drug class prescribed to lower cholesterol levels in patients, act by inhibiting the mevalonate pathway, and it is therefore possible that some of their undesirable side effects could be alleviated by activating the UPR(mt). Here, we screened a chemical library and identified 4 compounds that specifically activated the UPR(mt). One of these compounds, methacycline hydrochloride (a tetracycline antibiotic) also protected C. elegans and mammalian cells from statin toxicity. Methacycline hydrochloride and ethidium bromide, a known UPR(mt) activator, were also tested in mice: only ethidium bromide significantly activate the UPR(mt) in skeletal muscles. PMID:27123370

  4. Antibiotic pigment from desert soil actinomycetes; biological activity, purification and chemical screening

    Directory of Open Access Journals (Sweden)

    Selvameenal L

    2009-01-01

    Full Text Available An actinomycete strain, Streptomyces hygroscopicus subsp. ossamyceticus (strain D10 was isolated from Thar Desert soil, Rajasthan during the year 2006 and found to produce a yellow color pigment with antibiotic activity. Crude pigment was produced from strain D10 by solid state fermentation using wheat bran medium followed by extraction with ethyl acetate. The antimicrobial activity of the crude pigment was evaluated against drug resistant pathogens such as methicillin-resistant Staphylococcus aureus, vancomycin-resistant Staphylococcus aureus, extended spectrum b-lactamase producing cultures of Escherichia coli, Pseudomonas aeruginosa and Klebsiella sp. About 420 mg of crude pigment was produced per 10 g of wheat bran medium. In the disc diffusion method the crude ethyl acetate extract showed a minimum of 10 mm inhibition against Klebsiella sp. and maximum of 19 mm of inhibition against Escherichia coli. The crude pigment was partially purified using thin layer chromatography with the solvent system chloroform:methanol (30:70 and the Rf value was calculated as 0.768. Antimicrobial activity of the partially purified compound from thin layer chromatography was determined using the bioautography method. The purified pigment showed minimum of 15 mm inhibition against Klebsiella sp. and a maximum of 23 mm of inhibition against vancomycin-resistant Staphylococcus aureus in the disc diffusion method. Based on the results of chemical screening, the pigment was tentatively identified as group of sugar containing molecules.

  5. "I'm Healthy, It's Not Going To Be Me": Exploring experiences of carriers identified through a population reproductive genetic carrier screening panel in Australia.

    Science.gov (United States)

    Beard, Catherine A; Amor, David J; Di Pietro, Louisa; Archibald, Alison D

    2016-08-01

    Advancing genetic testing technologies mean that population-based carrier screening for multiple inherited conditions is now available. As the number of genetic conditions being screened increases, there is a need for research into how people experience these screening programs. This research aimed to explore how women experience simultaneous carrier screening for three inherited conditions: cystic fibrosis (CF), spinal muscular atrophy (SMA), and fragile X syndrome (FXS). A qualitative approach was adopted using in-depth semi-structured interviews to explore the experiences of ten female participants: five SMA carriers, three CF carriers, and two FXS premutation carriers. Eight participants were pregnant when offered screening by their general practitioner or obstetrician and the decision to have screening was described as straightforward. Participants reported experiencing emotional responses such as anxiety and stress while waiting for either their partner's carrier screen result (CF or SMA carriers) or the pregnancy's CVS result (FXS carrier) and sought additional information about the relevant condition during this time. Most participants were in favor of population carrier screening for these conditions, preferably prior to conception. Genetic counselors played an essential role in supporting couples after they received a carrier result given the variable consent processes undertaken when screening was offered. Further research should focus on the development of reliable online information tailored to people receiving carrier results and strategies for raising awareness of the availability of population carrier screening within the community. © 2016 Wiley Periodicals, Inc. PMID:27150953

  6. Costs and effects of genetic screening with application to cystic fibrosis and fragile X syndrome

    NARCIS (Netherlands)

    M.F. Wildhagen (Mark)

    1999-01-01

    textabstractTwo to six percent of all newborn children have a disorder with a genetic cause (1-3). For an increasing number of these diseases, the precise genetic cause is known and this can lead to new treatment opportunities (see Appendix A for a basic description of the mechanisms of genetic inhe

  7. Impact of human genome initiative-derived technology on genetic testing, screening and counseling: Cultural, ethical and legal issues

    Energy Technology Data Exchange (ETDEWEB)

    Trottier, R.W.; Hodgin, F.C.; Imara, M.; Phoenix, D.; Lybrook, S. (Morehouse Coll., Atlanta, GA (United States). School of Medicine); Crandall, L.A.; Moseley, R.E.; Armotrading, D. (Florida Univ., Gainesville, FL (United States). Coll. of Medicine)

    1993-01-01

    Genetic medical services provided by the Georgia Division of Public Health in two northern and two central districts are compared to services provided in a district in which a tertiary care facility is located. Genetics outreach public health nurses play key roles in Georgia's system of Children's Health Services Genetics Program, including significant roles as counselors and information sources on special needs social services and support organizations. Unique features of individual health districts, (e.g., the changing face of some rural communities in ethnocultural diversity and socioeconomic character), present new challenges to current and future genetics services delivery. Preparedness as to educational needs of both health professionals and the lay population is of foremost concern in light of the ever expanding knowledge and technology in medical genetics. Perspectives on genetics and an overview of services offered by a local private sector counselor are included for comparison to state supported services. The nature of the interactions which transpire between private and public genetic services resources in Georgia will be described. A special focus of this research includes issues associated with sickle cell disease newborn screening service delivery process in Georgia, with particular attention paid to patient follow-up and transition to primary care. Of particular interest to this focus is the problem of loss to follow-up in the current system. Critical factors in education and counseling of sickle cell patients and the expectations of expanding roles of primary care physicians are discussed. The Florida approach to the delivery of genetic services contrasts to the Georgia model by placing more emphasis on a consultant-specialist team approach.

  8. In vitro OECD test methods applied to screen the estrogenic effect of chemicals, used in Korea.

    Science.gov (United States)

    Lee, Hee-Seok; Park, Eun-Jung; Han, Songyi; Oh, Gyeong-Yong; Kim, Min-Hee; Kang, Hui-Seung; Suh, Jin-Hyang; Oh, Jae-Ho; Lee, Kwang-Soo; Hwang, Myung-Sil; Moon, Guiim; Hong, Jin-Hwan; Hwang, In-Gyun

    2016-09-01

    In this study, 27 chemicals found in household products, which became an issue in Korea were screened for the agonistoc and antagonistic effects against human estrogen receptor using official Organization for Economic Cooperation and Development (OECD) in vitro assays, STTA assay using ERα-HeLa-9903 cell line and BG1Luc ER TA assay. In the case of human ER agonist screening by two assays, all tested chemicals did not show agonist effect against ER. In ER antagonist test by BG1Luc ER TA assay, five surfactants α-dodecyl-ω-hydroxypoly(oxyethylene), alcohols C16-18 ethoxylated, nonylphenol, ethoxylated, 3,6,9,12,15,18,21-heptaoxatritriacontan-1-ol, and α-dodecyl-ω-hydroxypoly(oxy-1,2-ethanediyl)) were found to exhibit weak antagonistic activities. The agonist/antagonist effects against human estrogen receptor of various chemicals, used in Korea by OECD test guideline are reported in this study. These results indicated that two OECD in vitro assays will can be applied in Korea by screening of agonistic/antagonistic effects against human ER of various chemicals. PMID:27317829

  9. SYBR® Green qPCR Screening Methods for Detection of Anti-herbicide Genes in Genetically Modiifed Processed Products

    Institute of Scientific and Technical Information of China (English)

    Zhen Zhen; Lv Wei; Tang Zhi-fen; Liu Ying; Ao Jin-xia; Yuan Xiao-han; Zhang Ming-hui; Qiu You-wen; Gao Xue-jun

    2016-01-01

    The use of genetically modified organisms (GMOs) as food products becomes more and more widespread. The European Union has implemented a set of very strict procedures for the approval to grow, import and/or utilize GMOs as food or food ingredients. Thus, analytical methods for detection of GMOs are necessary in order to verify compliance with labelling requirements. There are few effective screening methods for processed GM (genetically modified) products. Three anti-herbicide genes (CP4-EPSPS,BAR andPAT) are common exogenous genes used in commercialized transgenic soybean, maize and rice. In the present study, a new SYBR® Green qPCR screening method was developed to simultaneously detect the three exogenous anti-herbicide genes and one endogenous gene in a run. We tested seven samples of representative processed products (soya lecithin, soya protein powder, chocolate beverage, infant rice cereal, maize protein powder, maize starch, and maize jam) using the developed method, and amplicons of endogenous gene and transgenic fragments were obtained from all the processed products, and the sensitivity was 0.1%. These results indicated that SYBR® Green qPCR screening method was appropriate for qualitative detection of transgenic soybean, maize and rice in processed products.

  10. Risk-based high-throughput chemical screening and prioritization using exposure models and in vitro bioactivity assays

    International Nuclear Information System (INIS)

    We present a risk-based high-throughput screening (HTS) method to identify chemicals for potential health concerns or for which additional information is needed. The method is applied to 180 organic chemicals as a case study. We first obtain information on how the chemical is used and identify relevant use scenarios (e.g., dermal application, indoor emissions). For each chemical and use scenario, exposure models are then used to calculate a chemical intake fraction, or a product intake fraction, accounting for chemical properties and the exposed population. We then combine these intake fractions with use scenario-specific estimates of chemical quantity to calculate daily intake rates (iR; mg/kg/day). These intake rates are compared to oral equivalent doses (OED; mg/kg/day), calculated from a suite of ToxCast in vitro bioactivity assays using in vitro-to-in vivo extrapolation and reverse dosimetry. Bioactivity quotients (BQs) are calculated as iR/OED to obtain estimates of potential impact associated with each relevant use scenario. Of the 180 chemicals considered, 38 had maximum iRs exceeding minimum OEDs (i.e., BQs > 1). For most of these compounds, exposures are associated with direct intake, food/oral contact, or dermal exposure. The method provides high-throughput estimates of exposure and important input for decision makers to identify chemicals of concern for further evaluation with additional information or more refined models

  11. Investigating the viability of genetic screening/testing for RA susceptibility using combinations of five confirmed risk loci

    Science.gov (United States)

    McClure, Annie; Lunt, Mark; Eyre, Steve; Ke, Xiayi; Thomson, Wendy; Hinks, Anne; Bowes, John; Gibbons, Laura; Plant, Darren; Wilson, Anthony G.; Marinou, Ioanna; Morgan, Ann W.; Emery, Paul; Steer, Sophia; Hocking, Lynne J.; Reid, David M.; Wordsworth, Paul; Harrison, Pille; Worthington, Jane

    2009-01-01

    Objective. Five loci—the shared epitope (SE) of HLA-DRB1, the PTPN22 gene, a locus on 6q23, the STAT4 gene and a locus mapping to the TRAF1/C5 genetic region—have now been unequivocally confirmed as conferring susceptibility to RA. The largest single effect is conferred by SE. We hypothesized that combinations of susceptibility alleles may increase risk over and above that of any individual locus alone. Methods. We analysed data from 4238 RA cases and 1811 controls, for which genotypes were available at all five loci. Results. Statistical analysis identified eight high-risk combinations conferring an odds ratio >6 compared with carriage of no susceptibility variants and, interestingly, 10% population controls carried a combination conferring high risk. All high-risk combinations included SE, and all but one contained PTPN22. Statistical modelling showed that a model containing only these two loci could achieve comparable sensitivity and specificity to a model including all five. Furthermore, replacing SE (which requires full subtyping at the HLA-DRB1 gene) with DRB1*1/4/10 carriage resulted in little further loss of information (correlation coefficient between models = 0.93). Conclusions. This represents the first exploration of the viability of population screening for RA and identifies several high-risk genetic combinations. However, given the population incidence of RA, genetic screening based on these loci alone is neither sufficiently sensitive nor specific at the current time. PMID:19741008

  12. Maine Coon renal screening: ultrasonographical characterisation and preliminary genetic analysis for common genes in cats with renal cysts.

    Science.gov (United States)

    Gendron, Karine; Owczarek-Lipska, Marta; Lang, Johann; Leeb, Tosso

    2013-12-01

    The objective of this study was to assess the prevalence of renal cysts and other renal abnormalities in purebred Maine Coon cats, and to characterise these through genetic typing. Voluntary pre-breeding screening programmes for polycystic kidney disease (PKD) are offered for this breed throughout Switzerland, Germany and other northern European countries. We performed a retrospective evaluation of Maine Coon screening for renal disease at one institution over an 8-year period. Renal ultrasonography was performed in 187 healthy Maine Coon cats. Renal changes were observed in 27 of these cats. Renal cysts were found in seven cats, and were mostly single and unilateral (6/7, 85.7%), small (mean 3.6 mm) and located at the corticomedullary junction (4/6, 66.7%). Sonographical changes indicating chronic kidney disease (CKD) were observed in 10/187 (5.3%) cats and changes of unknown significance were documented in 11/187 (5.9%) cats. All six cats genetically tested for PKD1 were negative for the mutation, and gene sequencing of these cats did not demonstrate any common genetic sequences. Cystic renal disease occurs with a low prevalence in Maine Coons and is unrelated to the PKD observed in Persians and related breeds. Ultrasonographical findings compatible with CKD are not uncommon in juvenile Maine Coons. PMID:23735675

  13. Maine Coon renal screening: ultrasonographical characterisation and preliminary genetic analysis for common genes in cats with renal cysts.

    Science.gov (United States)

    Gendron, Karine; Owczarek-Lipska, Marta; Lang, Johann; Leeb, Tosso

    2013-12-01

    The objective of this study was to assess the prevalence of renal cysts and other renal abnormalities in purebred Maine Coon cats, and to characterise these through genetic typing. Voluntary pre-breeding screening programmes for polycystic kidney disease (PKD) are offered for this breed throughout Switzerland, Germany and other northern European countries. We performed a retrospective evaluation of Maine Coon screening for renal disease at one institution over an 8-year period. Renal ultrasonography was performed in 187 healthy Maine Coon cats. Renal changes were observed in 27 of these cats. Renal cysts were found in seven cats, and were mostly single and unilateral (6/7, 85.7%), small (mean 3.6 mm) and located at the corticomedullary junction (4/6, 66.7%). Sonographical changes indicating chronic kidney disease (CKD) were observed in 10/187 (5.3%) cats and changes of unknown significance were documented in 11/187 (5.9%) cats. All six cats genetically tested for PKD1 were negative for the mutation, and gene sequencing of these cats did not demonstrate any common genetic sequences. Cystic renal disease occurs with a low prevalence in Maine Coons and is unrelated to the PKD observed in Persians and related breeds. Ultrasonographical findings compatible with CKD are not uncommon in juvenile Maine Coons.

  14. Epigenome: A Biomarker or Screening Tool to Evaluate Health Impact of Cumulative Exposure to Chemical and Non-Chemical Stressors.

    Science.gov (United States)

    Olden, Kenneth; Lin, Yu-Sheng; Bussard, David

    2016-06-01

    Current risk assessment practices and toxicity information are hard to utilize for assessing the health impact of combined or cumulative exposure to multiple chemical and non-chemical stressors encountered in the "real world" environment. Non-chemical stressors such as heat, radiation, noise, humidity, bacterial and viral agents, and social factors, like stress related to violence and socioeconomic position generally cannot be currently incorporated into the risk assessment paradigm. The Science and Decisions report released by the National Research Council (NRC) in 2009 emphasized the need to characterize the effects of multiple stressors, both chemical and non-chemical exposures. One impediment to developing information relating such non-chemical stressors to health effects and incorporating them into cumulative assessment has been the lack of analytical tools to easily and quantitatively monitor the cumulative exposure to combined effects of stressors over the life course.

  15. Epigenome: A Biomarker or Screening Tool to Evaluate Health Impact of Cumulative Exposure to Chemical and Non-Chemical Stressors.

    Science.gov (United States)

    Olden, Kenneth; Lin, Yu-Sheng; Bussard, David

    2016-06-01

    Current risk assessment practices and toxicity information are hard to utilize for assessing the health impact of combined or cumulative exposure to multiple chemical and non-chemical stressors encountered in the "real world" environment. Non-chemical stressors such as heat, radiation, noise, humidity, bacterial and viral agents, and social factors, like stress related to violence and socioeconomic position generally cannot be currently incorporated into the risk assessment paradigm. The Science and Decisions report released by the National Research Council (NRC) in 2009 emphasized the need to characterize the effects of multiple stressors, both chemical and non-chemical exposures. One impediment to developing information relating such non-chemical stressors to health effects and incorporating them into cumulative assessment has been the lack of analytical tools to easily and quantitatively monitor the cumulative exposure to combined effects of stressors over the life course. PMID:27534725

  16. Effects of genetic screening on perceptions of health: a pilot study.

    OpenAIRE

    Marteau, T M; Duijn, M.; Ellis, I

    1992-01-01

    The aim of the current study was to determine how carriers of a recessive gene, which confers no risk to their own health, perceive their health, relative to non-carriers. Perceptions of health in three groups were compared: those screened and found to carry the gene for Tay-Sachs disease, those screened and not found to carry the gene for Tay-Sachs disease, and a community based sample who, it was presumed, had not undergone screening. The groups did not differ in their perceptions of their ...

  17. Cultural Concerns when Counseling Orthodox Jewish Couples for Genetic Screening and PGD.

    Science.gov (United States)

    Grazi, Richard V; Wolowelsky, Joel B

    2015-12-01

    There is a spectrum of attitudes within the Orthodox Jewish community towards genetic testing and PGD. Increased understanding of the belief systems of the Orthodox Jewish population will enhance the genetic counselors' ability to better serve this unique group of patients. By improving cultural competence, genetic counselors can help patients choose the testing options that they deem appropriate, while simultaneously respecting the patient's belief system.

  18. Practical experiences with an extended screening strategy for genetically modified organisms (GMOs) in real-life samples.

    Science.gov (United States)

    Scholtens, Ingrid; Laurensse, Emile; Molenaar, Bonnie; Zaaijer, Stephanie; Gaballo, Heidi; Boleij, Peter; Bak, Arno; Kok, Esther

    2013-09-25

    Nowadays most animal feed products imported into Europe have a GMO (genetically modified organism) label. This means that they contain European Union (EU)-authorized GMOs. For enforcement of these labeling requirements, it is necessary, with the rising number of EU-authorized GMOs, to perform an increasing number of analyses. In addition to this, it is necessary to test products for the potential presence of EU-unauthorized GMOs. Analysis for EU-authorized and -unauthorized GMOs in animal feed has thus become laborious and expensive. Initial screening steps may reduce the number of GMO identification methods that need to be applied, but with the increasing diversity also screening with GMO elements has become more complex. For the present study, the application of an informative detailed 24-element screening and subsequent identification strategy was applied in 50 animal feed samples. Almost all feed samples were labeled as containing GMO-derived materials. The main goal of the study was therefore to investigate if a detailed screening strategy would reduce the number of subsequent identification analyses. An additional goal was to test the samples in this way for the potential presence of EU-unauthorized GMOs. Finally, to test the robustness of the approach, eight of the samples were tested in a concise interlaboratory study. No significant differences were found between the results of the two laboratories.

  19. The introduction history of invasive garden ants in Europe: Integrating genetic, chemical and behavioural approaches

    OpenAIRE

    Boomsma Jacobus J; Kronauer Daniel JC; Drijfhout Falko P; Ugelvig Line V; Pedersen Jes S; Cremer Sylvia

    2008-01-01

    Abstract Background The invasive garden ant, Lasius neglectus, is the most recently detected pest ant and the first known invasive ant able to become established and thrive in the temperate regions of Eurasia. In this study, we aim to reconstruct the invasion history of this ant in Europe analysing 14 populations with three complementary approaches: genetic microsatellite analysis, chemical analysis of cuticular hydrocarbon profiles and behavioural observations of aggression behaviour. We eva...

  20. The genetic code degeneracy and the amino acids chemical composition are connected

    OpenAIRE

    Negadi, Tidjani

    2009-01-01

    We show that our recently published Arithmetic Model of the genetic code based on Godel Encoding is robust against symmetry transformations, specially Rumer s one U > G, A > C, and constitutes a link between the degeneracy structure and the chemical composition of the 20 canonical amino acids. As a result, several remarkable atomic patterns involving hydrogen, carbon, nucleon and atom numbers are derived. This study has no obvious practical application(s) but could, we hope, add some new know...

  1. Big Data in Chemical Toxicity Research: The Use of High-Throughput Screening Assays To Identify Potential Toxicants

    OpenAIRE

    Zhu, Hao; Zhang, Jun; Kim, Marlene T.; Boison, Abena; Sedykh, Alexander; Moran, Kimberlee

    2014-01-01

    High-throughput screening (HTS) assays that measure the in vitro toxicity of environmental compounds have been widely applied as an alternative to in vivo animal tests of chemical toxicity. Current HTS studies provide the community with rich toxicology information that has the potential to be integrated into toxicity research. The available in vitro toxicity data is updated daily in structured formats (e.g., deposited into PubChem and other data-sharing web portals) or in an unstructured way ...

  2. The introduction history of invasive garden ants in Europe: Integrating genetic, chemical and behavioural approaches

    Directory of Open Access Journals (Sweden)

    Boomsma Jacobus J

    2008-02-01

    Full Text Available Abstract Background The invasive garden ant, Lasius neglectus, is the most recently detected pest ant and the first known invasive ant able to become established and thrive in the temperate regions of Eurasia. In this study, we aim to reconstruct the invasion history of this ant in Europe analysing 14 populations with three complementary approaches: genetic microsatellite analysis, chemical analysis of cuticular hydrocarbon profiles and behavioural observations of aggression behaviour. We evaluate the relative informative power of the three methodological approaches and estimate both the number of independent introduction events from a yet unknown native range somewhere in the Black Sea area, and the invasive potential of the existing introduced populations. Results Three clusters of genetically similar populations were detected, and all but one population had a similar chemical profile. Aggression between populations could be predicted from their genetic and chemical distance, and two major clusters of non-aggressive groups of populations were found. However, populations of L. neglectus did not separate into clear supercolonial associations, as is typical for other invasive ants. Conclusion The three methodological approaches gave consistent and complementary results. All joint evidence supports the inference that the 14 introduced populations of L. neglectus in Europe likely arose from only very few independent introductions from the native range, and that new infestations were typically started through introductions from other invasive populations. This indicates that existing introduced populations have a very high invasive potential when the ants are inadvertently spread by human transport.

  3. Combined Rational Design and a High Throughput Screening Platform for Identifying Chemical Inhibitors of a Ras-activating Enzyme*

    Science.gov (United States)

    Evelyn, Chris R.; Biesiada, Jacek; Duan, Xin; Tang, Hong; Shang, Xun; Papoian, Ruben; Seibel, William L.; Nelson, Sandra; Meller, Jaroslaw; Zheng, Yi

    2015-01-01

    The Ras family small GTPases regulate multiple cellular processes, including cell growth, survival, movement, and gene expression, and are intimately involved in cancer pathogenesis. Activation of these small GTPases is catalyzed by a special class of enzymes, termed guanine nucleotide exchange factors (GEFs). Herein, we developed a small molecule screening platform for identifying lead hits targeting a Ras GEF enzyme, SOS1. We employed an ensemble structure-based virtual screening approach in combination with a multiple tier high throughput experimental screen utilizing two complementary fluorescent guanine nucleotide exchange assays to identify small molecule inhibitors of GEF catalytic activity toward Ras. From a library of 350,000 compounds, we selected a set of 418 candidate compounds predicted to disrupt the GEF-Ras interaction, of which dual wavelength GDP dissociation and GTP-loading experimental screening identified two chemically distinct small molecule inhibitors. Subsequent biochemical validations indicate that they are capable of dose-dependently inhibiting GEF catalytic activity, binding to SOS1 with micromolar affinity, and disrupting GEF-Ras interaction. Mutagenesis studies in conjunction with structure-activity relationship studies mapped both compounds to different sites in the catalytic pocket, and both inhibited Ras signaling in cells. The unique screening platform established here for targeting Ras GEF enzymes could be broadly useful for identifying lead inhibitors for a variety of small GTPase-activating GEF reactions. PMID:25825487

  4. [The discussion of the infiltrative model of chemical knowledge stepping into genetics teaching in agricultural institute or university].

    Science.gov (United States)

    Zou, Ping; Luo, Pei-Gao

    2010-05-01

    Chemistry is an important group of basic courses, while genetics is one of the important major-basic courses in curriculum of many majors in agricultural institutes or universities. In order to establish the linkage between the major course and the basic course, the ability of application of the chemical knowledge previously learned in understanding genetic knowledge in genetics teaching is worthy of discussion for genetics teachers. In this paper, the authors advocate to apply some chemical knowledge previously learned to understand genetic knowledge in genetics teaching with infiltrative model, which could help students learn and understand genetic knowledge more deeply. Analysis of the intrinsic logistic relationship among the knowledge of different courses and construction of the integral knowledge network are useful for students to improve their analytic, comprehensive and logistic abilities. By this way, we could explore a new teaching model to develop the talents with new ideas and comprehensive competence in agricultural fields.

  5. Optimization of a Reduced Chemical Kinetic Model for HCCI Engine Simulations by Micro-Genetic Algorithm

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    A reduced chemical kinetic model (44 species and 72 reactions) for the homogeneous charge compression ignition (HCCI) combustion of n-heptane was optimized to improve its autoignition predictions under different engine operating conditions. The seven kinetic parameters of the optimized model were determined by using the combination of a micro-genetic algorithm optimization methodology and the SENKIN program of CHEMKIN chemical kinetics software package. The optimization was performed within the range of equivalence ratios 0.2-1.2, initial temperature 310-375 K and initial pressure 0.1-0.3 MPa. The engine simulations show that the optimized model agrees better with the detailed chemical kinetic model (544 species and 2 446 reactions) than the original model does.

  6. Hazard screening of chemical releases and environmental equity analysis of populations proximate to toxic release inventory facilities in Oregon.

    Science.gov (United States)

    Neumann, C M; Forman, D L; Rothlein, J E

    1998-04-01

    A comprehensive approach using hazard screening, demographic analysis, and a geographic information system (GIS) for mapping is employed to address environmental equity issues in Oregon. A media-specific chronic toxicity index [or chronic index (CI)] was used to compare environmental chemical releases reported in the EPA's Toxic Chemical Release Inventory (TRI) database. In 1992, 254 facilities reportedly released more than 40 million pounds of toxic chemicals directly into the environment on-site or transferred them to sewage treatment plants or other off-site facilities for disposal and recycling. For each reported on-site TRI chemical release, a CI based on oral toxicity factors and total mass was calculated. CIs were aggregated on a media-, facility-, and chemical-specific basis. Glycol ethers, nickel, trichloroethylene, chloroform, and manganese were ranked as the top five chemicals released statewide based on total CI. In contrast, based on total mass, methanol, nickel, ammonia, acetone, and toluene were identified as the top five TRI chemicals released in Oregon. TRI facility rankings were related to the demographics and household income of surrounding neighborhoods using bivariate GIS mapping and statistical analysis. TRI facilities were disproportionately located in racial and ethnic minority neighborhoods. They were also located in areas with lower incomes compared to those in the surrounding county. No relationship was observed between the hazard ranking of the TRI facilities overall and socioeconomic characteristics of the community in which they were located.

  7. Accelerating forward genetics for cell wall deconstruction

    OpenAIRE

    Vidaurre, Danielle; Bonetta, Dario

    2012-01-01

    The elucidation of the genes involved in cell wall synthesis and assembly remains one of the biggest challenges of cell wall biology. Although traditional genetic approaches, using simple yet elegant screens, have identified components of the cell wall, many unknowns remain. Exhausting the genetic toolbox by performing sensitized screens, adopting chemical genetics or combining these with improved cell wall imaging, hold the promise of new gene discovery and function. With the recent introduc...

  8. RNA sequencing of Sleeping Beauty transposon-induced tumors detects transposon-RNA fusions in forward genetic cancer screens

    Science.gov (United States)

    Temiz, Nuri A.; Moriarity, Branden S.; Wolf, Natalie K.; Riordan, Jesse D.; Dupuy, Adam J.; Largaespada, David A.; Sarver, Aaron L.

    2016-01-01

    Forward genetic screens using Sleeping Beauty (SB)-mobilized T2/Onc transposons have been used to identify common insertion sites (CISs) associated with tumor formation. Recurrent sites of transposon insertion are commonly identified using ligation-mediated PCR (LM-PCR). Here, we use RNA sequencing (RNA-seq) data to directly identify transcriptional events mediated by T2/Onc. Surprisingly, the majority (∼80%) of LM-PCR identified junction fragments do not lead to observable changes in RNA transcripts. However, in CIS regions, direct transcriptional effects of transposon insertions are observed. We developed an automated method to systematically identify T2/Onc-genome RNA fusion sequences in RNA-seq data. RNA fusion-based CISs were identified corresponding to both DNA-based CISs (Cdkn2a, Mycl1, Nf2, Pten, Sema6d, and Rere) and additional regions strongly associated with cancer that were not observed by LM-PCR (Myc, Akt1, Pth, Csf1r, Fgfr2, Wisp1, Map3k5, and Map4k3). In addition to calculating recurrent CISs, we also present complementary methods to identify potential driver events via determination of strongly supported fusions and fusions with large transcript level changes in the absence of multitumor recurrence. These methods independently identify CIS regions and also point to cancer-associated genes like Braf. We anticipate RNA-seq analyses of tumors from forward genetic screens will become an efficient tool to identify causal events. PMID:26553456

  9. RNA sequencing of Sleeping Beauty transposon-induced tumors detects transposon-RNA fusions in forward genetic cancer screens.

    Science.gov (United States)

    Temiz, Nuri A; Moriarity, Branden S; Wolf, Natalie K; Riordan, Jesse D; Dupuy, Adam J; Largaespada, David A; Sarver, Aaron L

    2016-01-01

    Forward genetic screens using Sleeping Beauty (SB)-mobilized T2/Onc transposons have been used to identify common insertion sites (CISs) associated with tumor formation. Recurrent sites of transposon insertion are commonly identified using ligation-mediated PCR (LM-PCR). Here, we use RNA sequencing (RNA-seq) data to directly identify transcriptional events mediated by T2/Onc. Surprisingly, the majority (∼80%) of LM-PCR identified junction fragments do not lead to observable changes in RNA transcripts. However, in CIS regions, direct transcriptional effects of transposon insertions are observed. We developed an automated method to systematically identify T2/Onc-genome RNA fusion sequences in RNA-seq data. RNA fusion-based CISs were identified corresponding to both DNA-based CISs (Cdkn2a, Mycl1, Nf2, Pten, Sema6d, and Rere) and additional regions strongly associated with cancer that were not observed by LM-PCR (Myc, Akt1, Pth, Csf1r, Fgfr2, Wisp1, Map3k5, and Map4k3). In addition to calculating recurrent CISs, we also present complementary methods to identify potential driver events via determination of strongly supported fusions and fusions with large transcript level changes in the absence of multitumor recurrence. These methods independently identify CIS regions and also point to cancer-associated genes like Braf. We anticipate RNA-seq analyses of tumors from forward genetic screens will become an efficient tool to identify causal events. PMID:26553456

  10. RNA sequencing of Sleeping Beauty transposon-induced tumors detects transposon-RNA fusions in forward genetic cancer screens.

    Science.gov (United States)

    Temiz, Nuri A; Moriarity, Branden S; Wolf, Natalie K; Riordan, Jesse D; Dupuy, Adam J; Largaespada, David A; Sarver, Aaron L

    2016-01-01

    Forward genetic screens using Sleeping Beauty (SB)-mobilized T2/Onc transposons have been used to identify common insertion sites (CISs) associated with tumor formation. Recurrent sites of transposon insertion are commonly identified using ligation-mediated PCR (LM-PCR). Here, we use RNA sequencing (RNA-seq) data to directly identify transcriptional events mediated by T2/Onc. Surprisingly, the majority (∼80%) of LM-PCR identified junction fragments do not lead to observable changes in RNA transcripts. However, in CIS regions, direct transcriptional effects of transposon insertions are observed. We developed an automated method to systematically identify T2/Onc-genome RNA fusion sequences in RNA-seq data. RNA fusion-based CISs were identified corresponding to both DNA-based CISs (Cdkn2a, Mycl1, Nf2, Pten, Sema6d, and Rere) and additional regions strongly associated with cancer that were not observed by LM-PCR (Myc, Akt1, Pth, Csf1r, Fgfr2, Wisp1, Map3k5, and Map4k3). In addition to calculating recurrent CISs, we also present complementary methods to identify potential driver events via determination of strongly supported fusions and fusions with large transcript level changes in the absence of multitumor recurrence. These methods independently identify CIS regions and also point to cancer-associated genes like Braf. We anticipate RNA-seq analyses of tumors from forward genetic screens will become an efficient tool to identify causal events.

  11. Screening of CACNA1A and ATP1A2 genes in hemiplegic migraine: clinical, genetic, and functional studies

    Science.gov (United States)

    Carreño, Oriel; Corominas, Roser; Serra, Selma Angèlica; Sintas, Cèlia; Fernández-Castillo, Noèlia; Vila-Pueyo, Marta; Toma, Claudio; Gené, Gemma G; Pons, Roser; Llaneza, Miguel; Sobrido, María-Jesús; Grinberg, Daniel; Valverde, Miguel Ángel; Fernández-Fernández, José Manuel; Macaya, Alfons; Cormand, Bru

    2013-01-01

    Hemiplegic migraine (HM) is a rare and severe subtype of autosomal dominant migraine, characterized by a complex aura including some degree of motor weakness. Mutations in four genes (CACNA1A, ATP1A2, SCN1A and PRRT2) have been detected in familial and in sporadic cases. This genetically and clinically heterogeneous disorder is often accompanied by permanent ataxia, epileptic seizures, mental retardation, and chronic progressive cerebellar atrophy. Here we report a mutation screening in the CACNA1A and ATP1A2 genes in 18 patients with HM. Furthermore, intragenic copy number variant (CNV) analysis was performed in CACNA1A using quantitative approaches. We identified four previously described missense CACNA1A mutations (p.Ser218Leu, p.Thr501Met, p.Arg583Gln, and p.Thr666Met) and two missense changes in the ATP1A2 gene, the previously described p.Ala606Thr and the novel variant p.Glu825Lys. No structural variants were found. This genetic screening allowed the identification of more than 30% of the disease alleles, all present in a heterozygous state. Functional consequences of the CACNA1A-p.Thr501Met mutation, previously described only in association with episodic ataxia, and ATP1A2-p.Glu825Lys, were investigated by means of electrophysiological studies, cell viability assays or Western blot analysis. Our data suggest that both these variants are disease-causing. PMID:24498617

  12. A Genetic Animal Model of Alcoholism for Screening Medications to Treat Addiction

    OpenAIRE

    Bell, Richard L.; Hauser, Sheketha; Rodd, Zachary A.; Liang, Tiebing; Sari, Youssef; McClintick, Jeanette; Rahman, Shafiqur; Engleman, Eric A.

    2016-01-01

    The purpose of this review is to present up-to-date pharmacological, genetic and behavioral findings from the alcohol-preferring P rat and summarize similar past work. Behaviorally, the focus will be on how the P rat meets criteria put forth for a valid animal model of alcoholism with a highlight on its use as an animal model of polysubstance abuse, including alcohol, nicotine and psychostimulants. Pharmacologically and genetically, the focus will be on the neurotransmitter and neuropeptide s...

  13. Bypassing antibiotic selection: positive screening of genetically modified cells with an antigen-dependent proliferation switch

    OpenAIRE

    Kawahara, Masahiro; Ueda, Hiroshi; Morita, Sumiyo; Tsumoto, Kouhei; Kumagai, Izumi; Nagamune, Teruyuki

    2003-01-01

    While antibiotic selection has been routinely used for the selection of genetically modified cells, administration of cytotoxic drugs often leads to deleterious effects not only to inert cells but also to transfected or transduced ones. In this study, we propose an Antigen-MEdiated Genetically modified cell Amplification (AMEGA) system employing antibody/receptor chimeras without antibiotic selection. Based on a rational design where the extracellular domains of dimeric erythropoietin recepto...

  14. Genetic and chemical knockdown: a complementary strategy for evaluating an anti-infective target

    Directory of Open Access Journals (Sweden)

    Ramachandran V

    2013-02-01

    Full Text Available Vasanthi Ramachandran,1,* Ragini Singh,2,* Xiaoyu Yang,1 Ragadeepthi Tunduguru,1 Subrat Mohapatra,2 Swati Khandelwal,2 Sanjana Patel,2 Santanu Datta21AstraZeneca India R&D, Bangalore, India; 2Cellworks India, Bangalore, India *These authors contributed equally to this workAbstract: The equity of a drug target is principally evaluated by its genetic vulnerability with tools ranging from antisense- and microRNA-driven knockdowns to induced expression of the target protein. In order to upgrade the process of antibacterial target identification and discern its most effective type of inhibition, an in silico toolbox that evaluates its genetic and chemical vulnerability leading either to stasis or cidal outcome was constructed and validated. By precise simulation and careful experimentation using enolpyruvyl shikimate-3-phosphate synthase and its specific inhibitor glyphosate, it was shown that genetic knockdown is distinct from chemical knockdown. It was also observed that depending on the particular mechanism of inhibition, viz competitive, uncompetitive, and noncompetitive, the antimicrobial potency of an inhibitor could be orders of magnitude different. Susceptibility of Escherichia coli to glyphosate and the lack of it in Mycobacterium tuberculosis could be predicted by the in silico platform. Finally, as predicted and simulated in the in silico platform, the translation of growth inhibition to a cidal effect was able to be demonstrated experimentally by altering the carbon source from sorbitol to glucose.Keywords: knockdown, inhibition, in silico, vulnerability

  15. Genetical studies of resistance to Phytophthora porri in Allium porrum, using a new early screening method.

    NARCIS (Netherlands)

    Smilde, W.D.; Nes, van M.; Reinink, K.; Kik, C.

    1997-01-01

    A new screening method was developed to evaluate resistance of leek (Allium porrum) to Phytophthora porri, based on inoculation by 24 h-immersion of leek plantlets in the 3–6 leaf stage in a suspension of ca. 100 zoospores.ml-1. The immersion test was used for identifying new sources of resistance a

  16. Sensitivity of neuroprogenitor cells to chemical-induced apoptosis using a multiplexed assay suitable for high-throughput screening

    International Nuclear Information System (INIS)

    High-throughput methods are useful for rapidly screening large numbers of chemicals for biological activity, including the perturbation of pathways that may lead to adverse cellular effects. In vitro assays for the key events of neurodevelopment, including apoptosis, may be used in a battery of tests for detecting chemicals that could result in developmental neurotoxicity. Apoptosis contributes to nervous system development by regulating the size of the neuroprogenitor cell pool, and the balance between cellular proliferation and apoptosis during neuroprogenitor cell proliferation helps to determine the size and shape of the nervous system. Therefore, chemicals that affect apoptosis during neuronal development can have deleterious effects on the developing brain. The present study examined the utility of a high-throughput assay to detect chemical-induced apoptosis in mouse or human neuroprogenitor cells, as well as differentiated human neurons derived from induced pluripotent stem cells. Apoptosis was assessed using an assay that measures enzymatic activity of caspase-3/7 in a rapid and cost efficient manner. The results show that all three commercially available models generated a robust source of proliferating neuroprogenitor cells, and that the assay was sensitive and reproducible when used in a multi-well plate format. There were differences in the response of rodent and human neuroprogenitor cells to a set of chemicals previously shown to induce apoptosis in vitro. Neuroprogenitor cells were more sensitive to chemical-induced apoptosis than differentiated neurons, suggesting that neuroprogenitor cells are one of the cell models that should be considered for use in a developmental neurotoxicity screening battery

  17. Bio-inspired Silicification of Silica-binding Peptide-Silk Protein Chimeras: Comparison of Chemically and Genetically Produced Proteins

    OpenAIRE

    Canabady-Rochelle, Laetitia L.S.; Belton, David J.; Deschaume, Olivier; Currie, Heather A.; Kaplan, David L; Perry, Carole C.

    2012-01-01

    Novel protein chimeras constituted of ‘silk’ and a silica-binding peptide (KSLSRHDHIHHH) were synthesized by genetic or chemical approaches and their influence on silica-silk based chimera composite formation evaluated. Genetic chimeras were constructed from 6 or 15 repeats of the 32 amino acid consensus sequence of Nephila clavipes spider silk ([SGRGGLGGQG AGAAAAAGGA GQGGYGGLGSQG]n) to which one silica binding peptide was fused at the N terminus. For the chemical chimera, 25 equivalents of t...

  18. Development of a qualitative, multiplex real-time PCR kit for screening of genetically modified organisms (GMOs).

    Science.gov (United States)

    Dörries, Hans-Henno; Remus, Ivonne; Grönewald, Astrid; Grönewald, Cordt; Berghof-Jäger, Kornelia

    2010-03-01

    The number of commercially available genetically modified organisms (GMOs) and therefore the diversity of possible target sequences for molecular detection techniques are constantly increasing. As a result, GMO laboratories and the food production industry currently are forced to apply many different methods to reliably test raw material and complex processed food products. Screening methods have become more and more relevant to minimize the analytical effort and to make a preselection for further analysis (e.g., specific identification or quantification of the GMO). A multiplex real-time PCR kit was developed to detect the 35S promoter of the cauliflower mosaic virus, the terminator of the nopaline synthase gene of Agrobacterium tumefaciens, the 35S promoter from the figwort mosaic virus, and the bar gene of the soil bacterium Streptomyces hygroscopicus as the most widely used sequences in GMOs. The kit contains a second assay for the detection of plant-derived DNA to control the quality of the often processed and refined sample material. Additionally, the plant-specific assay comprises a homologous internal amplification control for inhibition control. The determined limits of detection for the five assays were 10 target copies/reaction. No amplification products were observed with DNAs of 26 bacterial species, 25 yeasts, 13 molds, and 41 not genetically modified plants. The specificity of the assays was further demonstrated to be 100% by the specific amplification of DNA derived from reference material from 22 genetically modified crops. The applicability of the kit in routine laboratory use was verified by testing of 50 spiked and unspiked food products. The herein described kit represents a simple and sensitive GMO screening method for the reliable detection of multiple GMO-specific target sequences in a multiplex real-time PCR reaction.

  19. Data from Tiered High-Throughput Screening Approach to Identify Thyroperoxidase Inhibitors within the ToxCast Phase I and II Chemical Libraries

    Data.gov (United States)

    U.S. Environmental Protection Agency — High-throughput screening for potential thyroid-disrupting chemicals requires a system of assays to capture multiple molecular-initiating events (MIEs) that...

  20. Chemical Stimulants and Genetic Sexing Boost the SIT: Evidence from Ceratitis capitata and Bactrocera Dorsalis in Hawaii

    Science.gov (United States)

    Genetic and chemical means have been developed to significantly improve the effectiveness of the sterile insect technique against tephritid fruit flies in recent years. Beginning with the development of genetic sexing techniques some 25 years ago, all-male strains of several species of fruit flies h...

  1. Problems and solutions in the estimation of genetic risks from radiation and chemicals

    International Nuclear Information System (INIS)

    Extensive investigations with mice on the effects of various physical and biological factors, such as dose rate, sex and cell stage, on radiation-induced mutation have provided an evaluation of the genetics hazards of radiation in man. The mutational results obtained in both sexes with progressive lowering of the radiation dose rate have permitted estimation of the mutation frequency expected under the low-level radiation conditions of most human exposure. Supplementing the studies on mutation frequency are investigations on the phenotypic effects of mutations in mice, particularly anatomical disorders of the skeleton, which allow an estimation of the degree of human handicap associated with the occurrence of parallel defects in man. Estimation of the genetic risk from chemical mutagens is much more difficult, and the research is much less advanced. Results on transmitted mutations in mice indicate a poor correlation with mutation induction in non-mammalian organisms

  2. Chemical and genetic relationships among sage ( Salvia officinalis L.) cultivars and Judean sage ( Salvia judaica Boiss.).

    Science.gov (United States)

    Böszörményi, Andrea; Héthelyi, Eva; Farkas, Agnes; Horváth, Györgyi; Papp, Nóra; Lemberkovics, Eva; Szoke, Eva

    2009-06-10

    The essential oil composition and genetic variability of common sage ( Salvia officinalis L.) and its three ornamental cultivars ('Purpurascens', 'Tricolor', and 'Kew Gold') as well as Judean sage ( Salvia judaica Boiss.) were analyzed by GC-FID, GC-MS, and random amplified polymorphic DNA (RAPD). Common sage and its cultivars contained the same volatile compounds; only the ratio of compounds differed. The main compounds were the sesquiterpene alpha-humulene and the monoterpenes beta-pinene, eucalyptol, and camphor. Judean sage contained mainly the sesquiterpenes beta-cubebene and ledol. All of the samples exhibited characteristic RAPD patterns that allowed their identification. Cluster analyses based on oil composition and RAPD markers corresponded very well to each other, suggesting that there is a strong relationship between the chemical profile and the genetic variability.

  3. Production and Characterization of Chemically Inactivated Genetically Engineered Clostridium difficile Toxoids.

    Science.gov (United States)

    Vidunas, Eugene; Mathews, Antony; Weaver, Michele; Cai, Ping; Koh, Eun Hee; Patel-Brown, Sujata; Yuan, Hailey; Zheng, Zi-Rong; Carriere, Marjolaine; Johnson, J Erik; Lotvin, Jason; Moran, Justin

    2016-07-01

    A recombinant Clostridium difficile expression system was used to produce genetically engineered toxoids A and B as immunogens for a prophylactic vaccine against C. difficile-associated disease. Although all known enzymatic activities responsible for cytotoxicity were genetically abrogated, the toxoids exhibited residual cytotoxic activity as measured in an in vitro cell-based cytotoxicity assay. The residual cytotoxicity was eliminated by treating the toxoids with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide. Mass spectrometry and amino acid analysis of the EDC-inactivated toxoids identified crosslinks, glycine adducts, and β-alanine adducts. Surface plasmon resonance analysis demonstrated that modifications resulting from the chemical treatment did not appreciably affect recognition of epitopes by both toxin A- and B-specific neutralizing monoclonal antibodies. Compared to formaldehyde-inactivated toxoids, the EDC/N-hydroxysuccinimide-inactivated toxoids exhibited superior stability in solution with respect to reversion of cytotoxic activity. PMID:27233688

  4. Screening Genetic Resources of Capsicum Peppers in Their Primary Center of Diversity in Bolivia and Peru.

    Directory of Open Access Journals (Sweden)

    Maarten van Zonneveld

    Full Text Available For most crops, like Capsicum, their diversity remains under-researched for traits of interest for food, nutrition and other purposes. A small investment in screening this diversity for a wide range of traits is likely to reveal many traditional varieties with distinguished values. One objective of this study was to demonstrate, with Capsicum as model crop, the application of indicators of phenotypic and geographic diversity as effective criteria for selecting promising genebank accessions for multiple uses from crop centers of diversity. A second objective was to evaluate the expression of biochemical and agromorphological properties of the selected Capsicum accessions in different conditions. Four steps were involved: 1 Develop the necessary diversity by expanding genebank collections in Bolivia and Peru; 2 Establish representative subsets of ~100 accessions for biochemical screening of Capsicum fruits; 3 Select promising accessions for different uses after screening; and 4 Examine how these promising accessions express biochemical and agromorphological properties when grown in different environmental conditions. The Peruvian Capsicum collection now contains 712 accessions encompassing all five domesticated species (C. annuum, C. chinense, C. frutescens, C. baccatum, and C. pubescens. The collection in Bolivia now contains 487 accessions, representing all five domesticates plus four wild taxa (C. baccatum var. baccatum, C. caballeroi, C. cardenasii, and C. eximium. Following the biochemical screening, 44 Bolivian and 39 Peruvian accessions were selected as promising, representing wide variation in levels of antioxidant capacity, capsaicinoids, fat, flavonoids, polyphenols, quercetins, tocopherols, and color. In Peru, 23 promising accessions performed well in different environments, while each of the promising Bolivian accessions only performed well in a certain environment. Differences in Capsicum diversity and local contexts led to distinct

  5. Screening Genetic Resources of Capsicum Peppers in Their Primary Center of Diversity in Bolivia and Peru

    OpenAIRE

    Maarten van Zonneveld; Marleni Ramirez; Williams, David E; Michael Petz; Sven Meckelmann; Teresa Avila; Carlos Bejarano; Llermé Ríos; Karla Peña; Matthias Jäger; Dimary Libreros; Karen Amaya; Xavier Scheldeman

    2015-01-01

    For most crops, like Capsicum, their diversity remains under-researched for traits of interest for food, nutrition and other purposes. A small investment in screening this diversity for a wide range of traits is likely to reveal many traditional varieties with distinguished values. One objective of this study was to demonstrate, with Capsicum as model crop, the application of indicators of phenotypic and geographic diversity as effective criteria for selecting promising genebank accessions fo...

