Characterization of expressed resistance gene analogs (RGAs) from peanut expressed sequence tags (ESTs)

USDA-ARS?s Scientific Manuscript database

Cultivated peanut (Arachis hypogaea L.) is one of the most important food legume crops grown worldwide, and is a major source for edible oil and protein. However, due to low genetic variation, peanut is very vulnerable to a variety of pathogens, such as early leaf spot, late leaf spot, rust and Toma...

Monitoring volatilization products using Residual Gas Analyzers during MeV ion beam irradiations

NASA Astrophysics Data System (ADS)

Wetteland, C. J.; Kriewaldt, K.; Taylor, L. A.; McSween, H. Y.; Sickafus, K. E.

2018-03-01

The use of Residual Gas Analyzers (RGAs) during irradiation experiments can provide valuable information when incorporated into experimental end-stations. The instruments can track the volatilization products of beam-sensitive materials, which may ultimately aid researchers in selecting appropriate flux values for conducting experiments. Furthermore, the type of gaseous species released during an irradiation can be monitored directly, which may lead to new insights into the radiolysis and/or heating mechanisms responsible for gas evolution. A survey of several classes of materials exposed to extremes in particle flux is presented to show how RGA instrumentation can be incorporated to qualitatively assess ion-solid interactions in a variety of fields.

De Novo Foliar Transcriptome of Chenopodium amaranticolor and Analysis of Its Gene Expression During Virus-Induced Hypersensitive Response

PubMed Central

Zhang, Yongqiang; Pei, Xinwu; Zhang, Chao; Lu, Zifeng; Wang, Zhixing; Jia, Shirong; Li, Weimin

2012-01-01

Background The hypersensitive response (HR) system of Chenopodium spp. confers broad-spectrum virus resistance. However, little knowledge exists at the genomic level for Chenopodium, thus impeding the advanced molecular research of this attractive feature. Hence, we took advantage of RNA-seq to survey the foliar transcriptome of C. amaranticolor, a Chenopodium species widely used as laboratory indicator for pathogenic viruses, in order to facilitate the characterization of the HR-type of virus resistance. Methodology and Principal Findings Using Illumina HiSeq™ 2000 platform, we obtained 39,868,984 reads with 3,588,208,560 bp, which were assembled into 112,452 unigenes (3,847 clusters and 108,605 singletons). BlastX search against the NCBI NR database identified 61,698 sequences with a cut-off E-value above 10−5. Assembled sequences were annotated with gene descriptions, GO, COG and KEGG terms, respectively. A total number of 738 resistance gene analogs (RGAs) and homology sequences of 6 key signaling proteins within the R proteins-directed signaling pathway were identified. Based on this transcriptome data, we investigated the gene expression profiles over the stage of HR induced by Tobacco mosaic virus and Cucumber mosaic virus by using digital gene expression analysis. Numerous candidate genes specifically or commonly regulated by these two distinct viruses at early and late stages of the HR were identified, and the dynamic changes of the differently expressed genes enriched in the pathway of plant-pathogen interaction were particularly emphasized. Conclusions To our knowledge, this study is the first description of the genetic makeup of C. amaranticolor, providing deep insight into the comprehensive gene expression information at transcriptional level in this species. The 738 RGAs as well as the differentially regulated genes, particularly the common genes regulated by both TMV and CMV, are suitable candidates which merit further functional characterization

Identification of RFLP and NBS/PK profiling markers for disease resistance loci in genetic maps of oats.