  6. Screening Genetic Resources of Capsicum Peppers in Their Primary Center of Diversity in Bolivia and Peru.

    Science.gov (United States)

    van Zonneveld, Maarten; Ramirez, Marleni; Williams, David E; Petz, Michael; Meckelmann, Sven; Avila, Teresa; Bejarano, Carlos; Ríos, Llermé; Peña, Karla; Jäger, Matthias; Libreros, Dimary; Amaya, Karen; Scheldeman, Xavier

    2015-01-01

    For most crops, like Capsicum, their diversity remains under-researched for traits of interest for food, nutrition and other purposes. A small investment in screening this diversity for a wide range of traits is likely to reveal many traditional varieties with distinguished values. One objective of this study was to demonstrate, with Capsicum as model crop, the application of indicators of phenotypic and geographic diversity as effective criteria for selecting promising genebank accessions for multiple uses from crop centers of diversity. A second objective was to evaluate the expression of biochemical and agromorphological properties of the selected Capsicum accessions in different conditions. Four steps were involved: 1) Develop the necessary diversity by expanding genebank collections in Bolivia and Peru; 2) Establish representative subsets of ~100 accessions for biochemical screening of Capsicum fruits; 3) Select promising accessions for different uses after screening; and 4) Examine how these promising accessions express biochemical and agromorphological properties when grown in different environmental conditions. The Peruvian Capsicum collection now contains 712 accessions encompassing all five domesticated species (C. annuum, C. chinense, C. frutescens, C. baccatum, and C. pubescens). The collection in Bolivia now contains 487 accessions, representing all five domesticates plus four wild taxa (C. baccatum var. baccatum, C. caballeroi, C. cardenasii, and C. eximium). Following the biochemical screening, 44 Bolivian and 39 Peruvian accessions were selected as promising, representing wide variation in levels of antioxidant capacity, capsaicinoids, fat, flavonoids, polyphenols, quercetins, tocopherols, and color. In Peru, 23 promising accessions performed well in different environments, while each of the promising Bolivian accessions only performed well in a certain environment. Differences in Capsicum diversity and local contexts led to distinct outcomes in

  7. A microfluidic-based genetic screen to identify microbial virulence factors that inhibit dendritic cell migration

    OpenAIRE

    McLaughlin, Laura M.; Xu, Hui; Carden, Sarah E.; Fisher, Samantha; Reyes, Monique; Heilshorn, Sarah C.; Monack, Denise M.

    2014-01-01

    Microbial pathogens are able to modulate host cells and evade the immune system by multiple mechanisms. For example, Salmonella injects effector proteins into host cells and evades the host immune system in part by inhibiting dendritic cell (DC) migration. The identification of microbial factors that modulate normal host functions should lead to the development of new classes of therapeutics that target these pathways. Current screening methods to identify either host or pathogen genes involv...

  8. Development of quantitative duplex real-time PCR method for screening analysis of genetically modified maize.

    Science.gov (United States)

    Oguchi, Taichi; Onishi, Mari; Minegishi, Yasutaka; Kurosawa, Yasunori; Kasahara, Masaki; Akiyama, Hiroshi; Teshima, Reiko; Futo, Satoshi; Furui, Satoshi; Hino, Akihiro; Kitta, Kazumi

    2009-06-01

    A duplex real-time PCR method was developed for quantitative screening analysis of GM maize. The duplex real-time PCR simultaneously detected two GM-specific segments, namely the cauliflower mosaic virus (CaMV) 35S promoter (P35S) segment and an event-specific segment for GA21 maize which does not contain P35S. Calibration was performed with a plasmid calibrant specially designed for the duplex PCR. The result of an in-house evaluation suggested that the analytical precision of the developed method was almost equivalent to those of simplex real-time PCR methods, which have been adopted as ISO standard methods for the analysis of GMOs in foodstuffs and have also been employed for the analysis of GMOs in Japan. In addition, this method will reduce both the cost and time requirement of routine GMO analysis by half. The high analytical performance demonstrated in the current study would be useful for the quantitative screening analysis of GM maize. We believe the developed method will be useful for practical screening analysis of GM maize, although interlaboratory collaborative studies should be conducted to confirm this.

  9. An efficient platform for genetic selection and screening of gene switches in Escherichia coli.

    Science.gov (United States)

    Muranaka, Norihito; Sharma, Vandana; Nomura, Yoko; Yokobayashi, Yohei

    2009-04-01

    Engineered gene switches and circuits that can sense various biochemical and physical signals, perform computation, and produce predictable outputs are expected to greatly advance our ability to program complex cellular behaviors. However, rational design of gene switches and circuits that function in living cells is challenging due to the complex intracellular milieu. Consequently, most successful designs of gene switches and circuits have relied, to some extent, on high-throughput screening and/or selection from combinatorial libraries of gene switch and circuit variants. In this study, we describe a generic and efficient platform for selection and screening of gene switches and circuits in Escherichia coli from large libraries. The single-gene dual selection marker tetA was translationally fused to green fluorescent protein (gfpuv) via a flexible peptide linker and used as a dual selection and screening marker for laboratory evolution of gene switches. Single-cycle (sequential positive and negative selections) enrichment efficiencies of >7000 were observed in mock selections of model libraries containing functional riboswitches in liquid culture. The technique was applied to optimize various parameters affecting the selection outcome, and to isolate novel thiamine pyrophosphate riboswitches from a complex library. Artificial riboswitches with excellent characteristics were isolated that exhibit up to 58-fold activation as measured by fluorescent reporter gene assay. PMID:19190095

  10. At the Interface of Chemical and Biological Synthesis: An Expanded Genetic Code.

    Science.gov (United States)

    Xiao, Han; Schultz, Peter G

    2016-01-01

    The ability to site-specifically incorporate noncanonical amino acids (ncAAs) with novel structures into proteins in living cells affords a powerful tool to investigate and manipulate protein structure and function. More than 200 ncAAs with diverse biological, chemical, and physical properties have been genetically encoded in response to nonsense or frameshift codons in both prokaryotic and eukaryotic organisms with high fidelity and efficiency. In this review, recent advances in the technology and its application to problems in protein biochemistry, cellular biology, and medicine are highlighted. PMID:27413101

  11. High acceptance of an early dyslexia screening test involving genetic analyses in Germany.

    Science.gov (United States)

    Wilcke, Arndt; Müller, Bent; Schaadt, Gesa; Kirsten, Holger; Boltze, Johannes

    2016-02-01

    Dyslexia is a developmental disorder characterized by severe problems in the acquisition of reading and writing skills. It has a strong neurobiological basis. Genetic influence is estimated at 50-70%. One of the central problems with dyslexia is its late diagnosis, normally not before the end of the 2nd grade, resulting in the loss of several years for early therapy. Currently, research is focusing on the development of early tests for dyslexia, which may be based on EEG and genetics. Our aim was to determine the acceptance of such a future test among parents. We conducted a representative survey in Germany with 1000 parents of children aged 3-7 years, with and without experience of dyslexia. 88.7% of the parents supported the introduction of an early test for dyslexia based on EEG and genetics; 82.8% would have their own children tested, and 57.9% were willing to pay for the test if health insurance did not cover the costs. Test acceptance was significantly higher if parents had prior experience with dyslexia. The perceived benefits of such a test were early recognition and remediation and, preventing deficits. Concerns regarded the precision of the test, its potentially stigmatizing effect and its costs. The high overall support for the test leads to the conclusion that parents would accept a test for dyslexia based on EEG and genetics. PMID:26036858

  12. High acceptance of an early dyslexia screening test involving genetic analyses in Germany.

    Science.gov (United States)

    Wilcke, Arndt; Müller, Bent; Schaadt, Gesa; Kirsten, Holger; Boltze, Johannes

    2016-02-01

    Dyslexia is a developmental disorder characterized by severe problems in the acquisition of reading and writing skills. It has a strong neurobiological basis. Genetic influence is estimated at 50-70%. One of the central problems with dyslexia is its late diagnosis, normally not before the end of the 2nd grade, resulting in the loss of several years for early therapy. Currently, research is focusing on the development of early tests for dyslexia, which may be based on EEG and genetics. Our aim was to determine the acceptance of such a future test among parents. We conducted a representative survey in Germany with 1000 parents of children aged 3-7 years, with and without experience of dyslexia. 88.7% of the parents supported the introduction of an early test for dyslexia based on EEG and genetics; 82.8% would have their own children tested, and 57.9% were willing to pay for the test if health insurance did not cover the costs. Test acceptance was significantly higher if parents had prior experience with dyslexia. The perceived benefits of such a test were early recognition and remediation and, preventing deficits. Concerns regarded the precision of the test, its potentially stigmatizing effect and its costs. The high overall support for the test leads to the conclusion that parents would accept a test for dyslexia based on EEG and genetics.

  13. Using the pea aphid Acrythociphon pisum as a tool for screening biological responses to chemicals and drugs

    Directory of Open Access Journals (Sweden)

    Ledger Terence

    2009-09-01

    Full Text Available Abstract Background Though the biological process of aphid feeding is well documented, no one to date has sought to apply it as a tool to screen the biological responses to chemicals and drugs, in ecotoxicology, genotoxicology and/or for interactions in the cascade of sequential molecular events of embryogenesis. Parthenogenetic insect species present the advantage of an anatomical system composed of multiple germarium/ovarioles in the same mother with all the intermediate maturation stages of embryos from oocyte to first instar larva birth. This could be used as an interesting model to visualize at which step drugs interact with the cell signalling pathway during the ordered developmental process. Findings We designed a simple test for screening drugs by investigating simultaneously zygote mitotic division, the progression of embryo development, cell differentiation at early developmental stages and finally organogenesis and population growth rate. We aimed to analyze the toxicology effects of compounds and/or their interference on cellular signalling by examining at which step of the cascade, from zygote to mature embryo, the developmental process is perturbed. We reasoned that a parthenogenetic founder insect, in which the ovarioles shelter numerous embryos at different developmental stages, would allow us to precisely pinpoint the step of embryogenesis in which chemicals act through specific molecular targets as the known ordered homeobox genes. Conclusion Using this method we report the results of a genotoxicological and demographic analysis of three compound models bearing in common a bromo group: one is integrated as a base analog in DNA synthesis, two others activate permanently kinases. We report that one compound (Br-du altered drastically embryogenesis, which argues in favor of this simple technique as a cheap first screening of chemicals or drugs to be used in a number of genotoxicology applications.

  14. The ChemScreen project to design a pragmatic alternative approachto predict reproductive toxicity of chemicals

    NARCIS (Netherlands)

    Van der Burg, B.; Wedebye, E.B.; Dietrich, D.R.; Jaworska, J.; Mangelsdorf, I.; Paune, E.; Schwarz, M.; Piersma, A.H.; Kroese, E.D.

    2015-01-01

    There is a great need for rapid testing strategies for reproductive toxicity testing, avoiding animal use.The EU Framework program 7 project ChemScreen aimed to fill this gap in a pragmatic manner prefer-ably using validated existing tools and place them in an innovative alternative testing strategy

  15. Skin Cancer Screening

    Science.gov (United States)

    ... Genetics of Skin Cancer Skin Cancer Screening Research Skin Cancer Screening (PDQ®)–Patient Version What is screening? Go ... These are called diagnostic tests . General Information About Skin Cancer Key Points Skin cancer is a disease in ...

  16. Identification of putative estrogen receptor-mediated endocrine disrupting chemicals using QSAR- and structure-based virtual screening approaches

    International Nuclear Information System (INIS)

    Identification of endocrine disrupting chemicals is one of the important goals of environmental chemical hazard screening. We report on the development of validated in silico predictors of chemicals likely to cause estrogen receptor (ER)-mediated endocrine disruption to facilitate their prioritization for future screening. A database of relative binding affinity of a large number of ERα and/or ERβ ligands was assembled (546 for ERα and 137 for ERβ). Both single-task learning (STL) and multi-task learning (MTL) continuous quantitative structure–activity relationship (QSAR) models were developed for predicting ligand binding affinity to ERα or ERβ. High predictive accuracy was achieved for ERα binding affinity (MTL R2 = 0.71, STL R2 = 0.73). For ERβ binding affinity, MTL models were significantly more predictive (R2 = 0.53, p < 0.05) than STL models. In addition, docking studies were performed on a set of ER agonists/antagonists (67 agonists and 39 antagonists for ERα, 48 agonists and 32 antagonists for ERβ, supplemented by putative decoys/non-binders) using the following ER structures (in complexes with respective ligands) retrieved from the Protein Data Bank: ERα agonist (PDB ID: 1L2I), ERα antagonist (PDB ID: 3DT3), ERβ agonist (PDB ID: 2NV7), and ERβ antagonist (PDB ID: 1L2J). We found that all four ER conformations discriminated their corresponding ligands from presumed non-binders. Finally, both QSAR models and ER structures were employed in parallel to virtually screen several large libraries of environmental chemicals to derive a ligand- and structure-based prioritized list of putative estrogenic compounds to be used for in vitro and in vivo experimental validation. - Highlights: • This is the largest curated dataset inclusive of ERα and β (the latter is unique). • New methodology that for the first time affords acceptable ERβ models. • A combination of QSAR and docking enables prediction of affinity and function. • The results have

  17. Identification of putative estrogen receptor-mediated endocrine disrupting chemicals using QSAR- and structure-based virtual screening approaches

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Liying; Sedykh, Alexander; Tripathi, Ashutosh [Eshelman School of Pharmacy, University of North Carolina, Chapel Hill, NC (United States); Zhu, Hao [The Rutgers Center for Computational and Integrative Biology, Rutgers University, Camden, NJ (United States); Department of Chemistry, Rutgers University, Camden, NJ (United States); Afantitis, Antreas; Mouchlis, Varnavas D.; Melagraki, Georgia [NovaMechanics Ltd., Nicosia (Cyprus); Rusyn, Ivan, E-mail: iir@unc.edu [Department of Environmental Sciences and Engineering, University of North Carolina, Chapel Hill, NC (United States); Tropsha, Alexander, E-mail: alex_tropsha@unc.edu [Eshelman School of Pharmacy, University of North Carolina, Chapel Hill, NC (United States)

    2013-10-01

    Identification of endocrine disrupting chemicals is one of the important goals of environmental chemical hazard screening. We report on the development of validated in silico predictors of chemicals likely to cause estrogen receptor (ER)-mediated endocrine disruption to facilitate their prioritization for future screening. A database of relative binding affinity of a large number of ERα and/or ERβ ligands was assembled (546 for ERα and 137 for ERβ). Both single-task learning (STL) and multi-task learning (MTL) continuous quantitative structure–activity relationship (QSAR) models were developed for predicting ligand binding affinity to ERα or ERβ. High predictive accuracy was achieved for ERα binding affinity (MTL R{sup 2} = 0.71, STL R{sup 2} = 0.73). For ERβ binding affinity, MTL models were significantly more predictive (R{sup 2} = 0.53, p < 0.05) than STL models. In addition, docking studies were performed on a set of ER agonists/antagonists (67 agonists and 39 antagonists for ERα, 48 agonists and 32 antagonists for ERβ, supplemented by putative decoys/non-binders) using the following ER structures (in complexes with respective ligands) retrieved from the Protein Data Bank: ERα agonist (PDB ID: 1L2I), ERα antagonist (PDB ID: 3DT3), ERβ agonist (PDB ID: 2NV7), and ERβ antagonist (PDB ID: 1L2J). We found that all four ER conformations discriminated their corresponding ligands from presumed non-binders. Finally, both QSAR models and ER structures were employed in parallel to virtually screen several large libraries of environmental chemicals to derive a ligand- and structure-based prioritized list of putative estrogenic compounds to be used for in vitro and in vivo experimental validation. - Highlights: • This is the largest curated dataset inclusive of ERα and β (the latter is unique). • New methodology that for the first time affords acceptable ERβ models. • A combination of QSAR and docking enables prediction of affinity and function.

  18. HIGH-THROUGHPUT CHEMICAL SCREENING USING PROTEIN PROFILING OF FISH PLASMA

    Science.gov (United States)

    Compounds that affect the hormone system, referred to as "endocrine-disrupting chemicals" (EDCs), cause human and animal health problems. It is necessary to test putative EDC chemicals for such deleterious effects, though current testing methodologies are time/animal intensive an...

  19. Identification of Chemical-Genetic Interactions via Parallel Analysis of Barcoded Yeast Strains.

    Science.gov (United States)

    Suresh, Sundari; Schlecht, Ulrich; Xu, Weihong; Miranda, Molly; Davis, Ronald W; Nislow, Corey; Giaever, Guri; St Onge, Robert P

    2016-01-01

    The Yeast Knockout Collection is a complete set of gene deletion strains for the budding yeast, Saccharomyces cerevisiae In each strain, one of approximately 6000 open-reading frames is replaced with a dominant selectable marker flanked by two DNA barcodes. These barcodes, which are unique to each gene, allow the growth of thousands of strains to be individually measured from a single pooled culture. The collection, and other resources that followed, has ushered in a new era in chemical biology, enabling unbiased and systematic identification of chemical-genetic interactions (CGIs) with remarkable ease. CGIs link bioactive compounds to biological processes, and hence can reveal the mechanism of action of growth-inhibitory compounds in vivo, including those of antifungal, antibiotic, and anticancer drugs. The chemogenomic profiling method described here measures the sensitivity induced in yeast heterozygous and homozygous deletion strains in the presence of a chemical inhibitor of growth (termed haploinsufficiency profiling and homozygous profiling, respectively, or HIPHOP). The protocol is both scalable and amenable to automation. After competitive growth of yeast knockout collection cultures, with and without chemical inhibitors, CGIs can be identified and quantified using either array- or sequencing-based approaches as described here. PMID:27587778

  20. Environmental Application of Reporter-Genes Based Biosensors for Chemical Contamination Screening

    Directory of Open Access Journals (Sweden)

    Matejczyk Marzena

    2014-12-01

    Full Text Available The paper presents results of research concerning possibilities of applications of reporter-genes based microorganisms, including the selective presentation of defects and advantages of different new scientific achievements of methodical solutions in genetic system constructions of biosensing elements for environmental research. The most robust and popular genetic fusion and new trends in reporter genes technology – such as LacZ (β-galactosidase, xylE (catechol 2,3-dioxygenase, gfp (green fluorescent proteins and its mutated forms, lux (prokaryotic luciferase, luc (eukaryotic luciferase, phoA (alkaline phosphatase, gusA and gurA (β-glucuronidase, antibiotics and heavy metals resistance are described. Reporter-genes based biosensors with use of genetically modified bacteria and yeast successfully work for genotoxicity, bioavailability and oxidative stress assessment for detection and monitoring of toxic compounds in drinking water and different environmental samples, surface water, soil, sediments.

  1. Root Glucosinolate Profiles for Screening of Radish (Raphanus sativus L.) Genetic Resources.

    Science.gov (United States)

    Yi, Gibum; Lim, Sooyeon; Chae, Won Byoung; Park, Jeong Eun; Park, Hye Rang; Lee, Eun Jin; Huh, Jin Hoe

    2016-01-13

    Radish (Raphanus sativus L.), a root vegetable, is rich in glucosinolates (GLs), which are beneficial secondary metabolites for human health. To investigate the genetic variations in GL content in radish roots and the relationship with other root phenotypes, we analyzed 71 accessions from 23 different countries for GLs using HPLC. The most abundant GL in radish roots was glucoraphasatin, a GL with four-carbon aliphatic side chain. The content of glucoraphasatin represented at least 84.5% of the total GL content. Indolyl GL represented only 3.1% of the total GL at its maximum. The principal component analysis of GL profiles with various root phenotypes showed that four different genotypes exist in the 71 accessions. Although no strong correlation with GL content and root phenotype was observed, the varied GL content levels demonstrate the genetic diversity of GL content, and the amount that GLs could be potentially improved by breeding in radishes.

  2. [Rare diseases: specific ethical and legal aspects of genetic counseling and screening].

    Science.gov (United States)

    Sánchez-Caro, Javier

    2011-01-01

    This article analyses the specific rights of patients with rare diseases from a dual perspective. On the one hand, they concern a new generation of patients' rights that arise once the consolidation of basic rights has occurred, fundamentally after the application of Law 41/2002 (on Regulating Patient Autonomy and Rights and Obligations in the Field of Health Documentation and Information) and its development by the autonomous communities. On the other hand, the fundamental question raises a serious issue related to these patients, which involves the principles of equality, equity, non-discrimination and solidarity. This is aimed at promoting legislative measures to protect patients' equality of access to health and social services, with the ultimate aim of improving their quality of life. The author has given special relevance in his study to the treatment of rare diseases that are genetic in origin, and to the importance of adequate genetic counseling.

  3. Root Glucosinolate Profiles for Screening of Radish (Raphanus sativus L.) Genetic Resources.

    Science.gov (United States)

    Yi, Gibum; Lim, Sooyeon; Chae, Won Byoung; Park, Jeong Eun; Park, Hye Rang; Lee, Eun Jin; Huh, Jin Hoe

    2016-01-13

    Radish (Raphanus sativus L.), a root vegetable, is rich in glucosinolates (GLs), which are beneficial secondary metabolites for human health. To investigate the genetic variations in GL content in radish roots and the relationship with other root phenotypes, we analyzed 71 accessions from 23 different countries for GLs using HPLC. The most abundant GL in radish roots was glucoraphasatin, a GL with four-carbon aliphatic side chain. The content of glucoraphasatin represented at least 84.5% of the total GL content. Indolyl GL represented only 3.1% of the total GL at its maximum. The principal component analysis of GL profiles with various root phenotypes showed that four different genotypes exist in the 71 accessions. Although no strong correlation with GL content and root phenotype was observed, the varied GL content levels demonstrate the genetic diversity of GL content, and the amount that GLs could be potentially improved by breeding in radishes. PMID:26672790

  4. BIOCHEMICAL GENETIC STUDIES ON CUTTLEFISH SEPIELLA MAINDRONI (CEPHALOPODA: SEPIIDAE)- ACTIVE LOCI SCREENING OF ISOZYME

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Screening of 46 putative enzyme-coding loci and 4 different kinds of tissues of Sepiella maindroni de Rochebrone, 1884 for enzymatic activities using starch gel electrophoretic technique proved that the 21 enzymes such as AAT, AK, ALP, AP, CK, DIA, ES, FBP, G3PDH, GPI, GRS,IDH, LDH, MDH, MEP, MPI, NP, PGDH, PGM, SOD and XO* , were active to Sepiella maindroni after being stained. The tissue exhibiting stable and clear bands was also determined. Among tissues tested, mantle muscle tissue was the best for electrophoretic survey of isozymes. Buccal bulb muscle, eye and liver were fairly good for some special enzymes, such as DIA, ES, MPI, NP, etc.

  5. Genetic Screening Identifies Cyanogenesis-Deficient Mutants of Lotus japonicus and Reveals Enzymatic Specificity in Hydroxynitrile Glucoside Metabolism

    DEFF Research Database (Denmark)

    Takos, A.; Lai, D.; Mikkelsen, L.;

    2010-01-01

    . We developed a high-throughput screening method and used it to identify cyanogenesis deficient (cyd) mutants in the model legume Lotus japonicus. Mutants in both biosynthesis and catabolism of cyanogenic glucosides were isolated and classified following metabolic profiling of cyanogenic glucoside....... Catabolic mutants could be placed in two complementation groups, one of which, cyd2, encoded the beta-glucosidase BGD2. Despite the identification of nine independent cyd2 alleles, no mutants involving the gene encoding a closely related beta-glucosidase, BGD4, were identified. This indicated that BGD4...... glucosides and rhodiocyanosides. Our genetic analysis demonstrated specificity in the catabolic pathways for hydroxynitrile glucosides and implied specificity in their biosynthetic pathways as well. In addition, it has provided important tools for elucidating and potentially modifying cyanogenesis pathways...

  6. IN SILICO SCREENING OF CHEMICAL COMPOUNDS FROM SWEET FLAG (ARACUS CALAMUS L AS α-GLUCOSIDASE INHIBITOR

    Directory of Open Access Journals (Sweden)

    Dewi Yuliana

    2013-03-01

    Full Text Available Research have been conducted screening in silico chemical compound inhibitor α-glucosidase from plants dringo (Acorus calamus L based on the binding site (binding site are owned by some of the compounds obtained respectively from the inhibition of enzyme / receptor (docking using the program Argus Lab. Model of the enzyme α-glucosidase was obtained through the protein data bank with the code 1lwj in the donwload NCBI website. Models of chemical compounds contained in dringo (A. Calamus L obtained through the site Take out "jamu" Knapsack and made in the formula structures of 2D and 3D using the program ACD / Chemsketch. Docking results showed activity in the compound 1-ethenyl-1-methyl-2,4-at (prop-1-en-2-yl Cyclohexane with free energy - 8.04385 kcal / mol, and the compound Isocaespitol with a free energy - 8.28388 kcal / mol.

  7. Dermal permeation data and models for the prioritization and screening-level exposure assessment of organic chemicals.

    Science.gov (United States)

    Brown, Trevor N; Armitage, James M; Egeghy, Peter; Kircanski, Ida; Arnot, Jon A

    2016-09-01

    High-throughput screening (HTS) models are being developed and applied to prioritize chemicals for more comprehensive exposure and risk assessment. Dermal pathways are possible exposure routes to humans for thousands of chemicals found in personal care products and the indoor environment. HTS exposure models rely on skin permeability coefficient (KP; cm/h) models for exposure predictions. An initial database of approximately 1000 entries for empirically-based KP data was compiled from the literature and a subset of 480 data points for 245 organic chemicals derived from testing with human skin only and using only water as a vehicle was selected. The selected dataset includes chemicals with log octanol-water partition coefficients (KOW) ranging from -6.8 to 7.6 (median=1.8; 95% of the data range from -2.5 to 4.6) and molecular weight (MW) ranging from 18 to 765g/mol (median=180); only 3% >500g/mol. Approximately 53% of the chemicals in the database have functional groups which are ionizable in the pH range of 6 to 7.4, with 31% being appreciably ionized. The compiled log KP values ranged from -5.8 to 0.1cm/h (median=-2.6). The selected subset of the KP data was then used to evaluate eight representative KP models that can be readily applied for HTS assessments, i.e., parameterized with KOW and MW. The analysis indicates that a version of the SKINPERM model performs the best against the selected dataset. Comparisons of representative KP models against model input parameter property ranges (sensitivity analysis) and against chemical datasets requiring human health assessment were conducted to identify regions of chemical properties that should be tested to address uncertainty in KP models and HTS exposure assessments. PMID:27282209

  8. A tree based method for the rapid screening of chemical fingerprints

    DEFF Research Database (Denmark)

    Kristensen, Thomas Greve; Nielsen, Jesper; Pedersen, Christian Nørgaard Storm

    2009-01-01

    The fingerprint of a molecule is a bitstring based on its structure, constructed such that structurally similar molecules will have similar fingerprints. Molecular fingerprints can be used in an initial phase for identifying novel drug candidates by screening large databases for molecules...... with fingerprints similar to a query fingerprint. In this paper, we present a method which efficiently finds all fingerprints in a database with Tanimoto coefficient to the query fingerprint above a user defined threshold. The method is based on two novel data structures for rapid screening of large databases......: the kD grid and the Multibit tree. The kD grid is based on splitting the fingerprints into k shorter bitstrings and utilising these to compute bounds on the similarity of the complete bitstrings. The Multibit tree uses hierarchical clustering and similarity within each cluster to compute similar bounds...

  9. Genetic Screen Reveals the Role of Purine Metabolism in Staphylococcus aureus Persistence to Rifampicin

    Directory of Open Access Journals (Sweden)

    Rebecca Yee

    2015-12-01

    Full Text Available Chronic infections with Staphylococcus aureus such as septicemia, osteomyelitis, endocarditis, and biofilm infections are difficult to treat because of persisters. Despite many efforts in understanding bacterial persistence, the mechanisms of persister formation in S. aureus remain elusive. Here, we performed a genome-wide screen of a transposon mutant library to study the molecular mechanisms involved in persistence of community-acquired S. aureus. Screening of the library for mutants defective in persistence or tolerance to rifampicin revealed many genes involved in metabolic pathways that are important for antibiotic persistence. In particular, the identified mutants belonged to metabolic pathways involved in carbohydrate, amino acid, lipid, vitamin and purine biosynthesis. Five mutants played a role in purine biosynthesis and two mutants, purB, an adenylosuccinate lyase, and purM, a phosphoribosylaminoimidazole synthetase, were selected for further confirmation. Mutants purB and purM showed defective persistence compared to the parental strain USA300 in multiple stress conditions including various antibiotics, low pH, and heat stress. The defect in persistence was restored by complementation with the wildtype purB and purM gene in the respective mutants. These findings provide new insights into the mechanisms of persistence in S. aureus and provide novel therapeutic targets for developing more effective treatment for persistent infections due to S. aureus.

  10. Quantitative Phenotyping-Based In Vivo Chemical Screening in a Zebrafish Model of Leukemia Stem Cell Xenotransplantation

    Science.gov (United States)

    Zhang, Beibei; Shimada, Yasuhito; Kuroyanagi, Junya; Umemoto, Noriko; Nishimura, Yuhei; Tanaka, Toshio

    2014-01-01

    Zebrafish-based chemical screening has recently emerged as a rapid and efficient method to identify important compounds that modulate specific biological processes and to test the therapeutic efficacy in disease models, including cancer. In leukemia, the ablation of leukemia stem cells (LSCs) is necessary to permanently eradicate the leukemia cell population. However, because of the very small number of LSCs in leukemia cell populations, their use in xenotransplantation studies (in vivo) and the difficulties in functionally and pathophysiologically replicating clinical conditions in cell culture experiments (in vitro), the progress of drug discovery for LSC inhibitors has been painfully slow. In this study, we developed a novel phenotype-based in vivo screening method using LSCs xenotransplanted into zebrafish. Aldehyde dehydrogenase-positive (ALDH+) cells were purified from chronic myelogenous leukemia K562 cells tagged with a fluorescent protein (Kusabira-orange) and then implanted in young zebrafish at 48 hours post-fertilization. Twenty-four hours after transplantation, the animals were treated with one of eight different therapeutic agents (imatinib, dasatinib, parthenolide, TDZD-8, arsenic trioxide, niclosamide, salinomycin, and thioridazine). Cancer cell proliferation, and cell migration were determined by high-content imaging. Of the eight compounds that were tested, all except imatinib and dasatinib selectively inhibited ALDH+ cell proliferation in zebrafish. In addition, these anti-LSC agents suppressed tumor cell migration in LSC-xenotransplants. Our approach offers a simple, rapid, and reliable in vivo screening system that facilitates the phenotype-driven discovery of drugs effective in suppressing LSCs. PMID:24454867

  11. High-content chemical and RNAi screens for suppressors of neurotoxicity in a Huntington's disease model.

    Directory of Open Access Journals (Sweden)

    Joost Schulte

    Full Text Available To identify Huntington's Disease therapeutics, we conducted high-content small molecule and RNAi suppressor screens using a Drosophila primary neural culture Huntingtin model. Drosophila primary neurons offer a sensitive readout for neurotoxicty, as their neurites develop dysmorphic features in the presence of mutant polyglutamine-expanded Huntingtin compared to nonpathogenic Huntingtin. By tracking the subcellular distribution of mRFP-tagged pathogenic Huntingtin and assaying neurite branch morphology via live-imaging, we identified suppressors that could reduce Huntingtin aggregation and/or prevent the formation of dystrophic neurites. The custom algorithms we used to quantify neurite morphologies in complex cultures provide a useful tool for future high-content screening approaches focused on neurodegenerative disease models. Compounds previously found to be effective aggregation inhibitors in mammalian systems were also effective in Drosophila primary cultures, suggesting translational capacity between these models. However, we did not observe a direct correlation between the ability of a compound or gene knockdown to suppress aggregate formation and its ability to rescue dysmorphic neurites. Only a subset of aggregation inhibitors could revert dysmorphic cellular profiles. We identified lkb1, an upstream kinase in the mTOR/Insulin pathway, and four novel drugs, Camptothecin, OH-Camptothecin, 18β-Glycyrrhetinic acid, and Carbenoxolone, that were strong suppressors of mutant Huntingtin-induced neurotoxicity. Huntingtin neurotoxicity suppressors identified through our screen also restored viability in an in vivo Drosophila Huntington's Disease model, making them attractive candidates for further therapeutic evaluation.

  12. Interlaboratory validation of quantitative duplex real-time PCR method for screening analysis of genetically modified maize.

    Science.gov (United States)

    Takabatake, Reona; Koiwa, Tomohiro; Kasahara, Masaki; Takashima, Kaori; Futo, Satoshi; Minegishi, Yasutaka; Akiyama, Hiroshi; Teshima, Reiko; Oguchi, Taichi; Mano, Junichi; Furui, Satoshi; Kitta, Kazumi

    2011-01-01

    To reduce the cost and time required to routinely perform the genetically modified organism (GMO) test, we developed a duplex quantitative real-time PCR method for a screening analysis simultaneously targeting an event-specific segment for GA21 and Cauliflower Mosaic Virus 35S promoter (P35S) segment [Oguchi et al., J. Food Hyg. Soc. Japan, 50, 117-125 (2009)]. To confirm the validity of the method, an interlaboratory collaborative study was conducted. In the collaborative study, conversion factors (Cfs), which are required to calculate the GMO amount (%), were first determined for two real-time PCR instruments, the ABI PRISM 7900HT and the ABI PRISM 7500. A blind test was then conducted. The limit of quantitation for both GA21 and P35S was estimated to be 0.5% or less. The trueness and precision were evaluated as the bias and reproducibility of the relative standard deviation (RSD(R)). The determined bias and RSD(R) were each less than 25%. We believe the developed method would be useful for the practical screening analysis of GM maize.

  13. A highly sensitive and specific method for the screening detection of genetically modified organisms based on digital PCR without pretreatment.

    Science.gov (United States)

    Fu, Wei; Zhu, Pengyu; Wang, Chenguang; Huang, Kunlun; Du, Zhixin; Tian, Wenying; Wang, Qin; Wang, Huiyu; Xu, Wentao; Zhu, Shuifang

    2015-08-04

    Digital PCR has developed rapidly since it was first reported in the 1990 s. It was recently reported that an improved method facilitated the detection of genetically modified organisms (GMOs). However, to use this improved method, the samples must be pretreated, which could introduce inaccuracy into the results. In our study, we explored a pretreatment-free digital PCR detection method for the screening for GMOs. We chose the CaMV35s promoter and the NOS terminator as the templates in our assay. To determine the specificity of our method, 9 events of GMOs were collected, including MON810, MON863, TC1507, MIR604, MIR162, GA21, T25, NK603 and Bt176. Moreover, the sensitivity, intra-laboratory and inter-laboratory reproducibility of our detection method were assessed. The results showed that the limit of detection of our method was 0.1%, which was lower than the labeling threshold level of the EU. The specificity and stability among the 9 events were consistent, respectively. The intra-laboratory and inter-laboratory reproducibility were both good. Finally, the perfect fitness for the detection of eight double-blind samples indicated the good practicability of our method. In conclusion, the method in our study would allow more sensitive, specific and stable screening detection of the GMO content of international trading products.

  14. A highly sensitive and specific method for the screening detection of genetically modified organisms based on digital PCR without pretreatment.

    Science.gov (United States)

    Fu, Wei; Zhu, Pengyu; Wang, Chenguang; Huang, Kunlun; Du, Zhixin; Tian, Wenying; Wang, Qin; Wang, Huiyu; Xu, Wentao; Zhu, Shuifang

    2015-01-01

    Digital PCR has developed rapidly since it was first reported in the 1990 s. It was recently reported that an improved method facilitated the detection of genetically modified organisms (GMOs). However, to use this improved method, the samples must be pretreated, which could introduce inaccuracy into the results. In our study, we explored a pretreatment-free digital PCR detection method for the screening for GMOs. We chose the CaMV35s promoter and the NOS terminator as the templates in our assay. To determine the specificity of our method, 9 events of GMOs were collected, including MON810, MON863, TC1507, MIR604, MIR162, GA21, T25, NK603 and Bt176. Moreover, the sensitivity, intra-laboratory and inter-laboratory reproducibility of our detection method were assessed. The results showed that the limit of detection of our method was 0.1%, which was lower than the labeling threshold level of the EU. The specificity and stability among the 9 events were consistent, respectively. The intra-laboratory and inter-laboratory reproducibility were both good. Finally, the perfect fitness for the detection of eight double-blind samples indicated the good practicability of our method. In conclusion, the method in our study would allow more sensitive, specific and stable screening detection of the GMO content of international trading products. PMID:26239916

  15. Identification of small molecule and genetic modulators of AON-induced dystrophin exon skipping by high-throughput screening.

    Directory of Open Access Journals (Sweden)

    Debra A O'Leary

    Full Text Available One therapeutic approach to Duchenne Muscular Dystrophy (DMD recently entering clinical trials aims to convert DMD phenotypes to that of a milder disease variant, Becker Muscular Dystrophy (BMD, by employing antisense oligonucleotides (AONs targeting splice sites, to induce exon skipping and restore partial dystrophin function. In order to search for small molecule and genetic modulators of AON-dependent and independent exon skipping, we screened approximately 10,000 known small molecule drugs, >17,000 cDNA clones, and >2,000 kinase- targeted siRNAs against a 5.6 kb luciferase minigene construct, encompassing exon 71 to exon 73 of human dystrophin. As a result, we identified several enhancers of exon skipping, acting on both the reporter construct as well as endogenous dystrophin in mdx cells. Multiple mechanisms of action were identified, including histone deacetylase inhibition, tubulin modulation and pre-mRNA processing. Among others, the nucleolar protein NOL8 and staufen RNA binding protein homolog 2 (Stau2 were found to induce endogenous exon skipping in mdx cells in an AON-dependent fashion. An unexpected but recurrent theme observed in our screening efforts was the apparent link between the inhibition of cell cycle progression and the induction of exon skipping.

  16. Genetic and chemical modifiers of a CUG toxicity model in Drosophila.

    Directory of Open Access Journals (Sweden)

    Amparo Garcia-Lopez

    Full Text Available Non-coding CUG repeat expansions interfere with the activity of human Muscleblind-like (MBNL proteins contributing to myotonic dystrophy 1 (DM1. To understand this toxic RNA gain-of-function mechanism we developed a Drosophila model expressing 60 pure and 480 interrupted CUG repeats in the context of a non-translatable RNA. These flies reproduced aspects of the DM1 pathology, most notably nuclear accumulation of CUG transcripts, muscle degeneration, splicing misregulation, and diminished Muscleblind function in vivo. Reduced Muscleblind activity was evident from the sensitivity of CUG-induced phenotypes to a decrease in muscleblind genetic dosage and rescue by MBNL1 expression, and further supported by the co-localization of Muscleblind and CUG repeat RNA in ribonuclear foci. Targeted expression of CUG repeats to the developing eye and brain mushroom bodies was toxic leading to rough eyes and semilethality, respectively. These phenotypes were utilized to identify genetic and chemical modifiers of the CUG-induced toxicity. 15 genetic modifiers of the rough eye phenotype were isolated. These genes identify putative cellular processes unknown to be altered by CUG repeat RNA, and they include mRNA export factor Aly, apoptosis inhibitor Thread, chromatin remodelling factor Nurf-38, and extracellular matrix structural component Viking. Ten chemical compounds suppressed the semilethal phenotype. These compounds significantly improved viability of CUG expressing flies and included non-steroidal anti-inflammatory agents (ketoprofen, muscarinic, cholinergic and histamine receptor inhibitors (orphenadrine, and drugs that can affect sodium and calcium metabolism such as clenbuterol and spironolactone. These findings provide new insights into the DM1 phenotype, and suggest novel candidates for DM1 treatments.

  17. A genetic screen identifies Tor as an interactor of VAPB in a Drosophila model of amyotrophic lateral sclerosis.

    Science.gov (United States)

    Deivasigamani, Senthilkumar; Verma, Hemant Kumar; Ueda, Ryu; Ratnaparkhi, Anuradha; Ratnaparkhi, Girish S

    2014-10-31

    Amyotrophic Lateral Sclerosis (ALS) is a progressive neurodegenerative disorder characterized by selective death of motor neurons. In 5-10% of the familial cases, the disease is inherited because of mutations. One such mutation, P56S, was identified in human VAPB that behaves in a dominant negative manner, sequestering wild type protein into cytoplasmic inclusions. We have conducted a reverse genetic screen to identify interactors of Drosophila VAPB. We screened 2635 genes and identified 103 interactors, of which 45 were enhancers and 58 were suppressors of VAPB function. Interestingly, the screen identified known ALS loci - TBPH, alsin2 and SOD1. Also identified were genes involved in cellular energetics and homeostasis which were used to build a gene regulatory network of VAPB modifiers. One key modifier identified was Tor, whose knockdown reversed the large bouton phenotype associated with VAP(P58S) expression in neurons. A similar reversal was seen by over-expressing Tuberous Sclerosis Complex (Tsc1,2) that negatively regulates TOR signaling as also by reduction of S6K activity. In comparison, the small bouton phenotype associated with VAP(wt) expression was reversed with Tsc1 knock down as well as S6K-CA expression. Tor therefore interacts with both VAP(wt) and VAP(P58S), but in a contrasting manner. Reversal of VAP(P58S) bouton phenotypes in larvae fed with the TOR inhibitor Rapamycin suggests upregulation of TOR signaling in response to VAP(P58S) expression. The VAPB network and further mechanistic understanding of interactions with key pathways, such as the TOR cassette, will pave the way for a better understanding of the mechanisms of onset and progression of motor neuron disease.

  18. A genetic screen identifies Tor as an interactor of VAPB in a Drosophila model of amyotrophic lateral sclerosis

    Directory of Open Access Journals (Sweden)

    Senthilkumar Deivasigamani

    2014-10-01

    Full Text Available Amyotrophic Lateral Sclerosis (ALS is a progressive neurodegenerative disorder characterized by selective death of motor neurons. In 5–10% of the familial cases, the disease is inherited because of mutations. One such mutation, P56S, was identified in human VAPB that behaves in a dominant negative manner, sequestering wild type protein into cytoplasmic inclusions. We have conducted a reverse genetic screen to identify interactors of Drosophila VAPB. We screened 2635 genes and identified 103 interactors, of which 45 were enhancers and 58 were suppressors of VAPB function. Interestingly, the screen identified known ALS loci – TBPH, alsin2 and SOD1. Also identified were genes involved in cellular energetics and homeostasis which were used to build a gene regulatory network of VAPB modifiers. One key modifier identified was Tor, whose knockdown reversed the large bouton phenotype associated with VAP(P58S expression in neurons. A similar reversal was seen by over-expressing Tuberous Sclerosis Complex (Tsc1,2 that negatively regulates TOR signaling as also by reduction of S6K activity. In comparison, the small bouton phenotype associated with VAP(wt expression was reversed with Tsc1 knock down as well as S6K-CA expression. Tor therefore interacts with both VAP(wt and VAP(P58S, but in a contrasting manner. Reversal of VAP(P58S bouton phenotypes in larvae fed with the TOR inhibitor Rapamycin suggests upregulation of TOR signaling in response to VAP(P58S expression. The VAPB network and further mechanistic understanding of interactions with key pathways, such as the TOR cassette, will pave the way for a better understanding of the mechanisms of onset and progression of motor neuron disease.

  19. Evaluation of food-relevant chemicals in the ToxCast high-throughput screening program

    Data.gov (United States)

    U.S. Environmental Protection Agency — Thousands of chemicals are directly added to or come in contact with food, many of which have undergone little to no toxicological evaluation. The landscape of the...

  20. SCREENING OF CHEMICAL COMPOSITIONS OF CRUDE WATER EXTRACT OF DIFFERENT CASSAVA VARIETIES

    OpenAIRE

    Olajumoke Oke FAYINMINNU; Olubunmi Omowunmi FADINA; Alex Adeoluwa ADEDAPO

    2013-01-01

    Chemical composition of three sources of crude cassava water extract (CCWE) was evaluated in different varieties of cassava (MS6 Manihot Selection (local variety), TMS 30555 Tropical Manihot Selection (Improved variety) and Bulk (crude cassava water from cassava processing site). Crude cassava water extract from the pulp of cassava fresh roots was prepared and the chemical composition was determined in the analytical laboratory. The result of the analysis showed that, hydrocyanic acid (HCN) ...

  1. Sulfonamides identified as plant immune-priming compounds in high-throughput chemical screening increase disease resistance in Arabidopsis thaliana

    Directory of Open Access Journals (Sweden)

    Yoshiteru eNoutoshi

    2012-10-01

    Full Text Available Plant activators are agrochemicals that protect crops from diseases by activating the plant immune system. To isolate lead compounds for use as practical plant activators, we screened 2 different chemical libraries composed of various bioactive substances by using an established screening procedure that can selectively identify immune-priming compounds. We identified and characterized a group of sulfonamide compounds—sulfameter, sulfamethoxypyridazine, sulfabenzamide, and sulfachloropyridazine—among the various isolated candidate molecules. These sulfonamide compounds enhanced the avirulent Pseudomonas-induced cell death of Arabidopsis suspension cell cultures and increased disease resistance in Arabidopsis plants against both avirulent and virulent strains of the bacterium. These compounds did not prevent the growth of pathogenic bacteria in minimal liquid media at 200 µM. They also did not induce the expression of defense-related genes in Arabidopsis seedlings, at least not at 24 and 48 h after treatment, suggesting that they do not act as salicylic acid analogs. In addition, although sulfonamides are known to be folate biosynthesis inhibitors, the application of folate did not restore the potentiation effects of the sulfonamides on pathogen-induced cell death. Our data suggest that sulfonamides potentiate Arabidopsis disease resistance by their novel chemical properties.

  2. The role of genetic and chemical variation of Pinus sylvestris seedlings in influencing slug herbivory.

    Science.gov (United States)

    O'Reilly-Wapstra, Julianne M; Iason, Glenn R; Thoss, Vera

    2007-05-01

    This study investigated the genetic and chemical basis of resistance of Pinus sylvestris seedlings to herbivory by a generalist mollusc, Arion ater. Using feeding trials with captive animals, we examined selective herbivory by A. ater of young P. sylvestris seedlings of different genotypes and correlated preferences with seedling monoterpene levels. We also investigated the feeding responses of A. ater to artificial diets laced with two monoterpenes, Delta(3)-carene and alpha-pinene. Logistic regression indicated that two factors were the best predictors of whether seedlings in the trial would be consumed. Individual slug variation (replicates) was the most significant factor in the model; however, alpha-pinene concentration (also representing beta-pinene, Delta(3)-carene and total monoterpenes due to multicollinearity) of needles was also a significant factor. While A. ater did not select seedlings on the basis of family, seedlings not eaten were significantly higher in levels of alpha-pinene compared to seedlings that were consumed. We also demonstrated significant genetic variation in alpha-pinene concentration of seedlings between different families of P. sylvestris. Nitrogen and three morphological seedling characteristics (stem length, needle length and stem diameter) also showed significant genetic variation between P. sylvestris families. Artificial diets laced with high (5 mg g(-1) dry matter) quantities of either Delta(3)-carene or alpha-pinene, were eaten significantly less than control diets with no added monoterpenes, supporting the results of the seedling feeding trial. This study demonstrates that A. ater selectively feed on P. sylvestris seedlings and that this selection is based, in part, on the monoterpene concentration of seedlings. These results, coupled with significant genetic variation in alpha-pinene concentration of seedlings and evidence that slug herbivory is detrimental to P. sylvestris fitness, are discussed as possible evidence for A

  3. The use of isolated natural products as scaffolds for the generation of chemically diverse screening libraries for drug discovery.

    Science.gov (United States)

    Barnes, Emma C; Kumar, Rohitesh; Davis, Rohan A

    2016-03-01

    A diverse range of strategies leading to natural product derived or inspired screening libraries aims to increase the number of new chemical entities emerging per year. However, the use of isolated natural products as scaffolds for the semi-synthesis of larger biological screening libraries remains rare. This particular method avoids the time-consuming and resource intensive de novo synthetic strategy for scaffold production, and has become more feasible through improvements to synthetic and isolation methodologies. This Highlight examines the increasing popularity of small- to large-sized screening libraries generated directly from isolated natural products. Several of the examples detailed herein show how this strategy can lead to improvements in not only potency but also other important (and often forgotten) drug discovery parameters such as toxicity, selectivity, lipophilicity and bioavailability. However, there are still improvements to be made to this method, particularly in the choice of the natural product scaffold and the derivatising reagents used. Avoidance of known nuisance compounds or structural alert motifs (e.g. PAINS) that interfere with bioactivity screens, and impact downstream drug development will play a significant role in the future success of this methodology. Incorporation of rational design strategies that take into account the physicochemical parameters (e.g. log P, MW, HBA, HBD) of the final semi-synthetic library analogues will also facilitate the discovery and development of leads and drugs. A multi-pronged approach to drug discovery that incorporates the use of isolated natural product scaffolds for library generation will surely be beneficial. PMID:26739749

  4. Fabry disease mimicking hypertrophic cardiomyopathy: genetic screening needed for establishing the diagnosis in women

    DEFF Research Database (Denmark)

    Havndrup, Ole; Christiansen, Michael; Stoevring, Birgitte;

    2010-01-01

    AIMS: Fabry disease, an X-linked storage disorder caused by defective lysosomal enzyme alpha-galactosidase A activity, may resemble sarcomere-gene-associated hypertrophic cardiomyopathy (HCM). The 'cardiac variant' of Fabry disease which only affects the heart may be missed unless specifically....... In the remaining 59 probands, 3 (5%) had GLA mutations as follows. The first proband, a female with asymmetric septal hypertrophy (ASH), a significant left ventricular outflow tract gradient, and chronic obstructive pulmonary disease, was heterozygous for a novel missense mutation (p.N139S). The second proband...... therapy, supports systematic testing for Fabry disease. Enzyme measurements are sufficient in men, but genetic testing is needed in women....

  5. The genetics of multiple sclerosis: principles, background and updated results of the United Kingdom systematic genome screen.

    Science.gov (United States)

    Chataway, J; Feakes, R; Coraddu, F; Gray, J; Deans, J; Fraser, M; Robertson, N; Broadley, S; Jones, H; Clayton, D; Goodfellow, P; Sawcer, S; Compston, A

    1998-10-01

    Genetic susceptibility to multiple sclerosis is implicated on the basis of classical family studies and phenotype analyses. The only reproducible legacy from the candidate gene approach has been the discovery of population associations with alleles of the major histocompatibility complex. Systematic genome scanning has since been applied using a panel of anonymous markers to identify areas of linkage in co-affected siblings. Here, we describe the principles of genome screening and update the UK survey of multiple sclerosis. This identified 20 regions of potential interest, but in none was there unequivocal linkage. In theory, attempting to replicate these findings in a second set of sibling pair families is the most appropriate way to distinguish true from false positives, but unfortunately the number of families required to do this reliably is prohibitively large. We used three approaches to increase the definition achieved by the screen: (i) the number of sibling pairs typed in an identified region of potential linkage was extended; (ii) the information extraction was increased in an identified region; and (iii) a search was made for missed regions of potential linkage. Each of these approaches has considerable limitations. A chromosome-by-chromosome account is given to direct future searches. Although an additional marker placed distal to the 'hit' on chromosome 14q increased linkage in this area, and typing extra sibling pairs increased linkage on chromosomes 6p and 17q, evidence for linkage was more commonly reduced and no additional regions of interest were found. A further refinement of the genome screen was undertaken by conditioning for the presence of HLA-DR15. This produced a surprising degree of segregation among the regions of interest, which divided into two distinct groups depending on DR15 sharing: the DR15-sharing cohort comprised loci on chromosomal areas 1p, 17q and X; and the DR15-non-sharing cohort was made up of loci on 1cen, 3p, 7p, 14q and

  6. Ecological consequences, genetic and chemical variations in fragmented populations of a medicinal plant, justicia adhatoda and implications for its conservation

    International Nuclear Information System (INIS)

    Justicia adhatoda from Kohat Plateau was selected for genetic diversity studies, due to its fragmented habitat, importance in traditional and pharmaceutical medicine and a lack of population structure studies. We had two hypotheses: that habitat loss posed a greater threat to populations than loss of genetic diversity, and that chemical diversity would be higher among different populations than within populations. Genetic diversity within and among populations was evaluated using PBA (P450 based analogue) markers. AMOVA analysis revealed that there was higher genetic diversity within populations (90%) than among populations (10%). No genetic drift was observed, i.e., genetic diversity within populations was maintained despite fewer numbers of individuals in fragmented populations. Surveys of J. adhatoda populations revealed that they were growing in harsh conditions and were imperiled due to extensive harvesting for commercial and domestic purposes. Chemical diversity was evaluated by GC-MS (Gas Chromatograph-Mass Spectrometry) analysis of 90% methanol and 1:2 chloroform:methanol extracts. GC-MS analysis of both the extracts showed nine and 18 chemical compounds, respectively, with higher chemical variations among populations. It is therefore recommended that efforts for the conservation of severely fragmented populations of J. adhatoda must be carried out along with sustainable harvesting. (author)

  7. Screening and genetic improvement of pectinolytic fungi for degumming of textile fibers

    Directory of Open Access Journals (Sweden)

    Molina Silvia M.G.

    2001-01-01

    Full Text Available Aiming at contributing to technological improvements in plant fiber processing methods, this paper reports research work on the obtainment of more efficient pectinase-producing fungi strains. More specifically, this work reports the analysis of 18 strains of filamentous fungi, with the purpose of obtaining enzymes for textile fibers degumming. The strains were evaluated for production of pectinolytic enzymes under several growth conditions (culture medium and growth temperature. Production of pectinases was measured by an enzymatic index (EI in solid pectin medium. Among the tested strains, Penicillium chrysogenum IFO 4626 (Q 176 showed the best performance. Genetic improvement of this strain was carried out to increase its pectinase production, while keeping cellulase activity down to a negligible level, since cellulases are known to decrease the resistance of the fiber. Variability was induced through several cycles of mutation and selection by exposing conidea to ultra-violet light (UV. We selected 39 out of 390 isolated colonies. Resulting mutants produced nine times more pectin lyase (PL than the original strain in terms of PL specific activity, and five times more in terms of PL activity (i.e. mmoles liberated per minute of reaction per mL of medium. Periodically, mutant performance was evaluated in solid pectin medium. Genetic stability was maintained for four years after isolation.

  8. Mutation screening of three Chinese families with genetic epilepsy with febrile seizures plus.

    Science.gov (United States)

    Lin, Hua; Li, Jingyun; Wang, Mengyang; Wang, Zheng; Wang, Yuping; Wu, Liwen

    2011-08-15

    Genetic epilepsy with febrile seizures plus (GEFS+) is a familial autosomal dominant condition characterized by genetic heterogeneity. Five genes for GEFS+ identified in large families account for only a small proportion of families. Mutation in the majority of families with GEFS+ has not identified yet. The aim of our study is to search for the gene responsible for GEFS+ in three Chinese families by linkage analyses and a sequencing approach and to investigate the importance of coding and noncoding regions variations of four known GEFS+ genes (SCN1A, SCN1B, GABRG2 and SCN2A) in Chinese families. Results showed that a 6-cM candidate interval at 5q33-34 with a maximum LOD scores of 2.043 was identified in families B. Sequencing candidate gene GABRG2 and GABRA1 in this region did not identify a causative mutation. Moreover, no mutation was found in coding and noncoding regions of the four genes in three Chinese families. Besides excluding coding regions of four known GEFS+ genes, we also excluded the possibility of a mutation in the promoter, exon-intron boundaries, 5' untranslated regions (5' UTRs), and 3' UTRs of four known GEFS+ genes in three Chinese families. In conclusion, the present study demonstrates the heterogeneity of the etiologies of GEFS+. There are as yet undiscovered mechanisms underlying GEFS+. PMID:21704126

  9. Gametocytocidal screen identifies novel chemical classes with Plasmodium falciparum transmission blocking activity.

    Directory of Open Access Journals (Sweden)

    Natalie G Sanders

    Full Text Available Discovery of transmission blocking compounds is an important intervention strategy necessary to eliminate and eradicate malaria. To date only a small number of drugs that inhibit gametocyte development and thereby transmission from the mosquito to the human host exist. This limitation is largely due to a lack of screening assays easily adaptable to high throughput because of multiple incubation steps or the requirement for high gametocytemia. Here we report the discovery of new compounds with gametocytocidal activity using a simple and robust SYBR Green I- based DNA assay. Our assay utilizes the exflagellation step in male gametocytes and a background suppressor, which masks the staining of dead cells to achieve healthy signal to noise ratio by increasing signal of viable parasites and subtracting signal from dead parasites. By determining the contribution of exflagellation to fluorescent signal and using appropriate cutoff values, we were able to screen for gametocytocidal compounds. After assay validation and optimization, we screened an FDA approved drug library of approximately 1500 compounds, as well as the 400 compound MMV malaria box and identified 44 gametocytocidal compounds with sub to low micromolar IC50s. Major classes of compounds with gametocytocidal activity included quaternary ammonium compounds with structural similarity to choline, acridine-like compounds similar to quinacrine and pyronaridine, as well as antidepressant, antineoplastic, and anthelminthic compounds. Top drug candidates showed near complete transmission blocking in membrane feeding assays. This assay is simple, reproducible and demonstrated robust Z-factor values at low gametocytemia levels, making it amenable to HTS for identification of novel and potent gametocytocidal compounds.

  10. Metabolic and Genetic Screening of Electromagnetic Hypersensitive Subjects as a Feasible Tool for Diagnostics and Intervention

    OpenAIRE

    Chiara De Luca; Jeffrey Chung Sheun Thai; Desanka Raskovic; Eleonora Cesareo; Daniela Caccamo; Arseny Trukhanov; Liudmila Korkina

    2014-01-01

    Growing numbers of “electromagnetic hypersensitive” (EHS) people worldwide self-report severely disabling, multiorgan, non-specific symptoms when exposed to low-dose electromagnetic radiations, often associated with symptoms of multiple chemical sensitivity (MCS) and/or other environmental “sensitivity-related illnesses” (SRI). This cluster of chronic inflammatory disorders still lacks validated pathogenetic mechanism, diagnostic biomarkers, and management guidelines. We hypothesized that SRI...

  11. Phyto chemical Screening, Antibacterial, Antifungal and Anthelmintic Activity of Morinda citrifolia stem

    Directory of Open Access Journals (Sweden)

    Dr. D. Gopala Krishna

    2013-05-01

    Full Text Available In the present study, the Petroleum Ether and Alcoholic extract of Morinda citrifolia L. (Noni stem were subjected to preliminary screening for Antimicrobial and Aanthelmintic activity. The alcoholic extract exhibited significant Anti bacterial, Antifungal activity, comparable to the standard drug Tetracycline. The Petroleum Ether and Alcoholic extract were evaluated for Anthelmintic activity on adult Indian Earthworms, ‘Pheretima posithuma’. The Alcoholic extract produced more significant Anthelmintic activity than Petroleum ether extract and the activities are comparable with the reference drug Piperazine citrate

  12. Genetic and Chemical Profiling of Gymnema sylvestre Accessions from Central India: Its Implication for Quality Control and Therapeutic Potential of Plant

    Science.gov (United States)

    Verma, Ashutosh Kumar; Dhawan, Sunita Singh; Singh, Seema; Bharati, Kumar Avinash; Jyotsana

    2016-01-01

    Background: Gymnema sylvestre, a vulnerable plant species, is mentioned in Indian Pharmacopeia as an antidiabetic drug Objective: Study of genetic and chemical diversity and its implications in accessions of G. sylvestre Materials and Methods: Fourteen accessions of G. sylvestre collected from Central India and assessment of their genetic and chemical diversity were carried out using ISSR (inter simple sequence repeat) and HPLC (high performance liquid chromatography) fingerprinting methods Results: Among the screened 40 ISSR primers, 15 were found polymorphic and collectively produced nine unique accession-specific bands. The maximum and minimum numbers of amplicones were noted for ISSR-15 and ISSR-11, respectively. The ISSR -11 and ISSR-13 revealed 100% polymorphism. HPLC chromatograms showed that accessions possess the secondary metabolites of mid-polarity with considerable variability. Unknown peaks with retention time 2.63, 3.41, 23.83, 24.50, and 44.67 were found universal type. Comparative hierarchical clustering analysis based on foresaid fingerprints indicates that both techniques have equal potential to discriminate accessions according to percentage gymnemic acid in their leaf tissue. Second approach was noted more efficiently for separation of accessions according to their agro-climatic/collection site Conclusion: Highly polymorphic ISSRs could be utilized as molecular probes for further selection of high gymnemic acid yielding accessions. Observed accession specific bands may be used as a descriptor for plant accessions protection and converted into sequence tagged sites markers. Identified five universal type peaks could be helpful in identification of G. sylvestre-based various herbal preparations. SUMMARY Nine accession specific unique bandsFive marker peaks for G. sylvestre.Suitability of genetic and chemical fingerprinting Abbreviations used: HPLC: High Performance Liquid Chromatography, ISSR: Inter Simple Sequence Repeats, CTAB: Cetyl

  13. Pregnancy outcome after preimplantation genetic screening or natural conception in couples with unexplained recurrent miscarriage: a systematic review of the best available evidence.

    NARCIS (Netherlands)

    Musters, A.M.; Repping, S.; Korevaar, J.C.; Mastenbroek, S.; Limpens, J.; Veen, F. van der; Goddijn, M.

    2011-01-01

    The objective of this systematic review was to assess live birth rates and miscarriage rates after preimplantation genetic screening or natural conception for unexplained recurrent miscarriage. There were no randomized controlled trials or comparative studies found on this topic. Until data from ran

  14. Mental, psychomotor, neurologic, and behavioral outcomes of 2-year-old children born after preimplantation genetic screening: follow-up of a randomized controlled trial

    NARCIS (Netherlands)

    K.J. Middelburg; M. van der Heide; B. Houtzager; M. Jongbloed-Pereboom; V. Fidler; A.F. Bos; J. de Kok; M. Hadders-Algra

    2011-01-01

    Objective: To evaluate the effect of preimplantation genetic screening (PGS) on neurodevelopmental outcomes in children. Design: Prospective, assessor-blinded, follow-up study of children born to women randomly assigned to in vitro fertilization or intracytoplasmic sperm injection (IVF/ICSI) with or

  15. Genetics

    Science.gov (United States)

    ... Inheritance; Heterozygous; Inheritance patterns; Heredity and disease; Heritable; Genetic markers ... The chromosomes are made up of strands of genetic information called DNA. Each chromosome contains sections of ...

  16. Chemical Genetics of Rapamycin-Insensitive TORC2 in S. cerevisiae

    Directory of Open Access Journals (Sweden)

    Joseph I. Kliegman

    2013-12-01

    Full Text Available Current approaches for identifying synergistic targets use cell culture models to see if the combined effect of clinically available drugs is better than predicted by their individual efficacy. New techniques are needed to systematically and rationally identify targets and pathways that may be synergistic targets. Here, we created a tool to screen and identify molecular targets that may synergize with new inhibitors of target of rapamycin (TOR, a conserved protein that is a major integrator of cell proliferation signals in the nutrient-signaling pathway. Although clinical results from TOR complex 1 (TORC1-specific inhibition using rapamycin analogs have been disappointing, trials using inhibitors that also target TORC2 have been promising. To understand this increased therapeutic efficacy and to discover secondary targets for combination therapy, we engineered Tor2 in S. cerevisiae to accept an orthogonal inhibitor. We used this tool to create a chemical epistasis miniarray profile (ChE-MAP by measuring interactions between the chemically inhibited Tor2 kinase and a diverse library of deletion mutants. The ChE-MAP identified known TOR components and distinguished between TORC1- and TORC2-dependent functions. The results showed a TORC2-specific interaction with the pentose phosphate pathway, a previously unappreciated TORC2 function that suggests a role for the complex in balancing the high energy demand required for ribosome biogenesis.

  17. Current issues involving screening and identification of chemical contaminants in foods by mass spectrometry

    NARCIS (Netherlands)

    Lehotay, S.J.; Sapozhnikova, Y.; Mol, J.G.J.

    2015-01-01

    Although quantitative analytical methods must be empirically validated prior to their use in a variety of applications, including regulatory monitoring of chemical adulterants in foods, validation of qualitative method performance for the analytes and matrices of interest is frequently ignored, or g

  18. Current issues involving screening and identification of chemical contaminants in foods by mass spectrometry

    Science.gov (United States)

    Although quantitative analytical methods must be empirically validated prior to their actual use in a variety of applications, including regulatory monitoring of chemical adulterants in foods, validation of qualitative method performance for the analytes and matrices of interest is frequently ignore...

  19. Developing, Applying, and Evaluating Models for Rapid Screening of Chemical Exposures

    DEFF Research Database (Denmark)

    Arnot, J.; Shin, H.; Ernstoff, Alexi;

    2015-01-01

    to limited exposure data there is limited information on chemical use patterns and production and emission quantities. These data gaps require the application of mass balance, statistical and quantitative structure-activity relationship (QSAR) models to predict exposure and exposure potential for humans...

  20. A loss-of-function genetic screening identifies novel mediators of thyroid cancer cell viability

    Science.gov (United States)

    Cantisani, Maria Carmela; Parascandolo, Alessia; Perälä, Merja; Allocca, Chiara; Fey, Vidal; Sahlberg, Niko; Merolla, Francesco; Basolo, Fulvio; Laukkanen, Mikko O.; Kallioniemi, Olli Pekka; Santoro, Massimo; Castellone, Maria Domenica

    2016-01-01

    RET, BRAF and other protein kinases have been identified as major molecular players in thyroid cancer. To identify novel kinases required for the viability of thyroid carcinoma cells, we performed a RNA interference screening in the RET/PTC1(CCDC6-RET)-positive papillary thyroid cancer cell line TPC1 using a library of synthetic small interfering RNAs (siRNAs) targeting the human kinome and related proteins. We identified 14 hits whose silencing was able to significantly reduce the viability and the proliferation of TPC1 cells; most of them were active also in BRAF-mutant BCPAP (papillary thyroid cancer) and 8505C (anaplastic thyroid cancer) and in RAS-mutant CAL62 (anaplastic thyroid cancer) cells. These included members of EPH receptor tyrosine kinase family as well as SRC and MAPK (mitogen activated protein kinases) families. Importantly, silencing of the identified hits did not affect significantly the viability of Nthy-ori 3-1 (hereafter referred to as NTHY) cells derived from normal thyroid tissue, suggesting cancer cell specificity. The identified proteins are worth exploring as potential novel druggable thyroid cancer targets. PMID:27058903

  1. Screening-level models to estimate partition ratios of organic chemicals between polymeric materials, air and water.

    Science.gov (United States)

    Reppas-Chrysovitsinos, Efstathios; Sobek, Anna; MacLeod, Matthew

    2016-06-15

    Polymeric materials flowing through the technosphere are repositories of organic chemicals throughout their life cycle. Equilibrium partition ratios of organic chemicals between these materials and air (KMA) or water (KMW) are required for models of fate and transport, high-throughput exposure assessment and passive sampling. KMA and KMW have been measured for a growing number of chemical/material combinations, but significant data gaps still exist. We assembled a database of 363 KMA and 910 KMW measurements for 446 individual compounds and nearly 40 individual polymers and biopolymers, collected from 29 studies. We used the EPI Suite and ABSOLV software packages to estimate physicochemical properties of the compounds and we employed an empirical correlation based on Trouton's rule to adjust the measured KMA and KMW values to a standard reference temperature of 298 K. Then, we used a thermodynamic triangle with Henry's law constant to calculate a complete set of 1273 KMA and KMW values. Using simple linear regression, we developed a suite of single parameter linear free energy relationship (spLFER) models to estimate KMA from the EPI Suite-estimated octanol-air partition ratio (KOA) and KMW from the EPI Suite-estimated octanol-water (KOW) partition ratio. Similarly, using multiple linear regression, we developed a set of polyparameter linear free energy relationship (ppLFER) models to estimate KMA and KMW from ABSOLV-estimated Abraham solvation parameters. We explored the two LFER approaches to investigate (1) their performance in estimating partition ratios, and (2) uncertainties associated with treating all different polymers as a single "bulk" polymeric material compartment. The models we have developed are suitable for screening assessments of the tendency for organic chemicals to be emitted from materials, and for use in multimedia models of the fate of organic chemicals in the indoor environment. In screening applications we recommend that KMA and KMW be

  2. Development of a stable cell line with an intact PGC-1α/ERRα axis for screening environmental chemicals

    Energy Technology Data Exchange (ETDEWEB)

    Teng, Christina T., E-mail: teng1@niehs.nih.gov [DNTP, BioMolecular Screening Branch, Division, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, NC 27709 (United States); Beames, Burton; Alex Merrick, B. [DNTP, BioMolecular Screening Branch, Division, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, NC 27709 (United States); Martin, Negin; Romeo, Charles [DIR, Viral Core Lab, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, NC 27709 (United States); Jetten, Anton M. [DIR Laboratory of Respiratory Biology, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, NC 27709 (United States)

    2014-02-07

    Highlights: • We developed a stable cell line with intact PGC-1α/ERRα axis. • The ERRα repressor, XCT790, down regulates this pathway. • Phytoestrogen, genisten stimulates this pathway. - Abstract: The estrogen-related receptor α (ERRα) and the peroxisome proliferator-activated receptor γ (PPARγ) coactivator 1α (PGC-1α) play critical roles in the control of several physiological functions, including the regulation of genes involved in energy homeostasis. However, little is known about the ability of environmental chemicals to disrupt or modulate this important bioenergetics pathway in humans. The goal of this study was to develop a cell-based assay system with an intact PGC-1α/ERRα axis that could be used as a screening assay for detecting such chemicals. To this end, we successfully generated several stable cell lines expressing PGC-1α and showed that the reporter driven by the native ERRα hormone response unit (AAB-Luc) is active in these cell lines and that the activation is PGC-1α-dependent. Furthermore, we show that this activation can be blocked by the ERRα selective inverse agonist, XCT790. In addition, we find that genistein and bisphenol A further stimulate the reporter activity, while kaempferol has minimal effect. These cell lines will be useful for identifying environmental chemicals that modulate this important pathway.

  3. Big data in chemical toxicity research: the use of high-throughput screening assays to identify potential toxicants.

    Science.gov (United States)

    Zhu, Hao; Zhang, Jun; Kim, Marlene T; Boison, Abena; Sedykh, Alexander; Moran, Kimberlee

    2014-10-20

    High-throughput screening (HTS) assays that measure the in vitro toxicity of environmental compounds have been widely applied as an alternative to in vivo animal tests of chemical toxicity. Current HTS studies provide the community with rich toxicology information that has the potential to be integrated into toxicity research. The available in vitro toxicity data is updated daily in structured formats (e.g., deposited into PubChem and other data-sharing web portals) or in an unstructured way (papers, laboratory reports, toxicity Web site updates, etc.). The information derived from the current toxicity data is so large and complex that it becomes difficult to process using available database management tools or traditional data processing applications. For this reason, it is necessary to develop a big data approach when conducting modern chemical toxicity research. In vitro data for a compound, obtained from meaningful bioassays, can be viewed as a response profile that gives detailed information about the compound's ability to affect relevant biological proteins/receptors. This information is critical for the evaluation of complex bioactivities (e.g., animal toxicities) and grows rapidly as big data in toxicology communities. This review focuses mainly on the existing structured in vitro data (e.g., PubChem data sets) as response profiles for compounds of environmental interest (e.g., potential human/animal toxicants). Potential modeling and mining tools to use the current big data pool in chemical toxicity research are also described. PMID:25195622

  4. Bioinspired silicification of silica-binding peptide-silk protein chimeras: comparison of chemically and genetically produced proteins.

    Science.gov (United States)

    Canabady-Rochelle, Laetitia L S; Belton, David J; Deschaume, Olivier; Currie, Heather A; Kaplan, David L; Perry, Carole C

    2012-03-12

    Novel protein chimeras constituted of "silk" and a silica-binding peptide (KSLSRHDHIHHH) were synthesized by genetic or chemical approaches and their influence on silica-silk based chimera composite formation evaluated. Genetic chimeras were constructed from 6 or 15 repeats of the 32 amino acid consensus sequence of Nephila clavipes spider silk ([SGRGGLGGQG AGAAAAAGGA GQGGYGGLGSQG](n)) to which one silica binding peptide was fused at the N terminus. For the chemical chimera, 28 equiv of the silica binding peptide were chemically coupled to natural Bombyx mori silk after modification of tyrosine groups by diazonium coupling and EDC/NHS activation of all acid groups. After silica formation under mild, biomaterial-compatible conditions, the effect of peptide addition on the properties of the silk and chimeric silk-silica composite materials was explored. The composite biomaterial properties could be related to the extent of silica condensation and to the higher number of silica binding sites in the chemical chimera as compared with the genetically derived variants. In all cases, the structure of the protein/chimera in solution dictated the type of composite structure that formed with the silica deposition process having little effect on the secondary structural composition of the silk-based materials. Similarly to our study of genetic silk based chimeras containing the R5 peptide (SSKKSGSYSGSKGSKRRIL), the role of the chimeras (genetic and chemical) used in the present study resided more in aggregation and scaffolding than in the catalysis of condensation. The variables of peptide identity, silk construct (number of consensus repeats or silk source), and approach to synthesis (genetic or chemical) can be used to "tune" the properties of the composite materials formed and is a general approach that can be used to prepare a range of materials for biomedical and sensor-based applications. PMID:22229696

  5. Protection against genetic hazards from environmental chemical mutagens: experience with ionizing radiation

    International Nuclear Information System (INIS)

    In radiation protection, the recurrent theme is, and always has been, dose limitation whether it is for occupational workers, individual members of the public or the population as a whole. The key words are 'dose' and 'limitation'. The quantitative system of dose limitation has been achieved because of a number of conceptual developments in our understanding of the mechanism of radiation action, development of radiation dosimetry, the accumulation of a vast body of quantitative information on dose-effect relationships and the effects of various biological and physical variables that affect these relationships of data on patterns and levels of exposures likely to be encountered to make estimates of the effects expected to result from such exposures, and balancing of risks to society against the benefits derived, the latter a matter of informed judgement. The philosophy has always been to avoid all unnecessary exposures and to limit the necessary exposures (justified by the benefits expected) to as low a level as reasonably achievable, social and economic factors being taken into acccount. The introduction of the concept that the system of dose limitation to the population should be based on genetic risks has stressed the need for careful planning to ensure that our genetic heritage is not endangered. Transfer of this knowledge to the field of chemical protection is discussed. (Auth.)

  6. A microarray-based genetic screen for yeast chronological aging factors.

    Directory of Open Access Journals (Sweden)

    Mirela Matecic

    2010-04-01

    Full Text Available Model organisms have played an important role in the elucidation of multiple genes and cellular processes that regulate aging. In this study we utilized the budding yeast, Saccharomyces cerevisiae, in a large-scale screen for genes that function in the regulation of chronological lifespan, which is defined by the number of days that non-dividing cells remain viable. A pooled collection of viable haploid gene deletion mutants, each tagged with unique identifying DNA "bar-code" sequences was chronologically aged in liquid culture. Viable mutants in the aging population were selected at several time points and then detected using a microarray DNA hybridization technique that quantifies abundance of the barcode tags. Multiple short- and long-lived mutants were identified using this approach. Among the confirmed short-lived mutants were those defective for autophagy, indicating a key requirement for the recycling of cellular organelles in longevity. Defects in autophagy also prevented lifespan extension induced by limitation of amino acids in the growth media. Among the confirmed long-lived mutants were those defective in the highly conserved de novo purine biosynthesis pathway (the ADE genes, which ultimately produces IMP and AMP. Blocking this pathway extended lifespan to the same degree as calorie (glucose restriction. A recently discovered cell-extrinsic mechanism of chronological aging involving acetic acid secretion and toxicity was suppressed in a long-lived ade4Delta mutant and exacerbated by a short-lived atg16Delta autophagy mutant. The identification of multiple novel effectors of yeast chronological lifespan will greatly aid in the elucidation of mechanisms that cells and organisms utilize in slowing down the aging process.

  7. Genetic barcode sequencing for screening altered population dynamics of hematopoietic stem cells transduced with lentivirus

    Science.gov (United States)

    Zanatta, Daniela B; Tsujita, Maristela; Borelli, Primavera; Aguiar, Rodrigo B; Ferrari, Daniel G; Strauss, Bryan E

    2014-01-01

    Insertional mutagenesis has been associated with malignant cell transformation in gene therapy protocols, leading to discussions about vector security. Therefore, clonal analysis is important for the assessment of vector safety and its impact on patient health. Here, we report a unique approach to assess dynamic changes in clonality of lentivirus transduced cells upon Sanger sequence analysis of a specially designed genetic barcode. In our approach, changes in the electropherogram peaks are measured and compared between successive time points, revealing alteration in the cell population. After in vitro validation, barcoded lentiviral libraries carrying IL2RG or LMO2 transgenes, or empty vector were used to transduce mouse hematopoietic (ckit+) stem cells, which were subsequently transplanted in recipient mice. We found that neither the empty nor IL2RG encoding vector had an effect on cell dynamics. In sharp contrast, the LMO2 oncogene was associated with altered cell dynamics even though hematologic counts remained unchanged, suggesting that the barcode could reveal changes in cell populations not observed by the frontline clinical assay. We describe a simple and sensitive method for the analysis of clonality, which could be easily used by any laboratory for the assessment of cellular behavior upon lentiviral transduction. PMID:26052520

  8. Genetic screening and functional characterization of PDGFRB mutations associated with Basal Ganglia Calcification of Unknown Etiology

    Science.gov (United States)

    Sanchez-Contreras, Monica; Baker, Matthew C.; Finch, NiCole A.; Nicholson, Alexandra; Wojtas, Aleksandra; Wszolek, Zbigniew K.; Ross, Owen A.; Dickson, Dennis W.; Rademakers, Rosa

    2014-01-01

    Three causal genes for Idiopathic Basal Ganglia Calcification (IBGC) have been identified. Most recently, mutations in PDGFRB, encoding a member of the platelet-derived growth factor receptor family type β, and PDGFB, encoding PDGF-B, the specific ligand of PDGFRβ, were found implicating the PDGF-B/PDGFRβ pathway in abnormal brain calcification. In this study we aimed to identify and study mutations in PDGFRB and PDGFB in a series of 26 patients from the Mayo Clinic Florida Brain Bank with moderate to severe basal ganglia calcification (BCG) of unknown etiology. No mutations in PDGFB were found. However, we identified one mutation in PDGFRB, p.R695C located in the tyrosine kinase domain, in one BGC patient. We further studied the function of p.R695C mutant PDGFRβ and two previously reported mutants, p.L658P and p.R987W PDGFRβ in cell culture. We show that, in response to PDGF-BB stimulation, the p.L658P mutation completely suppresses PDGFRβ autophosphorylation whereas the p.R695C mutation results in partial loss of autophosphorylation. For the p.R987W mutation, our data suggest a different mechanism involving reduced protein levels. These genetic and functional studies provide the first insight into the pathogenic mechanisms associated with PDGFRB mutations and provide further support for a pathogenic role of PDGFRB mutations in BGC. PMID:24796542

  9. A screen for genetic suppressor elements of hepatitis C virus identifies a supercharged protein inhibitor of viral replication.

    Science.gov (United States)

    Simeon, Rudo L; Chen, Zhilei

    2013-01-01

    Genetic suppressor elements (GSEs) are biomolecules derived from a gene or genome of interest that act as transdominant inhibitors of biological functions presumably by disruption of critical biological interfaces. We exploited a cell death reporter cell line for hepatitis C virus (HCV) infection, n4mBid, to develop an iterative selection/enrichment strategy for the identification of anti-HCV GSEs. Using this approach, a library of fragments of an HCV genome was screened for sequences that suppress HCV infection. A 244 amino acid gene fragment, B1, was strongly enriched after 5 rounds of selection. B1 derives from a single-base frameshift of the enhanced green fluorescent protein (eGFP) which was used as a filler during fragment cloning. B1 has a very high net positive charge of 43 at neutral pH and a high charge-to-mass (kDa) ratio of 1.5. We show that B1 expression specifically inhibits HCV replication. In addition, five highly positively charged B1 fragments produced from progressive truncation at the C-terminus all retain the ability to inhibit HCV, suggesting that a high positive charge, rather than a particular motif in B1, likely accounts for B1's anti-HCV activity. Another supercharged protein, +36GFP, was also found to strongly inhibit HCV replication when added to cells at the time of infection. This study reports a new methodology for HCV inhibitor screening and points to the anti-HCV potential of positively charged proteins/peptides.

  10. A screen for genetic suppressor elements of hepatitis C virus identifies a supercharged protein inhibitor of viral replication.

    Directory of Open Access Journals (Sweden)

    Rudo L Simeon

    Full Text Available Genetic suppressor elements (GSEs are biomolecules derived from a gene or genome of interest that act as transdominant inhibitors of biological functions presumably by disruption of critical biological interfaces. We exploited a cell death reporter cell line for hepatitis C virus (HCV infection, n4mBid, to develop an iterative selection/enrichment strategy for the identification of anti-HCV GSEs. Using this approach, a library of fragments of an HCV genome was screened for sequences that suppress HCV infection. A 244 amino acid gene fragment, B1, was strongly enriched after 5 rounds of selection. B1 derives from a single-base frameshift of the enhanced green fluorescent protein (eGFP which was used as a filler during fragment cloning. B1 has a very high net positive charge of 43 at neutral pH and a high charge-to-mass (kDa ratio of 1.5. We show that B1 expression specifically inhibits HCV replication. In addition, five highly positively charged B1 fragments produced from progressive truncation at the C-terminus all retain the ability to inhibit HCV, suggesting that a high positive charge, rather than a particular motif in B1, likely accounts for B1's anti-HCV activity. Another supercharged protein, +36GFP, was also found to strongly inhibit HCV replication when added to cells at the time of infection. This study reports a new methodology for HCV inhibitor screening and points to the anti-HCV potential of positively charged proteins/peptides.

  11. Quantitative phenotyping-based in vivo chemical screening in a zebrafish model of leukemia stem cell xenotransplantation.

    Directory of Open Access Journals (Sweden)

    Beibei Zhang

    Full Text Available Zebrafish-based chemical screening has recently emerged as a rapid and efficient method to identify important compounds that modulate specific biological processes and to test the therapeutic efficacy in disease models, including cancer. In leukemia, the ablation of leukemia stem cells (LSCs is necessary to permanently eradicate the leukemia cell population. However, because of the very small number of LSCs in leukemia cell populations, their use in xenotransplantation studies (in vivo and the difficulties in functionally and pathophysiologically replicating clinical conditions in cell culture experiments (in vitro, the progress of drug discovery for LSC inhibitors has been painfully slow. In this study, we developed a novel phenotype-based in vivo screening method using LSCs xenotransplanted into zebrafish. Aldehyde dehydrogenase-positive (ALDH+ cells were purified from chronic myelogenous leukemia K562 cells tagged with a fluorescent protein (Kusabira-orange and then implanted in young zebrafish at 48 hours post-fertilization. Twenty-four hours after transplantation, the animals were treated with one of eight different therapeutic agents (imatinib, dasatinib, parthenolide, TDZD-8, arsenic trioxide, niclosamide, salinomycin, and thioridazine. Cancer cell proliferation, and cell migration were determined by high-content imaging. Of the eight compounds that were tested, all except imatinib and dasatinib selectively inhibited ALDH+ cell proliferation in zebrafish. In addition, these anti-LSC agents suppressed tumor cell migration in LSC-xenotransplants. Our approach offers a simple, rapid, and reliable in vivo screening system that facilitates the phenotype-driven discovery of drugs effective in suppressing LSCs.

  12. A high-throughput strategy for screening of bacterial artificial chromosome libraries and anchoring of clones on a genetic map constructed with single nucleotide polymorphisms

    Directory of Open Access Journals (Sweden)

    Deal Karin R

    2009-01-01

    Full Text Available Abstract Background Current techniques of screening bacterial artificial chromosome (BAC libraries for molecular markers during the construction of physical maps are slow, laborious and often assign multiple BAC contigs to a single locus on a genetic map. These limitations are the principal impediment in the construction of physical maps of large eukaryotic genomes. It is hypothesized that this impediment can be overcome by screening multidimensional pools of BAC clones using the highly parallel Illumina GoldenGate™ assay. Results To test the efficacy of the Golden Gate assay in BAC library screening, multidimensional pools involving 302976 Aegilops tauschii BAC clones were genotyped for the presence/absence of specific gene sequences with multiplexed Illumina GoldenGate oligonucleotide assays previously used to place single nucleotide polymorphisms on an Ae. tauschii genetic map. Of 1384 allele-informative oligonucleotide assays, 87.6% successfully clustered BAC pools into those positive for a BAC clone harboring a specific gene locus and those negative for it. The location of the positive BAC clones within contigs assembled from 199190 fingerprinted Ae. tauschii BAC clones was used to evaluate the precision of anchoring of BAC clones and contigs on the Ae. tauschii genetic map. For 41 (95% assays, positive BAC clones were neighbors in single contigs. Those contigs could be unequivocally assigned to loci on the genetic map. For two (5% assays, positive clones were in two different contigs and the relationships of these contigs to loci on the Ae. tauschii genetic map were equivocal. Screening of BAC libraries with a simple five-dimensional BAC pooling strategy was evaluated and shown to allow direct detection of positive BAC clones without the need for manual deconvolution of BAC clone pools. Conclusion The highly parallel Illumina oligonucleotide assay is shown here to be an efficient tool for screening BAC libraries and a strategy for high

  13. Development and validation of duplex, triplex, and pentaplex real-time PCR screening assays for the detection of genetically modified organisms in food and feed.

    Science.gov (United States)

    Huber, Ingrid; Block, Annette; Sebah, Daniela; Debode, Frédéric; Morisset, Dany; Grohmann, Lutz; Berben, Gilbert; Stebih, Dejan; Milavec, Mojca; Zel, Jana; Busch, Ulrich

    2013-10-30

    Worldwide, qualitative methods based on PCR are most commonly used as screening tools for genetically modified material in food and feed. However, the increasing number and diversity of genetically modified organisms (GMO) require effective methods for simultaneously detecting several genetic elements marking the presence of transgenic events. Herein we describe the development and validation of a pentaplex, as well as complementary triplex and duplex real-time PCR assays, for the detection of the most common screening elements found in commercialized GMOs: P-35S, T-nos, ctp2-cp4-epsps, bar, and pat. The use of these screening assays allows the coverage of many GMO events globally approved for commercialization. Each multiplex real-time PCR assay shows high specificity and sensitivity with an absolute limit of detection below 20 copies for the targeted sequences. We demonstrate by intra- and interlaboratory tests that the assays are robust as well as cost- and time-effective for GMO screening if applied in routine GMO analysis.

  14. Studies of Resurgent Bed Bugs: Population Genetic Structure, Impact of Aggregation on Development and Molecular Screening for Bartonella

    Science.gov (United States)

    Saenz, Virna Lisa

    The recent resurgence of bed bugs (Cimex lectularius L.) has created an unprecedented demand for research on its biology. The main objectives of this dissertation research were to investigate several aspects of bed bug biology: infestation and dispersal dynamics at a large and small geographical scale using molecular markers, to determine the impact of aggregation on bed bug development and to screen bed bug populations for a re-emergent pathogen. First, we studied the infestation and dispersal dynamics of bed bugs at large geographical scale (e.g., across cities, states). Although bed bug infestations are on the rise, there is a poor understanding of their dispersal patterns and sources of infestation. We conducted a genetic study of 21 bed bug infestations from the eastern United States. We genotyped samples comprised of 8 - 10 individuals per infestation at nine polymorphic microsatellite loci. Despite high genetic diversity across all infestations, with 5 -- 17 alleles per locus (mean = 10.3), we found low genetic diversity (1 -- 4 alleles per locus) within all but one of the infestations. These results suggest that nearly all the studied infestations were started by a small propagule possibly consisting of a singly mated female and/or her progeny. All infestations were strongly genetically differentiated from each other (mean pairwise FST between populations = 0.68) and we did not find strong evidence of a geographic pattern of structuring. The high level of genetic diversity across infestations from the eastern United States together with the lack of geographically organized structure is consistent with multiple introductions into the United States from foreign sources. This work is described in Chapter 2 and was published in the Journal of Medical Entomology in 2012. Second, we investigated dispersal and infestation dynamics of bed bugs at a fine geographical scale within three multistory apartment buildings: one from Raleigh, NC and two from Jersey City, NJ

  15. DART-MS as a preliminary screening method for "herbal incense": chemical analysis of synthetic cannabinoids.

    Science.gov (United States)

    Lesiak, Ashton D; Musah, Rabi A; Domin, Marek A; Shepard, Jason R E

    2014-03-01

    Direct analysis in real time mass spectrometry (DART-MS) served as a method for rapid high-throughput screening of six commercially available "Spice" products, detecting various combinations of five synthetic cannabinoids. Direct analysis in real time is an ambient ionization process that, along with high mass accuracy time-of-flight (TOF)-MS to 0.0001 Da, was employed to establish the presence of cannabinoids. Mass spectra were acquired by simply suspending a small portion of sample between the ion source and the mass spectrometer inlet. The ability to test minute amounts of sample is a major advantage when very limited amounts of evidentiary material are available. In addition, reports are widespread regarding the testing backlogs that now exist because of the large influx of designer drugs. This method circumvents time-consuming sample extraction, derivatization, chromatographic, and other sample preparative steps required for analysis by more conventional mass spectrometric methods. Accordingly, the synthetic cannabinoids AM-2201, JWH-122, JWH-203, JWH-210, and RCS-4 were identified in commercially available herbal Spice products, singly and in tandem, at concentrations within the range of 4-141 mg/g of material. Direct analysis in real time mass spectrometry decreases the time necessary to triage analytical evidence, and therefore, it has the potential to contribute to backlog reduction and more timely criminal prosecution.

  16. A chemical screen probing the relationship between mitochondrial content and cell size.

    Directory of Open Access Journals (Sweden)

    Toshimori Kitami

    Full Text Available The cellular content of mitochondria changes dynamically during development and in response to external stimuli, but the underlying mechanisms remain obscure. To systematically identify molecular probes and pathways that control mitochondrial abundance, we developed a high-throughput imaging assay that tracks both the per cell mitochondrial content and the cell size in confluent human umbilical vein endothelial cells. We screened 28,786 small molecules and observed that hundreds of small molecules are capable of increasing or decreasing the cellular content of mitochondria in a manner proportionate to cell size, revealing stereotyped control of these parameters. However, only a handful of compounds dissociate this relationship. We focus on one such compound, BRD6897, and demonstrate through secondary assays that it increases the cellular content of mitochondria as evidenced by fluorescence microscopy, mitochondrial protein content, and respiration, even after rigorous correction for cell size, cell volume, or total protein content. BRD6897 increases uncoupled respiration 1.6-fold in two different, non-dividing cell types. Based on electron microscopy, BRD6897 does not alter the percent of cytoplasmic area occupied by mitochondria, but instead, induces a striking increase in the electron density of existing mitochondria. The mechanism is independent of known transcriptional programs and is likely to be related to a blockade in the turnover of mitochondrial proteins. At present the molecular target of BRD6897 remains to be elucidated, but if identified, could reveal an important additional mechanism that governs mitochondrial biogenesis and turnover.

  17. Screening of phyto-chemical constituents, trace metals and antimicrobial efficiency of Cissus vitiginea

    Directory of Open Access Journals (Sweden)

    V. Subramani

    2014-06-01

    Full Text Available The present study focused on the phytochemical constituents, antimicrobial activity and trace metal concentrations of the Cissus vitiginea plant leaves which were collected from the Tiruchirappalli district, southern India. Preliminary phytochemical screening of leaves extracts revealed the presence of the bioactive compounds, such as steroids, triterpenoids, glycosides, sugar, alkaloids, flavonoids, tannins, amino acid, and coumarin in the leaves. The bacterial and fungal strains were tested for antimicrobial sensitivity against C. vitiginea using the disc diffusion method. The methanol extracts of the plant leaves exhibited the higher zone of inhibition against bacterial strains than fungal strains. The trace metal concentrations were analyzed form the powered plant leaves by 797 VA Computrace voltametry, Metrohm. The average concentrations of Cd, Cr, Cu, Fe, Ni, Pb and Zn were 0.05, BDL, 018, 0.38, BDL, BDL and 0.48 mg kg-1, respectively. The bioactive compounds responsible for these antimicrobial activities could be isolated and identified to develop a new drug of pharmaceutical interest.

  18. Identification of siRNA delivery enhancers by a chemical library screen

    Science.gov (United States)

    Gilleron, Jerome; Paramasivam, Prasath; Zeigerer, Anja; Querbes, William; Marsico, Giovanni; Andree, Cordula; Seifert, Sarah; Amaya, Pablo; Stöter, Martin; Koteliansky, Victor; Waldmann, Herbert; Fitzgerald, Kevin; Kalaidzidis, Yannis; Akinc, Akin; Maier, Martin A.; Manoharan, Muthiah; Bickle, Marc; Zerial, Marino

    2015-01-01

    Most delivery systems for small interfering RNA therapeutics depend on endocytosis and release from endo-lysosomal compartments. One approach to improve delivery is to identify small molecules enhancing these steps. It is unclear to what extent such enhancers can be universally applied to different delivery systems and cell types. Here, we performed a compound library screen on two well-established siRNA delivery systems, lipid nanoparticles and cholesterol conjugated-siRNAs. We identified fifty-one enhancers improving gene silencing 2–5 fold. Strikingly, most enhancers displayed specificity for one delivery system only. By a combination of quantitative fluorescence and electron microscopy we found that the enhancers substantially differed in their mechanism of action, increasing either endocytic uptake or release of siRNAs from endosomes. Furthermore, they acted either on the delivery system itself or the cell, by modulating the endocytic system via distinct mechanisms. Interestingly, several compounds displayed activity on different cell types. As proof of principle, we showed that one compound enhanced siRNA delivery in primary endothelial cells in vitro and in the endocardium in the mouse heart. This study suggests that a pharmacological approach can improve the delivery of siRNAs in a system-specific fashion, by exploiting distinct mechanisms and acting upon multiple cell types. PMID:26220182

  19. Integrating mechanistic and polymorphism data to characterize human genetic susceptibility for environmental chemical risk assessment in the 21st century

    Energy Technology Data Exchange (ETDEWEB)

    Mortensen, Holly M., E-mail: mortensen.holly@epa.gov [Office of Research and Development, US Environmental Protection Agency, National Center for Computational Toxicology, US EPA, 109 TW Alexander Dr., Mailcode B205-01, Research Triangle Park, NC 27711 (United States); Euling, Susan Y. [Office of Research and Development, US Environmental Protection Agency, National Center for Environmental Assessment, US EPA, 1200 Pennsylvania Ave., NW, Mail Code 8623P, Washington, DC 20460 (United States)

    2013-09-15

    Response to environmental chemicals can vary widely among individuals and between population groups. In human health risk assessment, data on susceptibility can be utilized by deriving risk levels based on a study of a susceptible population and/or an uncertainty factor may be applied to account for the lack of information about susceptibility. Defining genetic susceptibility in response to environmental chemicals across human populations is an area of interest in the NAS' new paradigm of toxicity pathway-based risk assessment. Data from high-throughput/high content (HT/HC), including -omics (e.g., genomics, transcriptomics, proteomics, metabolomics) technologies, have been integral to the identification and characterization of drug target and disease loci, and have been successfully utilized to inform the mechanism of action for numerous environmental chemicals. Large-scale population genotyping studies may help to characterize levels of variability across human populations at identified target loci implicated in response to environmental chemicals. By combining mechanistic data for a given environmental chemical with next generation sequencing data that provides human population variation information, one can begin to characterize differential susceptibility due to genetic variability to environmental chemicals within and across genetically heterogeneous human populations. The integration of such data sources will be informative to human health risk assessment.

  20. Integrating mechanistic and polymorphism data to characterize human genetic susceptibility for environmental chemical risk assessment in the 21st century

    International Nuclear Information System (INIS)

    Response to environmental chemicals can vary widely among individuals and between population groups. In human health risk assessment, data on susceptibility can be utilized by deriving risk levels based on a study of a susceptible population and/or an uncertainty factor may be applied to account for the lack of information about susceptibility. Defining genetic susceptibility in response to environmental chemicals across human populations is an area of interest in the NAS' new paradigm of toxicity pathway-based risk assessment. Data from high-throughput/high content (HT/HC), including -omics (e.g., genomics, transcriptomics, proteomics, metabolomics) technologies, have been integral to the identification and characterization of drug target and disease loci, and have been successfully utilized to inform the mechanism of action for numerous environmental chemicals. Large-scale population genotyping studies may help to characterize levels of variability across human populations at identified target loci implicated in response to environmental chemicals. By combining mechanistic data for a given environmental chemical with next generation sequencing data that provides human population variation information, one can begin to characterize differential susceptibility due to genetic variability to environmental chemicals within and across genetically heterogeneous human populations. The integration of such data sources will be informative to human health risk assessment

  1. Screening of adulterants in powdered foods and ingredients using line-scan Raman chemical imaging

    Science.gov (United States)

    Qin, Jianwei; Chao, Kuanglin; Kim, Moon S.

    2015-05-01

    A newly developed line-scan Raman imaging system using a 785 nm line laser was used to authenticate powdered foods and ingredients. The system was used to collect hyperspectral Raman images in a wavenumber range of 102-2865 cm-1 from three representative food powders mixed with selected adulterants with a concentration of 0.5%, including milk and melamine, flour and benzoyl peroxide, and starch and maleic anhydride. An acoustic mixer was used to create food adulterant mixtures. All the mixed samples were placed in sample holders with a surface area of 50 mm×50 mm. Spectral and image processing algorithms were developed based on single-band images at unique Raman peaks of the individual adulterants. Chemical images were created to show identification, spatial distribution, and morphological features of the adulterant particles mixed in the food powders. The potential of estimating mass concentrations of the adulterants using the percentages of the adulterant pixels in the chemical images was also demonstrated.

  2. Genetic risk factors for cerebrovascular disease in children with sickle cell disease: design of a case-control association study and genomewide screen

    Directory of Open Access Journals (Sweden)

    Brambilla Donald

    2003-07-01

    Full Text Available Abstract Background The phenotypic heterogeneity of sickle cell disease is likely the result of multiple genetic factors and their interaction with the sickle mutation. High transcranial doppler (TCD velocities define a subgroup of children with sickle cell disease who are at increased risk for developing ischemic stroke. The genetic factors leading to the development of a high TCD velocity (i.e. cerebrovascular disease and ultimately to stroke are not well characterized. Methods We have designed a case-control association study to elucidate the role of genetic polymorphisms as risk factors for cerebrovascular disease as measured by a high TCD velocity in children with sickle cell disease. The study will consist of two parts: a candidate gene study and a genomewide screen and will be performed in 230 cases and 400 controls. Cases will include 130 patients (TCD ≥ 200 cm/s randomized in the Stroke Prevention Trial in Sickle Cell Anemia (STOP study as well as 100 other patients found to have high TCD in STOP II screening. Four hundred sickle cell disease patients with a normal TCD velocity (TCD Discussion It is expected that this study will yield important information on genetic risk factors for the cerebrovascular disease phenotype in sickle cell disease by clarifying the role of candidate genes in the development of high TCD. The genomewide screen for a large number of SNPs may uncover the association of novel polymorphisms with cerebrovascular disease and stroke in sickle cell disease.

  3. A genetic screen reveals Arabidopsis stomatal and/or apoplastic defenses against Pseudomonas syringae pv. tomato DC3000.

    Directory of Open Access Journals (Sweden)

    Weiqing Zeng

    2011-10-01

    Full Text Available Bacterial infection of plants often begins with colonization of the plant surface, followed by entry into the plant through wounds and natural openings (such as stomata, multiplication in the intercellular space (apoplast of the infected tissues, and dissemination of bacteria to other plants. Historically, most studies assess bacterial infection based on final outcomes of disease and/or pathogen growth using whole infected tissues; few studies have genetically distinguished the contribution of different host cell types in response to an infection. The phytotoxin coronatine (COR is produced by several pathovars of Pseudomonas syringae. COR-deficient mutants of P. s. tomato (Pst DC3000 are severely compromised in virulence, especially when inoculated onto the plant surface. We report here a genetic screen to identify Arabidopsis mutants that could rescue the virulence of COR-deficient mutant bacteria. Among the susceptible to coronatine-deficient Pst DC3000 (scord mutants were two that were defective in stomatal closure response, two that were defective in apoplast defense, and four that were defective in both stomatal and apoplast defense. Isolation of these three classes of mutants suggests that stomatal and apoplastic defenses are integrated in plants, but are genetically separable, and that COR is important for Pst DC3000 to overcome both stomatal guard cell- and apoplastic mesophyll cell-based defenses. Of the six mutants defective in bacterium-triggered stomatal closure, three are defective in salicylic acid (SA-induced stomatal closure, but exhibit normal stomatal closure in response to abscisic acid (ABA, and scord7 is compromised in both SA- and ABA-induced stomatal closure. We have cloned SCORD3, which is required for salicylic acid (SA biosynthesis, and SCORD5, which encodes an ATP-binding cassette (ABC protein, AtGCN20/AtABCF3, predicted to be involved in stress-associated protein translation control. Identification of SCORD5 begins to

  4. A PCR-based forward genetics screening, using expression domain-specific markers, identifies mutants in endosperm transfer cell development

    Directory of Open Access Journals (Sweden)

    Luis Miguel Muñiz

    2014-04-01

    Full Text Available Mutant collections are an invaluable source of material on which forward genetic approaches allow the identification of genes affecting a wide variety of biological processes. However, some particular developmental stages and morphological structures may resist analysis due to their physical inaccessibility or to deleterious effects associated to their modification. Furthermore, lethal mutations acting early in development may escape detection. We have approached the characterisation of 101 maize seed mutants, selected from a collection of 27500 visually screened Mu-insertion lines, using a molecular marker approach based on a set of genes previously ascribed to different tissue compartments within the early developing kernel. A streamlined combination of qRT-PCR assays has allowed us to preliminary pinpoint the affected compartment, establish developmental comparisons to WT siblings and select mutant lines with alterations in the different compartments. Furthermore, clusters of markers co-affected by the underlying mutation were identified. We have analysed more extensively a set of lines presenting significant variation in transfer cell-associated expression markers, and have performed morphological observations, and immunolocalization experiments to confirm the results, validating this approach as an efficient mutant description tool.

  5. Possibilities of chemical weed control in Lupinus albus and Lupinus luteus-screening of herbicides.

    Science.gov (United States)

    Dewitte, K; Latré, J; Haesaert, G

    2006-01-01

    Weed control in sweet lupins is still a problem. Especially the phytotoxicity of herbicides in sweet lupins is not enough studied. Therefore a screening with 16 selected herbicides and 4 lupin varieties has been set up. During the growing season 2005, 10 of the tested herbicides were applied in pre-emergence, 6 in post-emergence. Pre-emergence: Most of the active matters tested in pre-emergence were not phytotoxic for lupins. Pendimethalin (1000 g/ha), linuron (500 g/ha), chlorotoluron (1500 g/ha), prosulfocarb (2400 g/ha), clomazone (72 g/ha), isoxaben (100 g/ha), metamitron (1050 g/ha) and dimethenamid-P (720 g/ha) were applied without causing any significant phytotoxic symptoms. Only the lupins treated with aclonifen (1200 g/ha) showed a significant growth inhibition, 3 weeks after treatment. Significantly more chlorosis was noticed when the lupins were treated with aclonifen or with diflufenican, in preemergence. Post-emergence: In post-emergence, diflufenican (50 g/ha) did not cause any crop damage. Florasulam (5 g/ha) caused almost 100% necrosis in L. albus as well as in L. luteus. Bentazon (652 g/ha), thifensulfuron-methyl (15 g/ha) and metribuzin (175 g/ha) caused obvious necrosis and growth inhibition of the crop. The growth inhibition was significantly more severe for lupins treated with bentazon than if they were treated with thifensulfuron-methyl or metribuzin. Three weeks after treatment, clomazone (90 g/ha) and diflufenican (50 g/ha), did not cause any crop injury at all. The results indicated an interesting range of active matters which can be applied in pre-emergence, but weed control in post-emergence stays difficult.

  6. Preliminary Phytochemical Screening and Physico-Chemical Parameters of Artemisia absinthium and Artemisia annua.

    Directory of Open Access Journals (Sweden)

    Praveen Kumar Ashok

    2013-03-01

    Full Text Available The family Asteraceae or compositae known as the ester, daisy or sunflower family is the largest family of flowering plants. Artemisia is a large diverse genus of plants with between 100 to 150 species belonging to the family asteraceae (compositae. It comprises hardy herbs and shrubs known for their volatile oils. They grow in temperate climate of the northern hemisphere and southern hemisphere usually in dry or, semidry habitats. The collected herbs were authenticated, dried and extracted to calculate the percentage of yield. Phytochemical studies of the Hexane and alcoholic extracts showed the presence of various phytoconstituents i.e. carbohydrate, saponins, phytosterol, proteins and amino acid, tannin, phenolic compounds and flavonoids. It was observed that all the extracts show more important chemical constituents for various pharmacological activities. The determination of these characters will aid future investigators in their Pharmacological analysis of this species.

  7. At the brink of supercoloniality: genetic, behavioral and chemical assessments of population structure of the desert ant Cataglyphis niger

    Directory of Open Access Journals (Sweden)

    Maya eSaar

    2014-05-01

    Full Text Available The nesting habits of ants play an important role in structuring ant populations. They vary from monodomy, a colony occupies a single nest, via polydomy, a colony occupies multiple adjacent nests, to supercoloniality, a colony spans over large territories comprising dozen to thousands nests without having any boundaries. The population structure of the desert ant Cataglyphis niger, previously considered to form supercolonies, was studied using genetic, chemical and behavioral tools in plots of 50x50 meters at two distinct populations. At the Palmahim site, the plot comprised 15 nests that according to the genetic analysis constituted three colonies. Likewise at the Rishon Leziyyon site 14 nests constituted 5 genetic colonies. In both sites, both chemical analysis and the behavioral (aggression tests confirmed the colony genetic architecture. The behavioral tests also revealed that aggression between colonies within a population was higher than that exhibited between colonies of different populations, suggesting the occurrence of the nasty neighbor phenomenon. In contrast to supercolony structure previously reported in another population of this species, the presently studied populations were composed of polydomous colonies. However, both the genetic and chemical data revealed that the inter-colonial differences between sites were larger than those within site, suggesting some within-site population viscosity. Thus, C. niger exhibits flexible nesting characteristics, from polydomy to supercoloniality, and can be considered at the brink of supercoloniality. We attribute the differences in population structure among sites to the intensity of intraspecific competition.

  8. Characterization of the onset asphaltenes by focused-beam laser reflectance : a tool for chemical additives screening

    Energy Technology Data Exchange (ETDEWEB)

    Marugan, J.; Calles, J.A.; Dufour, J.; Gimenez-Aguirre, R. [Univ. Rey Juan Carlos, Madrid (Spain). URJC-Repsol-YPF Flow Assurance Laboratory, Dept. of Chemical and Environmental Technology; Pena, J.L. [Univ. Rey Juan Carlos, Madrid (Spain). URJC-Repsol-YPF Flow Assurance Laboratory, Dept. of Chemical and Environmental Technology; Centro Tecnologico Repsol-YPF, Madrid (Spain); Merino-Garcia, D. [Centro Tecnologico Repsol-YPF, Madrid (Spain)

    2008-07-01

    The deposition of asphaltenes in crude oil can cause flow assurance problems. In this study, a laser reflectance technique known as Focused-Beam Reflectance Measurement (FBRM) was used to study the kinetics of asphaltenes aggregation near onset. The FBRM tool provides a very sensitive way of determining the onset n-alkane/oil mass ratio. The influence of the n-alkane solvent and temperature on the solvent/oil threshold ratio of 2 South American crude oils with 21 and 27 API were investigated. The FBRM technique provided kinetic information about the evolution with time of the size distribution of asphaltenes flocs. Additional FBRM experiments of asphaltene redissolution and reprecipitation were also performed for a comparative evaluation, beginning with the solids recovered following the IP- 143 standard, which were fractionated into 4 different polarity groups using n-pentane - chloroform mixtures. The objective was to find correlations between polarity of the asphaltenes and its instability near the onset. Metal content was determined through atomic emission spectroscopy. The solids were characterized by 1H NMR, FT-IR spectroscopy, and vapour-pressure osmometry in order to determine the chemical and structural features of the most unstable asphaltenes. The FBRM probe was used to screen commercial chemical additives to prevent asphaltenes deposits. This technique was shown to be a very powerful tool for examining the influence of additives on the aggregation kinetics and the particle size distribution of the first asphaltene solids.

  9. Germ cell toxicity: significance in genetic and fertility effects of radiation and chemicals

    International Nuclear Information System (INIS)

    The primordial germ cells originate in the region of the caudal end of the primitive streak, root of the allantois, and yolk sac splanchnopleure, and migrate to the gonadal ridges where they divide to form the oogonia of the female and gonocytes of the male. In the female, the transition to oocytes occurs in utero, and the female mammal is born with a finite number of oocytes that cannot be replaced. By contrast, the gonocytes of the male initiate divisions soon after birth to form the spermatogonial stem cells, which persist throughout reproductive life of the male and are capable of regenerating the seminiferous epithelium after injury. As a result of these basic differences in gametogenesis, the response of the male and female to radiation and chemicals is different. The response of both the male and female changes with development of the embryonic to the adult gonad, and with differentiation and maturation in the adult. The primordial germ cells, early oocytes, and differentiating spermatogonia of the adult male are unusually sensitive to the cytotoxic action of noxious agents, but each agent elicits a specific response owing to the intricate biochemical and physiological changes associated with development and maturation of the gametes. The relationship of germ cell killing to fertility is direct, and long-term fertility effects can be predicted from histological analysis of the gonads. The relationship to genetic effects, on the other hand, is indirect, and acts primarily by limiting the cell stages available for testing, by affecting the distribution of mitotically active stem cells among the different stages of the mitotic cycle, and thereby changing both the type and frequency of genetic effects observed

  10. Potential of gas chromatography-atmospheric pressure chemical ionization-tandem mass spectrometry for screening and quantification of hexabromocyclododecane.

    Science.gov (United States)

    Sales, Carlos; Portolés, Tania; Sancho, Juan Vicente; Abad, Esteban; Ábalos, Manuela; Sauló, Jordi; Fiedler, Heidelore; Gómara, Belén; Beltrán, Joaquim

    2016-01-01

    A fast method for the screening and quantification of hexabromocyclododecane (sum of all isomers) by gas chromatography using a triple quadrupole mass spectrometer with atmospheric pressure chemical ionization (GC-APCI-QqQ) is proposed. This novel procedure makes use of the soft atmospheric pressure chemical ionization source, which results in less fragmentation of the analyte than by conventional electron impact (EI) and chemical ionization (CI) sources, favoring the formation of the [M - Br](+) ion and, thus, enhancing sensitivity and selectivity. Detection was based on the consecutive loses of HBr from the [M - Br](+) ion to form the specific [M - H5Br6](+) and [M - H4Br5](+) ions, which were selected as quantitation (Q) and qualification (q) transitions, respectively. Parameters affecting ionization and MS/MS detection were studied. Method performance was also evaluated; calibration curves were found linear from 1 pg/μL to 100 pg/μL for the total HBCD concentration; instrumental detection limit was estimated to be 0.10 pg/μL; repeatability and reproducibility, expressed as relative standard deviation, were better than 7% in both cases. The application to different real samples [polyurethane foam disks (PUFs), food, and marine samples] pointed out a rapid way to identify and allow quantification of this compound together with a number of polybrominated diphenyl ethers (BDE congeners 28, 47, 66, 85, 99, 100, 153, 154, 183, 184, 191, 196, 197, and 209) and two other novel brominated flame retardants [i.e., decabromodiphenyl ethane (DBDPE) and 1,2-bis(2,4,6-tribromophenoxy)ethane (BTBPE)] because of their presence in the same fraction when performing the usual sample treatment. PMID:26554601

  11. Potential of gas chromatography-atmospheric pressure chemical ionization-tandem mass spectrometry for screening and quantification of hexabromocyclododecane.

    Science.gov (United States)

    Sales, Carlos; Portolés, Tania; Sancho, Juan Vicente; Abad, Esteban; Ábalos, Manuela; Sauló, Jordi; Fiedler, Heidelore; Gómara, Belén; Beltrán, Joaquim

    2016-01-01

    A fast method for the screening and quantification of hexabromocyclododecane (sum of all isomers) by gas chromatography using a triple quadrupole mass spectrometer with atmospheric pressure chemical ionization (GC-APCI-QqQ) is proposed. This novel procedure makes use of the soft atmospheric pressure chemical ionization source, which results in less fragmentation of the analyte than by conventional electron impact (EI) and chemical ionization (CI) sources, favoring the formation of the [M - Br](+) ion and, thus, enhancing sensitivity and selectivity. Detection was based on the consecutive loses of HBr from the [M - Br](+) ion to form the specific [M - H5Br6](+) and [M - H4Br5](+) ions, which were selected as quantitation (Q) and qualification (q) transitions, respectively. Parameters affecting ionization and MS/MS detection were studied. Method performance was also evaluated; calibration curves were found linear from 1 pg/μL to 100 pg/μL for the total HBCD concentration; instrumental detection limit was estimated to be 0.10 pg/μL; repeatability and reproducibility, expressed as relative standard deviation, were better than 7% in both cases. The application to different real samples [polyurethane foam disks (PUFs), food, and marine samples] pointed out a rapid way to identify and allow quantification of this compound together with a number of polybrominated diphenyl ethers (BDE congeners 28, 47, 66, 85, 99, 100, 153, 154, 183, 184, 191, 196, 197, and 209) and two other novel brominated flame retardants [i.e., decabromodiphenyl ethane (DBDPE) and 1,2-bis(2,4,6-tribromophenoxy)ethane (BTBPE)] because of their presence in the same fraction when performing the usual sample treatment.

  12. Screening of chemical composition, antimicrobial and antioxidant activities of Artemisia essential oils.

    Science.gov (United States)

    Lopes-Lutz, Daíse; Alviano, Daniela S; Alviano, Celuta S; Kolodziejczyk, Paul P

    2008-05-01

    The chemical composition of essential oils isolated from aerial parts of seven wild sages from Western Canada -Artemisia absinthium L., Artemisia biennis Willd., Artemisia cana Pursh, Artemisia dracunculus L., Artemisia frigida Willd., Artemisia longifolia Nutt. and Artemisia ludoviciana Nutt., was investigated by GC-MS. A total of 110 components were identified accounting for 71.0-98.8% of the oil composition. High contents of 1,8-cineole (21.5-27.6%) and camphor (15.9-37.3%) were found in Artemisia cana, A. frigida, A. longifolia and A. ludoviciana oils. The oil of A. ludoviciana was also characterized by a high content of oxygenated sesquiterpenes with a 5-ethenyltetrahydro-5-methyl-2-furanyl moiety, of which davanone (11.5%) was the main component identified. A. absinthium oil was characterized by high amounts of myrcene (10.8%), trans-thujone (10.1%) and trans-sabinyl acetate (26.4%). A. biennis yielded an oil rich in (Z)-beta-ocimene (34.7%), (E)-beta-farnesene (40.0%) and the acetylenes (11.0%) (Z)- and (E)-en-yn-dicycloethers. A. dracunculus oil contained predominantly phenylpropanoids such as methyl chavicol (16.2%) and methyl eugenol (35.8%). Artemisia oils had inhibitory effects on the growth of bacteria (Escherichia coli, Staphylococcus aureus, and Staphylococcus epidermidis), yeasts (Candida albicans, Cryptococcus neoformans), dermatophytes (Trichophyton rubrum, Microsporum canis, and Microsporum gypseum), Fonsecaea pedrosoi and Aspergillus niger. A. biennis oil was the most active against dermatophytes, Cryptococcus neoformans, Fonsecaea pedrosoi and Aspergillus niger, and A. absinthium oil the most active against Staphylococcus strains. In addition, antioxidant (beta-carotene/linoleate model) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activities were determined, and weak activities were found for these oils. PMID:18417176

  13. Impact of Gene Patents and Licensing Practices on Access to Genetic Testing and Carrier Screening for Tay-Sachs and Canavan Disease

    OpenAIRE

    Colaianni, Alessandra; Chandrasekharan, Subhashini; Cook-Deegan, Robert

    2010-01-01

    Genetic testing for Tay-Sachs and Canavan disease is particularly important for Ashkenazi Jews, as both conditions are more frequent in that population. This comparative case study was possible because of different patenting and licensing practices. The role of DNA testing differs between Tay-Sachs and Canavan diseases. The first-line screening test for Tay-Sachs remains an enzyme activity test, rather than genotyping. Genotyping is used for preimplantation diagnosis and confirmatory testing....

  14. Validation of a qualitative screening method for pesticides in fruits and vegetables by gas chromatography quadrupole-time of flight mass spectrometry with atmospheric pressure chemical ionization

    NARCIS (Netherlands)

    Portoles, T.; Mol, J.G.J.; Sancho, J.V.; Lopez, F.J.; Hernandez, F.

    2014-01-01

    A wide-scope screening method was developed for the detection of pesticides in fruit and vegetables. The method was based on gas chromatography coupled to a hybrid quadrupole time-of-flight mass spectrometer with an atmospheric pressure chemical ionization source (GC-(APCI)QTOF MS). A non-target acq

  15. Development of a bioassay to screen for chemicals mimicking the anti-aging effects of calorie restriction

    International Nuclear Information System (INIS)

    Research highlights: → We identified four sequence motifs lying upstream of putative pro-longevity genes. → One of these motifs binds to HNF-4α. → HNF-4α/PGC-1α could up-regulate the transcription of a reporter gene linked to this motif. → The reporter system described here could be used to screen candidate anti-aging molecules. -- Abstract: Suppression of the growth hormone/insulin-like growth factor-I pathway in Ames dwarf (DF) mice, and caloric restriction (CR) in normal mice extends lifespan and delays the onset of age-related disorders. In combination, these interventions have an additive effect on lifespan in Ames DF mice. Therefore, common signaling pathways regulated by DF and CR could have additive effects on longevity. In this study, we tried to identity the signaling mechanism and develop a system to assess pro-longevity status in cells and mice. We previously identified genes up-regulated in the liver of DF and CR mice by DNA microarray analysis. Motif analysis of the upstream sequences of those genes revealed four major consensus sequence motifs, which have been named dwarfism and calorie restriction-responsive elements (DFCR-REs). One of the synthesized sequences bound to hepatocyte nuclear factor-4α (HNF-4α), an important transcription factor involved in liver metabolism. Furthermore, using this sequence information, we developed a highly sensitive bioassay to identify chemicals mimicking the anti-aging effects of CR. When the reporter construct, containing an element upstream of a secreted alkaline phosphatase (SEAP) gene, was co-transfected with HNF-4α and its regulator peroxisome proliferator-activated receptor (PPAR) γ coactivator-1α (PGC-1α), SEAP activity was increased compared with untransfected controls. Moreover, transient transgenic mice established using this construct showed increased SEAP activity in CR mice compared with ad libitum-fed mice. These data suggest that because of its rapidity, ease of use, and specificity

  16. Development of a bioassay to screen for chemicals mimicking the anti-aging effects of calorie restriction

    Energy Technology Data Exchange (ETDEWEB)

    Chiba, Takuya, E-mail: takuya@nagasaki-u.ac.jp [Department of Investigative Pathology, Graduate School of Biomedical Sciences, Nagasaki University, 1-12-4 Sakamoto, Nagasaki 852-8523 (Japan); Tsuchiya, Tomoshi [Division of Surgical Oncology, Graduate School of Biomedical Sciences, Nagasaki University, 1-7-1 Sakamoto, Nagasaki 852-8501 (Japan); Komatsu, Toshimitsu; Mori, Ryoichi; Hayashi, Hiroko [Department of Investigative Pathology, Graduate School of Biomedical Sciences, Nagasaki University, 1-12-4 Sakamoto, Nagasaki 852-8523 (Japan); Shimano, Hitoshi [Department of Internal Medicine (Endocrinology and Metabolism), Graduate School of Comprehensive Human Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba Ibaraki 305-8575 (Japan); Spindler, Stephen R. [Department of Biochemistry, Room 5478, Boyce Hall, University of California - Riverside, Riverside, CA 92521 (United States); Shimokawa, Isao [Department of Investigative Pathology, Graduate School of Biomedical Sciences, Nagasaki University, 1-12-4 Sakamoto, Nagasaki 852-8523 (Japan)

    2010-10-15

    Research highlights: {yields} We identified four sequence motifs lying upstream of putative pro-longevity genes. {yields} One of these motifs binds to HNF-4{alpha}. {yields} HNF-4{alpha}/PGC-1{alpha} could up-regulate the transcription of a reporter gene linked to this motif. {yields} The reporter system described here could be used to screen candidate anti-aging molecules. -- Abstract: Suppression of the growth hormone/insulin-like growth factor-I pathway in Ames dwarf (DF) mice, and caloric restriction (CR) in normal mice extends lifespan and delays the onset of age-related disorders. In combination, these interventions have an additive effect on lifespan in Ames DF mice. Therefore, common signaling pathways regulated by DF and CR could have additive effects on longevity. In this study, we tried to identity the signaling mechanism and develop a system to assess pro-longevity status in cells and mice. We previously identified genes up-regulated in the liver of DF and CR mice by DNA microarray analysis. Motif analysis of the upstream sequences of those genes revealed four major consensus sequence motifs, which have been named dwarfism and calorie restriction-responsive elements (DFCR-REs). One of the synthesized sequences bound to hepatocyte nuclear factor-4{alpha} (HNF-4{alpha}), an important transcription factor involved in liver metabolism. Furthermore, using this sequence information, we developed a highly sensitive bioassay to identify chemicals mimicking the anti-aging effects of CR. When the reporter construct, containing an element upstream of a secreted alkaline phosphatase (SEAP) gene, was co-transfected with HNF-4{alpha} and its regulator peroxisome proliferator-activated receptor (PPAR) {gamma} coactivator-1{alpha} (PGC-1{alpha}), SEAP activity was increased compared with untransfected controls. Moreover, transient transgenic mice established using this construct showed increased SEAP activity in CR mice compared with ad libitum-fed mice. These data

  17. Suppressor Screens in Arabidopsis.

    Science.gov (United States)

    Li, Xin; Zhang, Yuelin

    2016-01-01

    Genetic screens have proven to be a useful tool in the dissection of biological processes in plants. Specifically, suppressor screens have been widely used to study signal transduction pathways. Here we provide a detailed protocol for ethyl methanesulfonate (EMS) mutagenesis used in our suppressor screens in Arabidopsis and discuss the basic principles behind suppressor screen design and downstream analyses. PMID:26577776

  18. Haploid genetic screens identify an essential role for PLP2 in the downregulation of novel plasma membrane targets by viral E3 ubiquitin ligases.

    Directory of Open Access Journals (Sweden)

    Richard T Timms

    Full Text Available The Kaposi's sarcoma-associated herpesvirus gene products K3 and K5 are viral ubiquitin E3 ligases which downregulate MHC-I and additional cell surface immunoreceptors. To identify novel cellular genes required for K5 function we performed a forward genetic screen in near-haploid human KBM7 cells. The screen identified proteolipid protein 2 (PLP2, a MARVEL domain protein of unknown function, as essential for K5 activity. Genetic loss of PLP2 traps the viral ligase in the endoplasmic reticulum, where it is unable to ubiquitinate and degrade its substrates. Subsequent analysis of the plasma membrane proteome of K5-expressing KBM7 cells in the presence and absence of PLP2 revealed a wide range of novel K5 targets, all of which required PLP2 for their K5-mediated downregulation. This work ascribes a critical function to PLP2 for viral ligase activity and underlines the power of non-lethal haploid genetic screens in human cells to identify the genes involved in pathogen manipulation of the host immune system.

  19. Risks of Skin Cancer Screening

    Science.gov (United States)

    ... Genetics of Skin Cancer Skin Cancer Screening Research Skin Cancer Screening (PDQ®)–Patient Version What is screening? Go ... These are called diagnostic tests . General Information About Skin Cancer Key Points Skin cancer is a disease in ...

  20. Phenotypic chemical screening using a zebrafish neural crest EMT reporter identifies retinoic acid as an inhibitor of epithelial morphogenesis.

    Science.gov (United States)

    Jimenez, Laura; Wang, Jindong; Morrison, Monique A; Whatcott, Clifford; Soh, Katherine K; Warner, Steven; Bearss, David; Jette, Cicely A; Stewart, Rodney A

    2016-04-01

    The epithelial-to-mesenchymal transition (EMT) is a highly conserved morphogenetic program essential for embryogenesis, regeneration and cancer metastasis. In cancer cells, EMT also triggers cellular reprogramming and chemoresistance, which underlie disease relapse and decreased survival. Hence, identifying compounds that block EMT is essential to prevent or eradicate disseminated tumor cells. Here, we establish a whole-animal-based EMT reporter in zebrafish for rapid drug screening, calledTg(snai1b:GFP), which labels epithelial cells undergoing EMT to producesox10-positive neural crest (NC) cells. Time-lapse and lineage analysis ofTg(snai1b:GFP)embryos reveal that cranial NC cells delaminate from two regions: an early population delaminates adjacent to the neural plate, whereas a later population delaminates from within the dorsal neural tube. TreatingTg(snai1b:GFP)embryos with candidate small-molecule EMT-inhibiting compounds identified TP-0903, a multi-kinase inhibitor that blocked cranial NC cell delamination in both the lateral and medial populations. RNA sequencing (RNA-Seq) analysis and chemical rescue experiments show that TP-0903 acts through stimulating retinoic acid (RA) biosynthesis and RA-dependent transcription. These studies identify TP-0903 as a new therapeutic for activating RAin vivoand raise the possibility that RA-dependent inhibition of EMT contributes to its prior success in eliminating disseminated cancer cells.

  1. Phenotypic chemical screening using a zebrafish neural crest EMT reporter identifies retinoic acid as an inhibitor of epithelial morphogenesis

    Directory of Open Access Journals (Sweden)

    Laura Jimenez

    2016-04-01

    Full Text Available The epithelial-to-mesenchymal transition (EMT is a highly conserved morphogenetic program essential for embryogenesis, regeneration and cancer metastasis. In cancer cells, EMT also triggers cellular reprogramming and chemoresistance, which underlie disease relapse and decreased survival. Hence, identifying compounds that block EMT is essential to prevent or eradicate disseminated tumor cells. Here, we establish a whole-animal-based EMT reporter in zebrafish for rapid drug screening, called Tg(snai1b:GFP, which labels epithelial cells undergoing EMT to produce sox10-positive neural crest (NC cells. Time-lapse and lineage analysis of Tg(snai1b:GFP embryos reveal that cranial NC cells delaminate from two regions: an early population delaminates adjacent to the neural plate, whereas a later population delaminates from within the dorsal neural tube. Treating Tg(snai1b:GFP embryos with candidate small-molecule EMT-inhibiting compounds identified TP-0903, a multi-kinase inhibitor that blocked cranial NC cell delamination in both the lateral and medial populations. RNA sequencing (RNA-Seq analysis and chemical rescue experiments show that TP-0903 acts through stimulating retinoic acid (RA biosynthesis and RA-dependent transcription. These studies identify TP-0903 as a new therapeutic for activating RA in vivo and raise the possibility that RA-dependent inhibition of EMT contributes to its prior success in eliminating disseminated cancer cells.

  2. Screening estrogenic activities of chemicals or mixtures in vivo using transgenic (cyp19a1b-GFP zebrafish embryos.

    Directory of Open Access Journals (Sweden)

    François Brion

    Full Text Available The tg(cyp19a1b-GFP transgenic zebrafish expresses GFP (green fluorescent protein under the control of the cyp19a1b gene, encoding brain aromatase. This gene has two major characteristics: (i it is only expressed in radial glial progenitors in the brain of fish and (ii it is exquisitely sensitive to estrogens. Based on these properties, we demonstrate that natural or synthetic hormones (alone or in binary mixture, including androgens or progestagens, and industrial chemicals induce a concentration-dependent GFP expression in radial glial progenitors. As GFP expression can be quantified by in vivo imaging, this model presents a very powerful tool to screen and characterize compounds potentially acting as estrogen mimics either directly or after metabolization by the zebrafish embryo. This study also shows that radial glial cells that act as stem cells are direct targets for a large panel of endocrine disruptors, calling for more attention regarding the impact of environmental estrogens and/or certain pharmaceuticals on brain development. Altogether these data identify this in vivo bioassay as an interesting alternative to detect estrogen mimics in hazard and risk assessment perspective.

  3. Twenty-year outcome analysis of genetic screening programs for Tay-Sachs and beta-thalassemia disease carriers in high schools.

    Science.gov (United States)

    Mitchell, J J; Capua, A; Clow, C; Scriver, C R

    1996-10-01

    Programs for education, screening, and counseling of senior-high-school students, in populations at high risk for Tay-Sachs and beta-thalassemia diseases, have existed for >20 years in Montreal. Four process and outcome variables are reported here: (i) voluntary participation rates in the high-school cohort; (ii) uptake rates for the screening test; (iii) origin of carrier couples seeking the prenatal diagnosis option in the programs; and (iv) change in incidence of the two diseases. Between 1972 and 1992, we screened 14,844 Ashkenazi-Jewish students, identified 521 HexA-deficient carriers (frequency 1:28), reached 89% of the demographic cohort in the educational component of the program, and achieved 67% voluntary participation in the subsequent screening phase. The corresponding data for the beta-thalassemia program are 25,274 students (mainly of Mediterranean origin) representing 67% of the cohort with 61% voluntary participation in the screening phase (693 carriers; frequency 1:36). From demographic data, we deduce that virtually all the carriers identified in the high-school screening program remembered their status, had their partner tested if they did not already know they were a carrier couple, and took up the options for reproductive counseling/prenatal diagnosis. In Montreal, the current origin of all couples using prenatal diagnosis for Tay-Sachs and beta-thalassemia diseases is the corresponding genetic screening/testing program, whereas, at the beginning of the programs, it was always because there was a history of an affected person in the family. Incidence of the two diseases has fallen by 90%-95% over 20 years; the rare new cases are born (with two exceptions) outside the target communities or to nonscreened couples. PMID:8808593

  4. Chip-based mtDNA mutation screening enables fast and reliable genetic diagnosis of OXPHOS patients

    NARCIS (Netherlands)

    R.G.E. van Eijsden (Rudy); E. Briem (Egill); V. Tiranti (Valeria); H.J.M. Smeets (Hubert); M. Gerards (Mike); L.M.T. Eijssen (Lars); A. Hendrickx (Alexandra); R.J.E. Jongbloed (Roselie); J.H.J. Wokke (John); R.Q. Hintzen (Rogier); M.E. Rubio-Gozalbo (Estela); I.F.M. de Coo (René)

    2006-01-01

    textabstractPURPOSE: Oxidative phosphorylation is under dual genetic control of the nuclear and the mitochondrial DNA (mtDNA). Oxidative phosphorylation disorders are clinically and genetically heterogeneous, which makes it difficult to determine the genetic defect, and symptom-based protocols which

  5. Using Ambystoma mexicanum (Mexican axolotl) embryos, chemical genetics, and microarray analysis to identify signaling pathways associated with tissue regeneration.

    Science.gov (United States)

    Ponomareva, Larissa V; Athippozhy, Antony; Thorson, Jon S; Voss, S Randal

    2015-12-01

    Amphibian vertebrates are important models in regenerative biology because they present exceptional regenerative capabilities throughout life. However, it takes considerable effort to rear amphibians to juvenile and adult stages for regeneration studies, and the relatively large sizes that frogs and salamanders achieve during development make them difficult to use in chemical screens. Here, we introduce a new tail regeneration model using late stage Mexican axolotl embryos. We show that axolotl embryos completely regenerate amputated tails in 7days before they exhaust their yolk supply and begin to feed. Further, we show that axolotl embryos can be efficiently reared in microtiter plates to achieve moderate throughput screening of soluble chemicals to investigate toxicity and identify molecules that alter regenerative outcome. As proof of principle, we identified integration 1 / wingless (Wnt), transforming growth factor beta (Tgf-β), and fibroblast growth factor (Fgf) pathway antagonists that completely block tail regeneration and additional chemicals that significantly affected tail outgrowth. Furthermore, we used microarray analysis to show that inhibition of Wnt signaling broadly affects transcription of genes associated with Wnt, Fgf, Tgf-β, epidermal growth factor (Egf), Notch, nerve growth factor (Ngf), homeotic gene (Hox), rat sarcoma/mitogen-activated protein kinase (Ras/Mapk), myelocytomatosis viral oncogene (Myc), tumor protein 53 (p53), and retinoic acid (RA) pathways. Punctuated changes in the expression of genes known to regulate vertebrate development were observed; this suggests the tail regeneration transcriptional program is hierarchically structured and temporally ordered. Our study establishes the axolotl as a chemical screening model to investigate signaling pathways associated with tissue regeneration. PMID:26092703

  6. Using Ambystoma mexicanum (Mexican axolotl) embryos, chemical genetics, and microarray analysis to identify signaling pathways associated with tissue regeneration.

    Science.gov (United States)

    Ponomareva, Larissa V; Athippozhy, Antony; Thorson, Jon S; Voss, S Randal

    2015-12-01

    Amphibian vertebrates are important models in regenerative biology because they present exceptional regenerative capabilities throughout life. However, it takes considerable effort to rear amphibians to juvenile and adult stages for regeneration studies, and the relatively large sizes that frogs and salamanders achieve during development make them difficult to use in chemical screens. Here, we introduce a new tail regeneration model using late stage Mexican axolotl embryos. We show that axolotl embryos completely regenerate amputated tails in 7days before they exhaust their yolk supply and begin to feed. Further, we show that axolotl embryos can be efficiently reared in microtiter plates to achieve moderate throughput screening of soluble chemicals to investigate toxicity and identify molecules that alter regenerative outcome. As proof of principle, we identified integration 1 / wingless (Wnt), transforming growth factor beta (Tgf-β), and fibroblast growth factor (Fgf) pathway antagonists that completely block tail regeneration and additional chemicals that significantly affected tail outgrowth. Furthermore, we used microarray analysis to show that inhibition of Wnt signaling broadly affects transcription of genes associated with Wnt, Fgf, Tgf-β, epidermal growth factor (Egf), Notch, nerve growth factor (Ngf), homeotic gene (Hox), rat sarcoma/mitogen-activated protein kinase (Ras/Mapk), myelocytomatosis viral oncogene (Myc), tumor protein 53 (p53), and retinoic acid (RA) pathways. Punctuated changes in the expression of genes known to regulate vertebrate development were observed; this suggests the tail regeneration transcriptional program is hierarchically structured and temporally ordered. Our study establishes the axolotl as a chemical screening model to investigate signaling pathways associated with tissue regeneration.

  7. Chemical-genetic disruption of clathrin function spares adaptor complex 3–dependent endosome vesicle biogenesis

    OpenAIRE

    Zlatic, Stephanie A.; Grossniklaus, Emily J.; Ryder, Pearl V.; Salazar, Gloria; Mattheyses, Alexa L.; Peden, Andrew A.; Faundez, Victor

    2013-01-01

    A role for clathrin in AP-3–dependent vesicle biogenesis has been inferred from biochemical interactions and colocalization between this adaptor and clathrin. The functionality of these molecular associations, however, is controversial. We comprehensively explore the role of clathrin in AP-3–dependent vesicle budding, using rapid chemical-genetic perturbation of clathrin function with a clathrin light chain–FKBP chimera oligomerizable by the drug AP20187. We find that AP-3 interacts and coloc...

  8. The Chemical and Genetic Characteristics of Szechuan Pepper (Zanthoxylum bungeanum and Z. armatum) Cultivars and Their Suitable Habitat

    OpenAIRE

    Xiang, Li; Liu, Yue; Xie, Caixiang; Li, Xiwen; Yu, Yadong; Ye, Meng; Chen, Shilin

    2016-01-01

    Szechuan peppers, famous for their unique sensation and flavor, are widely used as a food additive and traditional herbal medicine. Zanthoxylum bungeanum and Z. armatum are both commonly recognized as Szechuan peppers, but they have different tastes and effects. The chemical components, genetic characteristics, and suitable habitat of six cultivars were analyzed in this study. The results indicated that Z. armatum contained a larger proportion of volatile oil, whereas Z. bungeanum produced a ...

  9. The chemical and genetic characteristics of Szechuan pepper (Zanthoxylum bungeanum and Z. armatum) cultivars and their suitable habitat

    OpenAIRE

    Li eXiang; Yue eLiu; Caixiang eXie; Xiwen eLi; Yadong eYu; Meng eYe; Shilin eChen

    2016-01-01

    Szechuan peppers, famous for their unique sensation and flavor, are widely used as a food additive and traditional herbal medicine. Zanthoxylum bungeanum and Z. armatum are both commonly recognized as Szechuan peppers, but they have different tastes and effects. The chemical components, genetic characteristics, and suitable habitat of six cultivars were analyzed in this study. The results indicated that Z. armatum contained a larger proportion of volatile oil, whereas Z. bungeanum produced a ...

  10. Impact of human genome initiative-derived technology on genetic testing, screening and counseling: Cultural, ethical and legal issues. Progress report

    Energy Technology Data Exchange (ETDEWEB)

    Trottier, R.W.; Hodgin, F.C.; Imara, M.; Phoenix, D.; Lybrook, S. [Morehouse Coll., Atlanta, GA (United States). School of Medicine; Crandall, L.A.; Moseley, R.E.; Armotrading, D. [Florida Univ., Gainesville, FL (United States). Coll. of Medicine

    1993-03-01

    Genetic medical services provided by the Georgia Division of Public Health in two northern and two central districts are compared to services provided in a district in which a tertiary care facility is located. Genetics outreach public health nurses play key roles in Georgia`s system of Children`s Health Services Genetics Program, including significant roles as counselors and information sources on special needs social services and support organizations. Unique features of individual health districts, (e.g., the changing face of some rural communities in ethnocultural diversity and socioeconomic character), present new challenges to current and future genetics services delivery. Preparedness as to educational needs of both health professionals and the lay population is of foremost concern in light of the ever expanding knowledge and technology in medical genetics. Perspectives on genetics and an overview of services offered by a local private sector counselor are included for comparison to state supported services. The nature of the interactions which transpire between private and public genetic services resources in Georgia will be described. A special focus of this research includes issues associated with sickle cell disease newborn screening service delivery process in Georgia, with particular attention paid to patient follow-up and transition to primary care. Of particular interest to this focus is the problem of loss to follow-up in the current system. Critical factors in education and counseling of sickle cell patients and the expectations of expanding roles of primary care physicians are discussed. The Florida approach to the delivery of genetic services contrasts to the Georgia model by placing more emphasis on a consultant-specialist team approach.

  11. MRI screening for breast cancer in women with familial or genetic predisposition : design of the Dutch National Study (MRISC)

    NARCIS (Netherlands)

    Kriege, M; Brekelmans, C T; Boetes, C; Rutgers, E J; Oosterwijk, J C; Tollenaar, R A; Manoliu, R A; Holland, R; de Koning, H J; Klijn, J G

    2001-01-01

    Mammography screening of women aged 50-70 years for breast cancer has proven to be effective in reducing breast cancer mortality. There is no consensus about the value of breast cancer screening in women aged 40-49 years. Five to ten per cent of all breast cancers are hereditary. One of the options

  12. Establishment of IL-7 Expression Reporter Human Cell Lines, and Their Feasibility for High-Throughput Screening of IL-7-Upregulating Chemicals

    Science.gov (United States)

    Sim, Chan Kyu; Kim, Inki

    2016-01-01

    Interleukin-7 (IL-7) is a cytokine essential for T cell homeostasis, and is clinically important. However, the regulatory mechanism of IL-7 gene expression is not well known, and a systematic approach to screen chemicals that regulate IL-7 expression has not yet been developed. In this study, we attempted to develop human reporter cell lines using CRISPR/Cas9-mediated genome editing technology. For this purpose, we designed donor DNA that contains an enhanced green fluorescent protein (eGFP) gene, drug selection cassette, and modified homologous arms which are considered to enhance the translation of the eGFP reporter transcript, and also a highly efficient single-guide RNA with a minimal off-target effect to target the IL-7 start codon region. By applying this system, we established IL-7 eGFP reporter cell lines that could report IL-7 gene transcription based on the eGFP protein signal. Furthermore, we utilized the cells to run a pilot screen campaign for IL-7-upregulating chemicals in a high-throughput format, and identified a chemical that can up-regulate IL-7 gene transcription. Collectively, these results suggest that our IL-7 reporter system can be utilized in large-scale chemical library screening to reveal novel IL-7 regulatory pathways and to identify potential drugs for development of new treatments in immunodeficiency disease. PMID:27589392

  13. Establishment of IL-7 Expression Reporter Human Cell Lines, and Their Feasibility for High-Throughput Screening of IL-7-Upregulating Chemicals.

    Science.gov (United States)

    Cho, Yeon Sook; Kim, Byung Soo; Sim, Chan Kyu; Kim, Inki; Lee, Myeong Sup

    2016-01-01

    Interleukin-7 (IL-7) is a cytokine essential for T cell homeostasis, and is clinically important. However, the regulatory mechanism of IL-7 gene expression is not well known, and a systematic approach to screen chemicals that regulate IL-7 expression has not yet been developed. In this study, we attempted to develop human reporter cell lines using CRISPR/Cas9-mediated genome editing technology. For this purpose, we designed donor DNA that contains an enhanced green fluorescent protein (eGFP) gene, drug selection cassette, and modified homologous arms which are considered to enhance the translation of the eGFP reporter transcript, and also a highly efficient single-guide RNA with a minimal off-target effect to target the IL-7 start codon region. By applying this system, we established IL-7 eGFP reporter cell lines that could report IL-7 gene transcription based on the eGFP protein signal. Furthermore, we utilized the cells to run a pilot screen campaign for IL-7-upregulating chemicals in a high-throughput format, and identified a chemical that can up-regulate IL-7 gene transcription. Collectively, these results suggest that our IL-7 reporter system can be utilized in large-scale chemical library screening to reveal novel IL-7 regulatory pathways and to identify potential drugs for development of new treatments in immunodeficiency disease. PMID:27589392

  14. Identification of hepta-histidine as a candidate drug for Huntington’s disease by in silico-in vitro- in vivo-integrated screens of chemical libraries

    Science.gov (United States)

    Imamura, Tomomi; Fujita, Kyota; Tagawa, Kazuhiko; Ikura, Teikichi; Chen, Xigui; Homma, Hidenori; Tamura, Takuya; Mao, Ying; Taniguchi, Juliana Bosso; Motoki, Kazumi; Nakabayashi, Makoto; Ito, Nobutoshi; Yamada, Kazunori; Tomii, Kentaro; Okano, Hideyuki; Kaye, Julia; Finkbeiner, Steven; Okazawa, Hitoshi

    2016-01-01

    We identified drug seeds for treating Huntington’s disease (HD) by combining in vitro single molecule fluorescence spectroscopy, in silico molecular docking simulations, and in vivo fly and mouse HD models to screen for inhibitors of abnormal interactions between mutant Htt and physiological Ku70, an essential DNA damage repair protein in neurons whose function is known to be impaired by mutant Htt. From 19,468 and 3,010,321 chemicals in actual and virtual libraries, fifty-six chemicals were selected from combined in vitro-in silico screens; six of these were further confirmed to have an in vivo effect on lifespan in a fly HD model, and two chemicals exerted an in vivo effect on the lifespan, body weight and motor function in a mouse HD model. Two oligopeptides, hepta-histidine (7H) and Angiotensin III, rescued the morphological abnormalities of primary neurons differentiated from iPS cells of human HD patients. For these selected drug seeds, we proposed a possible common structure. Unexpectedly, the selected chemicals enhanced rather than inhibited Htt aggregation, as indicated by dynamic light scattering analysis. Taken together, these integrated screens revealed a new pathway for the molecular targeted therapy of HD. PMID:27653664

  15. Impact of gene patents and licensing practices on access to genetic testing and carrier screening for Tay-Sachs and Canavan disease.

    Science.gov (United States)

    Colaianni, Alessandra; Chandrasekharan, Subhashini; Cook-Deegan, Robert

    2010-04-01

    Genetic testing for Tay-Sachs and Canavan disease is particularly important for Ashkenazi Jews, because both conditions are more frequent in that population. This comparative case study was possible because of different patenting and licensing practices. The role of DNA testing differs between Tay-Sachs and Canavan diseases. The first-line screening test for Tay-Sachs remains an enzyme activity test rather than genotyping. Genotyping is used for preimplantation diagnosis and confirmatory testing. In contrast, DNA-based testing is the basis for Canavan screening and diagnosis. The HEXA gene for Tay-Sachs was cloned at the National Institutes of Health, and the gene was patented but has not been licensed. The ASPA gene for Canavan disease was cloned and patented by Miami Children's Hospital. Miami Children's Hospital did not inform family members and patient groups that had contributed to the gene discovery that it was applying for a patent, and pursued restrictive licensing practices when a patent issued in 1997. This led to intense controversy, litigation, and a sealed, nonpublic 2003 settlement that apparently allowed for nonexclusive licensing. A survey of laboratories revealed a possible price premium for ASPA testing, with per-unit costs higher than for other genetic tests in the Secretary's Advisory Committee on Genetics, Health, and Society case studies. The main conclusion from comparing genetic testing for Tay-Sachs and Canavan diseases, however, is that patenting and licensing conducted without communication with patients and advocates cause mistrust and can lead to controversy and litigation, a negative model to contrast with the positive model of patenting and licensing for genetic testing of cystic fibrosis. PMID:20393311

  16. Evaluating the impact of a fluoropolymer plant on a river macrobenthic community by a combined chemical, ecological and genetic approach.

    Science.gov (United States)

    Rusconi, Marianna; Marziali, Laura; Stefani, Fabrizio; Valsecchi, Sara; Bettinetti, Roberta; Mazzoni, Michela; Rosignoli, Federica; Polesello, Stefano

    2015-12-15

    Effect-based monitoring is a recommended approach suggested in European Guidelines to assess the response of ecosystem affected by a pollution source, considering the effects at community, population, individual but also at suborganism level. A combined chemical, ecological and genetic approach was applied in order to assess the impact of a fluoropolymer plant on the macrobenthic community of the Northern Italian river Bormida (Piedmont region). The macrobenthic community living downstream of the industrial discharge was chronically exposed to a mixture of perfluoroalkyl substances (PFAS), with perfluorooctanoic acid as the main compound, at concentrations up to several μgL(-1). Ecological assessment proved that the downstream community was not substantially different from that living upstream of the pollution source. The impact on community is not quantifiable with the traditional monitoring methods used for ecological classification under European regulation because macrobenthic communities showed only slight differences in their structure. In order to highlight effects on genetic variability of the native population, a subcellular analysis by using the AFLP (Amplified Fragment Length Polymorphism) genetic technique was applied to genotype of individuals of a selected species (Hydropsyche modesta, Trichoptera) collected in the two sampling sites. Percentage of variation between the two populations was 6.8%, a threshold compatible with a genetic drift induced in the downstream population. The genetic study carried out in field identified a significant divergence between exposed and non-exposed populations, but at present it is not possible to associate this divergence to a specific effect induced by PFAS.

  17. Development and Interlaboratory Validation of a Simple Screening Method for Genetically Modified Maize Using a ΔΔC(q)-Based Multiplex Real-Time PCR Assay.

    Science.gov (United States)

    Noguchi, Akio; Nakamura, Kosuke; Sakata, Kozue; Sato-Fukuda, Nozomi; Ishigaki, Takumi; Mano, Junichi; Takabatake, Reona; Kitta, Kazumi; Teshima, Reiko; Kondo, Kazunari; Nishimaki-Mogami, Tomoko

    2016-04-19

    A number of genetically modified (GM) maize events have been developed and approved worldwide for commercial cultivation. A screening method is needed to monitor GM maize approved for commercialization in countries that mandate the labeling of foods containing a specified threshold level of GM crops. In Japan, a screening method has been implemented to monitor approved GM maize since 2001. However, the screening method currently used in Japan is time-consuming and requires generation of a calibration curve and experimental conversion factor (C(f)) value. We developed a simple screening method that avoids the need for a calibration curve and C(f) value. In this method, ΔC(q) values between the target sequences and the endogenous gene are calculated using multiplex real-time PCR, and the ΔΔC(q) value between the analytical and control samples is used as the criterion for determining analytical samples in which the GM organism content is below the threshold level for labeling of GM crops. An interlaboratory study indicated that the method is applicable independently with at least two models of PCR instruments used in this study. PMID:27010783

  18. Societies drifting apart? Behavioural, genetic and chemical differentiation between supercolonies in the yellow crazy ant Anoplolepis gracilipes.

    Directory of Open Access Journals (Sweden)

    Jochen Drescher

    Full Text Available BACKGROUND: In populations of most social insects, gene flow is maintained through mating between reproductive individuals from different colonies in periodic nuptial flights followed by dispersal of the fertilized foundresses. Some ant species, however, form large polygynous supercolonies, in which mating takes place within the maternal nest (intranidal mating and fertilized queens disperse within or along the boundary of the supercolony, leading to supercolony growth (colony budding. As a consequence, gene flow is largely confined within supercolonies. Over time, such supercolonies may diverge genetically and, thus, also in recognition cues (cuticular hydrocarbons, CHC's by a combination of genetic drift and accumulation of colony-specific, neutral mutations. METHODOLOGY/PRINCIPAL FINDINGS: We tested this hypothesis for six supercolonies of the invasive ant Anoplolepis gracilipes in north-east Borneo. Within supercolonies, workers from different nests tolerated each other, were closely related and showed highly similar CHC profiles. Between supercolonies, aggression ranged from tolerance to mortal encounters and was negatively correlated with relatedness and CHC profile similarity. Supercolonies were genetically and chemically distinct, with mutually aggressive supercolony pairs sharing only 33.1%±17.5% (mean ± SD of their alleles across six microsatellite loci and 73.8%±11.6% of the compounds in their CHC profile. Moreover, the proportion of alleles that differed between supercolony pairs was positively correlated to the proportion of qualitatively different CHC compounds. These qualitatively differing CHC compounds were found across various substance classes including alkanes, alkenes and mono-, di- and trimethyl-branched alkanes. CONCLUSIONS: We conclude that positive feedback between genetic, chemical and behavioural traits may further enhance supercolony differentiation through genetic drift and neutral evolution, and may drive

  19. A large-scale complex haploinsufficiency-based genetic interaction screen in Candida albicans: analysis of the RAM network during morphogenesis.

    Directory of Open Access Journals (Sweden)

    Nike Bharucha

    2011-04-01

    Full Text Available The morphogenetic transition between yeast and filamentous forms of the human fungal pathogen Candida albicans is regulated by a variety of signaling pathways. How these pathways interact to orchestrate morphogenesis, however, has not been as well characterized. To address this question and to identify genes that interact with the Regulation of Ace2 and Morphogenesis (RAM pathway during filamentation, we report the first large-scale genetic interaction screen in C. albicans.Our strategy for this screen was based on the concept of complex haploinsufficiency (CHI. A heterozygous mutant of CBK1(cbk1Δ/CBK1, a key RAM pathway protein kinase, was subjected to transposon-mediated, insertional mutagenesis. The resulting double heterozygous mutants (6,528 independent strains were screened for decreased filamentation on SpiderMedium (SM. From the 441 mutants showing altered filamentation, 139 transposon insertion sites were sequenced,yielding 41 unique CBK1-interacting genes. This gene set was enriched in transcriptional targets of Ace2 and, strikingly, the cAMP-dependent protein kinase A (PKA pathway, suggesting an interaction between these two pathways. Further analysis indicates that the RAM and PKA pathways co-regulate a common set of genes during morphogenesis and that hyperactivation of the PKA pathway may compensate for loss of RAM pathway function. Our data also indicate that the PKA–regulated transcription factor Efg1 primarily localizes to yeast phase cells while the RAM–pathway regulated transcription factor Ace2 localizes to daughter nuclei of filamentous cells, suggesting that Efg1 and Ace2 regulate a common set of genes at separate stages of morphogenesis. Taken together, our observations indicate that CHI–based screening is a useful approach to genetic interaction analysis in C. albicans and support a model in which these two pathways regulate a common set of genes at different stages of filamentation.

  20. Description of the type of chemical restraint used by French veterinarians to perform hip dysplasia screening radiographs. A retrospective study based on 3302 radiographs.

    Science.gov (United States)

    Maitre, P; Genevois, J-P; Remy, D; Carozzo, C; Arnault, F; Buttin, P; Viguier, E; Fau, D

    2010-01-01

    The objective of this study was to investigate the type of chemical restraint used by French practitioners to perform official hip screening radiographs, to determine the proportion of dogs under general anesthesia versus dogs under sedation, and to search for a difference in hip dysplasia (HD) prevalence between the two groups. From September 2005 to August 2008, 3302 conventional ventrodorsal hip extended radiographs sent for official scoring to the same panellist were selected because information related to the type of chemical restraint had been provided. There were 2825 dogs under general anesthesia and 477 were sedated. Chemical restraint used by French veterinarians to perform HD screening radiographs is mainly based on general intra-venous anesthesia with an alpha2 agonist associated with ketamine. A single injection of alpha2 agonist is also mostly used for dogs which are radiographed while under sedation. A very low (1.7%) difference in HD prevalence was noted between the anesthetized and the sedated group. Except for acepromazine, which has been demonstrated to provide insufficient muscle relaxation to show evidence of hip laxity, the protocols seem acceptable as regards the Federation Cynologique Internationale requirements for HD screening. PMID:20585709

  1. Genetic screening for Niemann–Pick disease type C in adults with neurological and psychiatric symptoms: findings from the ZOOM study

    OpenAIRE

    Bauer, Peter; Balding, David J.; Klünemann, Hans H.; Linden, David E. J.; Ory, Daniel S.; Pineda, Mercè; Priller, Josef; Sedel, Frederic; Muller, Audrey; Chadha-Boreham, Harbajan; Richard W D Welford; Strasser, Daniel S.; Patterson, Marc C.

    2013-01-01

    Niemann–Pick disease type C (NP-C) is a rare, autosomal-recessive, progressive neurological disease caused by mutations in either the NPC1 gene (in 95% of cases) or the NPC2 gene. This observational, multicentre genetic screening study evaluated the frequency and phenotypes of NP-C in consecutive adult patients with neurological and psychiatric symptoms. Diagnostic testing for NP-C involved NPC1 and NPC2 exonic gene sequencing and gene dosage analysis. When available, results of filipin stain...

  2. A Screen for Modifiers of Cilia Phenotypes Reveals Novel MKS Alleles and Uncovers a Specific Genetic Interaction between osm-3 and nphp-4

    OpenAIRE

    Masyukova, Svetlana V.; Dawn E Landis; Henke, Scott J.; Williams, Corey L.; Pieczynski, Jay N.; Roszczynialski, Kelly N.; Jannese E Covington; Malarkey, Erik B.; Yoder, Bradley K.

    2016-01-01

    Nephronophthisis (NPHP) is a ciliopathy in which genetic modifiers may underlie the variable penetrance of clinical features. To identify modifiers, a screen was conducted on C. elegans nphp-4(tm925) mutants. Mutations in ten loci exacerbating nphp-4(tm925) ciliary defects were obtained. Four loci have been identified, three of which are established ciliopathy genes mks-1, mks-2, and mks-5. The fourth allele (yhw66) is a missense mutation (S316F) in OSM-3, a kinesin required for cilia distal ...

  3. Use of a Two-Color Genetic Screen To Identify a Domain of the Global Regulator Lrp That Is Specifically Required for pap Phase Variation

    OpenAIRE

    Kaltenbach, Linda; Braaten, Bruce; Tucker, Julie; Krabbe, Margareta; Low, David

    1998-01-01

    The global regulator Lrp plays a central role as both a repressor and an activator in Pap phase variation. Unlike most other members of the Lrp regulon such as ilvIH, activation of papBA transcription requires the coregulator PapI and is methylation dependent. We developed a two-color genetic screen to identify Lrp mutations that inhibit Pap phase variation but still activate ilvIH transcription, reasoning that such mutations might identify PapI binding or methylation-responsive domains. Amin...

  4. Human Genetics in Rheumatoid Arthritis Guides a High-Throughput Drug Screen of the CD40 Signaling Pathway

    NARCIS (Netherlands)

    Li, Gang; Diogo, Dorothee; Wu, Di; Spoonamore, Jim; Dancik, Vlado; Franke, Lude; Kurreeman, Fina; Rossin, Elizabeth J.; Duclos, Grant; Hartland, Cathy; Zhou, Xuezhong; Li, Kejie; Liu, Jun; De Jager, Philip L.; Siminovitch, Katherine A.; Zhernakova, Alexandra; Raychaudhuri, Soumya; Bowes, John; Eyre, Steve; Padyukov, Leonid; Gregersen, Peter K.; Worthington, Jane; Gupta, Namrata; Clemons, Paul A.; Stahl, Eli; Tolliday, Nicola; Plenge, Robert M.

    2013-01-01

    Although genetic and non-genetic studies in mouse and human implicate the CD40 pathway in rheumatoid arthritis (RA), there are no approved drugs that inhibit CD40 signaling for clinical care in RA or any other disease. Here, we sought to understand the biological consequences of a CD40 risk variant

  5. Screening for Functional Non-coding Genetic Variants Using Electrophoretic Mobility Shift Assay (EMSA) and DNA-affinity Precipitation Assay (DAPA).

    Science.gov (United States)

    Miller, Daniel E; Patel, Zubin H; Lu, Xiaoming; Lynch, Arthur T; Weirauch, Matthew T; Kottyan, Leah C

    2016-01-01

    Population and family-based genetic studies typically result in the identification of genetic variants that are statistically associated with a clinical disease or phenotype. For many diseases and traits, most variants are non-coding, and are thus likely to act by impacting subtle, comparatively hard to predict mechanisms controlling gene expression. Here, we describe a general strategic approach to prioritize non-coding variants, and screen them for their function. This approach involves computational prioritization using functional genomic databases followed by experimental analysis of differential binding of transcription factors (TFs) to risk and non-risk alleles. For both electrophoretic mobility shift assay (EMSA) and DNA affinity precipitation assay (DAPA) analysis of genetic variants, a synthetic DNA oligonucleotide (oligo) is used to identify factors in the nuclear lysate of disease or phenotype-relevant cells. For EMSA, the oligonucleotides with or without bound nuclear factors (often TFs) are analyzed by non-denaturing electrophoresis on a tris-borate-EDTA (TBE) polyacrylamide gel. For DAPA, the oligonucleotides are bound to a magnetic column and the nuclear factors that specifically bind the DNA sequence are eluted and analyzed through mass spectrometry or with a reducing sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) followed by Western blot analysis. This general approach can be widely used to study the function of non-coding genetic variants associated with any disease, trait, or phenotype. PMID:27585267

  6. An optimized chemical kinetic mechanism for HCCI combustion of PRFs using multi-zone model and genetic algorithm

    International Nuclear Information System (INIS)

    Highlights: • A new chemical kinetic mechanism for PRFs HCCI combustion is developed. • New mechanism optimization is performed using genetic algorithm and multi-zone model. • Engine-related combustion and performance parameters are predicted accurately. • Engine unburned HC and CO emissions are predicted by the model properly. - Abstract: Development of comprehensive chemical kinetic mechanisms is required for HCCI combustion and emissions prediction to be used in engine development. The main purpose of this study is development of a new chemical kinetic mechanism for primary reference fuels (PRFs) HCCI combustion, which can be applied to combustion models to predict in-cylinder pressure and exhaust CO and UHC emissions, accurately. Hence, a multi-zone model is developed for HCCI engine simulation. Two semi-detailed chemical kinetic mechanisms those are suitable for premixed combustion are used for n-heptane and iso-octane HCCI combustion simulation. The iso-octane mechanism contains 84 species and 484 reactions and the n-heptane mechanism contains 57 species and 296 reactions. A simple interaction between iso-octane and n-heptane is considered in new mechanism. The multi-zone model is validated using experimental data for pure n-heptane and iso-octane. A new mechanism is prepared by combination of these two mechanisms for n-heptane and iso-octane blended fuel, which includes 101 species and 594 reactions. New mechanism optimization is performed using genetic algorithm and multi-zone model. Mechanism contains low temperature heat release region, which decreases with increasing octane number. The results showed that the optimized chemical kinetic mechanism is capable of predicting engine-related combustion and performance parameters. Also after implementing the optimized mechanism, engine unburned HC and CO emissions predicted by the model are in good agreement with the corresponding experimental data

  7. Screening of antibiotics and chemical analysis of penicillin residue in fresh milk and traditional dairy products in Oyo state, Nigeria

    Science.gov (United States)

    Olatoye, Isaac Olufemi; Daniel, Oluwayemisi Folashade; Ishola, Sunday Ayobami

    2016-01-01

    Background and Aim: There are global public health and economic concerns on chemical residues in food of animal origin. The use of antibiotics in dairy cattle for the treatment of diseases such as mastitis has contributed to the presence of residues in dairy products. Penicillin residues as low as 1 ppb can lead to allergic reactions and shift of resistance patterns in microbial population as well as interfere with the processing of several dairy products. Antibiotic monitoring is an essential quality control measure in safe milk production. This study was aimed at determining antibiotic residue contamination and the level of penicillin in dairy products from Fulani cattle herds in Oyo State. Materials and Methods: The presence of antibiotic residues in 328 samples of fresh milk, 180 local cheese (wara), and 90 fermented milk (nono) from Southwest, Nigeria were determined using Premi® test kit (R-Biopharm AG, Germany) followed by high-performance liquid chromatography analysis of penicillin-G residue. Results: Antibiotic residues were obtained in 40.8%, 24.4% and 62.3% fresh milk, wara and nono, respectively. Penicillin-G residue was also detected in 41.1% fresh milk, 40.2% nono and 24.4% wara at mean concentrations of 15.22±0.61, 8.24±0.50 and 7.6±0.60 μg/L with 39.3%, 36.7% and 21.1%, respectively, containing penicillin residue above recommended Codex maximum residue limit (MRL) of 5 μg/L in dairy. There was no significant difference between the mean penicillin residues in all the dairy products in this study. Conclusion: The results are of food safety concern since the bulk of the samples and substantial quantities of dairy products in Oyo state contained violative levels of antibiotic residues including penicillin residues in concentrations above the MRL. This could be due to indiscriminate and unregulated administration of antibiotics to dairy cattle. Regulatory control of antibiotic use, rapid screening of milk and dairy farmers’ extension education

  8. Integration of genetic virtual screening patterns and latent multivariate modeling techniques for QSAR optimization based on combinations and/or interactions between peptides and proteins

    Institute of Scientific and Technical Information of China (English)

    LI ZhiLiang; CHEN Gang; LI GenRong; TIAN FeiFei; WU ShiRong; YANG ShanBin; YANG ShengXi; ZHOU Yuan; ZHANG QiaoXia; QIN RenHui; MEI Hu

    2008-01-01

    Both the concept and the model of snug quantitative structure-activity relationship (QSAR) were pro-posed and developed for molecular design through constructing QSAR based on some known mode of receptor/ligand interactions. Many disadvantages of traditional models can be avoided by using the proposed method because the traditional models only determined upon molecular structural features in sample sets themselves. A genetic virtual screening of peptide/protein combinations (GVSPPC) is proposed for the first time by utilizing this idea to examine peptide/protein affinity activities. A genetic algorithm (GA) was developed for screening combinative targets with an interaction mode for virtual receptors. GVSPPC succeeds in disposing difficulties in rational QSAR, in order to search for the ligand/receptor interactions on conditions of unknown structures. Some bioactive oligo-/poly-peptide systems covering 58 angiotensin converting enzyme (ACE) inhibitors and 18 double site mutation residues in camel antibody protein cAb-Lys3 were investigated by GVSPPC with satisfactory results (Rcu2 > 0.91, Qcv2 0.86, ERMS = 0.19-0.95), respectively, which demonstrates that GVSPPC is more inter-pretable in the ligand-receptor interaction than the traditional QSAR method.

  9. Screening for JH1 genetic defect carriers in Jersey cattle by a polymerase chain reaction and restriction fragment length polymorphism assay.

    Science.gov (United States)

    Zhang, Yi; Guo, Gang; Huang, Hetian; Lu, Lu; Wang, Lijie; Fang, Lingzhao; Liu, Lin; Wang, Yachun; Zhang, Shengli

    2015-09-01

    An autosomal recessive genetic defect termed JH1 has been associated with early embryonic loss in the Jersey cattle breed. The genetic basis has been identified as a cytosine to thymine mutation in the CWC15 gene that changes an amino acid from arginine to a stop code. To screen for JH1 carriers in an imported Jersey population in China, a method based on a polymerase chain reaction amplification followed by a restriction fragment length polymorphism assay (PCR-RFLP) was developed for the accurate diagnosis of the JH1 allele. A total of 449 randomly chosen cows were examined with the PCR-RFLP assay, and 31 were identified as JH1 carriers, corresponding to a carrier frequency of 6.9%. The PCR-RFLP method was validated by DNA sequencing of 8 positive and 13 negative samples, with all 21 samples giving the expected DNA sequence. In addition, 3 negative and 3 positive samples were confirmed by a commercial microarray-based single nucleotide polymorphism assay. Finally, samples from 9 bulls in the United States of known status were correctly identified as carriers (5 bulls) or noncarriers (4 bulls). As the JH1 defect has most likely spread worldwide, implementing routine screening is necessary to avoid the risk of carrier-to-carrier matings and to gradually eradicate the deleterious gene.

  10. Integration of genetic virtual screening patterns and latent multivariate modeling techniques for QSAR optimization based on combinations and/or interactions between peptides and proteins

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Both the concept and the model of snug quantitative structure-activity relationship (QSAR) were pro-posed and developed for molecular design through constructing QSAR based on some known mode of receptor/ligand interactions. Many disadvantages of traditional models can be avoided by using the proposed method because the traditional models only determined upon molecular structural features in sample sets themselves. A genetic virtual screening of peptide/protein combinations (GVSPPC) is proposed for the first time by utilizing this idea to examine peptide/protein affinity activities. A genetic algorithm (GA) was developed for screening combinative targets with an interaction mode for virtual receptors. GVSPPC succeeds in disposing difficulties in rational QSAR,in order to search for the ligand/receptor interactions on conditions of unknown structures. Some bioactive oligo-/poly-peptide systems covering 58 angiotensin converting enzyme (ACE) inhibitors and 18 double site mutation residues in camel antibody protein cAb-Lys3 were investigated by GVSPPC with satisfactory results (R 2 cu>0.91,Q 2 cv > 0.86,ERMS=0.19-0.95),respectively,which demonstrates that GVSPPC is more inter-pretable in the ligand-receptor interaction than the traditional QSAR method.

  11. Dynamic phosphoregulation of the cortical actin cytoskeleton and endocytic machinery revealed by real-time chemical genetic analysis

    OpenAIRE

    Sekiya-Kawasaki, Mariko; Groen, Aaron Chris; Cope, M. Jamie T.V.; Kaksonen, Marko; Watson, Hadiya A.; Zhang, Chao; Shokat, Kevan M.; Wendland, Beverly; McDonald, Kent L.; McCaffery, J. Michael; Drubin, David G.

    2003-01-01

    We used chemical genetics to control the activity of budding yeast Prk1p, which is a protein kinase that is related to mammalian GAK and AAK1, and which targets several actin regulatory proteins implicated in endocytosis. In vivo Prk1p inhibition blocked pheromone receptor endocytosis, and caused cortical actin patches to rapidly aggregate into large clumps that contained Abp1p, Sla2p, Pan1p, Sla1p, and Ent1p. Clump formation depended on Arp2p, suggesting that this phenotype might result from...

  12. Genetic Counseling

    Science.gov (United States)

    ... go for genetic counseling, such as: A family history of a genetic condition To learn about genetic screening for diseases that are more common in certain ethnic groups (e.g., sickle cell disease in African Americans and Tay-Sachs disease in Ashkenazi Jews) To discuss abnormal results ...

  13. A forward-genetic screen and dynamic analysis of lambda phage host-dependencies reveals an extensive interaction network and a new anti-viral strategy.

    Directory of Open Access Journals (Sweden)

    Nathaniel D Maynard

    2010-07-01

    Full Text Available Latently infecting viruses are an important class of virus that plays a key role in viral evolution and human health. Here we report a genome-scale forward-genetics screen for host-dependencies of the latently-infecting bacteriophage lambda. This screen identified 57 Escherichia coli (E. coli genes--over half of which have not been previously associated with infection--that when knocked out inhibited lambda phage's ability to replicate. Our results demonstrate a highly integrated network between lambda and its host, in striking contrast to the results from a similar screen using the lytic-only infecting T7 virus. We then measured the growth of E. coli under normal and infected conditions, using wild-type and knockout strains deficient in one of the identified host genes, and found that genes from the same pathway often exhibited similar growth dynamics. This observation, combined with further computational and experimental analysis, led us to identify a previously unannotated gene, yneJ, as a novel regulator of lamB gene expression. A surprising result of this work was the identification of two highly conserved pathways involved in tRNA thiolation-one pathway is required for efficient lambda replication, while the other has anti-viral properties inhibiting lambda replication. Based on our data, it appears that 2-thiouridine modification of tRNAGlu, tRNAGln, and tRNALys is particularly important for the efficient production of infectious lambda phage particles.

  14. Screening of Phage-Resistant Tryptophanase Genetic Engineering Strains%抗噬菌体色氨酸酶基因工程菌的筛选

    Institute of Scientific and Technical Information of China (English)

    梅运军; 牟艳华; 石德太; 胡纯; 王文清

    2012-01-01

    利用自发突变、紫外诱变、协同进化、紫外耦合协同进化4种方法对色氨酸酶基因工程菌WD0801进行了抗噬菌体筛选.结果表明紫外耦合协同进化法具有较好的筛选效果,筛选的42株稳定遗传的抗性菌株中有23株的长势高于出发株,11株的酶活优于出发株.%4 methods, spontaneous mutation,UV mutagenesis,co-evolution,UV coupled co-evolution, were employed to screen EscheTichia coli tryptophanase genetic engineering strains. The results showed that the screening efficiency of UV-coupled co-evolution method was the best. Among the 42 stable phage—resistant strains screened out, the growth vigor of 23 strains, and enzyme activity of 11 strains were higher than the original strain.

  15. Anabolic implant effects on visceral organ mass, chemical body composition, and estimated energetic efficiency in cloned (genetically identical) beef steers.

    Science.gov (United States)

    Hutcheson, J P; Johnson, D E; Gerken, C L; Morgan, J B; Tatum, J D

    1997-10-01

    Six sets of four genetically identical Brangus steers (n = 24; X BW 409 kg) were used to determine the effect of different anabolic implants on visceral organ mass, chemical body composition, estimated tissue deposition, and energetic efficiency. Steers within a clone set were randomly assigned to one of the following implant treatments: C, no implant; E, estrogenic; A, androgenic, or AE, androgenic + estrogenic. Steers were slaughtered 112 d after implanting; visceral organs were weighed and final body composition determined by mechanical grinding and chemical analysis of the empty body. Mass of the empty gastrointestinal tract (GIT) was reduced approximately 9% (P .10) the efficiency of ME utilization. In general, estrogenic implants decreased GIT, androgenic implants increased liver, and all implants increased hide mass. Steers implanted with an AE combination had additive effects on protein deposition compared with either implant alone. The NEg requirements for body gain are estimated to be reduced 19% by estrogenic or combination implants. PMID:9331863

  16. A Sleeping Beauty DNA transposon-based genetic sensor for functional screening of vitamin D3 analogues

    DEFF Research Database (Denmark)

    Staunstrup, Nicklas Heine; Sharma, Nynne; Bak, Rasmus Otkjær;

    2011-01-01

    Analogues of vitamin D3 are extensively used in the treatment of various illnesses, such as osteoporosis, inflammatory skin diseases, and cancer. Functional testing of new vitamin D3 analogues and formulations for improved systemic and topical administration is supported by sensitive screening me...

  17. After the Introduction into the National Newborn Screening Program : Who Is Receiving Genetic Counseling for Hemoglobinopathies in The Netherlands?

    NARCIS (Netherlands)

    Kaufmann, J. O.; Krapels, I. P. C.; Van Brussel, B. T. J.; Zekveld-Vroon, R. C.; Oosterwijk, J. C.; van Erp, F.; van Echtelt, J.; Zwijnenburg, P. J. G.; Petrij, F.; Bakker, E.; Giordano, P. C.

    2014-01-01

    OBJECTIVE: Universal newborn screening for hemoglobinopathies started in The Netherlands in 2007. Herewith severe conditions, such as sickle cell disease, β-thalassemia major and hemoglobin H disease are putatively identified. Additionally, at least 1,800 carriers of hemoglobin variants associated w

  18. Rat reverse genetics : generation and characterization of chemically induced rat mutants

    NARCIS (Netherlands)

    van Boxtel, R.

    2010-01-01

    The use of animal models has been crucial for studying the function of genetic elements in the human genome. Embryonic stem (ES) cell-based homologous recombination (HR) has proven a very efficient technique for gene manipulation. However, this technique is not (yet) available for all model organism

  19. [Research progress in chemical communication among insect-resistant genetically modified plants, insect pests and natural enemies].

    Science.gov (United States)

    Liu, Qing-Song; Li, Yun-He; Chen, Xiu-Ping; Peng, Yu-Fa

    2014-08-01

    Semiochemicals released by plants or insects play an important role in the communication among plants, phytophagous insects and their natural enemies. They thus form a chemical information network which regulates intra- and inter-specific behaviors and sustains the composition and structure of plant and insect communities. The application of insect-resistant genetically modified (IRGM) crops may affect the chemical communication within and among the tritrophic levels, and thus cause disturbances to the biotic community structure and the stability of the farmland ecosystem. This has raised concerns about the environmental safety of IRGM crops and triggered research worldwide. In the current article we provided a brief summary of the chemical communication among plants, herbivores and natural enemies; analyzed the potential of IRGM crops to affect the chemical communication between plants and arthropods and the related mechanisms; and discussed the current research progress and the future prospects in this field. We hope that this will promote the research in this field by Chinese scientists and increase our understanding of the potential effects of growing of IRGM crops on the arthropod community structure.

  20. Analysis of mutants from a genetic screening reveals the control of intestine and liver development by many common genes in zebrafish.

    Science.gov (United States)

    Jiang, Faming; Chen, Jiehui; Ma, Xirui; Huang, Chao; Zhu, Shicheng; Wang, Fei; Li, Li; Luo, Lingfei; Ruan, Hua; Huang, Honghui

    2015-05-01

    Both the intestine and liver develop from the endoderm, yet little is known how these two digestive organs share and differ in their developmental programs, at the molecular level. A classical forward genetic screen, with no gene bias, is an effective way to address this question by examining the defects of the intestine and liver in obtained mutants to assess mutated genes responsible for the development of either organ or both. We report here such a screen in zebrafish. ENU was used as the mutagen because of its high mutagenic efficiency and no site preference. Embryos were collected at 3.5 dpf for RNA whole mount in situ hybridization with a cocktail probe of the intestine marker ifabp and the liver marker lfabp to check phenotypes and determine their parental heterozygosis. A total of 52 F2 putative mutants were identified, and those with general developmental defects were aborted. To rule out non-inheritable phenotypes caused by high mutation background, F2 putative mutants were outcrossed with wild type fish and a re-screen in F3 generations was performed. After complementation tests between F3 mutants with similar phenotypes originating from the same F2 families, a total of 37 F3 mutant lines originated from 22 F2 families were identified after screening 78 mutagenized genomes. Classification of mutant phenotypes indicated that 31 out of the 37 mutants showed defects in both the intestine and liver. In addition, four "intestine specific mutants" and two "liver specific mutants" showed selectively more severe phenotype in the intestine and liver respectively. These results suggested that the intestine and liver share a substantial number of essential genes during both organs development in zebrafish. Further studies of the mutants are likely to shed more insights into the molecular basis of the digestive system development in the zebrafish and vertebrate. PMID:25824031

  1. Analysis of mutants from a genetic screening reveals the control of intestine and liver development by many common genes in zebrafish.

    Science.gov (United States)

    Jiang, Faming; Chen, Jiehui; Ma, Xirui; Huang, Chao; Zhu, Shicheng; Wang, Fei; Li, Li; Luo, Lingfei; Ruan, Hua; Huang, Honghui

    2015-05-01

    Both the intestine and liver develop from the endoderm, yet little is known how these two digestive organs share and differ in their developmental programs, at the molecular level. A classical forward genetic screen, with no gene bias, is an effective way to address this question by examining the defects of the intestine and liver in obtained mutants to assess mutated genes responsible for the development of either organ or both. We report here such a screen in zebrafish. ENU was used as the mutagen because of its high mutagenic efficiency and no site preference. Embryos were collected at 3.5 dpf for RNA whole mount in situ hybridization with a cocktail probe of the intestine marker ifabp and the liver marker lfabp to check phenotypes and determine their parental heterozygosis. A total of 52 F2 putative mutants were identified, and those with general developmental defects were aborted. To rule out non-inheritable phenotypes caused by high mutation background, F2 putative mutants were outcrossed with wild type fish and a re-screen in F3 generations was performed. After complementation tests between F3 mutants with similar phenotypes originating from the same F2 families, a total of 37 F3 mutant lines originated from 22 F2 families were identified after screening 78 mutagenized genomes. Classification of mutant phenotypes indicated that 31 out of the 37 mutants showed defects in both the intestine and liver. In addition, four "intestine specific mutants" and two "liver specific mutants" showed selectively more severe phenotype in the intestine and liver respectively. These results suggested that the intestine and liver share a substantial number of essential genes during both organs development in zebrafish. Further studies of the mutants are likely to shed more insights into the molecular basis of the digestive system development in the zebrafish and vertebrate.

  2. The role of acoustic screens in distribution of technogenic magnetic particles and chemical pollution in roadside soil

    Science.gov (United States)

    Wawer, Małgorzata; Magiera, Tadeusz; Szuszkiewicz, Marcin

    2015-04-01

    Roads constructed nowadays should by all means be functional for their motorized users but at the same time their effect on the environment ought to be limited to the minimum. Despite the existence of various methods for preventing from negative influence of roads on the environment, there is still lack of adequate techniques to monitor and reduce the spreading of roadside pollution in the air and soils. The aim of the study was to assess the influence of acoustic screens on spreading and deposition of solid pollutants deriving from car emissions, based on their quantitative and qualitative analysis. During this study, measurements of magnetic susceptibility and analyses of heavy metals as well as Pt and Rh contents in soil and plant samples (Taraxacum officinale, Plantago major, Parthenocissus quinquefolia) collected near different kinds of acoustic screens ("green walls", Plexiglass, sawdust concrete, steel panels and earth embankments) have been done. Previous investigations showed showed that most of traffic emission is deposited in the close vicinity of the roads (up to 10 m) and the level of contamination decreased with increasing distance from the road edge. However, the results of this project indicate that, in the area where the acoustic screens are located, this distribution is disturbed and the additional enrichment of heavy metals in soil about 10 - 15 m behind screens is observed. Spatial distribution of heavy metal contents in soil samples corresponds to its magnetic susceptibility values. High contents of Fe, Zn, Mn and Pb was observed next to acoustic screens made of sawdust concrete and steel panels. Additionally, concentration of Zn in soil samples collected close to these screens exceeded threshold value. Analyses of plants showed that the highest content of examined elements and highest values of magnetic susceptibility were recorded near road edge. What is more, samples of Parthenocissus quinquefolia collected at height 0.2 m were characterized

  3. A chemical-genetic strategy reveals distinct temporal requirements for SAD-1 kinase in neuronal polarization and synapse formation

    Directory of Open Access Journals (Sweden)

    Shokat Kevan M

    2008-09-01

    Full Text Available Abstract Background Neurons assemble into a functional network through a sequence of developmental processes including neuronal polarization and synapse formation. In Caenorhabditis elegans, the serine/threonine SAD-1 kinase is essential for proper neuronal polarity and synaptic organization. To determine if SAD-1 activity regulates the establishment or maintenance of these neuronal structures, we examined its temporal requirements using a chemical-genetic method that allows for selective and reversible inactivation of its kinase activity in vivo. Results We generated a PP1 analog-sensitive variant of SAD-1. Through temporal inhibition of SAD-1 kinase activity we show that its activity is required for the establishment of both neuronal polarity and synaptic organization. However, while SAD-1 activity is needed strictly when neurons are polarizing, the temporal requirement for SAD-1 is less stringent in synaptic organization, which can also be re-established during maintenance. Conclusion This study reports the first temporal analysis of a neural kinase activity using the chemical-genetic system. It reveals that neuronal polarity and synaptic organization have distinct temporal requirements for SAD-1.

  4. Synthetic dual-input mammalian genetic circuits enable tunable and stringent transcription control by chemical and light.

    Science.gov (United States)

    Chen, Xianjun; Li, Ting; Wang, Xue; Du, Zengmin; Liu, Renmei; Yang, Yi

    2016-04-01

    Programmable transcription factors can enable precise control of gene expression triggered by a chemical inducer or light. To obtain versatile transgene system with combined benefits of a chemical inducer and light inducer, we created various chimeric promoters through the assembly of different copies of the tet operator and Gal4 operator module, which simultaneously responded to a tetracycline-responsive transcription factor and a light-switchable transactivator. The activities of these chimeric promoters can be regulated by tetracycline and blue light synergistically or antagonistically. Further studies of the antagonistic genetic circuit exhibited high spatiotemporal resolution and extremely low leaky expression, which therefore could be used to spatially and stringently control the expression of highly toxic protein Diphtheria toxin A for light regulated gene therapy. When transferring plasmids engineered for the gene switch-driven expression of a firefly luciferase (Fluc) into mice, the Fluc expression levels of the treated animals directly correlated with the tetracycline and light input program. We suggest that dual-input genetic circuits using TET and light that serve as triggers to achieve expression profiles may enable the design of robust therapeutic gene circuits for gene- and cell-based therapies.

  5. A Class of Diacylglycerol Acyltransferase 1 Inhibitors Identified by a Combination of Phenotypic High-throughput Screening, Genomics, and Genetics

    Directory of Open Access Journals (Sweden)

    Kirsten Tschapalda

    2016-06-01

    Full Text Available Excess lipid storage is an epidemic problem in human populations. Thus, the identification of small molecules to treat or prevent lipid storage-related metabolic complications is of great interest. Here we screened >320.000 compounds for their ability to prevent a cellular lipid accumulation phenotype. We used fly cells because the multifarious tools available for this organism should facilitate unraveling the mechanism-of-action of active small molecules. Of the several hundred lipid storage inhibitors identified in the primary screen we concentrated on three structurally diverse and potent compound classes active in cells of multiple species (including human and negligible cytotoxicity. Together with Drosophila in vivo epistasis experiments, RNA-Seq expression profiles suggested that the target of one of the small molecules was diacylglycerol acyltransferase 1 (DGAT1, a key enzyme in the production of triacylglycerols and prominent human drug target. We confirmed this prediction by biochemical and enzymatic activity tests.

  6. Top Value Added Chemicals from Biomass - Volume I, Results of Screening for Potential Candidates from Sugars and Synthesis Gas

    Energy Technology Data Exchange (ETDEWEB)

    None

    2004-08-01

    This report identifies twelve building block chemicals that can be produced from sugars via biological or chemical conversions. The twelve building blocks can be subsequently converted to a number of high-value bio-based chemicals or materials. Building block chemicals, as considered for this analysis, are molecules with multiple functional groups that possess the potential to be transformed into new families of useful molecules. The twelve sugar-based building blocks are 1,4-diacids (succinic, fumaric and malic), 2,5-furan dicarboxylic acid, 3-hydroxy propionic acid, aspartic acid, glucaric acid, glutamic acid, itaconic acid, levulinic acid, 3-hydroxybutyrolactone, glycerol, sorbitol, and xylitol/arabinitol.

  7. Comparison of Pyrolysis Mass Spectrometry and Near Infrared Spectroscopy for Genetic Analysis of Lignocellulose Chemical Composition in Populus

    Directory of Open Access Journals (Sweden)

    Jianxing Zhang

    2014-03-01

    Full Text Available Genetic analysis of wood chemical composition is often limited by the cost and throughput of direct analytical methods. The speed and low cost of Fourier transform near infrared (FT-NIR overcomes many of these limitations, but it is an indirect method relying on calibration models that are typically developed and validated with small sample sets. In this study, we used >1500 young greenhouse grown trees from a clonally propagated single Populus family, grown at low and high nitrogen, and compared FT-NIR calibration sample sizes of 150, 250, 500 and 750 on calibration and prediction model statistics, and heritability estimates developed with pyrolysis molecular beam mass spectrometry (pyMBMS wood chemical composition. As calibration sample size increased from 150 to 750, predictive model statistics improved slightly. Overall, stronger calibration and prediction statistics were obtained with lignin, S-lignin, S/G ratio, and m/z 144 (an ion from cellulose, than with C5 and C6 carbohydrates, and m/z 114 (an ion from xylan. Although small differences in model statistics were observed between the 250 and 500 sample calibration sets, when predicted values were used for calculating genetic control, the 500 sample set gave substantially more similar results to those obtained with the pyMBMS data. With the 500 sample calibration models, genetic correlations obtained with FT-NIR and pyMBMS methods were similar. Quantitative trait loci (QTL analysis with pyMBMS and FT-NIR predictions identified only three common loci for lignin traits. FT-NIR identified four QTLs that were not found with pyMBMS data, and these QTLs were for the less well predicted carbohydrate traits.

  8. Chemical Variation in a Dominant Tree Species: Population Divergence, Selection and Genetic Stability across Environments

    OpenAIRE

    Julianne M O'Reilly-Wapstra; Miller, Alison M.; Hamilton, Matthew G.; Dean Williams; Naomi Glancy-Dean; Potts, Brad M.

    2013-01-01

    Understanding among and within population genetic variation of ecologically important plant traits provides insight into the potential evolutionary processes affecting those traits. The strength and consistency of selection driving variability in traits would be affected by plasticity in differences among genotypes across environments (G×E). We investigated population divergence, selection and environmental plasticity of foliar plant secondary metabolites (PSMs) in a dominant tree species, Eu...

  9. Design and development of genetically encoded fluorescent sensors to monitor intracellular chemical and physical parameters

    OpenAIRE

    Germond, Arno; Fujita, Hideaki; Ichimura, Taro; Watanabe, Tomonobu M.

    2016-01-01

    Over the past decades many researchers have made major contributions towards the development of genetically encoded (GE) fluorescent sensors derived from fluorescent proteins. GE sensors are now used to study biological phenomena by facilitating the measurement of biochemical behaviors at various scales, ranging from single molecules to single cells or even whole animals. Here, we review the historical development of GE fluorescent sensors and report on their current status. We specifically f...

  10. A Bow-Tie Genetic Architecture for Morphogenesis Suggested by a Genome-Wide RNAi Screen in Caenorhabditis elegans

    OpenAIRE

    Nelson, Matthew D.; Elinor Zhou; Karin Kiontke; Hélène Fradin; Grayson Maldonado; Daniel Martin; Khushbu Shah; Fitch, David H. A.

    2011-01-01

    During animal development, cellular morphogenesis plays a fundamental role in determining the shape and function of tissues and organs. Identifying the components that regulate and drive morphogenesis is thus a major goal of developmental biology. The four-celled tip of the Caenorhabditis elegans male tail is a simple but powerful model for studying the mechanism of morphogenesis and its spatiotemporal regulation. Here, through a genome-wide post-embryonic RNAi-feeding screen, we identified 2...

  11. Profiling Environmental Chemicals in the Antioxidant Response Element Pathway using Quantitative High Throughput Screening (qHTS)

    Science.gov (United States)

    The antioxidant response element (ARE) signaling pathway plays an important role in the amelioration of oxidative stress, which can contribute to a number of diseases, including cancer. We screened 1408 NTP-provided substances in 1536-well qHTS format at concentrations ranging fr...

  12. [Genetics and genetic counseling].

    Science.gov (United States)

    Izzi, Claudia; Liut, Francesca; Dallera, Nadia; Mazza, Cinzia; Magistroni, Riccardo; Savoldi, Gianfranco; Scolari, Francesco

    2016-01-01

    Autosomal Dominant Polycystic Kidney Disease (ADPKD) is the most frequent genetic disease, characterized by progressive development of bilateral renal cysts. Two causative genes have been identified: PKD1 and PKD2. ADPKD phenotype is highly variable. Typically, ADPKD is an adult onset disease. However, occasionally, ADPKD manifests as very early onset disease. The phenotypic variability of ADPKD can be explained at three genetic levels: genic, allelic and gene modifier effects. Recent advances in molecular screening for PKD gene mutations and the introduction of the new next generation sequencing (NGS)- based genotyping approach have generated considerable improvement regarding the knowledge of genetic basis of ADPKD. The purpose of this article is to provide a comprehensive review of the genetics of ADPKD, focusing on new insights in genotype-phenotype correlation and exploring novel clinical approach to genetic testing. Evaluation of these new genetic information requires a multidisciplinary approach involving a nephrologist and a clinical geneticist. PMID:27067213

  13. PROLIFERATION AS A KEY EVENT IN DEVELOPMENTAL TOXICITY: "CHEMICAL SCREENING IN HUMAN NEURAL STEM CELLS USING HIGH CONTENT IMAGING

    Science.gov (United States)

    New toxicity testing approaches will rely on in vitro assays to assess chemical effects at the cellular and molecular level. Cell proliferation is imperative to normal development, and chemical disruption of this process can be detrimental to the organism. As part of an effort to...

  14. High-throughput screening of chemical effects on steroidogenesis using H295R human adrenocortical carcinoma cells

    Science.gov (United States)

    Disruption of steroidogenesis by environmental chemicals can result in altered hormone levels causing adverse reproductive and developmental effects. A high-throughput assay using H295R human adrenocortical carcinoma cells was used to evaluate the effect of 2,060 chemical samples...

  15. Prioritizing Environmental Chemicals for Obesity and Diabetes Outcomes Research: A Screening Approach Using ToxCast High Throughput Data

    Science.gov (United States)

    Background: Diabetes and obesity are major threats to public health in the US and abroad. Understanding the role chemicals in our environment play in the development of these conditions is an emerging issue in environmental health, although identifying and prioritizing chemicals ...

  16. Validation of a qualitative screening method for pesticides in fruits and vegetables by gas chromatography quadrupole-time of flight mass spectrometry with atmospheric pressure chemical ionization

    International Nuclear Information System (INIS)

    Highlights: • Applicability of GC-(APCI)QTOF MS as new tool for wide-scope screening of pesticides in fruits and vegetables demonstrated. • Validation of screening method according to SANCO/12571/2013. • Detection of the pesticides based on the presence of M+·/MH+ in most cases. • Screening detection limit 0.01 mg kg−1 for 77% of the pesticides investigated. • Successful identification at 0.01 mg kg−1 for 70% of the pesticides/matrix combinations. - Abstract: A wide-scope screening method was developed for the detection of pesticides in fruit and vegetables. The method was based on gas chromatography coupled to a hybrid quadrupole time-of-flight mass spectrometer with an atmospheric pressure chemical ionization source (GC-(APCI)QTOF MS). A non-target acquisition was performed through two alternating scan events: one at low collision energy and another at a higher collision energy ramp (MSE). In this way, both protonated molecule and/or molecular ion together with fragment ions were obtained in a single run. Validation was performed according to SANCO/12571/2013 by analysing 20 samples (10 different commodities in duplicate), fortified with a test set of 132 pesticides at 0.01, 0.05 and 0.20 mg kg−1. For screening, the detection was based on one diagnostic ion (in most cases the protonated molecule). Overall, at the 0.01 mg kg−1 level, 89% of the 2620 fortifications made were detected. The screening detection limit for individual pesticides was 0.01 mg kg−1 for 77% of the pesticides investigated. The possibilities for identification according to the SANCO criteria, requiring two ions with a mass accuracy ≤±5 ppm and an ion-ratio deviation ≤±30%, were investigated. At the 0.01 mg kg−1 level, identification was possible for 70% of the pesticides detected during screening. This increased to 87% and 93% at the 0.05 and 0.20 mg kg−1 level, respectively. Insufficient sensitivity for the second ion was the main reason for the inability to

  17. Validation of a qualitative screening method for pesticides in fruits and vegetables by gas chromatography quadrupole-time of flight mass spectrometry with atmospheric pressure chemical ionization

    Energy Technology Data Exchange (ETDEWEB)

    Portolés, T. [Research Institute for Pesticides and Water, University Jaume I, 12071 Castellón (Spain); RIKILT Institute of Food Safety, Wageningen University and Research Centre, Akkermaalsbos 2, 6708 WB Wageningen (Netherlands); Mol, J.G.J. [RIKILT Institute of Food Safety, Wageningen University and Research Centre, Akkermaalsbos 2, 6708 WB Wageningen (Netherlands); Sancho, J.V.; López, Francisco J. [Research Institute for Pesticides and Water, University Jaume I, 12071 Castellón (Spain); Hernández, F., E-mail: hernandf@uji.es [Research Institute for Pesticides and Water, University Jaume I, 12071 Castellón (Spain)

    2014-08-01

    Highlights: • Applicability of GC-(APCI)QTOF MS as new tool for wide-scope screening of pesticides in fruits and vegetables demonstrated. • Validation of screening method according to SANCO/12571/2013. • Detection of the pesticides based on the presence of M+·/MH+ in most cases. • Screening detection limit 0.01 mg kg{sup −1} for 77% of the pesticides investigated. • Successful identification at 0.01 mg kg{sup −1} for 70% of the pesticides/matrix combinations. - Abstract: A wide-scope screening method was developed for the detection of pesticides in fruit and vegetables. The method was based on gas chromatography coupled to a hybrid quadrupole time-of-flight mass spectrometer with an atmospheric pressure chemical ionization source (GC-(APCI)QTOF MS). A non-target acquisition was performed through two alternating scan events: one at low collision energy and another at a higher collision energy ramp (MS{sup E}). In this way, both protonated molecule and/or molecular ion together with fragment ions were obtained in a single run. Validation was performed according to SANCO/12571/2013 by analysing 20 samples (10 different commodities in duplicate), fortified with a test set of 132 pesticides at 0.01, 0.05 and 0.20 mg kg{sup −1}. For screening, the detection was based on one diagnostic ion (in most cases the protonated molecule). Overall, at the 0.01 mg kg{sup −1} level, 89% of the 2620 fortifications made were detected. The screening detection limit for individual pesticides was 0.01 mg kg{sup −1} for 77% of the pesticides investigated. The possibilities for identification according to the SANCO criteria, requiring two ions with a mass accuracy ≤±5 ppm and an ion-ratio deviation ≤±30%, were investigated. At the 0.01 mg kg{sup −1} level, identification was possible for 70% of the pesticides detected during screening. This increased to 87% and 93% at the 0.05 and 0.20 mg kg{sup −1} level, respectively. Insufficient sensitivity for the second

  18. Top Value Added Chemicals From Biomass: I. Results of Screening for Potential Candidates from Sugars and Synthesis Gas

    Energy Technology Data Exchange (ETDEWEB)

    Werpy, Todd A.; Holladay, John E.; White, James F.

    2004-11-01

    This report identifies twelve building block chemicals that can be produced from sugars via biological or chemical conversions. The twelve building blocks can be subsequently converted to a number of high-value bio-based chemicals or materials. Building block chemicals, as considered for this analysis, are molecules with multiple functional groups that possess the potential to be transformed into new families of useful molecules. The twelve sugar-based building blocks are 1,4-diacids (succinic, fumaric and malic), 2,5-furan dicarboxylic acid, 3-hydroxy propionic acid, aspartic acid, glucaric acid, glutamic acid, itaconic acid, levulinic acid, 3-hydroxybutyrolactone, glycerol, sorbitol, and xylitol/arabinitol. In addition to building blocks, the report outlines the central technical barriers that are preventing the widespread use of biomass for products and chemicals.

  19. CHEMICALS

    CERN Multimedia

    Medical Service

    2002-01-01

    It is reminded that all persons who use chemicals must inform CERN's Chemistry Service (TIS-GS-GC) and the CERN Medical Service (TIS-ME). Information concerning their toxicity or other hazards as well as the necessary individual and collective protection measures will be provided by these two services. Users must be in possession of a material safety data sheet (MSDS) for each chemical used. These can be obtained by one of several means : the manufacturer of the chemical (legally obliged to supply an MSDS for each chemical delivered) ; CERN's Chemistry Service of the General Safety Group of TIS ; for chemicals and gases available in the CERN Stores the MSDS has been made available via EDH either in pdf format or else via a link to the supplier's web site. Training courses in chemical safety are available for registration via HR-TD. CERN Medical Service : TIS-ME :73186 or service.medical@cern.ch Chemistry Service : TIS-GS-GC : 78546

  20. Acanthocephalan fish parasites (Rhadinorhynchidae Lühe, 1912) as potential biomarkers: Molecular-chemical screening by pyrolysis-field ionization mass spectrometry

    Science.gov (United States)

    Kleinertz, S.; Eckhardt, K.-U.; Theisen, S.; Palm, H. W.; Leinweber, P.

    2016-07-01

    The present study represents the first molecular-chemical screening by pyrolysis-field ionization mass spectrometry applied on fish parasites. A total of 71 fishes from Balinese fish markets, 36 Auxis rochei (Risso, 1810) and 35 A. thazard (Lacepède, 1800), were studied for their acanthocephalan parasites. This is the first record of Rhadinorhynchus zhukovi in Balinese waters, Indonesia, and we describe for the first time A. rochei and A. thazard as R. zhukovi hosts. Using this method, small scale variations within the chemical compounds of acanthocephalans could be detected. Using this methodology it will be possible to generate additional, pollutant specific information from aquatic habitats in future with the potential of a new bioindicator application for parasite/host origin and/or environmental pollution.

  1. QSAR model for human pregnane X receptor (PXR) binding: Screening of environmental chemicals and correlations with genotoxicity, endocrine disruption and teratogenicity

    Energy Technology Data Exchange (ETDEWEB)

    Dybdahl, Marianne, E-mail: mdyb@food.dtu.dk; Nikolov, Nikolai G.; Wedebye, Eva Bay; Jónsdóttir, Svava Ósk; Niemelä, Jay R.

    2012-08-01

    The pregnane X receptor (PXR) has a key role in regulating the metabolism and transport of structurally diverse endogenous and exogenous compounds. Activation of PXR has the potential to initiate adverse effects, causing drug–drug interactions, and perturbing normal physiological functions. Therefore, identification of PXR ligands would be valuable information for pharmaceutical and toxicological research. In the present study, we developed a quantitative structure–activity relationship (QSAR) model for the identification of PXR ligands using data based on a human PXR binding assay. A total of 631 molecules, representing a variety of chemical structures, constituted the training set of the model. Cross-validation of the model showed a sensitivity of 82%, a specificity of 85%, and a concordance of 84%. The developed model provided knowledge about molecular descriptors that may influence the binding of molecules to PXR. The model was used to screen a large inventory of environmental chemicals, of which 47% was found to be within domain of the model. Approximately 35% of the chemicals within domain were predicted to be PXR ligands. The predicted PXR ligands were found to be overrepresented among chemicals predicted to cause adverse effects, such as genotoxicity, teratogenicity, estrogen receptor activation and androgen receptor antagonism compared to chemicals not causing these effects. The developed model may be useful as a tool for predicting potential PXR ligands and for providing mechanistic information of toxic effects of chemicals. -- Highlights: ► Global QSAR model for the identification of PXR ligands was developed. ► Molecular descriptors that may influence PXR binding were identified. ► 35% of a large set of environmental chemicals were predicted to be PXR ligands. ► Predicted PXR binding was associated with various adverse effects.

  2. A genetic screen identifies BRCA2 and PALB2 as key regulators of G2 checkpoint maintenance

    DEFF Research Database (Denmark)

    Menzel, Tobias; Nähse-Kumpf, Viola; Kousholt, Arne Nedergaard;

    2011-01-01

    To identify key connections between DNA-damage repair and checkpoint pathways, we performed RNA interference screens for regulators of the ionizing radiation-induced G2 checkpoint, and we identified the breast cancer gene BRCA2. The checkpoint was also abrogated following depletion of PALB2......, an interaction partner of BRCA2. BRCA2 and PALB2 depletion led to premature checkpoint abrogation and earlier activation of the AURORA A-PLK1 checkpoint-recovery pathway. These results indicate that the breast cancer tumour suppressors and homologous recombination repair proteins BRCA2 and PALB2 are main...

  3. BIOCHEMICAL GENETIC STUDIES ON CUTYLEFISH SEPIELLA MAINDRONI (CEPHALOPODA: SEPIIDAE)——ACTIVE LOCI SCREENING OF ISOZYME

    Institute of Scientific and Technical Information of China (English)

    郑小东; YutakaNatsukari; 王如才; 王昭萍; 李云

    2001-01-01

    Screening of 46 putative enzyme-coding loci and 4 different kinds of tissues of Sepiella maindroni de Rochebrone, 1884 for enzymatic activities using starch gel electrophoretic technique proved that the 21 enzymes such as AAT, AK, ALP, AP, CK, DIA, ES, FBP, G3PDH, GPI, GRS,IDH, LDH, MDH, MEP, MPI, NP, PGDH, PGM, SOD and XO* , were active to Sepiella maindroni after being stained. The tissue exhibiting stable and clear bands was also determined. Among tissues tested, mantle muscle tissue was the best for electrophoretic survey of isozymes. Buccal bulb muscle, eye and liver were fairly good for some special enzymes, such as DIA, ES, MPI, NT, etc.

  4. Mycobacterium avium restriction fragment lenght polymorphism-IS IS1245 and the simple double repetitive element polymerase chain reaction typing method to screen genetic diversity in Brazilian strains

    Directory of Open Access Journals (Sweden)

    Patrícia Carvalho de Sequeira

    2005-11-01

    Full Text Available Simple double repetitive element polymerase chain reaction (MaDRE-PCR and Pvu II-IS1245 restriction fragment length polymorphism (RFLP typing methods were used to type 41 Mycobacterium avium isolates obtained from 14 Aids inpatients and 10 environment and animals specimens identified among 53 mycobacteria isolated from 237 food, chicken, and pig. All environmental and animals strains showed orphan patterns by both methods. By MaDRE-PCR four patients, with multiple isolates, showed different patterns, suggesting polyclonal infection that was confirmed by RFLP in two of them. This first evaluation of MaDRE-PCR on Brazilian M. avium strains demonstrated that the method seems to be useful as simple and less expensive typing method for screening genetic diversity in M. avium strains on selected epidemiological studies, although with limitation on analysis identical patterns except for one band.

  5. Application of DNA Hybridization Biosensor as a Screening Method for the Detection of Genetically Modified Food Components

    Directory of Open Access Journals (Sweden)

    Marian Filipiak

    2008-03-01

    Full Text Available An electrochemical biosensor for the detection of genetically modified food components is presented. The biosensor was based on 21-mer single-stranded oligonucleotide (ssDNA probe specific to either 35S promoter or nos terminator, which are frequently present in transgenic DNA cassettes. ssDNA probe was covalently attached by 5’-phosphate end to amino group of cysteamine self-assembled monolayer (SAM on gold electrode surface with the use of activating reagents – water soluble 1-ethyl-3(3’- dimethylaminopropyl-carbodiimide (EDC and N-hydroxy-sulfosuccinimide (NHS. The hybridization reaction on the electrode surface was detected via methylene blue (MB presenting higher affinity to ssDNA probe than to DNA duplex. The electrode modification procedure was optimized using 19-mer oligoG and oligoC nucleotides. The biosensor enabled distinction between DNA samples isolated from soybean RoundupReady® (RR soybean and non-genetically modified soybean. The frequent introduction of investigated DNA sequences in other genetically modified organisms (GMOs give a broad perspectives for analytical application of the biosensor.

  6. A Hybrid Improved Genetic Algorithm and Its Application in Dynamic Optimization Problems of Chemical Processes

    Institute of Scientific and Technical Information of China (English)

    SUN Fan; DU Wenli; QI Rongbin; QIAN Feng; ZHONG Weimin

    2013-01-01

    The solutions of dynamic optimization problems are usually very difficult due to their highly nonlinear and multidimensional nature.Genetic algorithm(GA)has been proved to be a feasible method when the gradient is difficult to calculate.Its advantage is that the control profiles at all time stages are optimized simultaneously,but its convergence is very slow in the later period of evolution and it is easily trapped in the local optimum.In this study,a hybrid improved genetic algorithm(HIGA)for solving dynamic optimization problems is proposed to overcome these defects.Simplex method(SM)is used to perform the local search in the neighborhood of the optimal solution.By using SM,the ideal searching direction of global optimal solution could be found as soon as possible and the convergence speed of the algorithm is improved.The hybrid algorithm presents some improvements,such as protecting the best individual,accepting immigrations,as well as employing adaptive crossover and Gaussian mutation operators.The efficiency of the proposed algorithm is demonstrated by solving several dynamic optimization problems.At last,HIGA is applied to the optimal production of secreted protein in a fed batch reactor and the optimal feed-rate found by HIGA is effective and relatively stable.

  7. A Genetic Screen Identifies a Critical Role for the WDR81-WDR91 Complex in the Trafficking and Degradation of Tetherin.

    Science.gov (United States)

    Rapiteanu, Radu; Davis, Luther J; Williamson, James C; Timms, Richard T; Paul Luzio, J; Lehner, Paul J

    2016-08-01

    Tetherin (BST2/CD317) is a viral restriction factor that anchors enveloped viruses to host cells and limits viral spread. The HIV-1 Vpu accessory protein counteracts tetherin by decreasing its cell surface expression and targeting it for ubiquitin-dependent endolysosomal degradation. Although the Vpu-mediated downregulation of tetherin has been extensively studied, the molecular details are not completely elucidated. We therefore used a forward genetic screen in human haploid KBM7 cells to identify novel genes required for tetherin trafficking. Our screen identified WDR81 as a novel gene required for tetherin trafficking and degradation in both the presence and absence of Vpu. WDR81 is a BEACH-domain containing protein that is also required for the degradation of EGF-stimulated epidermal growth factor receptor (EGFR) and functions in a complex with the WDR91 protein. In the absence of WDR81 the endolysosomal compartment appears swollen, with enlarged early and late endosomes and reduced delivery of endocytosed dextran to cathepsin-active lysosomes. Our data suggest a role for the WDR81-WDR91 complex in the fusion of endolysosomal compartments and the absence of WDR81 leads to impaired receptor trafficking and degradation. PMID:27126989

  8. Development and validation of real-time PCR screening methods for detection of cry1A.105 and cry2Ab2 genes in genetically modified organisms.

    Science.gov (United States)

    Dinon, Andréia Z; Prins, Theo W; van Dijk, Jeroen P; Arisi, Ana Carolina M; Scholtens, Ingrid M J; Kok, Esther J

    2011-05-01

    Primers and probes were developed for the element-specific detection of cry1A.105 and cry2Ab2 genes, based on their DNA sequence as present in GM maize MON89034. Cry genes are present in many genetically modified (GM) plants and they are important targets for developing GMO element-specific detection methods. Element-specific methods can be of use to screen for the presence of GMOs in food and feed supply chains. Moreover, a combination of GMO elements may indicate the potential presence of unapproved GMOs (UGMs). Primer-probe combinations were evaluated in terms of specificity, efficiency and limit of detection. Except for specificity, the complete experiment was performed in 9 PCR runs, on 9 different days and by testing 8 DNA concentrations. The results showed a high specificity and efficiency for cry1A.105 and cry2Ab2 detection. The limit of detection was between 0.05 and 0.01 ng DNA per PCR reaction for both assays. These data confirm the applicability of these new primer-probe combinations for element detection that can contribute to the screening for GM and UGM crops in food and feed samples.

  9. A genetic screen for modifiers of Drosophila caspase Dcp-1 reveals caspase involvement in autophagy and novel caspase-related genes

    Directory of Open Access Journals (Sweden)

    Ahnn Joohong

    2010-01-01

    Full Text Available Abstract Background Caspases are cysteine proteases with essential functions in the apoptotic pathway; their proteolytic activity toward various substrates is associated with the morphological changes of cells. Recent reports have described non-apoptotic functions of caspases, including autophagy. In this report, we searched for novel modifiers of the phenotype of Dcp-1 gain-of-function (GF animals by screening promoter element- inserted Drosophila melanogaster lines (EP lines. Results We screened ~15,000 EP lines and identified 72 Dcp-1-interacting genes that were classified into 10 groups based on their functions and pathways: 4 apoptosis signaling genes, 10 autophagy genes, 5 insulin/IGF and TOR signaling pathway genes, 6 MAP kinase and JNK signaling pathway genes, 4 ecdysone signaling genes, 6 ubiquitination genes, 11 various developmental signaling genes, 12 transcription factors, 3 translation factors, and 11 other unclassified genes including 5 functionally undefined genes. Among them, insulin/IGF and TOR signaling pathway, MAP kinase and JNK signaling pathway, and ecdysone signaling are known to be involved in autophagy. Together with the identification of autophagy genes, the results of our screen suggest that autophagy counteracts Dcp-1-induced apoptosis. Consistent with this idea, we show that expression of eGFP-Atg5 rescued the eye phenotype caused by Dcp-1 GF. Paradoxically, we found that over-expression of full-length Dcp-1 induced autophagy, as Atg8b-GFP, an indicator of autophagy, was increased in the eye imaginal discs and in the S2 cell line. Taken together, these data suggest that autophagy suppresses Dcp-1-mediated apoptotic cell death, whereas Dcp-1 positively regulates autophagy, possibly through feedback regulation. Conclusions We identified a number of Dcp-1 modifiers that genetically interact with Dcp-1-induced cell death. Our results showing that Dcp-1 and autophagy-related genes influence each other will aid future

  10. A haploid genetic screen identifies the G1/S regulatory machinery as a determinant of Wee1 inhibitor sensitivity

    NARCIS (Netherlands)

    Heijink, Anne Margriet; Blomen, Vincent A.; Bisteau, Xavier; Degener, Fabian; Matsushita, Felipe Yu; Kaldis, Philipp; Foijer, Floris; van Vugt, Marcel A. T. M.

    2015-01-01

    The Wee1 cell cycle checkpoint kinase prevents premature mitotic entry by inhibiting cyclin-dependent kinases. Chemical inhibitors of Wee1 are currently being tested clinically as targeted anticancer drugs. Wee1 inhibition is thought to be preferentially cytotoxic in p53-defective cancer cells. Howe

  11. A haploid genetic screen identifies the G(1)/S regulatory machinery as a determinant of Wee1 inhibitor sensitivity

    NARCIS (Netherlands)

    Heijink, Anne Margriet; Blomen, Vincent A.; Bisteau, Xavier; Degener, Fabian; Matsushita, Felipe Yu; Kaldis, Philipp; Foijer, Floris; van Vugt, Marcel A. T. M.

    2015-01-01

    The Wee1 cell cycle checkpoint kinase prevents premature mitotic entry by inhibiting cyclin-dependent kinases. Chemical inhibitors of Wee1 are currently being tested clinically as targeted anticancer drugs. Wee1 inhibition is thought to be preferentially cytotoxic in p53-defective cancer cells. Howe

  12. Neuroblastoma Screening

    Science.gov (United States)

    ... Health Professional Neuroblastoma Treatment Neuroblastoma Screening Research Neuroblastoma Screening (PDQ®)–Patient Version What is screening? Go to Health Professional Version Screening is looking ...

  13. Validation of a qualitative screening method for pesticides in fruits and vegetables by gas chromatography quadrupole-time of flight mass spectrometry with atmospheric pressure chemical ionization.

    Science.gov (United States)

    Portolés, T; Mol, J G J; Sancho, J V; López, Francisco J; Hernández, F

    2014-08-01

    A wide-scope screening method was developed for the detection of pesticides in fruit and vegetables. The method was based on gas chromatography coupled to a hybrid quadrupole time-of-flight mass spectrometer with an atmospheric pressure chemical ionization source (GC-(APCI)QTOF MS). A non-target acquisition was performed through two alternating scan events: one at low collision energy and another at a higher collision energy ramp (MS(E)). In this way, both protonated molecule and/or molecular ion together with fragment ions were obtained in a single run. Validation was performed according to SANCO/12571/2013 by analysing 20 samples (10 different commodities in duplicate), fortified with a test set of 132 pesticides at 0.01, 0.05 and 0.20mg kg(-1). For screening, the detection was based on one diagnostic ion (in most cases the protonated molecule). Overall, at the 0.01mg kg(-1) level, 89% of the 2620 fortifications made were detected. The screening detection limit for individual pesticides was 0.01mg kg(-1) for 77% of the pesticides investigated. The possibilities for identification according to the SANCO criteria, requiring two ions with a mass accuracy ≤±5ppm and an ion-ratio deviation ≤±30%, were investigated. At the 0.01mg kg(-1) level, identification was possible for 70% of the pesticides detected during screening. This increased to 87% and 93% at the 0.05 and 0.20mg kg(-1) level, respectively. Insufficient sensitivity for the second ion was the main reason for the inability to identify detected pesticides, followed by deviations in mass accuracy and ion ratios.

  14. Chemical and genetic diversity of Zataria multiflora Boiss. accessions growing wild in Iran.

    Science.gov (United States)

    Hadian, Javad; Ebrahimi, Samad Nejad; Mirjalili, Mohammad Hossein; Azizi, Ali; Ranjbar, Hamid; Friedt, Wolfgang

    2011-01-01

    Zataria multiflora Boiss. is an aromatic shrub belonging to the Lamiaceae family. Its aerial parts are used in the traditional medicine and in the pharmaceutical and food industries. The terpenoid and genetic profiles of 18 accessions of Z. multiflora, collected in different locations in Iran, have been analyzed by GC/FID and GC/MS or by AFLP (amplified fragment length polymorphism) analyses, respectively. Altogether, 56 compounds were identified in the essential oils, with the major constituents being thymol (6.0-54.9%), carvacrol (0.7-50.6%), linalool (1.2-46.8%), and p-cymene (1.6-14.8%). On the basis of the essential-oil composition, the 18 accessions were divided into four groups. The first group was characterized by a high content of carvacrol, thymol, and linalool, the second group was dominated by carvacrol, thymol, and p-cymene, the third group was characterized by a high concentration of thymol and a low content of carvacrol and p-cymene, and the forth group contained linalool and carvacrol as the main components. The AFLP results revealed that the average genetic similarity (GS) between the accessions was 0.61, ranging from 0.40 to 0.77. The UPGMA (unweighted pair-group method with arithmetic mean) cluster analysis divided all accessions into five groups at a similarity level of 0.60. The two clusters generated, the first based on the essential-oil compositions and the second on the AFLP data, showed a different pattern of relationships among the accessions. The knowledge of the Z. multiflora chemotype diversity, illustrated in this study, will allow an improvement of the homogeneity of the plant material for the production of different types of essential oils, depending on the demands of the pharmaceutical and food industries for specific uses. PMID:21259428

  15. Fungi as chemical industries and genetic engineering for the production of biologically active secondary metabolites

    Institute of Scientific and Technical Information of China (English)

    Abid; Ali; Khan; Nafees; Bacha; Bashir; Ahmad; Ghosia; Lutfullah; Umar; Farooq; Russell; John; Cox

    2014-01-01

    Fungi is somewhere in between the micro and macro organisms which is a good source of producing biologically active secondary metabolites.Fungi have been used as tool for producing different types of secondary metabolites by providing different nutrients at different laboratory conditions.The fungi have been engineered for the desired secondary metabolites by using different laboratory techniques,for example,homologous and heterologous expressions.This review reported how the fungi are used as chemical industry for the production of secondary metabolites and how they are engineered in laboratory for the production of desirable metabolites:also the biosynthetic pathways of the bio-organic-molecules were reported.

  16. Fungi as chemical industries and genetic engineering for the production of biologically active secondary metabolites

    Institute of Scientific and Technical Information of China (English)

    Abid Ali Khan; Nafees Bacha; Bashir Ahmad; Ghosia Lutfullah; Umar Farooq; Russell John Cox

    2014-01-01

    Fungi is somewhere in between the micro and macro organisms which is a good source of producing biologically active secondary metabolites. Fungi have been used as tool for producing different types of secondary metabolites by providing different nutrients at different laboratory conditions. The fungi have been engineered for the desired secondary metabolites by using different laboratory techniques, for example, homologous and heterologous expressions. This review reported how the fungi are used as chemical industry for the production of secondary metabolites and how they are engineered in laboratory for the production of desirable metabolites;also the biosynthetic pathways of the bio-organic-molecules were reported.

  17. Brush and Spray: A High-Throughput Systemic Acquired Resistance Assay Suitable for Large-Scale Genetic Screening1[W][OA

    Science.gov (United States)

    Jing, Beibei; Xu, Shaohua; Xu, Mo; Li, Yan; Li, Shuxin; Ding, Jinmei; Zhang, Yuelin

    2011-01-01

    Systemic acquired resistance (SAR) is a defense mechanism induced in the distal parts of plants after primary infection. It confers long-lasting protection against a broad spectrum of microbial pathogens. Lack of high-throughput assays has hampered the forward genetic analysis of SAR. Here, we report the development of an easy and efficient assay for SAR and its application in a forward genetic screen for SAR-deficient mutants in Arabidopsis (Arabidopsis thaliana). Using the new assay for SAR, we identified six flavin-dependent monooxygenase1, four AGD2-like defense response protein1, three salicylic acid induction-deficient2, one phytoalexin deficient4, and one avrPphB-susceptible3 alleles as well as a gain-of-function mutant of CALMODULIN-BINDING TRANSCRIPTION ACTIVATOR3 designated camta3-3D. Like transgenic plants overexpressing CAMTA3, camta3-3D mutant plants exhibit compromised SAR and enhanced susceptibility to virulent pathogens, suggesting that CAMTA3 is a critical regulator of both basal resistance and SAR. PMID:21900483

  18. Genome-wide genetic screen identified the link between dG9a and epidermal growth factor receptor signaling pathway in vivo.

    Science.gov (United States)

    Shimaji, Kouhei; Konishi, Takahiro; Yoshida, Hideki; Kimura, Hiroshi; Yamaguchi, Masamitsu

    2016-08-01

    G9a is one of the histone H3 Lys 9 (H3K9) specific methyltransferases first identified in mammals. Drosophila G9a (dG9a) has been reported to induce H3K9 dimethylation in vivo, and the target genes of dG9a were identified during embryonic and larval stages. Although dG9a is important for a variety of developmental processes, the link between dG9a and signaling pathways are not addressed yet. Here, by genome-wide genetic screen, taking advantage of the rough eye phenotype of flies that over-express dG9a in eye discs, we identified 16 genes that enhanced the rough eye phenotype induced by dG9a over-expression. These 16 genes included Star, anterior open, bereft and F-box and leucine-rich repeat protein 6 which are components of epidermal growth factor receptor (EGFR) signaling pathway. When dG9a over-expression was combined with mutation of Star, differentiation of R7 photoreceptors in eye imaginal discs as well as cone cells and pigment cells in pupal retinae was severely inhibited. Furthermore, the dG9a over-expression reduced the activated ERK signals in eye discs. These data demonstrate a strong genetic link between dG9a and the EGFR signaling pathway. PMID:27343629

  19. Application of DNA Hybridization Biosensor as a Screening Method for the Detection of Genetically Modified Food Components

    OpenAIRE

    Marian Filipiak; Marta Ligaj; Mariusz Tichoniuk

    2008-01-01

    An electrochemical biosensor for the detection of genetically modified food components is presented. The biosensor was based on 21-mer single-stranded oligonucleotide (ssDNA probe) specific to either 35S promoter or nos terminator, which are frequently present in transgenic DNA cassettes. ssDNA probe was covalently attached by 5’-phosphate end to amino group of cysteamine self-assembled monolayer (SAM) on gold electrode surface with the use of activating reagents – water soluble 1-e...

  20. Do recent US Supreme Court rulings on patenting of genes and genetic diagnostics affect the practice of genetic screening and diagnosis in prenatal and reproductive care?

    Science.gov (United States)

    Chandrasekharan, Subhashini; McGuire, Amy L; Van den Veyver, Ignatia B

    2014-10-01

    Thousands of patents have been awarded that claim human gene sequences and their uses, and some have been challenged in court. In a recent high-profile case, Association for Molecular Pathology, et al. v. Myriad Genetics, Inc., et al., the US Supreme Court ruled that genes are natural occurring substances and therefore not patentable through 'composition of matter' claims. The consequences of this ruling will extend well beyond ending Myriad's monopoly over BRCA testing and may affect similar monopolies of other commercial laboratories for tests involving other genes. It could also simplify intellectual property issues surrounding genome-wide clinical sequencing, which can generate results for genes covered by intellectual property. Non-invasive prenatal testing (NIPT) for common aneuploidies using cell-free fetal (cff) DNA in maternal blood is currently offered through commercial laboratories and is also the subject of ongoing patent litigation. The recent Supreme Court decision in the Myriad case has already been invoked by a lower district court in NIPT litigation and resulted in invalidation of primary claims in a patent on currently marketed cffDNA-based testing for chromosomal aneuploidies.

  1. Genetic and chemical characterization of ibuprofen degradation by Sphingomonas Ibu-2.

    Science.gov (United States)

    Murdoch, Robert W; Hay, Anthony G

    2013-03-01

    Sphingomonas Ibu-2 has the unusual ability to cleave the acid side chain from the pharmaceutical ibuprofen and related arylacetic acid derivatives to yield corresponding catechols under aerobic conditions via a previously uncharacterized mechanism. Screening a chromosomal library of Ibu-2 DNA in Escherichia coli EPI300 allowed us to identify one fosmid clone (pFOS3G7) that conferred the ability to metabolize ibuprofen to isobutylcatechol. Characterization of pFOS3G7 loss-of-function transposon mutants permitted identification of five ORFs, ipfABDEF, whose predicted amino acid sequences bore similarity to the large and small units of an aromatic dioxygenase (ipfAB), a sterol carrier protein X (SCPx) thiolase (ipfD), a domain of unknown function 35 (DUF35) protein (ipfE) and an aromatic CoA ligase (ipfF). Two additional ORFs, ipfH and ipfI, which encode putative ferredoxin reductase and ferredoxin components of an aromatic dioxygenase system, respectively, were also identified on pFOS3G7. Complementation of a markerless loss-of-function ipfD deletion mutant restored catechol production as did complementation of the ipfF Tn mutant. Expression of subcloned ipfABDEF alone in E. coli did not impart full metabolic activity unless coexpressed with ipfHI. CoA ligation followed by ring oxidation is common to phenylacetic acid pathways. However, the need for a putative SCPx thiolase (IpfD) and DUF35 protein (IpfE) in aerobic arylacetic acid degradation is unprecedented. This work provides preliminary insights into the mechanism behind this novel arylacetic acid-deacylating, catechol-generating activity.

  2. Comprehensive predictions of target proteins based on protein-chemical interaction using virtual screening and experimental verifications

    Directory of Open Access Journals (Sweden)

    Kobayashi Hiroki

    2012-04-01

    Full Text Available Abstract Background Identification of the target proteins of bioactive compounds is critical for elucidating the mode of action; however, target identification has been difficult in general, mostly due to the low sensitivity of detection using affinity chromatography followed by CBB staining and MS/MS analysis. Results We applied our protocol of predicting target proteins combining in silico screening and experimental verification for incednine, which inhibits the anti-apoptotic function of Bcl-xL by an unknown mechanism. One hundred eighty-two target protein candidates were computationally predicted to bind to incednine by the statistical prediction method, and the predictions were verified by in vitro binding of incednine to seven proteins, whose expression can be confirmed in our cell system. As a result, 40% accuracy of the computational predictions was achieved successfully, and we newly found 3 incednine-binding proteins. Conclusions This study revealed that our proposed protocol of predicting target protein combining in silico screening and experimental verification is useful, and provides new insight into a strategy for identifying target proteins of small molecules.

  3. Inhibitors of ROS production by the ubiquinone-binding site of mitochondrial complex I identified by chemical screening.

    Science.gov (United States)

    Orr, Adam L; Ashok, Deepthi; Sarantos, Melissa R; Shi, Tong; Hughes, Robert E; Brand, Martin D

    2013-12-01

    Mitochondrial production of reactive oxygen species is often considered an unavoidable consequence of aerobic metabolism and currently cannot be manipulated without perturbing oxidative phosphorylation. Antioxidants are widely used to suppress effects of reactive oxygen species after formation, but they can never fully prevent immediate effects at the sites of production. To identify site-selective inhibitors of mitochondrial superoxide/H2O2 production that do not interfere with mitochondrial energy metabolism, we developed a robust small-molecule screen and secondary profiling strategy. We describe the discovery and characterization of a compound (N-cyclohexyl-4-(4-nitrophenoxy)benzenesulfonamide; CN-POBS) that selectively inhibits superoxide/H2O2 production from the ubiquinone-binding site of complex I (site I(Q)) with no effects on superoxide/H2O2 production from other sites or on oxidative phosphorylation. Structure/activity studies identified a core structure that is important for potency and selectivity for site I(Q). By employing CN-POBS in mitochondria respiring on NADH-generating substrates, we show that site I(Q) does not produce significant amounts of superoxide/H2O2 during forward electron transport on glutamate plus malate. Our screening platform promises to facilitate further discovery of direct modulators of mitochondrially derived oxidative damage and advance our ability to understand and manipulate mitochondrial reactive oxygen species production under both normal and pathological conditions. PMID:23994103

  4. An Improved Hybrid Genetic Algorithm for Chemical Plant Layout Optimization with Novel Non-overlapping and Toxic Gas Dispersion Constraints

    Institute of Scientific and Technical Information of China (English)

    XU Yuan; WANG Zhenyu; ZHU Qunxiong

    2013-01-01

    New approaches for facility distribution in chemical plants are proposed including an improved non-overlapping constraint based on projection relationships of facilities and a novel toxic gas dispersion constraint.In consideration of the large number of variables in the plant layout model,our new method can significantly reduce the number of variables with their own projection relationships.Also,as toxic gas dispersion is a usual incident in a chemical plant,a simple approach to describe the gas leakage is proposed,which can clearly represent the constraints of potential emission source and sitting facilities.For solving the plant layout model,an improved genetic algorithm (GA) based on infeasible solution fix technique is proposed,which improves the globe search ability of GA.The case study and experiment show that a better layout plan can be obtained with our method,and the safety factors such as gas dispersion and minimum distances can be well handled in the solution.

  5. Profiling Environmental Chemicals for Activity in the Antioxidant Response Element Signaling Pathway Using a High-Throughput Screening Approach

    Science.gov (United States)

    1 ABSTRACT 2 3 BACKGROUND: Oxidative stress has been implicated in the pathogenesis of a variety 4 of diseases ranging from cancer to neurodegeneration, highlighti.ng the need to identify 5 chemicals that can induce this effect. The antioxidant response element (ARE)...

  6. 78 FR 35922 - Endocrine Disruptor Screening Program; Final Second List of Chemicals and Substances for Tier 1...

    Science.gov (United States)

    2013-06-14

    ... to identify substances that have the potential to interact with the endocrine system (specifically... interfere with the endocrine systems of humans or other species, and it would be inappropriate to do so. In... endocrine system. The determination that a chemical does or is not likely to have the potential to...

  7. Lactose intolerance genetic testing: is it useful as routine screening? Results on 1426 south-central Italy patients.

    Science.gov (United States)

    Santonocito, Concetta; Scapaticci, Margherita; Guarino, Donatella; Annicchiarico, Eleonora Brigida; Lisci, Rosalia; Penitente, Romina; Gasbarrini, Antonio; Zuppi, Cecilia; Capoluongo, Ettore

    2015-01-15

    Adult-type hypolactasia is a widespread condition throughout the world, causing lactose malabsorption. Several studies suggested that the identification of C/T-13910 and G/A-22018 mutations, located upstream the gene encoding the lactase-phlorizin hydrolase (LPH), is a useful tool for the differential diagnosis of hypolactasia. We evaluated the frequencies of C/T-13910 and G/A-22018 variants in a central-south Italian population and the usefulness of lactase deficiency genetic testing in the clinic practice. The genomic DNA of 1426 patients and 1000 healthy controls from central-south Italy was isolated from peripheral whole blood and genotyped for the C/T-13910 and G/A-22018 polymorphisms by high-resolution melting analysis (HRMA) and sequencing. The frequencies of genotypes in the 1426 patients analysed were as follows: 1077 CC/GG (75.5%), 287 CT/GA (20.1%), 24 TT/AA (1.7%), 38 CC/GA (2.7%). Only 64 out of 1426 (4.5%) performed also L-BHT test, 29 of which were negative for L-BHT also in presence of different genotypes. Among the 35 individuals with L-BHT positive, 34 were CC/GG and only one CT/GA. Although lactose genetic test is a good predictor of persistence/non-persistence lactase in specific population, its use in the central-south Italy population should be limited given the high prevalence of the CCGG diplotype in normal individuals. PMID:25281930

  8. Using the Drosophila Melanogaster Genetics Reference Panel to Identify Toxicity Pathways for Toluene

    Science.gov (United States)

    Mechanistic information is needed to link effects of chemicals at molecular targets in high­ throughput screening assays to adverse outcomes in whole organisms. This study was designed to use the Drosophila Genetic Reference Panel (DGRP), a set of genetically well...

  9. A bow-tie genetic architecture for morphogenesis suggested by a genome-wide RNAi screen in Caenorhabditis elegans.

    Directory of Open Access Journals (Sweden)

    Matthew D Nelson

    2011-03-01

    Full Text Available During animal development, cellular morphogenesis plays a fundamental role in determining the shape and function of tissues and organs. Identifying the components that regulate and drive morphogenesis is thus a major goal of developmental biology. The four-celled tip of the Caenorhabditis elegans male tail is a simple but powerful model for studying the mechanism of morphogenesis and its spatiotemporal regulation. Here, through a genome-wide post-embryonic RNAi-feeding screen, we identified 212 components that regulate or participate in male tail tip morphogenesis. We constructed a working hypothesis for a gene regulatory network of tail tip morphogenesis. We found regulatory roles for the posterior Hox genes nob-1 and php-3, the TGF-β pathway, nuclear hormone receptors (e.g. nhr-25, the heterochronic gene blmp-1, and the GATA transcription factors egl-18 and elt-6. The majority of the pathways converge at dmd-3 and mab-3. In addition, nhr-25 and dmd-3/mab-3 regulate each others' expression, thus placing these three genes at the center of a complex regulatory network. We also show that dmd-3 and mab-3 negatively regulate other signaling pathways and affect downstream cellular processes such as vesicular trafficking (e.g. arl-1, rme-8 and rearrangement of the cytoskeleton (e.g. cdc-42, nmy-1, and nmy-2. Based on these data, we suggest that male tail tip morphogenesis is governed by a gene regulatory network with a bow-tie architecture.

  10. Chemical and genetic characterization of Phlomis species and wild hybrids in Crete.

    Science.gov (United States)

    Georgescu, Luciana; Stefanakis, Michalis K; Kokkini, Stella; Katerinopoulos, Haralambos E; Pirintsos, Stergios A

    2016-02-01

    The genus Phlomis is represented in the island of Crete (Greece, Eastern Mediterranean) by three species Phlomis cretica C. Presl., Phlomis fruticosa L., the island endemic Phlomis lanata Willd. and three hybrids Phlomis x cytherea Rech.f. (P. cretica x P. fruticosa), Phlomis x commixta Rech.f. (P. cretica x P. lanata) and Phlomis x sieberi Vierh. (P. fruticosa x P. lanata). This work describes (a) the profile of hybrids and parental species concerning their volatile compounds, (b) the suitability of ribosomal nuclear (ITS region), chloroplast (trnH-psbA), and AFLP markers to identify hybrids and (c) their competence to characterize the different chemotypes of both hybrids and their parental species. The cluster analysis and PCA constructed from chemical data (volatile oils) suggest that there are three groups of taxa. Group IA includes P. cretica and P. fruticosa, group IB includes P. x cytherea, whereas group II consists of P. x commixta, P. x sieberi and P. lanata. Volatile compounds detected only in the hybrids P. x sieberi and P. x commixta correspond to the 3% of the total compounds, value that is much higher in P. x cytherea (21%). Neighbor-joining, statistical parsimony analysis and the observations drawn from ribotypes spectrum of ITS markers divided Phlomis species in two groups, P. lanata and the complex P. cretica/P. fruticosa. In contrast to the ITS region, the plastid DNA marker follows a geographically related pattern. Neighbor-Net, PCA and Bayesian assignment analysis performed for AFLP markers separated the genotypes into three groups corresponding to populations of P. cretica, P. fruticosa, and P. lanata, respectively, while populations of P. x commixta, P. x cytherea, and P. x sieberi presented admixed ancestry. Most of the P. x cytherea samples were identified as F1 hybrids by Bayesian assignment test, while those of P. x commixta and P. x sieberi were identified as F2 hybrids. Overall, high chemical differentiation is revealed in one of the

  11. Canine toys and training devices as sources of exposure to phthalates and bisphenol A: quantitation of chemicals in leachate and in vitro screening for endocrine activity.

    Science.gov (United States)

    Wooten, Kimberly J; Smith, Philip N

    2013-11-01

    Chewing and mouthing behaviors exhibited by pet dogs are likely to lead to oral exposures to a variety of environmental chemicals. Products intended for chewing and mouthing uses include toys and training devices that are often made of plastics. The goal of the current study was to determine if a subset of phthalates and bisphenol A (BPA), endocrine disrupting chemicals commonly found in plastics, leach out of dog toys and training devices (bumpers) into synthetic canine saliva. In vitro assays were used to screen leachates for endocrine activity. Bumper leachates were dominated by di-2-ethylhexyl phthalate (DEHP) and BPA, with concentrations reaching low μg mL(-1) following short immersions in synthetic saliva. Simulated chewing of bumpers during immersion in synthetic saliva increased concentrations of phthalates and BPA as compared to new bumpers, while outdoor storage had variable effects on concentrations (increased DEHP; decreased BPA). Toys leached substantially lower concentrations of phthalates and BPA, with the exception of one toy which leached considerable amounts of diethyl phthalate. In vitro assays indicated anti-androgenic activity of bumper leachates, and estrogenic activity of both bumper and toy leachates. These results confirm that toys and training devices are potential sources of exposure to endocrine disrupting chemicals in pet dogs. PMID:24007620

  12. Colorectal Cancer Screening

    Science.gov (United States)

    ... laxatives to clear the colon, shows polyps clearly. DNA stool test This test checks DNA in stool cells for genetic changes that may be a sign of colorectal cancer. Screening clinical trials are taking place in many parts of the ... Screening tests have risks. False-negative test results can occur. ...

  13. State-of-the-art of screening methods for the rapid identification of chemicals in drinking water Deliverable D1

    OpenAIRE

    Llorca, Marta; Rodríguez-Mozaz, Sara

    2013-01-01

    The contamination of drinking water is potentially harmful and poses a risk to public health. If any observation suggests a potential contamination of drinking water, such as consumer complaints about the alteration of the water’s organoleptic properties, the appearance of health problems or an alarm triggered by sensors, a rapid identification of the hazard causing the problem is necessary. With regards to chemical contamination, EU Member States have several strategies to deal with the pres...

  14. Human Pluripotent Stem Cell Based Developmental Toxicity Assays for Chemical Safety Screening and Systems Biology Data Generation.

    Science.gov (United States)

    Shinde, Vaibhav; Klima, Stefanie; Sureshkumar, Perumal Srinivasan; Meganathan, Kesavan; Jagtap, Smita; Rempel, Eugen; Rahnenführer, Jörg; Hengstler, Jan Georg; Waldmann, Tanja; Hescheler, Jürgen; Leist, Marcel; Sachinidis, Agapios

    2015-01-01

    Efficient protocols to differentiate human pluripotent stem cells to various tissues in combination with -omics technologies opened up new horizons for in vitro toxicity testing of potential drugs. To provide a solid scientific basis for such assays, it will be important to gain quantitative information on the time course of development and on the underlying regulatory mechanisms by systems biology approaches. Two assays have therefore been tuned here for these requirements. In the UKK test system, human embryonic stem cells (hESC) (or other pluripotent cells) are left to spontaneously differentiate for 14 days in embryoid bodies, to allow generation of cells of all three germ layers. This system recapitulates key steps of early human embryonic development, and it can predict human-specific early embryonic toxicity/teratogenicity, if cells are exposed to chemicals during differentiation. The UKN1 test system is based on hESC differentiating to a population of neuroectodermal progenitor (NEP) cells for 6 days. This system recapitulates early neural development and predicts early developmental neurotoxicity and epigenetic changes triggered by chemicals. Both systems, in combination with transcriptome microarray studies, are suitable for identifying toxicity biomarkers. Moreover, they may be used in combination to generate input data for systems biology analysis. These test systems have advantages over the traditional toxicological studies requiring large amounts of animals. The test systems may contribute to a reduction of the costs for drug development and chemical safety evaluation. Their combination sheds light especially on compounds that may influence neurodevelopment specifically. PMID:26132533

  15. Screening and genetic diagnosis of childhood thalassemia in Sanya city%三亚市儿童地中海贫血筛查及基因分析

    Institute of Scientific and Technical Information of China (English)

    陈爱华; 陈垂海; 陈求珠; 周祥敏; 陈垂婉; 陈垂任; 霍开明

    2015-01-01

    ObjectiveTo investigate the incidence and gene mutation spectrum of childhood thalassemia in Sanya city. MethodsThere were 938 children in our hospital screened by routine blood test, C-reactive protein test and hemoglobinelectro-phoresis. Based on the screening results in children with thalassemia and its subtypes, the alpha thalassaemia children were diag-nosed by gap - polymerase chain reaction, while the beta thalassaemia ones were diagnosed by polymerase chain reaction-reverse dot blot, and all the results were retrospectively analyzed.ResultsScreening positive rate of thalassemia anemia was 13.65% (128/938), genetic diagnosis positive rate was 11.41% (107/938). In 107 cases diagnosed with thalassemia by genetic test, 59 were alpha thalassaemia, and 46 were beta thalassaemia, and 2 cases were alpha thalassaemia combined with beta thalassaemia. In 59 cases of alpha thalassaemia, 31 had genotype of - SEA/alpha alpha , 13 cases with genotype alpha 4.2 / alpha alpha, and 6 were of HbH disease. In the 46 cases of beta thalassaemia children, six gene locus mutations were found : CD41-42 (CTTT) in 21 cases, TATAbox - 28 (A - G) in 13 cases, CDs14/15 (G) in 5 cases, IVS - Int 654 (C - > T) in 4 cases, CD17 (A - > T) in 2 cases, and 27-28 (C) in 1 case.ConclusionHigh prevalence of thalassemia in children from Sanya city warrants genetic counseling, premarital check-up and prenatal diagnosis.%目的:调查三亚市儿童地中海贫血的发病率和基因突变类型。方法对938例儿童进行血常规、CRP检测和血红蛋白电泳试验,筛查地中海贫血发生情况及其所属类型;对α-地中海贫血患儿用gap-PCR法、β-地中海贫血患儿用PCR-RDB法进行基因诊断。结果地中海贫血筛查阳性率为13.65%(128/938),基因诊断阳性率为11.41%(107/938)。在107例经基因诊断确诊为地中海贫血的患儿中,α-地中海贫血59例,β-地中海贫血46例,α合并β地中海贫血2例。59例α-地中海

  16. The Rules of Aggression: How Genetic, Chemical and Spatial Factors Affect Intercolony Fights in a Dominant Species, the Mediterranean Acrobat Ant Crematogaster scutellaris.

    Directory of Open Access Journals (Sweden)

    Filippo Frizzi

    Full Text Available Nest-mate recognition plays a key role in the biology of ants. Although individuals coming from a foreign nest are, in most cases, promptly rejected, the degree of aggressiveness towards non nest-mates may be highly variable among species and relies on genetic, chemical and environmental factors. We analyzed intraspecific relationships among neighboring colonies of the dominant Mediterranean acrobat ant Crematogaster scutellaris integrating genetic, chemical and behavioral analyses. Colony structure, parental relationships between nests, cuticular hydrocarbons profiles (CHCs and aggressive behavior against non nest-mates were studied in 34 nests located in olive tree trunks. Bayesian clustering analysis of allelic variation at nine species-specific microsatellite DNA markers pooled nests into 14 distinct clusters, each representing a single colony, confirming a polydomous arrangement of nests in this species. A marked genetic separation among colonies was also detected, probably due to long distance dispersion of queens and males during nuptial flights. CHCs profiles varied significantly among colonies and between nests of the same colony. No relationship between CHCs profiles and genetic distances was detected. The level of aggressiveness between colonies was inversely related to chemical and spatial distance, suggesting a 'nasty neighbor' effect. Our findings also suggest that CHCs profiles in C. scutellaris may be linked to external environmental factors rather than genetic relationships.

  17. A genetic screen in zebrafish identifies the mutants vps18, nf2 and foie gras as models of liver disease.

    Science.gov (United States)

    Sadler, Kirsten C; Amsterdam, Adam; Soroka, Carol; Boyer, James; Hopkins, Nancy

    2005-08-01

    Hepatomegaly is a sign of many liver disorders. To identify zebrafish mutants to serve as models for hepatic pathologies, we screened for hepatomegaly at day 5 of embryogenesis in 297 zebrafish lines bearing mutations in genes that are essential for embryonic development. Seven mutants were identified, and three have phenotypes resembling different liver diseases. Mutation of the class C vacuolar protein sorting gene vps18 results in hepatomegaly associated with large, vesicle-filled hepatocytes, which we attribute to the failure of endosomal-lysosomal trafficking. Additionally, these mutants develop defects in the bile canaliculi and have marked biliary paucity, suggesting that vps18 also functions to traffic vesicles to the hepatocyte apical membrane and may play a role in the development of the intrahepatic biliary tree. Similar findings have been reported for individuals with arthrogryposis-renal dysfunction-cholestasis (ARC) syndrome, which is due to mutation of another class C vps gene. A second mutant, resulting from disruption of the tumor suppressor gene nf2, develops extrahepatic choledochal cysts in the common bile duct, suggesting that this gene regulates division of biliary cells during development and that nf2 may play a role in the hyperplastic tendencies observed in biliary cells in individuals with choledochal cysts. The third mutant is in the novel gene foie gras, which develops large, lipid-filled hepatocytes, resembling those in individuals with fatty liver disease. These mutants illustrate the utility of zebrafish as a model for studying liver development and disease, and provide valuable tools for investigating the molecular pathogenesis of congenital biliary disorders and fatty liver disease.

  18. RET and EDNRB mutation screening in patients with Hirschsprung disease: Functional studies and its implications for genetic counseling.

    Science.gov (United States)

    Widowati, Titis; Melhem, Shamiram; Patria, Suryono Y; de Graaf, Bianca M; Sinke, Richard J; Viel, Martijn; Dijkhuis, Jos; Sadewa, Ahmad H; Purwohardjono, Rochadi; Soenarto, Yati; Hofstra, Robert Mw; Sribudiani, Yunia

    2016-06-01

    Hirschsprung disease (HSCR) is a major cause of chronic constipation in children. HSCR can be caused by germline mutations in RET and EDNRB. Defining causality of the mutations identified is difficult and almost exclusively based on in silico predictions. Therefore, the reported frequency of pathogenic mutations might be overestimated. We combined mutation analysis with functional assays to determine the frequencies of proven pathogenic RET and EDNRB mutations in HSCR. We sequenced RET and EDNRB in 57 HSCR patients. The identified RET-coding variants were introduced into RET constructs and these were transfected into HEK293 cells to determine RET phosphorylation and activation via ERK. An exon trap experiment was performed to check a possible splice-site mutation. We identified eight rare RET-coding variants, one possible splice-site variant, but no rare EDNRB variants. Western blotting showed that three coding variants p.(Pr270Leu), p.(Ala756Val) and p.(Tyr1062Cys) resulted in lower activation of RET. Moreover, only two RET variants (p.(Ala756Val) and p.(Tyr1062Cys)) resulted in reduced ERK activation. Splice-site assays on c.1880-11A>G could not confirm its pathogenicity. Our data suggest that indeed almost half of the identified rare variants are proven pathogenic and that, hence, functional studies are essential for proper genetic counseling.

  19. Intra-observer agreement in single and joint double readings of contrast-enhanced breast MRI screening for women with high genetic breast cancer risks

    Directory of Open Access Journals (Sweden)

    Hugo C

    2013-04-01

    Full Text Available Objectives: To examine intra-observer reliability (IR for lesion detection on contrast-enhanced breast magnetic resonance images (MRI for screening women at high risk of breast cancer in single and joint double readings, without case selection. Methods: Contrast-enhanced breast MRIs were interpreted twice by the same independent reader and twice in joint readings. IR was assessed for lesion detection, normal MRI identification, mass, non-mass like enhancements (NMLE and focus characterisation, and BI-RADS assessment. Results: MRI examinations for 124 breasts, 65 women (mean age 43.4y were retrospectively reviewed with 110 lesions identified. Abnormal BIRADS (3-5 classifications were found for 52.3% in single readings and 58.5% in joint readings. Seven biopsies were performed for 4 histologically confirmed cancers. IR for BI-RADS classifications was good for single (0.63, 95% CI: 0.49-0.77, and joint readings (0.77, 95% CI: 0.61-0.93. IR for background parenchymal enhancement (BPE was moderate across single (0.53, 95% CI: 0.40-0.65 and joint readings (0.44, 95% CI: 0.33-0.56. IR for BI-RADS category according to each enhancement was poor for single (0.27, 95% CI: 0.10-0.44, and higher for joint readings, (0.58, 95% CI: 0.43-0.72. Conclusions: IR in BI-RADS breast assessments or BI-RADS lesion assessments are better with joint reading in screening for women with high genetic risks, in particular for abnormal MRI (BI-RADS 3, 4 and 5.

  20. Chemical inducible promoter used to obtain transgenic plants with a silent marker and organisms and cells and methods of using same for screening for mutations

    Science.gov (United States)

    Zuo, Jianru; Chua, Nam-Hai

    2007-06-12

    Disclosed is a chemically inducible promoter for transforming plants or plant cells with genes which are regulatable by adding the plants or cells to a medium containing an inducer or by removing them from such medium. The promoter is inducible by a glucocorticoid, estrogen or inducer not endogenous to plants. Such promoters may be used with any plant genes that can promote shoot regeneration and development to induce shoot formation in the presence of a glucocorticoid, estrogen or inducer. The promoter may be used with antibiotic or herbicide resistance genes or other genes which are regulatable by the presence or absence of a given inducer. Also presented are organisms or cells comprising a gene wherein the natural promoter of the gene is disrupted and the gene is placed under the control of a transgenic inducible promoter. These organisms and cells and their progeny are useful for screening for conditional gain of function and loss of function mutations.

  1. Identification and quantification of estrogenic compounds in recycled and virgin paper for household use as determined by an in vitro yeast estrogen screen and chemical analysis

    DEFF Research Database (Denmark)

    Vinggaard, Anne; Körner, Wolfgang; Lund, Kirsten H.;

    2000-01-01

    The use of recycled paper for the manufacture of food contact materials is widespread, but very little is known about the presence of potential contaminants in the paper. The purpose of this study was to assess the worst-case migration of estrogenic active compounds using extracts of paper...... for household use. Twenty different brands of kitchen rolls, nine of which were made from recycled paper and the remainder from virgin paper, were obtained from retail shops. Paper extracts were subjected to (a) determination of the total estrogenic activity by using an in vitro estrogen screen based on yeast...... cells stably transfected with the human estrogen receptor alpha and (b) chemical analysis and quantification by GC/MS, GC/FTIR/MS, and GC/FID for detection of a variety of estrogenic compounds. A marked estrogenic response was observed in nine of the extracts, seven of which were made from recycled...

  2. Evaluation of an hPXR reporter gene assay for the detection of aquatic emerging pollutants: screening of chemicals and application to water samples

    Energy Technology Data Exchange (ETDEWEB)

    Creusot, Nicolas; Kinani, Said; Maillot-Marechal, Emmanuelle; Porcher, Jean-Marc; Ait-Aissa, Selim [Unite Ecotoxicologie, INERIS, Verneuil-en-Halatte (France); Balaguer, Patrick [IRCM-UM1-CRLC Val d' Aurelle, INSERM U896, Montpellier (France); Tapie, Nathalie; LeMenach, Karyn; Budzinski, Helene [ISM/LPTC-UMR 5255 CNRS Universite Bordeaux 1, Talence (France)

    2010-01-15

    Many environmental endocrine-disrupting compounds act as ligands for nuclear receptors. Among these receptors, the human pregnane X receptor (hPXR) is well described as a xenobiotic sensor to various classes of chemicals, including pharmaceuticals, pesticides, and steroids. To assess the potential use of PXR as a sensor for aquatic emerging pollutants, we employed an in vitro reporter gene assay (HG5LN-hPXR cells) to screen a panel of environmental chemicals and to assess PXR-active chemicals in (waste) water samples. Of the 57 compounds tested, 37 were active in the bioassay and 10 were identified as new PXR agonists: triazin pesticides (promethryn, terbuthryn, terbutylazine), pharmaceuticals (fenofibrate, bezafibrate, clonazepam, medazepam) and non co-planar polychlorobiphenyls (PCBs; PCB101, 138, 180). Furthermore, we detected potent PXR activity in two types of water samples: passive polar organic compounds integrative sampler (POCIS) extracts from a river moderately impacted by agricultural and urban inputs and three effluents from sewage treatment works (STW). Fractionation of POCIS samples showed the highest PXR activity in the less polar fraction, while in the effluents, PXR activity was mainly associated with the dissolved water phase. Chemical analyses quantified several PXR-active substances (i.e., alkylphenols, hormones, pharmaceuticals, pesticides, PCBs, bisphenol A) in POCIS fractions and effluent extracts. However, mass-balance calculations showed that the analyzed compounds explained only 0.03% and 1.4% of biological activity measured in POCIS and STW samples, respectively. In effluents, bisphenol A and 4-tert-octylphenol were identified as main contributors of instrumentally derived PXR activities. Finally, the PXR bioassay provided complementary information as compared to estrogenic, androgenic, and dioxin-like activity measured in these samples. This study shows the usefulness of HG5LN-hPXR cells to detect PXR-active compounds in water samples

  3. Targeting helicase-dependent amplification products with an electrochemical genosensor for reliable and sensitive screening of genetically modified organisms.

    Science.gov (United States)

    Moura-Melo, Suely; Miranda-Castro, Rebeca; de-Los-Santos-Álvarez, Noemí; Miranda-Ordieres, Arturo J; Dos Santos Junior, J Ribeiro; da Silva Fonseca, Rosana A; Lobo-Castañón, Maria Jesús

    2015-08-18

    Cultivation of genetically modified organisms (GMOs) and their use in food and feed is constantly expanding; thus, the question of informing consumers about their presence in food has proven of significant interest. The development of sensitive, rapid, robust, and reliable methods for the detection of GMOs is crucial for proper food labeling. In response, we have experimentally characterized the helicase-dependent isothermal amplification (HDA) and sequence-specific detection of a transgene from the Cauliflower Mosaic Virus 35S Promoter (CaMV35S), inserted into most transgenic plants. HDA is one of the simplest approaches for DNA amplification, emulating the bacterial replication machinery, and resembling PCR but under isothermal conditions. However, it usually suffers from a lack of selectivity, which is due to the accumulation of spurious amplification products. To improve the selectivity of HDA, which makes the detection of amplification products more reliable, we have developed an electrochemical platform targeting the central sequence of HDA copies of the transgene. A binary monolayer architecture is built onto a thin gold film where, upon the formation of perfect nucleic acid duplexes with the amplification products, these are enzyme-labeled and electrochemically transduced. The resulting combined system increases genosensor detectability up to 10(6)-fold, allowing Yes/No detection of GMOs with a limit of detection of ∼30 copies of the CaMV35S genomic DNA. A set of general utility rules in the design of genosensors for detection of HDA amplicons, which may assist in the development of point-of-care tests, is also included. The method provides a versatile tool for detecting nucleic acids with extremely low abundance not only for food safety control but also in the diagnostics and environmental control areas.

  4. Preliminary screening of alternative technologies to incineration for treatment of chemical-agent-contaminated soil, Rocky Mountain Arsenal

    Energy Technology Data Exchange (ETDEWEB)

    Shem, L.M.; Rosenblatt, D.H.; Smits, M.P.; Wilkey, P.L.; Ballou, S.W.

    1995-12-01

    In support of the U.S. Army`s efforts to determine the best technologies for remediation of soils, water, and structures contaminated with pesticides and chemical agents, Argonne National Laboratory has reviewed technologies for treating soils contaminated with mustard, lewisite, sarin, o-ethyl s-(2- (diisopropylamino)ethyl)methyl-phosphonothioate (VX), and their breakdown products. This report focuses on assessing alternatives to incineration for dealing with these contaminants. For each technology, a brief description is provided, its suitability and constraints on its use are identified, and its overall applicability for treating the agents of concern is summarized. Technologies that merit further investigation are identified.

  5. 胚胎植入前遗传学筛查的临床应用%Clinical Application of Preimplantation Genetic Screening

    Institute of Scientific and Technical Information of China (English)

    张倩; 郑叶; 颜军昊

    2014-01-01

    With the development of assisted reproductive technology and genetic analysis technology, the preimplantation genetic screening (PGS) is used to detect the numerical chromosomal abnormalities (aneuploid) in embryos, so as to improve pregnancy outcomes of in vitro fertilization-embryo transfer (IVF-ET). New methods and technologies have been used in PGS, such as blastocyst stage biopsy, comparative genomic hybridization, microarrays and next generation sequencing, which increase the diagnose accuracy and decrease the risk of misdiagnosis. At the same time, PGS is also facing many challenges. The clinical application of PGS was reviewed in this article.%随着辅助生殖技术和遗传学分析技术的发展,胚胎植入前遗传学筛查应用于胚胎染色体数目异常(非整倍体)检测,以期改善体外受精-胚胎移植的妊娠结局。新方法、新技术不断出现并应用于胚胎植入前遗传学筛查中,如囊胚期活检、比较基因组杂交技术、微阵列技术、第二代测序技术等,显著增加了诊断准确性,减少误诊风险。同时,胚胎植入前遗传学筛查的广泛应用也面临许多挑战。综述该领域的应用进展和面临的挑战。

  6. Formaldehyde in Alcoholic Beverages: Large Chemical Survey Using Purpald Screening Followed by Chromotropic Acid Spectrophotometry with Multivariate Curve Resolution

    Directory of Open Access Journals (Sweden)

    Julien A. Jendral

    2011-01-01

    Full Text Available A strategy for analyzing formaldehyde in beer, wine, spirits, and unrecorded alcohol was developed, and 508 samples from worldwide origin were analyzed. In the first step, samples are qualitatively screened using a simple colorimetric test with the purpald reagent, which is extremely sensitive for formaldehyde (detection limit 0.1 mg/L. 210 samples (41% gave a positive purpald reaction. In the second step, formaldehyde in positive samples is confirmed by quantitative spectrophotometry of the chromotropic acid-formaldehyde derivative combined with Multivariate Curve Resolution-Alternating Least Squares (MCR-ALS. Calculation of UV-VIS and 13C NMR spectra confirmed the monocationic dibenzoxanthylium structure as the product of the reaction and disproved the widely cited para,para-quinoidal structure. Method validation for the spectrophotometric procedure showed a detection limit of 0.09 mg/L and a precision of 4.2–8.2% CV. In total, 132 samples (26% contained formaldehyde with an average of 0.27 mg/L (range 0–14.4 mg/L. The highest incidence occurred in tequila (83%, Asian spirits (59%, grape marc (54%, and brandy (50%. Our survey showed that only 9 samples (1.8% had formaldehyde levels above the WHO IPCS tolerable concentration of 2.6 mg/L.

  7. Analysis of genetics and chemical contents relation compared to commonly used Cissus quadrangularis L. and barcode markers of some Thailand Cissus species.

    Science.gov (United States)

    Sudmoon, Runglawan; Chaveerach, Arunrat; Tanee, Tawatchai

    2016-01-01

    Several Cissus species are commonly used as traditional and modified medicines, and their chemical constituents are major point for precise usage. However, C. quadrangularis is the only species for which the usages and the chemical composition have been reported. These data should be investigated for other species in the genus. Eight species namely C. assamica, C. carnosa, C. elongata, C. hastata, C. javana, C. pteroclada, C. quadrangular is and C. repens were evaluated for genetic relationships and chemical composition. Constructed dendrogram shows high-powered efficiency of inter-simple sequence repeat (ISSR) data used which can clearly identify different and identical species. Genetic similarity (S) value of the identical species is 0.86, whereas for different species the value can vary from 0.53 to 0.75. Four highly related species (S=0.64-0.72), C. assamica, C. carnosa, C. hastata and C. repens were selected to undergo chemical study by gas chromatography-mass spectrometry (GC-MS) on the methanol crude extract. Only one compound, β-sitosterol, found in the four species is identical to the compound reported from C. quadrangular is, where there were five identical chemicals found in the selected species. Species-specific barcode with rbcL region was constructed. Nucleotide variation was evaluated indicating genetic distance value of 0.025 to 0.072. PMID:26826840

  8. Carbonado: Physical and chemical properties, a critical evaluation of proposed origins, and a revised genetic model

    Science.gov (United States)

    Haggerty, Stephen E.

    2014-03-01

    Carbonado-diamond is the most controversial of all diamond types and is found only in Brazil, and the Central African Republic (Bangui). Neither an affinity to Earth's mantle, nor an origin in the crust can be unequivocally established. Carbonado-diamond is at least 3.8 Ga old, an age about 0.5 Ga older than the oldest diamonds yet reported in kimberlites and lamproites on Earth. Derived from Neo- to Mid-Proterozoic meta-conglomerates, the primary magmatic host rock has not been identified. Discovered in 1841, the material is polycrystalline, robust and coke-like, and is best described as a strongly bonded micro-diamond ceramic. It is characteristically porous, which precludes an origin at high pressures and high temperatures in Earth's deep interior, yet it is also typically patinated, with a glass-like surface that resembles melting. With exotic inclusions of highly reduced metals, carbides, and nitrides the origin of carbonado-diamond is made even more challenging. But the challenge is important because a new diamondiferous host rock may be involved, and the development of a new physical process for generating diamond is possibly assured. The combination of micro-crystals and random crystal orientation leads to extreme mechanical toughness, and a predicable super-hardness. The physical and chemical properties of carbonado are described with a view to the development of a mimetic strategy to synthesize carbonado and to duplicate its extreme toughness and super-hardness. Textural variations are described with an emphasis on melt-like surface features, not previously discussed in the literature, but having a very clear bearing on the history and genesis of carbonado. Selected physical properties are presented and the proposed origins, diverse in character and imaginatively novel, are critically reviewed. From our present knowledge of the dynamic Earth, all indications are that carbonado is unlikely to be of terrestrial origin. A revised model for the origin of

  9. The Chemical and Genetic Characteristics of Szechuan Pepper (Zanthoxylum bungeanum and Z. armatum) Cultivars and Their Suitable Habitat

    Science.gov (United States)

    Xiang, Li; Liu, Yue; Xie, Caixiang; Li, Xiwen; Yu, Yadong; Ye, Meng; Chen, Shilin

    2016-01-01

    Szechuan peppers, famous for their unique sensation and flavor, are widely used as a food additive and traditional herbal medicine. Zanthoxylum bungeanum and Z. armatum are both commonly recognized as Szechuan peppers, but they have different tastes and effects. The chemical components, genetic characteristics, and suitable habitat of six cultivars were analyzed in this study. The results indicated that Z. armatum contained a larger proportion of volatile oil, whereas Z. bungeanum produced a more non-volatile ether extraction. The average content of volatile oil and non-volatile ether extract of Z. armatum were 11.84 and 11.63%, respectively, and the average content of volatile oil and non-volatile ether extract of Z. bungeanum were 6.46 and 14.23%, respectively. Combined with an internal transcribed spacer 2 (ITS2) sequence characters and chemical PCA results, six cultivars were classified into their own groups, for the two species in particular. The temperature in January and July were the most significant ecological factors influencing the contents of the Z. armatum volatile oil. However, annual precipitation, temperature in January and relevant humidity had a significant positive correlation with the content of non-volatile ether extract in Z. bungeanum. Thus, the most suitable areas for producing Z. bungeanum cultivars ranged from the Hengduan Mountains to the Ta-pa Mountains, and the regions suitable for Z. armatum cultivars were found to be in the Sichuan Basin and Dalou-Wu mountains. The predicted suitable habitat could be used as a preliminary test area for Szechuan pepper cultivar production. PMID:27148298

  10. The Chemical and Genetic Characteristics of Szechuan Pepper (Zanthoxylum bungeanum and Z. armatum) Cultivars and Their Suitable Habitat.

    Science.gov (United States)

    Xiang, Li; Liu, Yue; Xie, Caixiang; Li, Xiwen; Yu, Yadong; Ye, Meng; Chen, Shilin

    2016-01-01

    Szechuan peppers, famous for their unique sensation and flavor, are widely used as a food additive and traditional herbal medicine. Zanthoxylum bungeanum and Z. armatum are both commonly recognized as Szechuan peppers, but they have different tastes and effects. The chemical components, genetic characteristics, and suitable habitat of six cultivars were analyzed in this study. The results indicated that Z. armatum contained a larger proportion of volatile oil, whereas Z. bungeanum produced a more non-volatile ether extraction. The average content of volatile oil and non-volatile ether extract of Z. armatum were 11.84 and 11.63%, respectively, and the average content of volatile oil and non-volatile ether extract of Z. bungeanum were 6.46 and 14.23%, respectively. Combined with an internal transcribed spacer 2 (ITS2) sequence characters and chemical PCA results, six cultivars were classified into their own groups, for the two species in particular. The temperature in January and July were the most significant ecological factors influencing the contents of the Z. armatum volatile oil. However, annual precipitation, temperature in January and relevant humidity had a significant positive correlation with the content of non-volatile ether extract in Z. bungeanum. Thus, the most suitable areas for producing Z. bungeanum cultivars ranged from the Hengduan Mountains to the Ta-pa Mountains, and the regions suitable for Z. armatum cultivars were found to be in the Sichuan Basin and Dalou-Wu mountains. The predicted suitable habitat could be used as a preliminary test area for Szechuan pepper cultivar production.

  11. Alginate based 3D hydrogels as an in vitro co-culture model platform for the toxicity screening of new chemical entities

    International Nuclear Information System (INIS)

    Prediction of human response to potential therapeutic drugs is through conventional methods of in vitro cell culture assays and expensive in vivo animal testing. Alternatives to animal testing require sophisticated in vitro model systems that must replicate in vivo like function for reliable testing applications. Advancements in biomaterials have enabled the development of three-dimensional (3D) cell encapsulated hydrogels as in vitro drug screening tissue model systems. In this study, we have developed an in vitro platform to enable high density 3D culture of liver cells combined with a monolayer growth of target breast cancer cell line (MCF-7) in a static environment as a representative example of screening drug compounds for hepatotoxicity and drug efficacy. Alginate hydrogels encapsulated with serial cell densities of HepG2 cells (105-108 cells/ml) are supported by a porous poly-carbonate disc platform and co-cultured with MCF-7 cells within standard cell culture plates during a 3 day study period. The clearance rates of drug transformation by HepG2 cells are measured using a coumarin based pro-drug. The platform was used to test for HepG2 cytotoxicity 50% (CT50) using commercially available drugs which further correlated well with published in vivo LD50 values. The developed test platform allowed us to evaluate drug dose concentrations to predict hepatotoxicity and its effect on the target cells. The in vitro 3D co-culture platform provides a scalable and flexible approach to test multiple-cell types in a hybrid setting within standard cell culture plates which may open up novel 3D in vitro culture techniques to screen new chemical entity compounds. - Graphical abstract: Display Omitted Highlights: → A porous support disc design to support the culture of desired cells in 3D hydrogels. → Demonstrated the co-culture of two cell types within standard cell-culture plates. → A scalable, low cost approach to toxicity screening involving multiple cell types.

  12. Multiplex polymerase chain reaction-capillary gel electrophoresis: a promising tool for GMO screening--assay for simultaneous detection of five genetically modified cotton events and species.

    Science.gov (United States)

    Nadal, Anna; Esteve, Teresa; Pla, Maria

    2009-01-01

    A multiplex polymerase chain reaction assay coupled to capillary gel electrophoresis for amplicon identification by size and color (multiplex PCR-CGE-SC) was developed for simultaneous detection of cotton species and 5 events of genetically modified (GM) cotton. Validated real-time-PCR reactions targeting Bollgard, Bollgard II, Roundup Ready, 3006-210-23, and 281-24-236 junction sequences, and the cotton reference gene acp1 were adapted to detect more than half of the European Union-approved individual or stacked GM cotton events in one reaction. The assay was fully specific (PCR-CGE-SC assay to allow simultaneous detection of 6 cotton and 5 maize targets (two endogenous genes and 9 GM events) in two multiplex PCRs and a single CGE, making the approach more economic. Besides allowing simultaneous detection of many targets with adequate specificity and sensitivity, the multiplex PCR-CGE-SC approach has high throughput and automation capabilities, while keeping a very simple protocol, e.g., amplification and labeling in one step. Thus, it is an easy and inexpensive tool for initial screening, to be complemented with quantitative assays if necessary.

  13. BRCA Genetic Screening in Middle Eastern and North African: Mutational Spectrum and Founder BRCA1 Mutation (c.798_799delTT in North African

    Directory of Open Access Journals (Sweden)

    Abdelilah Laraqui

    2015-01-01

    Full Text Available Background. The contribution of BRCA1 mutations to both hereditary and sporadic breast and ovarian cancer (HBOC has not yet been thoroughly investigated in MENA. Methods. To establish the knowledge about BRCA1 mutations and their correlation with the clinical aspect in diagnosed cases of HBOC in MENA populations. A systematic review of studies examining BRCA1 in BC women in Cyprus, Jordan, Egypt, Lebanon, Morocco, Algeria, and Tunisia was conducted. Results. Thirteen relevant references were identified, including ten studies which performed DNA sequencing of all BRCA1 exons. For the latter, 31 mutations were detected in 57 of the 547 patients ascertained. Familial history of BC was present in 388 (71% patients, of whom 50 were mutation carriers. c.798_799delTT was identified in 11 North African families, accounting for 22% of total identified BRCA1 mutations, suggesting a founder allele. A broad spectrum of other mutations including c.68_69delAG, c.181T>G, c.5095C>T, and c.5266dupC, as well as sequence of unclassified variants and polymorphisms, was also detected. Conclusion. The knowledge of genetic structure of BRCA1 in MENA should contribute to the assessment of the necessity of preventive programs for mutation carriers and clinical management. The high prevalence of BC and the presence of frequent mutations of the BRCA1 gene emphasize the need for improving screening programs and individual testing/counseling.

  14. A haploid genetic screen identifies the major facilitator domain containing 2A (MFSD2A) transporter as a key mediator in the response to tunicamycin.

    Science.gov (United States)

    Reiling, Jan H; Clish, Clary B; Carette, Jan E; Varadarajan, Malini; Brummelkamp, Thijn R; Sabatini, David M

    2011-07-19

    Tunicamycin (TM) inhibits eukaryotic asparagine-linked glycosylation, protein palmitoylation, ganglioside production, proteoglycan synthesis, 3-hydroxy-3-methylglutaryl coenzyme-A reductase activity, and cell wall biosynthesis in bacteria. Treatment of cells with TM elicits endoplasmic reticulum stress and activates the unfolded protein response. Although widely used in laboratory settings for many years, it is unknown how TM enters cells. Here, we identify in an unbiased genetic screen a transporter of the major facilitator superfamily, major facilitator domain containing 2A (MFSD2A), as a critical mediator of TM toxicity. Cells without MFSD2A are TM-resistant, whereas MFSD2A-overexpressing cells are hypersensitive. Hypersensitivity is associated with increased cellular TM uptake concomitant with an enhanced endoplasmic reticulum stress response. Furthermore, MFSD2A mutant analysis reveals an important function of the C terminus for correct intracellular localization and protein stability, and it identifies transmembrane helical amino acid residues essential for mediating TM sensitivity. Overall, our data uncover a critical role for MFSD2A by acting as a putative TM transporter at the plasma membrane.

  15. Reverse genetic screen for loss-of-function mutations uncovers a frameshifting deletion in the melanophilin gene accountable for a distinctive coat color in Belgian Blue cattle.

    Science.gov (United States)

    Li, Wanbo; Sartelet, Arnaud; Tamma, Nico; Coppieters, Wouter; Georges, Michel; Charlier, Carole

    2016-02-01

    In the course of a reverse genetic screen in the Belgian Blue cattle breed, we uncovered a 10-bp deletion (c.87_96del) in the first coding exon of the melanophilin gene (MLPH), which introduces a premature stop codon (p.Glu32Aspfs*1) in the same exon, truncating 94% of the protein. Recessive damaging mutations in the MLPH gene are well known to cause skin, hair, coat or plumage color dilution phenotypes in numerous species, including human, mice, dog, cat, mink, rabbit, chicken and quail. Large-scale array genotyping undertaken to identify p.Glu32Aspfs*1 homozygous mutant animals revealed a mutation frequency of 5% in the breed and allowed for the identification of 10 homozygous mutants. As expression of a colored coat requires at least one wild-type allele at the co-dominant Roan locus encoded by the KIT ligand gene (KITLG), homozygous mutants for p.Ala227Asp corresponding with the missense mutation were excluded. The six remaining colored calves displayed a distinctive dilution phenotype as anticipated. This new coat color was named 'cool gray'. It is the first damaging mutation in the MLPH gene described in cattle and extends the already long list of species with diluted color due to recessive mutations in MLPH and broadens the color palette of gray in this breed. PMID:26582259

  16. Genetic Screen Reveals Link between the Maternal Effect Sterile Gene mes-1 and Pseudomonas aeruginosa-induced Neurodegeneration in Caenorhabditis elegans.

    Science.gov (United States)

    Wu, Qiuli; Cao, Xiou; Yan, Dong; Wang, Dayong; Aballay, Alejandro

    2015-12-01

    Increasing evidence indicates that immune responses to microbial infections may contribute to neurodegenerative diseases. Here, we show that Pseudomonas aeruginosa infection of Caenorhabditis elegans causes a number of neural changes that are hallmarks of neurodegeneration. Using an unbiased genetic screen to identify genes involved in the control of P. aeruginosa-induced neurodegeneration, we identified mes-1, which encodes a receptor tyrosine kinase-like protein that is required for unequal cell divisions in the early embryonic germ line. We showed that sterile but not fertile mes-1 animals were resistant to neurodegeneration induced by P. aeruginosa infection. Similar results were observed using animals carrying a mutation in the maternal effect gene pgl-1, which is required for postembryonic germ line development, and the germ line-deficient strains glp-1 and glp-4. Additional studies indicated that the FOXO transcription factor DAF-16 is required for resistance to P. aeruginosa-induced neurodegeneration in germ line-deficient strains. Thus, our results demonstrate that P. aeruginosa infection results in neurodegeneration phenotypes in C. elegans that are controlled by the germ line in a cell-nonautonomous manner.

  17. Reverse genetic screen for loss-of-function mutations uncovers a frameshifting deletion in the melanophilin gene accountable for a distinctive coat color in Belgian Blue cattle.

    Science.gov (United States)

    Li, Wanbo; Sartelet, Arnaud; Tamma, Nico; Coppieters, Wouter; Georges, Michel; Charlier, Carole

    2016-02-01

    In the course of a reverse genetic screen in the Belgian Blue cattle breed, we uncovered a 10-bp deletion (c.87_96del) in the first coding exon of the melanophilin gene (MLPH), which introduces a premature stop codon (p.Glu32Aspfs*1) in the same exon, truncating 94% of the protein. Recessive damaging mutations in the MLPH gene are well known to cause skin, hair, coat or plumage color dilution phenotypes in numerous species, including human, mice, dog, cat, mink, rabbit, chicken and quail. Large-scale array genotyping undertaken to identify p.Glu32Aspfs*1 homozygous mutant animals revealed a mutation frequency of 5% in the breed and allowed for the identification of 10 homozygous mutants. As expression of a colored coat requires at least one wild-type allele at the co-dominant Roan locus encoded by the KIT ligand gene (KITLG), homozygous mutants for p.Ala227Asp corresponding with the missense mutation were excluded. The six remaining colored calves displayed a distinctive dilution phenotype as anticipated. This new coat color was named 'cool gray'. It is the first damaging mutation in the MLPH gene described in cattle and extends the already long list of species with diluted color due to recessive mutations in MLPH and broadens the color palette of gray in this breed.

  18. Multiplex ligation-dependent probe amplification for genetic screening in autism spectrum disorders: Efficient identification of known microduplications and identification of a novel microduplication in ASMT

    Directory of Open Access Journals (Sweden)

    Reichert Jennifer G

    2008-10-01

    Full Text Available Abstract Background It has previously been shown that specific microdeletions and microduplications, many of which also associated with cognitive impairment (CI, can present with autism spectrum disorders (ASDs. Multiplex ligation-dependent probe amplification (MLPA represents an efficient method to screen for such recurrent microdeletions and microduplications. Methods In the current study, a total of 279 unrelated subjects ascertained for ASDs were screened for genomic disorders associated with CI using MLPA. Fluorescence in situ hybridization (FISH, quantitative polymerase chain reaction (Q-PCR and/or direct DNA sequencing were used to validate potential microdeletions and microduplications. Methylation-sensitive MLPA was used to characterize individuals with duplications in the Prader-Willi/Angelman (PWA region. Results MLPA showed two subjects with typical ASD-associated interstitial duplications of the 15q11-q13 PWA region of maternal origin. Two additional subjects showed smaller, de novo duplications of the PWA region that had not been previously characterized. Genes in these two novel duplications include GABRB3 and ATP10A in one case, and MKRN3, MAGEL2 and NDN in the other. In addition, two subjects showed duplications of the 22q11/DiGeorge syndrome region. One individual was found to carry a 12 kb deletion in one copy of the ASPA gene on 17p13, which when mutated in both alleles leads to Canavan disease. Two subjects showed partial duplication of the TM4SF2 gene on Xp11.4, previously implicated in X-linked non-specific mental retardation, but in our subsequent analyses such variants were also found in controls. A partial duplication in the ASMT gene, located in the pseudoautosomal region 1 (PAR1 of the sex chromosomes and previously suggested to be involved in ASD susceptibility, was observed in 6–7% of the cases but in only 2% of controls (P = 0.003. Conclusion MLPA proves to be an efficient method to screen for chromosomal

  19. Evaluation of genetic variability of wild hops (Humulus lupulus L.) in Canada and the Caucasus region by chemical and molecular methods.

    Science.gov (United States)

    Patzak, Josef; Nesvadba, Vladimír; Krofta, Karel; Henychova, Alena; Marzoev, Arkady Inalovic; Richards, Ken

    2010-07-01

    Wild hops (Humulus lupulus L.) are potential new germplasms to expand the variability of genetic resources for hop breeding. We evaluated Canadian (62 plants) and Caucasian (58 plants) wild hops by their chemical characteristics and with molecular genetic analyses using sequence-tagged site and simple sequence repeat markers, in comparison with European (104 plants) and North American (27 plants) wild hops. The contents of alpha and beta acids varied from 0.36% to 5.11% and from 0.43% to 6.66% in Canadian wild hops, and from 0.85% to 3.65% and from 1.22% to 4.81% in Caucasian wild hops, respectively. The contents of cohumulone and colupulone distinctly differed between European and North American wild hops: the cohumulone level in alpha acids was in the range 46.1%-68.4% among North American wild hops and in the range 13.6%-30.6% among European wild hops. The high content of myrcene and the low contents of humulene, farnesene, and selinenes were typical for wild hops from Canada, in contrast to wild hops from the Caucasus region. We compared the chemical characteristics with molecular genetic data. Chemical characteristics differentiated wild hops into North American and Eurasian groups. Molecular genetic analysis was able to separate Caucasian wild hops from European wild hops. We proved a hop phylogeny by means of wide molecular analysis. PMID:20616876

  20. Differences between first and subsequent rounds of the MRISC breast cancer screening program for women with a familial or genetic predisposition

    NARCIS (Netherlands)

    Kriege, M; Brekelmans, CTM; Boetes, C; Muller, SH; Zonderland, HM; Obdeijn, IM; Manoliu, RA; Kok, T; Rutgers, EJT; de Koning, HJ; Klijn, JGM

    2006-01-01

    BACKGROUND. within the Dutch MRI Screening (MRISC) study, a Dutch multicenter screening study for hereditary breast cancer, the authors investigated whether previously reported increased diagnostic accuracy of magnetic resonance imaging (MRI) compared with mammography would be maintained during subs

  1. A method for the amplification of chemically induced transformation in C3H/10T1/2 clone 8 cells: its use as a potential screening assay.

    Science.gov (United States)

    Schechtman, L M; Kiss, E; McCarvill, J; Nims, R; Kouri, R E; Lubet, R A

    1987-09-01

    dimethylnitrosamine, 2-naphthylamine, 2-aminoanthracene, and aflatoxin B1. With the use of this revised assay, 14 coded and 23 model compounds were tested. Agreement with in vivo results was observed to be over 85%. The marked sensitivity and discriminatory ability of this revised assay procedure suggest its usefulness as a screen for potential carcinogens of diverse chemical structure. PMID:3114533

  2. Vision Screening

    Science.gov (United States)

    ... offer vision screening programs for children. At what age should a child have his or her vision screened? Vision screening ... a child fails a vision screening at any age, the child should be referred for a comprehensive eye examination. ...

  3. In-bead screening

    DEFF Research Database (Denmark)

    2013-01-01

    The present invention relates to screening of one-bead-one-compound (OBOC) combinatorial libraries which is useful for the discovery of compounds displaying molecular interactions with a biological or a physicochemical system, such as substrates and inhibitors of enzymes and the like. The invention...... provides a method for screening a library of compounds for their interaction with a physico- chemical or biological system and a corresponding kit for performing the method of screening a one-bead-one-compound library of compounds....

  4. Small Molecule DFPM Derivative-Activated Plant Resistance Protein Signaling in Roots Is Unaffected by EDS1 Subcellular Targeting Signal and Chemical Genetic Isolation of victr R-Protein Mutants.

    Directory of Open Access Journals (Sweden)

    Hans-Henning Kunz

    Full Text Available The small molecule DFPM ([5-(3,4-dichlorophenylfuran-2-yl]-piperidine-1-ylmethanethione was recently shown to trigger signal transduction via early effector-triggered immunity signaling genes including EDS1 and PAD4 in Arabidopsis thaliana accession Col-0. Chemical genetic analyses of A. thaliana natural variants identified the plant Resistance protein-like Toll/Interleukin1 Receptor (TIR-Nucleotide Binding (NB-Leucine-Rich Repeat (LRR protein VICTR as required for DFPM-mediated root growth arrest. Here a chemical genetic screen for mutants which disrupt DFPM-mediated root growth arrest in the Col-0 accession identified new mutant alleles of the TIR-NB-LRR gene VICTR. One allele, victr-6, carries a Gly216-to-Asp mutation in the Walker A domain supporting an important function of the VICTR nucleotide binding domain in DFPM responses consistent with VICTR acting as a canonical Resistance protein. The essential nucleo-cytoplasmic regulator of TIR-NB-LRR-mediated effector-triggered immunity, EDS1, was reported to have both nuclear and cytoplasmic actions in pathogen resistance. DFPM was used to investigate the requirements for subcellular EDS1 localization in DFPM-mediated root growth arrest. EDS1-YFP fusions engineered to localize mainly in the cytoplasm or the nucleus by tagging with a nuclear export signal (NES or a nuclear localization signal (NLS, respectively, were tested. We found that wild-type EDS1-YFP and both the NES and NLS-tagged EDS1 variants were induced by DFPM treatments and fully complemented eds1 mutant plants in root responses to DFPM, suggesting that enrichment of EDS1 in either compartment could confer DFPM-mediated root growth arrest. We further found that a light and O2-dependent modification of DFPM is necessary to mediate DFPM signaling in roots. Chemical analyses including Liquid Chromatography-Mass Spectrometry and High-Resolution Atmospheric Pressure Chemical Ionization Mass Spectrometry identified a DFPM modification

  5. Small Molecule DFPM Derivative-Activated Plant Resistance Protein Signaling in Roots Is Unaffected by EDS1 Subcellular Targeting Signal and Chemical Genetic Isolation of victr R-Protein Mutants.

    Science.gov (United States)

    Kunz, Hans-Henning; Park, Jiyoung; Mevers, Emily; García, Ana V; Highhouse, Samantha; Gerwick, William H; Parker, Jane E; Schroeder, Julian I

    2016-01-01

    The small molecule DFPM ([5-(3,4-dichlorophenyl)furan-2-yl]-piperidine-1-ylmethanethione) was recently shown to trigger signal transduction via early effector-triggered immunity signaling genes including EDS1 and PAD4 in Arabidopsis thaliana accession Col-0. Chemical genetic analyses of A. thaliana natural variants identified the plant Resistance protein-like Toll/Interleukin1 Receptor (TIR)-Nucleotide Binding (NB)-Leucine-Rich Repeat (LRR) protein VICTR as required for DFPM-mediated root growth arrest. Here a chemical genetic screen for mutants which disrupt DFPM-mediated root growth arrest in the Col-0 accession identified new mutant alleles of the TIR-NB-LRR gene VICTR. One allele, victr-6, carries a Gly216-to-Asp mutation in the Walker A domain supporting an important function of the VICTR nucleotide binding domain in DFPM responses consistent with VICTR acting as a canonical Resistance protein. The essential nucleo-cytoplasmic regulator of TIR-NB-LRR-mediated effector-triggered immunity, EDS1, was reported to have both nuclear and cytoplasmic actions in pathogen resistance. DFPM was used to investigate the requirements for subcellular EDS1 localization in DFPM-mediated root growth arrest. EDS1-YFP fusions engineered to localize mainly in the cytoplasm or the nucleus by tagging with a nuclear export signal (NES) or a nuclear localization signal (NLS), respectively, were tested. We found that wild-type EDS1-YFP and both the NES and NLS-tagged EDS1 variants were induced by DFPM treatments and fully complemented eds1 mutant plants in root responses to DFPM, suggesting that enrichment of EDS1 in either compartment could confer DFPM-mediated root growth arrest. We further found that a light and O2-dependent modification of DFPM is necessary to mediate DFPM signaling in roots. Chemical analyses including Liquid Chromatography-Mass Spectrometry and High-Resolution Atmospheric Pressure Chemical Ionization Mass Spectrometry identified a DFPM modification product that is

  6. Screening in liver disease

    Institute of Scientific and Technical Information of China (English)

    Paolo Del Poggio; Marzio Mazzoleni

    2006-01-01

    A disease is suitable for screening if it is common, if the target population can be identified and reached and if both a good screening test and an effective therapy are available. Of the most common liver diseases only viral hepatitis and genetic hemochromatosis partially satisfy these conditions. Hepatitis C is common, the screening test is good and the therapy eliminates the virus in half of the cases, but problems arise in the definition of the target population. In fact generalized population screening is not endorsed by international guidelines,although some recommend screening immigrants from high prevalence countries. Opportunistic screening (case finding) of individuals with classic risk factors,such as transfusion before 1992 and drug addiction,is the most frequently used strategy, but there is disagreement whether prison inmates, individuals with a history of promiscuous or traumatic sex and health care workers should be screened. In a real practice setting the performance of opportunistic screening by general practitioners is low but can be ameliorated by training programs. Screening targeted to segments of the population or mass campaigns are expensive and therefore interventions should be aimed to improve opportunistic screening and the detection skills of general practitioners. Regarding genetic hemochromatosis there is insufficient evidence for population screening, but individual physicians can decide to screen racial groups with a high prevalence of the disease, such as people in early middle age and of northern European origin. In the other cases opportunistic screening of high risk individuals should be performed, with a high level of suspicion in case of unexplained liver disease, diabetes, juvenile artropathy, sexual dysfunction and skin pigmentation.

  7. Screening in liver disease.

    Science.gov (United States)

    Del Poggio, Paolo; Mazzoleni, Marzio

    2006-09-01

    A disease is suitable for screening if it is common, if the target population can be identified and reached and if both a good screening test and an effective therapy are available. Of the most common liver diseases only viral hepatitis and genetic hemochromatosis partially satisfy these conditions. Hepatitis C is common, the screening test is good and the therapy eliminates the virus in half of the cases, but problems arise in the definition of the target population. In fact generalized population screening is not endorsed by international guidelines, although some recommend screening immigrants from high prevalence countries. Opportunistic screening (case finding) of individuals with classic risk factors, such as transfusion before 1992 and drug addiction, is the most frequently used strategy, but there is disagreement whether prison inmates, individuals with a history of promiscuous or traumatic sex and health care workers should be screened. In a real practice setting the performance of opportunistic screening by general practitioners is low but can be ameliorated by training programs. Screening targeted to segments of the population or mass campaigns are expensive and therefore interventions should be aimed to improve opportunistic screening and the detection skills of general practitioners. Regarding genetic hemochromatosis there is insufficient evidence for population screening, but individual physicians can decide to screen racial groups with a high prevalence of the disease, such as people in early middle age and of northern European origin. In the other cases opportunistic screening of high risk individuals should be performed, with a high level of suspicion in case of unexplained liver disease, diabetes, juvenile artropathy, sexual dysfunction and skin pigmentation. PMID:16981254

  8. Antioxidant and Anti-Inflammatory Activity Determination of One Hundred Kinds of Pure Chemical Compounds Using Offline and Online Screening HPLC Assay

    Directory of Open Access Journals (Sweden)

    Kwang Jin Lee

    2015-01-01

    Full Text Available This study investigated the antioxidant activity of one hundred kinds of pure chemical compounds found within a number of natural substances and oriental medicinal herbs (OMH. Three different methods were used to evaluate the antioxidant activity of DPPH radical-scavenging activity, ABTS radical-scavenging activity, and online screening HPLC-ABTS assays. The results indicated that 17 compounds exhibited better inhibitory activity against ABTS radical than DPPH radical. The IC50 rate of a more practical substance is determined, and the ABTS assay IC50 values of gallic acid hydrate, (+-catechin hydrate, caffeic acid, rutin hydrate, hyperoside, quercetin, and kaempferol compounds were 1.03 ± 0.25, 3.12 ± 0.51, 1.59 ± 0.06, 4.68 ± 1.24, 3.54 ± 0.39, 1.89 ± 0.33, and 3.70 ± 0.15 μg/mL, respectively. The ABTS assay is more sensitive to identifying the antioxidant activity since it has faster reaction kinetics and a heightened response to antioxidants. In addition, there was a very small margin of error between the results of the offline-ABTS assay and those of the online screening HPLC-ABTS assay. We also evaluated the effects of 17 compounds on the NO secretion in LPS-stimulated RAW 264.7 cells and also investigated the cytotoxicity of 17 compounds using a cell counting kit (CCK in order to determine the optimal concentration that would provide an effective anti-inflammatory action with minimum toxicity. These results will be compiled into a database, and this method can be a powerful preselection tool for compounds intended to be studied for their potential bioactivity and antioxidant activity related to their radical-scavenging capacity.

  9. Use of electron ionization and atmospheric pressure chemical ionization in gas chromatography coupled to time-of-flight mass spetrometry for screening and identification of organic pollutants in waters

    NARCIS (Netherlands)

    Portoles, T.; Mol, J.G.J.; Sancho, J.V.; Hernandez, F.

    2014-01-01

    A new approach has been developed for multiclass screening of organic contaminants in water based on the use of gas chromatography coupled to hybrid quadrupole high-resolution time-of-flight mass spectrometry with atmospheric pressure chemical ionization (GC–(APCI)QTOF MS). The soft ionization promo

  10. Top Value-Added Chemicals from Biomass - Volume II—Results of Screening for Potential Candidates from Biorefinery Lignin

    Energy Technology Data Exchange (ETDEWEB)

    Holladay, John E.; White, James F.; Bozell, Joseph J.; Johnson, David

    2007-10-01

    roughly align with the three “product types.” From this analysis a list of technical barriers was developed which can be used to identify research needs. Lignin presents many challenges for use in the biorefinery. Chemically it differs from sugars having a complex aromatic substructure. Unlike cellulose, which has a relatively simple substructure of glucose subunits, lignin has a high degree of variability in its structure which differs both from biomass source and from the recovery process used. In addition to its variability lignin is also reactive and to some degree less stable thermally and oxidatively to other biomass streams. What this means is that integrating a lignin process stream within the biorefinery will require identifying the best method to separate lignin from biomass cost-effectively.

  11. The Functional Significance of Posttranslational Modifications on Polo-Like Kinase 1 Revealed by Chemical Genetic Complementation.

    Science.gov (United States)

    Lasek, Amber L; McPherson, Brittany M; Trueman, Natalie G; Burkard, Mark E

    2016-01-01

    Mitosis is coordinated by carefully controlled phosphorylation and ubiquitin-mediated proteolysis. Polo-like kinase 1 (Plk1) plays a central role in regulating mitosis and cytokinesis by phosphorylating target proteins. Yet, Plk1 is itself a target for posttranslational modification by phosphorylation and ubiquitination. We developed a chemical-genetic complementation assay to evaluate the functional significance of 34 posttranslational modifications (PTMs) on human Plk1. To do this, we used human cells that solely express a modified analog-sensitive Plk1 (Plk1AS) and complemented with wildtype Plk1. The wildtype Plk1 provides cells with a functional Plk1 allele in the presence of 3-MB-PP1, a bulky ATP-analog inhibitor that specifically inhibits Plk1AS. Using this approach, we evaluated the ability of 34 singly non-modifiable Plk1 mutants to complement Plk1AS in the presence of 3-MB-PP1. Mutation of the T-loop activating residue T210 and adjacent T214 are lethal, but surprisingly individual mutation of the remaining 32 posttranslational modification sites did not disrupt the essential functions of Plk1. To evaluate redundancy, we simultaneously mutated all phosphorylation sites in the kinase domain except for T210 and T214 or all sites in the C-terminal polo-box domain (PBD). We discovered that redundant phosphorylation events within the kinase domain are required for accurate chromosome segregation in anaphase but those in the PBD are dispensable. We conclude that PTMs within the T-loop of Plk1 are essential and nonredundant, additional modifications in the kinase domain provide redundant control of Plk1 function, and those in the PBD are dispensable for essential mitotic functions of Plk1. This comprehensive evaluation of Plk1 modifications demonstrates that although phosphorylation and ubiquitination are important for mitotic progression, many individual PTMs detected in human tissue may have redundant, subtle, or dispensable roles in gene function. PMID

  12. Development and Validation of a P-35S, T-nos, T-35S and P-FMV Tetraplex Real-time PCR Screening Method to Detect Regulatory Genes of Genetically Modified Organisms in Food.

    Science.gov (United States)

    Eugster, Albert; Murmann, Petra; Kaenzig, Andre; Breitenmoser, Alda

    2014-10-01

    In routine analysis screening methods based on real-time PCR (polymerase chain reaction) are most commonly used for the detection of genetically modified (GM) plant material in food and feed. Screening tests are based on sequences frequently used for GM development, allowing the detection of a large number of GMOs (genetically modified organisms). Here, we describe the development and validation of a tetraplex real-time PCR screening assay comprising detection systems for the regulatory genes Cauliflower Mosaic Virus 35S promoter, Agrobacterium tumefaciens nos terminator, Cauliflower Mosaic Virus 35S terminator and Figwort Mosaic Virus 34S promoter. Three of the four primer and probe combinations have already been published elsewhere, whereas primers and probe for the 35S terminator have been developed in-house. Adjustment of primer and probe concentrations revealed a high PCR sensitivity with insignificant physical cross-talk between the four detection channels. The sensitivity of each PCR-system is sufficient to detect a GMO concentration as low as 0.05% of the containing respective element. The specificity of the described tetraplex is high when tested on DNA from GM maize, soy, rapeseed and tomato. We also demonstrate the robustness of the system by inter-laboratory tests. In conclusion, this method provides a sensitive and reliable screening procedure for the detection of the most frequently used regulatory elements present in GM crops either authorised or unauthorised for food.

  13. Development and Validation of a P-35S, T-nos, T-35S and P-FMV Tetraplex Real-time PCR Screening Method to Detect Regulatory Genes of Genetically Modified Organisms in Food.

    Science.gov (United States)

    Eugster, Albert; Murmann, Petra; Kaenzig, Andre; Breitenmoser, Alda

    2014-10-01

    In routine analysis screening methods based on real-time PCR (polymerase chain reaction) are most commonly used for the detection of genetically modified (GM) plant material in food and feed. Screening tests are based on sequences frequently used for GM development, allowing the detection of a large number of GMOs (genetically modified organisms). Here, we describe the development and validation of a tetraplex real-time PCR screening assay comprising detection systems for the regulatory genes Cauliflower Mosaic Virus 35S promoter, Agrobacterium tumefaciens nos terminator, Cauliflower Mosaic Virus 35S terminator and Figwort Mosaic Virus 34S promoter. Three of the four primer and probe combinations have already been published elsewhere, whereas primers and probe for the 35S terminator have been developed in-house. Adjustment of primer and probe concentrations revealed a high PCR sensitivity with insignificant physical cross-talk between the four detection channels. The sensitivity of each PCR-system is sufficient to detect a GMO concentration as low as 0.05% of the containing respective element. The specificity of the described tetraplex is high when tested on DNA from GM maize, soy, rapeseed and tomato. We also demonstrate the robustness of the system by inter-laboratory tests. In conclusion, this method provides a sensitive and reliable screening procedure for the detection of the most frequently used regulatory elements present in GM crops either authorised or unauthorised for food. PMID:25437161

  14. Genetic screening for Niemann-Pick disease type C in adults with neurological and psychiatric symptoms: findings from the ZOOM study.

    Science.gov (United States)

    Bauer, Peter; Balding, David J; Klünemann, Hans H; Linden, David E J; Ory, Daniel S; Pineda, Mercè; Priller, Josef; Sedel, Frederic; Muller, Audrey; Chadha-Boreham, Harbajan; Welford, Richard W D; Strasser, Daniel S; Patterson, Marc C

    2013-11-01

    Niemann-Pick disease type C (NP-C) is a rare, autosomal-recessive, progressive neurological disease caused by mutations in either the NPC1 gene (in 95% of cases) or the NPC2 gene. This observational, multicentre genetic screening study evaluated the frequency and phenotypes of NP-C in consecutive adult patients with neurological and psychiatric symptoms. Diagnostic testing for NP-C involved NPC1 and NPC2 exonic gene sequencing and gene dosage analysis. When available, results of filipin staining, plasma cholestane-3β,5α,6β-triol assays and measurements of relevant sphingolipids were also collected. NPC1 and NPC2 gene sequencing was completed in 250/256 patients from 30 psychiatric and neurological reference centres across the EU and USA [median (range) age 38 (18-90) years]. Three patients had a confirmed diagnosis of NP-C; two based on gene sequencing alone (two known causal disease alleles) and one based on gene sequencing and positive filipin staining. A further 12 patients displayed either single mutant NP-C alleles (8 with NPC1 mutations and 3 with NPC2 mutations) or a known causal disease mutation and an unclassified NPC1 allele variant (1 patient). Notably, high plasma cholestane-3β,5α,6β-triol levels were observed for all NP-C cases (n = 3). Overall, the frequency of NP-C patients in this study [1.2% (95% CI; 0.3%, 3.5%)] suggests that there may be an underdiagnosed pool of NP-C patients among adults who share common neurological and psychiatric symptoms. PMID:23773996

  15. FINAL REPORT ON THE EVALUATION OF FOUR TOXIC CHEMICALS IN AN 'IN VIVO/IN VITRO' TOXICOLOGICAL SCREEN: ACRYLAMIDE, CHLORDECONE, CYCLOPHOSPHAMIDE, AND DIETHYLSTILBESTROL

    Science.gov (United States)

    An in vivo/in vitro Toxicological Screen (Tox Screen) has been developed for screening large numbers of wastes for biological activity. Emphasis is placed on identifying a wide range of potential toxic responses by employing diverse test methods with toxic endpoints in mutagenesi...

  16. A new in vitro screening bioassay for the ecotoxicological evaluation of the estrogenic responses of environmental chemicals using roach (Rutilus rutilus) liver explant culture.

    Science.gov (United States)

    Gerbron, Marie; Geraudie, Perrine; Rotchell, Jeanette; Minier, Christophe

    2010-10-01

    There is growing evidence that many chemicals released in the environment are able to disturb the normal endocrinology of organisms affecting the structure and function of their reproductive system. This has prompted the scientific community to develop appropriate testing methods to identify active compounds and elucidate mechanisms of action. Of particular interest are in vitro screening methods that can document the effects of these endocrine disrupting compounds on fish. In this study, an in vitro bioassay was developed in the roach (Rutilus rutilus) for evaluating the estrogenicity or antiestrogenicity potency of environmental pollutants by measuring vitellogenin (VTG) induction in cultured liver explants. The cell viability was assessed by the measurement of nonspecific esterase activity using a fluorescein diacetate hydrolysis assay. Results showed that explants could be cultured for 72 h without any significant loss of activity. Dose-dependent responses have been measured with estrogenic model compounds such as 17-β-estradiol (E2) and 17-α-ethynylestradiol (EE2) or antiestrogenic compounds such as tamoxifen. Lowest observable effective concentrations were 1 nM for E2, 1 nM for EE2, and 100 nM for tamoxifen, showing a good sensitivity of the test system. Estrogenicity of butyl 4-hydroxybenzoate, 4-nonylphenol, and bisphenol A was tested. bisphenol A (100 μM) or butylparaben induced a twofold increase in VTG production when compared with 100 nM E2, whereas this production was only 20% with 100 μM 4-nonylphenol. Overall, this study shows that the bioassay could provide valuable information on endocrine disrupting chemicals including metabolites and mixtures of compounds. PMID:20549626

  17. Dual genetic selection of synthetic riboswitches in Escherichia coli.

    Science.gov (United States)

    Nomura, Yoko; Yokobayashi, Yohei

    2014-01-01

    This chapter describes a genetic selection strategy to engineer synthetic riboswitches that can chemically regulate gene expression in Escherichia coli. Riboswitch libraries are constructed by randomizing the nucleotides that potentially comprise an expression platform and fused to the hybrid selection/screening marker tetA-gfpuv. Iterative ON and OFF selections are performed under appropriate conditions that favor the survival or the growth of the cells harboring the desired riboswitches. After the selection, rapid screening of individual riboswitch clones is performed by measuring GFPuv fluorescence without subcloning. This optimized dual genetic selection strategy can be used to rapidly develop synthetic riboswitches without detailed computational design or structural knowledge. PMID:24549616

  18. Effects of natural and chemical stressors on Enchytraeus albidus: can oxidative stress parameters be used as fast screening tools for the assessment of different stress impacts in soils?

    Science.gov (United States)

    Howcroft, C F; Amorim, M J B; Gravato, C; Guilhermino, L; Soares, A M V M

    2009-02-01

    Enchytraeids are important organisms of the soil biocenosis. They improve the soil pore structure and the degradation of organic matter. These organisms are used in standardized testing, using survival and reproduction (6 weeks) as endpoints. The use of biomarkers, linked to ecologically relevant alterations at higher levels of biological organization, is a promising tool for Environmental Risk Assessment. Here, enchytraeids were exposed for different time periods (two days and three weeks) to different soils (OECD artificial soil, different compositions in its organic matter, clay or pH value, and LUFA 2.2 natural soil) and different chemicals (Phenmedipham and copper). The main question addressed in the present study was if the effects of chemicals and different soil properties are preceded by alterations at the sub-cellular level, and if these endpoints may be used reliantly as faster screening tools for the assessment of different stress conditions in soils. The parameters measured in E. albidus whole body were: lipid peroxidation (LPO), total glutathione (TG), as well as the enzymatic activities of superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), glutathione peroxidase (GPx) and glutathione S-transferase (GST). The results showed that biomarker responses in E. albidus were significantly affected by the soil type (GST, CAT, GPx, GR and LPO) and the duration of exposure in OECD artificial soil (GST, GPx, GR, CAT and LPO) but not in LUFA 2.2 natural soil. For the abiotic factors studied, after 2 days, low pH decreased significantly the TG levels and the activities of CAT and GR,and low OM also significantly decreased CAT and GR activities. After 3 weeks, differences in soil properties caused a decrease in GR and GPx activities, whereas increased GST activity was observed due to low organic matter and pH. Copper significantly increased the activities of CAT, GPx and GR, and decreased the activity of GST after 2 days as well as inscreasing

  19. The Clinical Applications of Newborn Deafness Gene Screening and Genetic Counseling%新生儿聋病基因筛查及咨询的临床意义

    Institute of Scientific and Technical Information of China (English)

    高儒真; 陈晓巍; 历东东; 姜鸿

    2015-01-01

    目的:通过新生儿致聋基因突变筛查、遗传咨询及随访,分析新生儿常见遗传性耳聋基因的突变频率、类型及听力表型,探讨耳聋基因筛查联合新生儿听力筛查对漏筛风险的规避性及遗传咨询的应用范围和临床价值。方法分析北京市新生儿血样44200例,应用微阵列芯片法检测中国人常见的4个致聋基因的9个突变位点,对检出者进行随访并提供遗传咨询。结果本组新生儿耳聋基因突变检出率为4.26%(1884例);GJB2(1027例)与SLC26A4(617例)基因突变占比最高,总携带率为3.72%;致聋药物易感基因线粒体DNA基因突变率达0.25%(112例);明确单个耳聋基因纯合及复合性突变16例,同时携带不同基因突变个体21例;以可随访新生儿998例为进一步研究对象,建立并实践筛查阳性个体听力随访流程及遗传咨询规范。结论新生儿听力筛查联合耳聋基因筛查可有效提前确诊遗传性耳聋的时间;基因筛查可用于指导遗传咨询,通过建立病历追访和听力随访体系、掌握咨询要点,可避免、预防和减少耳聋的发生,并有效指导家族成员婚育。%Objective To determine the prevalence of deafness gene mutations in newborns and provide clinical experience for newborn genetic screening and effective genetic counseling.Methods The blood samples of 44200 newborns in Beijing were screened for hot spot mutations of GJB2,GJB3,SLC26A4 and MTRNR1 associated with hearing loss. The genetic screening results were comprehensively analyzed with hearing results in genetic counseling.Results 1 or 2 pathogenic mutations were identified in 1884(4.26%)individuals.Among them, mutations of GJB2 and SLC26A4 were commom with the rate of 3.72%.112 babies were found to carry the MTRNR1 mutation.16 compound heterozygotes and homozygotes were also identified. 21 newborns showed 2 mutations in 2 different genes.998 newborns visited

  20. Chemical processes of galvanized steel corrosion in the post-LOCA phase of a PWR and the prevention of sump screen clogging

    International Nuclear Information System (INIS)

    from those of the formation and deposition of corrosion products. Changes of the chemical composition of the circulating media were determined by chemical analysis (ICP-MS) of samples taken at defined times, and general parameters such as conductivity and pH were measured. Galvanized samples and fiber beds were examined after each experiment by means of photographic methods, light-microscopy and different kinds of chemical analysis. The chemical analyses of the depositions on fiber beds showed that the clogging is predominantly caused by the corrosion products of iron and lower amounts of zinc compounds. Thus, the corrosion of galvanized steel in boric acid is explained by a mechanism starting at the surface with fast Zn dissolution but without solid corrosion product formation. The Zn corrosion is mainly influenced by pH and concentration of zinc ions in the coolant. Since boric acid/borate acts as a buffer system, the pH value increases faster at the beginning and reaches up to 6.8 in case of sufficient Zn amount (dissolution of compact zinc or large zinc surface). A local (flow induced) corrosion occurs if a fast liquid flow strikes the top face of a horizontal galvanized coupon (small sample area). Precondition for this process is a sufficiently low pH of solution in connection with a high hydrodynamic impact of the liquid flow on the corroding surface. For a limited period, the risk of strainer clogging due to formation of corrosion products of galvanized steel may be reduced by an additional amount of submerged Zn or changing the coolant chemistry by alkaline additions. These two possibilities were investigated by test series using galvanized steel coupons. The addition of borax seems to be the most effective method to reduce the corrosion rate and the risk of sump screen clogging. The results were validated with galvanized gratings in a further test series since the flow conditions of a liquid jet on flat coupons significantly differ from those on gratings

  1. Health Screening

    Science.gov (United States)

    Screenings are tests that look for diseases before you have symptoms. Screening tests can find diseases early, when they're easier to treat. You can get some screenings in your doctor's office. Others need special equipment, ...

  2. A Syntenic Cross Species Aneuploidy Genetic Screen Links RCAN1 Expression to β-Cell Mitochondrial Dysfunction in Type 2 Diabetes

    Science.gov (United States)

    Peiris, Heshan; Duffield, Michael D.; Fadista, Joao; Kashmir, Vinder; Genders, Amanda J.; McGee, Sean L.; Martin, Alyce M.; Saiedi, Madiha; Morton, Nicholas; Carter, Roderick; Cousin, Michael A.; Oskolkov, Nikolay; Volkov, Petr; Hough, Tertius A.; Fisher, Elizabeth M. C.; Tybulewicz, Victor L. J.; Busciglio, Jorge; Coskun, Pinar E.; Becker, Ann; Belichenko, Pavel V.; Mobley, William C.; Ryan, Michael T.; Chan, Jeng Yie; Laybutt, D. Ross; Coates, P. Toby; Yang, Sijun; Ling, Charlotte; Groop, Leif; Pritchard, Melanie A.; Keating, Damien J.

    2016-01-01

    Type 2 diabetes (T2D) is a complex metabolic disease associated with obesity, insulin resistance and hypoinsulinemia due to pancreatic β-cell dysfunction. Reduced mitochondrial function is thought to be central to β-cell dysfunction. Mitochondrial dysfunction and reduced insulin secretion are also observed in β-cells of humans with the most common human genetic disorder, Down syndrome (DS, Trisomy 21). To identify regions of chromosome 21 that may be associated with perturbed glucose homeostasis we profiled the glycaemic status of different DS mouse models. The Ts65Dn and Dp16 DS mouse lines were hyperglycemic, while Tc1 and Ts1Rhr mice were not, providing us with a region of chromosome 21 containing genes that cause hyperglycemia. We then examined whether any of these genes were upregulated in a set of ~5,000 gene expression changes we had identified in a large gene expression analysis of human T2D β-cells. This approach produced a single gene, RCAN1, as a candidate gene linking hyperglycemia and functional changes in T2D β-cells. Further investigations demonstrated that RCAN1 methylation is reduced in human T2D islets at multiple sites, correlating with increased expression. RCAN1 protein expression was also increased in db/db mouse islets and in human and mouse islets exposed to high glucose. Mice overexpressing RCAN1 had reduced in vivo glucose-stimulated insulin secretion and their β-cells displayed mitochondrial dysfunction including hyperpolarised membrane potential, reduced oxidative phosphorylation and low ATP production. This lack of β-cell ATP had functional consequences by negatively affecting both glucose-stimulated membrane depolarisation and ATP-dependent insulin granule exocytosis. Thus, from amongst the myriad of gene expression changes occurring in T2D β-cells where we had little knowledge of which changes cause β-cell dysfunction, we applied a trisomy 21 screening approach which linked RCAN1 to β-cell mitochondrial dysfunction in T2D

  3. A Screen for Modifiers of Cilia Phenotypes Reveals Novel MKS Alleles and Uncovers a Specific Genetic Interaction between osm-3 and nphp-4.

    Science.gov (United States)

    Masyukova, Svetlana V; Landis, Dawn E; Henke, Scott J; Williams, Corey L; Pieczynski, Jay N; Roszczynialski, Kelly N; Covington, Jannese E; Malarkey, Erik B; Yoder, Bradley K

    2016-02-01

    Nephronophthisis (NPHP) is a ciliopathy in which genetic modifiers may underlie the variable penetrance of clinical features. To identify modifiers, a screen was conducted on C. elegans nphp-4(tm925) mutants. Mutations in ten loci exacerbating nphp-4(tm925) ciliary defects were obtained. Four loci have been identified, three of which are established ciliopathy genes mks-1, mks-2, and mks-5. The fourth allele (yhw66) is a missense mutation (S316F) in OSM-3, a kinesin required for cilia distal segment assembly. While osm-3(yhw66) mutants alone have no overt cilia phenotype, nphp-4(tm925);osm-3(yhw66) double mutants lack distal segments and are dye-filling (Dyf) and osmotic avoidance (Osm) defective, similar to osm-3(mn357) null mutants. In osm-3(yhw66) mutants anterograde intraflagellar transport (IFT) velocity is reduced. Furthermore, expression of OSM-3(S316F)::GFP reduced IFT velocities in nphp-4(tm925) mutants, but not in wild type animals. In silico analysis indicates the S316F mutation may affect a phosphorylation site. Putative phospho-null OSM-3(S316F) and phospho-mimetic OSM-3(S316D) proteins accumulate at the cilia base and tip respectively. FRAP analysis indicates that the cilia entry rate of OSM-3(S316F) is slower than OSM-3 and that in the presence of OSM-3(S316F), OSM-3 and OSM-3(S316D) rates decrease. In the presence OSM-3::GFP or OSM-3(S316D)::GFP, OSM-3(S316F)::tdTomato redistributes along the cilium and accumulates in the cilia tip. OSM-3(S316F) and OSM-3(S316D) are functional as they restore cilia distal segment formation in osm-3(mn357) null mutants; however, only OSM-3(S316F) rescues the osm-3(mn357) null Dyf phenotype. Despite rescue of cilia length in osm-3(mn357) null mutants, neither OSM-3(S316F) nor OSM-3(S316D) restores ciliary defects in nphp-4(tm925);osm-3(yhw66) double mutants. Thus, these OSM-3 mutations cause NPHP-4 dependent and independent phenotypes. These data indicate that in addition to regulating cilia protein entry or exit

  4. A Screen for Modifiers of Cilia Phenotypes Reveals Novel MKS Alleles and Uncovers a Specific Genetic Interaction between osm-3 and nphp-4.

    Directory of Open Access Journals (Sweden)

    Svetlana V Masyukova

    2016-02-01

    Full Text Available Nephronophthisis (NPHP is a ciliopathy in which genetic modifiers may underlie the variable penetrance of clinical features. To identify modifiers, a screen was conducted on C. elegans nphp-4(tm925 mutants. Mutations in ten loci exacerbating nphp-4(tm925 ciliary defects were obtained. Four loci have been identified, three of which are established ciliopathy genes mks-1, mks-2, and mks-5. The fourth allele (yhw66 is a missense mutation (S316F in OSM-3, a kinesin required for cilia distal segment assembly. While osm-3(yhw66 mutants alone have no overt cilia phenotype, nphp-4(tm925;osm-3(yhw66 double mutants lack distal segments and are dye-filling (Dyf and osmotic avoidance (Osm defective, similar to osm-3(mn357 null mutants. In osm-3(yhw66 mutants anterograde intraflagellar transport (IFT velocity is reduced. Furthermore, expression of OSM-3(S316F::GFP reduced IFT velocities in nphp-4(tm925 mutants, but not in wild type animals. In silico analysis indicates the S316F mutation may affect a phosphorylation site. Putative phospho-null OSM-3(S316F and phospho-mimetic OSM-3(S316D proteins accumulate at the cilia base and tip respectively. FRAP analysis indicates that the cilia entry rate of OSM-3(S316F is slower than OSM-3 and that in the presence of OSM-3(S316F, OSM-3 and OSM-3(S316D rates decrease. In the presence OSM-3::GFP or OSM-3(S316D::GFP, OSM-3(S316F::tdTomato redistributes along the cilium and accumulates in the cilia tip. OSM-3(S316F and OSM-3(S316D are functional as they restore cilia distal segment formation in osm-3(mn357 null mutants; however, only OSM-3(S316F rescues the osm-3(mn357 null Dyf phenotype. Despite rescue of cilia length in osm-3(mn357 null mutants, neither OSM-3(S316F nor OSM-3(S316D restores ciliary defects in nphp-4(tm925;osm-3(yhw66 double mutants. Thus, these OSM-3 mutations cause NPHP-4 dependent and independent phenotypes. These data indicate that in addition to regulating cilia protein entry or exit, NPHP-4

  5. A Syntenic Cross Species Aneuploidy Genetic Screen Links RCAN1 Expression to β-Cell Mitochondrial Dysfunction in Type 2 Diabetes.

    Directory of Open Access Journals (Sweden)

    Heshan Peiris

    2016-05-01

    Full Text Available Type 2 diabetes (T2D is a complex metabolic disease associated with obesity, insulin resistance and hypoinsulinemia due to pancreatic β-cell dysfunction. Reduced mitochondrial function is thought to be central to β-cell dysfunction. Mitochondrial dysfunction and reduced insulin secretion are also observed in β-cells of humans with the most common human genetic disorder, Down syndrome (DS, Trisomy 21. To identify regions of chromosome 21 that may be associated with perturbed glucose homeostasis we profiled the glycaemic status of different DS mouse models. The Ts65Dn and Dp16 DS mouse lines were hyperglycemic, while Tc1 and Ts1Rhr mice were not, providing us with a region of chromosome 21 containing genes that cause hyperglycemia. We then examined whether any of these genes were upregulated in a set of ~5,000 gene expression changes we had identified in a large gene expression analysis of human T2D β-cells. This approach produced a single gene, RCAN1, as a candidate gene linking hyperglycemia and functional changes in T2D β-cells. Further investigations demonstrated that RCAN1 methylation is reduced in human T2D islets at multiple sites, correlating with increased expression. RCAN1 protein expression was also increased in db/db mouse islets and in human and mouse islets exposed to high glucose. Mice overexpressing RCAN1 had reduced in vivo glucose-stimulated insulin secretion and their β-cells displayed mitochondrial dysfunction including hyperpolarised membrane potential, reduced oxidative phosphorylation and low ATP production. This lack of β-cell ATP had functional consequences by negatively affecting both glucose-stimulated membrane depolarisation and ATP-dependent insulin granule exocytosis. Thus, from amongst the myriad of gene expression changes occurring in T2D β-cells where we had little knowledge of which changes cause β-cell dysfunction, we applied a trisomy 21 screening approach which linked RCAN1 to β-cell mitochondrial

  6. A Syntenic Cross Species Aneuploidy Genetic Screen Links RCAN1 Expression to β-Cell Mitochondrial Dysfunction in Type 2 Diabetes.

    Science.gov (United States)

    Peiris, Heshan; Duffield, Michael D; Fadista, Joao; Jessup, Claire F; Kashmir, Vinder; Genders, Amanda J; McGee, Sean L; Martin, Alyce M; Saiedi, Madiha; Morton, Nicholas; Carter, Roderick; Cousin, Michael A; Kokotos, Alexandros C; Oskolkov, Nikolay; Volkov, Petr; Hough, Tertius A; Fisher, Elizabeth M C; Tybulewicz, Victor L J; Busciglio, Jorge; Coskun, Pinar E; Becker, Ann; Belichenko, Pavel V; Mobley, William C; Ryan, Michael T; Chan, Jeng Yie; Laybutt, D Ross; Coates, P Toby; Yang, Sijun; Ling, Charlotte; Groop, Leif; Pritchard, Melanie A; Keating, Damien J

    2016-05-01

    Type 2 diabetes (T2D) is a complex metabolic disease associated with obesity, insulin resistance and hypoinsulinemia due to pancreatic β-cell dysfunction. Reduced mitochondrial function is thought to be central to β-cell dysfunction. Mitochondrial dysfunction and reduced insulin secretion are also observed in β-cells of humans with the most common human genetic disorder, Down syndrome (DS, Trisomy 21). To identify regions of chromosome 21 that may be associated with perturbed glucose homeostasis we profiled the glycaemic status of different DS mouse models. The Ts65Dn and Dp16 DS mouse lines were hyperglycemic, while Tc1 and Ts1Rhr mice were not, providing us with a region of chromosome 21 containing genes that cause hyperglycemia. We then examined whether any of these genes were upregulated in a set of ~5,000 gene expression changes we had identified in a large gene expression analysis of human T2D β-cells. This approach produced a single gene, RCAN1, as a candidate gene linking hyperglycemia and functional changes in T2D β-cells. Further investigations demonstrated that RCAN1 methylation is reduced in human T2D islets at multiple sites, correlating with increased expression. RCAN1 protein expression was also increased in db/db mouse islets and in human and mouse islets exposed to high glucose. Mice overexpressing RCAN1 had reduced in vivo glucose-stimulated insulin secretion and their β-cells displayed mitochondrial dysfunction including hyperpolarised membrane potential, reduced oxidative phosphorylation and low ATP production. This lack of β-cell ATP had functional consequences by negatively affecting both glucose-stimulated membrane depolarisation and ATP-dependent insulin granule exocytosis. Thus, from amongst the myriad of gene expression changes occurring in T2D β-cells where we had little knowledge of which changes cause β-cell dysfunction, we applied a trisomy 21 screening approach which linked RCAN1 to β-cell mitochondrial dysfunction in T2D.

  7. A Syntenic Cross Species Aneuploidy Genetic Screen Links RCAN1 Expression to β-Cell Mitochondrial Dysfunction in Type 2 Diabetes.

    Science.gov (United States)

    Peiris, Heshan; Duffield, Michael D; Fadista, Joao; Jessup, Claire F; Kashmir, Vinder; Genders, Amanda J; McGee, Sean L; Martin, Alyce M; Saiedi, Madiha; Morton, Nicholas; Carter, Roderick; Cousin, Michael A; Kokotos, Alexandros C; Oskolkov, Nikolay; Volkov, Petr; Hough, Tertius A; Fisher, Elizabeth M C; Tybulewicz, Victor L J; Busciglio, Jorge; Coskun, Pinar E; Becker, Ann; Belichenko, Pavel V; Mobley, William C; Ryan, Michael T; Chan, Jeng Yie; Laybutt, D Ross; Coates, P Toby; Yang, Sijun; Ling, Charlotte; Groop, Leif; Pritchard, Melanie A; Keating, Damien J

    2016-05-01

    Type 2 diabetes (T2D) is a complex metabolic disease associated with obesity, insulin resistance and hypoinsulinemia due to pancreatic β-cell dysfunction. Reduced mitochondrial function is thought to be central to β-cell dysfunction. Mitochondrial dysfunction and reduced insulin secretion are also observed in β-cells of humans with the most common human genetic disorder, Down syndrome (DS, Trisomy 21). To identify regions of chromosome 21 that may be associated with perturbed glucose homeostasis we profiled the glycaemic status of different DS mouse models. The Ts65Dn and Dp16 DS mouse lines were hyperglycemic, while Tc1 and Ts1Rhr mice were not, providing us with a region of chromosome 21 containing genes that cause hyperglycemia. We then examined whether any of these genes were upregulated in a set of ~5,000 gene expression changes we had identified in a large gene expression analysis of human T2D β-cells. This approach produced a single gene, RCAN1, as a candidate gene linking hyperglycemia and functional changes in T2D β-cells. Further investigations demonstrated that RCAN1 methylation is reduced in human T2D islets at multiple sites, correlating with increased expression. RCAN1 protein expression was also increased in db/db mouse islets and in human and mouse islets exposed to high glucose. Mice overexpressing RCAN1 had reduced in vivo glucose-stimulated insulin secretion and their β-cells displayed mitochondrial dysfunction including hyperpolarised membrane potential, reduced oxidative phosphorylation and low ATP production. This lack of β-cell ATP had functional consequences by negatively affecting both glucose-stimulated membrane depolarisation and ATP-dependent insulin granule exocytosis. Thus, from amongst the myriad of gene expression changes occurring in T2D β-cells where we had little knowledge of which changes cause β-cell dysfunction, we applied a trisomy 21 screening approach which linked RCAN1 to β-cell mitochondrial dysfunction in T2D

  8. Morphological, chemical and genetic differentiation of two subspecies of Cistus creticus L. (C. creticus subsp. eriocephalus and C. creticus subsp. corsicus).

    Science.gov (United States)

    Paolini, Julien; Falchi, Alessandra; Quilichini, Yann; Desjobert, Jean-Marie; Cian, Marie-Cecile De; Varesi, Laurent; Costa, Jean

    2009-06-01

    Cistus creticus L., an aromatic species from the Mediterranean area, contains various diterpenes bearing the labdane skeleton. The production of essential oil from this species has potential economic value, but so far, it has not been optimized. In order to contribute to a better knowledge of this species and to its differentiation, the morphological characters, volatile chemical composition and genetic data of two subspecies (C. creticus subsp. eriocephalus and C. creticus subsp. corsicus) were investigated. The leaf trichomes were studied using scanning electron microscopy. The chemical composition of Corsican essential oil (C. creticus subsp. corsicus) has been reported using GC, GC/MS and 13C NMR; the main constituents were oxygenated labdane diterpenes (33.9%) such as 13-epi-manoyl oxide (18.5%). Using plant material (54 samples) collected from 18 geographically distinct areas of the islands of Corsica and Sardinia, the basis of variation in the headspace solid-phase microextraction volatile fraction and an inter-simple sequence repeat genetic analysis were also examined. It was shown that the two subspecies of C. creticus differed in morphology, essential oil production, volatile fraction composition and genetic data. PMID:19660770

  9. Potential of atmospheric pressure chemical ionization source in gas chromatography tandem mass spectrometry for the screening of urinary exogenous androgenic anabolic steroids.

    Science.gov (United States)

    Raro, M; Portolés, T; Pitarch, E; Sancho, J V; Hernández, F; Garrostas, L; Marcos, J; Ventura, R; Segura, J; Pozo, O J

    2016-02-01

    The atmospheric pressure chemical ionization (APCI) source for gas chromatography-mass spectrometry analysis has been evaluated for the screening of 16 exogenous androgenic anabolic steroids (AAS) in urine. The sample treatment is based on the strategy currently applied in doping control laboratories i.e. enzymatic hydrolysis, liquid-liquid extraction (LLE) and derivatization to form the trimethylsilyl ether-trimethylsilyl enol ether (TMS) derivatives. These TMS derivatives are then analyzed by gas chromatography tandem mass spectrometry using a triple quadrupole instrument (GC-QqQ MS/MS) under selected reaction monitoring (SRM) mode. The APCI promotes soft ionization with very little fragmentation resulting, in most cases, in abundant [M + H](+) or [M + H-2TMSOH](+) ions, which can be chosen as precursor ions for the SRM transitions, improving in this way the selectivity and sensitivity of the method. Specificity of the transitions is also of great relevance, as the presence of endogenous compounds can affect the measurements when using the most abundant ions. The method has been qualitatively validated by spiking six different urine samples at two concentration levels each. Precision was generally satisfactory with RSD values below 25 and 15% at the low and high concentration level, respectively. Most the limits of detection (LOD) were below 0.5 ng mL(-1). Validation results were compared with the commonly used method based on the electron ionization (EI) source. EI analysis was found to be slightly more repeatable whereas lower LODs were found for APCI. In addition, the applicability of the developed method has been tested in samples collected after the administration of 4-chloromethandienone. The highest sensitivity of the APCI method for this compound, allowed to increase the period in which its administration can be detected. PMID:26772132

  10. Potential of atmospheric pressure chemical ionization source in gas chromatography tandem mass spectrometry for the screening of urinary exogenous androgenic anabolic steroids.

    Science.gov (United States)

    Raro, M; Portolés, T; Pitarch, E; Sancho, J V; Hernández, F; Garrostas, L; Marcos, J; Ventura, R; Segura, J; Pozo, O J

    2016-02-01

    The atmospheric pressure chemical ionization (APCI) source for gas chromatography-mass spectrometry analysis has been evaluated for the screening of 16 exogenous androgenic anabolic steroids (AAS) in urine. The sample treatment is based on the strategy currently applied in doping control laboratories i.e. enzymatic hydrolysis, liquid-liquid extraction (LLE) and derivatization to form the trimethylsilyl ether-trimethylsilyl enol ether (TMS) derivatives. These TMS derivatives are then analyzed by gas chromatography tandem mass spectrometry using a triple quadrupole instrument (GC-QqQ MS/MS) under selected reaction monitoring (SRM) mode. The APCI promotes soft ionization with very little fragmentation resulting, in most cases, in abundant [M + H](+) or [M + H-2TMSOH](+) ions, which can be chosen as precursor ions for the SRM transitions, improving in this way the selectivity and sensitivity of the method. Specificity of the transitions is also of great relevance, as the presence of endogenous compounds can affect the measurements when using the most abundant ions. The method has been qualitatively validated by spiking six different urine samples at two concentration levels each. Precision was generally satisfactory with RSD values below 25 and 15% at the low and high concentration level, respectively. Most the limits of detection (LOD) were below 0.5 ng mL(-1). Validation results were compared with the commonly used method based on the electron ionization (EI) source. EI analysis was found to be slightly more repeatable whereas lower LODs were found for APCI. In addition, the applicability of the developed method has been tested in samples collected after the administration of 4-chloromethandienone. The highest sensitivity of the APCI method for this compound, allowed to increase the period in which its administration can be detected.

  11. Results from Screening Polyurethane Foam Based Consumer Products for Flame Retardant Chemicals: Assessing Impacts on the Change in the Furniture Flammability Standards

    Science.gov (United States)

    2016-01-01

    Flame retardant (FR) chemicals have often been added to polyurethane foam to meet required state and federal flammability standards. However, some FRs (e.g., PBDEs and TDCIPP) are associated with health hazards and are now restricted from use in some regions. In addition, California’s residential furniture flammability standard (TB-117) has undergone significant amendments over the past few years, and TDCIPP has been added to California’s Proposition 65 list. These events have likely led to shifts in the types of FRs used, and the products to which they are applied. To provide more information on the use of FRs in products containing polyurethane foam (PUF), we established a screening service for the general public. Participants residing in the US were allowed to submit up to 5 samples from their household for analysis, free of charge, and supplied information on the product category, labeling, and year and state of purchase. Between February 2014 and June 2016, we received 1141 PUF samples for analysis from various products including sofas, chairs, mattresses, car seats and pillows. Of these samples tested, 52% contained a FR at levels greater than 1% by weight. Tris(1,3-dichloroisopropyl)phosphate (TDCIPP) was the most common FR detected in PUF samples, and was the most common FR detected in all product categories. Analysis of the data by purchasing date suggests that the use of TDCIPP decreased in recent years, paralleled with an increase in the use of TCIPP and a nonhalogenated aryl phosphate mixture we call “TBPP.” In addition, we observed significant decreases in FR applications in furniture products and child car seats, suggesting the use of additive FRs in PUF may be declining, perhaps as a reflection of recent changes to TB-117 and Proposition 65. More studies are needed to determine how these changes in FR use relate to changes in exposure among the general population. PMID:27552529

  12. Saúde pública e ética na era da medicina genômica: rastreamentos genéticos Public health and ethics in the age of genomic medicine: genetic screening

    Directory of Open Access Journals (Sweden)

    Flavia Miranda Gomes de Constantino Bandeira

    2006-03-01

    Full Text Available O presente artigo tem como objetivo contextualizar o campo da saúde pública diante dos grandes avanços da biotecnologia e genética aplicada, destacando elementos para a problematização do tema tais como benefícios e questões éticas relacionados aos rastreamentos genéticos. O Projeto Genoma Humano gerou várias expectativas, dentre elas, a possibilidade de rastrear genes associados a doenças e comportamentos, e mais ainda, de intervir geneticamente no ser humano, levantando preocupações relativas ao renascimento da eugenia, ao aconselhamento genético, e ao uso da informação genética como critério de acesso aos planos de saúde e postos de trabalho. Uma discussão de todos esses tópicos é essencial para que a saúde pública seja beneficiada com as informações obtidas através da análise genômica das populações.This article has the objective to bring the field of public health into context in the face of the great advances of biotechnology and applied genetics, focusing on issues related to the theme such as benefits and ethics concerning genetic screening. The Human Genome Project has generated many expectations among which the possibility of screening genes associated to diseases and behaviors, moreover, the possibility of genetic interventions on humans, creating concerns related to the resurgence of Eugenia, of genetic counseling and the use of genetic information as a standard for access to healthcare clinics and jobs. The discussion of all these issues is essential to benefit public health with information obtained through population genomic analysis.

  13. The Future of Prenatal Diagnosis and Screening

    OpenAIRE

    Eugene Pergament

    2014-01-01

    The future of prenatal diagnosis and screening lies in developing clinical approaches and laboratory technologies applicable to genetic analyses and therapeutic interventions during embryonic development.

  14. Assessment of genetic response and character association for yield and yield components in Lentil (Lens culinaris L. population developed through chemical mutagenesis

    Directory of Open Access Journals (Sweden)

    Ruhul Amin

    2015-12-01

    Full Text Available Genetic variation is imperative to any plant improvement program. Therefore, this study was primarily based on this aspect of inducing desirable genetic variation for enhancement of the available lentil genetic diversity. The lentil seeds were treated with methyl methanesulfonate (MMS alone and in combination with dimethyl sulfoxide (DMSO for inducing polygenic variation as well as determining the impact of DMSO on mutagenecity of MMS. Comparative observations were recorded for bio-physiological damages, morphological variation, and quantitative traits to assess the genetic response of the lentil cultivar L 4076 toward the different concentrations of chemicals. Significant statistics suggested that the DMSO interfere with the extent of mutagenecity of MMS in lentil which could be attributed to either synergistic action of both or variation in MMS uptake. The outcome of mutagenesis on the character association study revealed that mutagenic treatments can modify significantly the manner of association between any two traits in lentil. The moderate doses of MMS in combination with 2% DMSO showed notable diminution in the biological damages while accelerating the rate of desirable high-yielding mutants had proved to be economical. The segregate of the selected mutants in future generations will definitely contribute to the improvement of Lentil genotype.

  15. Genetic counseling, prenatal screening and diagnosis of Down syndrome in the second trimester in women of advanced maternal age: a prospective study

    Institute of Scientific and Technical Information of China (English)

    QI Qing-wei; JIANG Yu-lin; ZHOU Xi-ya; LIU Jun-tao; YIN Jie; BIAN Xu-ming

    2013-01-01

    Background The incidence of autosomal trisomy in livebirths is strongly dependent on maternal age.Special consideration is given to the provision of prenatal screening and cytogenetic testing to women of advanced maternal age (AMA).The aim of this study was to evaluate the effectiveness of second trimester prenatal screening and amniocentesis for Down syndrome (DS) and compare the trends of choice of screening and amniocentesis among AMAwomen.Methods A total of 5404 AMA patients with natural singleton pregnancy were recruited for this prospective study from January 2008 to December 2010.The gestational weeks were from 15 weeks to 20+6 weeks.The patients referred were grouped into a screening group (2107 cases) and an amniocentesis group (3297 cases) by their own decision.The prevalence of DS was compared between the two groups by chi-square test.Choice rates for each maternal age with trends were compared by regression analysis.Results There were 18 cases of fetal DS detected in the screening group with a prevalence of 8.54‰ (18/2107).Twentyfive cases of fetal DS were diagnosed in the amniocentesis group with a prevalence of 7.58‰ (25/3297).No statistical difference was observed in the prevalence of DS between the screening and amniocentesis group (P=0.928).The invasive testing rate for DS in the amniocentesis group was 5.54 times higher than that of the screening group (1/131.88 vs.1/23.78).With the increase of the maternal age,the choice of amniocentesis increased while the choice of the screening showed an opposite trend.The choice of the AMA women between the screening and amniocantesis was significantly age relevant (P=0.012).Conclusions The second trimester serum screening in combination with maternal age was more effective than maternal age alone to screen for DS.We suggest educating the patients by recommending AMA women be informed of both screening and amniocentesis options.

  16. Genetic dissimilarity of ‘yellow’ and ‘sleep’ passion fruit accessions based on the fruits physical-chemical characteristics

    Directory of Open Access Journals (Sweden)

    Carlos Bernard Moreno Cerqueira-Silva

    2009-01-01

    Full Text Available The intra- and inter-specific genetic dissimilarity between 14 ‘yellow’ passion fruit plants and eight ‘sleep’passion fruit plants were evaluated through nine physical-chemical descriptors, whose measured values were submitted todescriptive (average, standard deviation and variation coefficient and inferential [univariated (ANOVA, averages andcorrelations tests and multivariated (Mahalanobis distance, hierarchical clustering, Singh coefficient and Mantel test]statistics. Intra- and, especially, inter-specific variability were found among the passion fruit plants (p < 0.001. The totalsoluble solid, equatorial diameter, total titratable acidity, and fruit weight descriptors presented the highest percentage ofrelative contribution, totaling 85.2% of the observed divergence. Preferential crossings among genotypes with physicalchemicalcharacteristics of desirable fruits and useful genetic dissimilarity in divergent and convergent crossings wereidentified.

  17. Chemical genetics and cereal starch metabolism: structural basis of the non-covalent and covalent inhibition of barley β-amylase.

    Science.gov (United States)

    Rejzek, Martin; Stevenson, Clare E; Southard, Andrew M; Stanley, Duncan; Denyer, Kay; Smith, Alison M; Naldrett, Mike J; Lawson, David M; Field, Robert A

    2011-03-01

    There are major issues regarding the proposed pathway for starch degradation in germinating cereal grain. Given the commercial importance but genetic intractability of the problem, we have embarked on a program of chemical genetics studies to identify and dissect the pathway and regulation of starch degradation in germinating barley grains. As a precursor to in vivo studies, here we report systematic analysis of the reversible and irreversible inhibition of the major β-amylase of the grain endosperm (BMY1). The molecular basis of inhibitor action was defined through high resolution X-ray crystallography studies of unliganded barley β-amylase, as well as its complexes with glycone site binder disaccharide iminosugar G1M, irreversible inhibitors α-epoxypropyl and α-epoxybutyl glucosides, which target the enzyme's catalytic residues, and the aglycone site binders acarbose and α-cyclodextrin.

  18. Twenty-year outcome analysis of genetic screening programs for Tay-Sachs and beta-thalassemia disease carriers in high schools.

    OpenAIRE

    Mitchell, J. J.; Capua, A; Clow, C.; Scriver, C R

    1996-01-01

    Programs for education, screening, and counseling of senior-high-school students, in populations at high risk for Tay-Sachs and beta-thalassemia diseases, have existed for >20 years in Montreal. Four process and outcome variables are reported here: (i) voluntary participation rates in the high-school cohort; (ii) uptake rates for the screening test; (iii) origin of carrier couples seeking the prenatal diagnosis option in the programs; and (iv) change in incidence of the two diseases. Between ...

  19. Genetic variation on 9p22 is associated with abnormal ovarian ultrasound results in the Prostate, Lung, Colorectal, and Ovarian Cancer Screening Trial.

    Directory of Open Access Journals (Sweden)

    Nicolas Wentzensen

    Full Text Available BACKGROUND: A recent ovarian cancer genome-wide association study (GWAS identified a locus on 9p22 associated with reduced ovarian cancer risk. The single nucleotide polymorphism (SNP markers localize to the BNC2 gene, which has been associated with ovarian development. METHODS: We analyzed the association of 9p22 SNPs with transvaginal ultrasound (TVU screening results and CA-125 blood levels from participants without ov