PubMed

Sanz, M J; Loarce, Y; Fominaya, A; Vossen, J H; Ferrer, E

2013-01-01

Two of the domains most widely shared among R genes are the nucleotide binding site (NBS) and protein kinase (PK) domains. The present study describes and maps a number of new oat resistance gene analogues (RGAs) with two purposes in mind: (1) to identify genetic regions that contain R genes and (2) to determine whether RGAs can be used as molecular markers for qualitative loci and for QTLs affording resistance to Puccinia coronata. Such genes have been mapped in the diploid A. strigosa × A. wiestii (Asw map) and the hexaploid MN841801-1 × Noble-2 (MN map). Genomic and cDNA NBS-RGA probes from oat, barley and wheat were used to produce RFLPs and to obtain markers by motif-directed profiling based on the NBS (NBS profiling) and PK (PK profiling) domains. The efficiency of primers used in NBS/PK profiling to amplify RGA fragments was assessed by sequencing individual marker bands derived from genomic and cDNA fragments. The positions of 184 markers were identified in the Asw map, while those for 99 were identified in the MN map. Large numbers of NBS and PK profiling markers were found in clusters across different linkage groups, with the PK profiling markers more evenly distributed. The location of markers throughout the genetic maps and the composition of marker clusters indicate that NBS- and PK-based markers cover partly complementary regions of oat genomes. Markers of the different classes obtained were found associated with the two resistance loci, PcA and R-284B-2, mapped on Asw, and with five out of eight QTLs for partial resistance in the MN map. 53 RGA-RFLPs and 187 NBS/PK profiling markers were also mapped on the hexaploid map A. byzantina cv. Kanota × A. sativa cv. Ogle. Significant co-localization was seen between the RGA markers in the KO map and other markers closely linked to resistance loci, such as those for P. coronata and barley yellow dwarf virus (Bydv) that were previously mapped in other segregating populations.

Highly Sensitive H2S Sensor Based on the Metal-Catalyzed SnO2 Nanocolumns Fabricated by Glancing Angle Deposition

PubMed Central

Yoo, Kwang Soo; Han, Soo Deok; Moon, Hi Gyu; Yoon, Seok-Jin; Kang, Chong-Yun

2015-01-01

As highly sensitive H2S gas sensors, Au- and Ag-catalyzed SnO2 thin films with morphology-controlled nanostructures were fabricated by using e-beam evaporation in combination with the glancing angle deposition (GAD) technique. After annealing at 500 °C for 40 h, the sensors showed a polycrystalline phase with a porous, tilted columnar nanostructure. The gas sensitivities (S = Rgas/Rair) of Au and Ag-catalyzed SnO2 sensors fabricated by the GAD process were 0.009 and 0.015, respectively, under 5 ppm H2S at 300 °C, and the 90% response time was approximately 5 s. These sensors showed excellent sensitivities compared with the SnO2 thin film sensors that were deposited normally (glancing angle = 0°, S = 0.48). PMID:26134105

THE USE OF KF STYLE FLANGES IN LOW PARTICULATE APPLICATIONS

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kendziora, K. R.; Angelo, J.; Baffes, C.

As SCRF particle accelerator technology advances the need for “low particulate” and “particle free” vacuum sys-tems becomes greater and greater. In the course of the op-eration of these systems, there comes a time when various instruments have to be temporarily attached for diagnostic purposes: RGAs, leak detectors, and additional pumps. In an effort to make the additions of these instruments easier and more time effective, we propose to use KF style flanges for these types of temporary diagnostic connec-tions. This document will describe the tests used to com-pare the particles generated using the assembly of the, widely accepted for “particlemore » free” use, conflat flange to the proposed KF style flange, and demonstrate that KF flanges produce comparable or even less particles« less

Removal of natural hormones in dairy farm wastewater using reactive and sorptive materials.

PubMed

Cai, Kai; Phillips, Debra H; Elliott, Christopher T; Muller, Marc; Scippo, Marie-Louise; Connolly, Lisa

2013-09-01

The objective of this study was to examine the oestrogen and androgen hormone removal efficiency of reactive (Connelly zero-valent iron (ZVI), Gotthart Maier ZVI) and sorptive (AquaSorb 101 granular activated carbon (GAC) and OrganoLoc PM-100 organoclay (OC)) materials from HPLC grade water and constructed wetland system (CWS) treated dairy farm wastewater. Batch test studies were performed and hormone concentration analysis carried out using highly sensitive reporter gene assays (RGAs). The results showed that hormonal interaction with these materials is selective for individual classes of hormones. Connelly ZVI and AquaSorb 101 GAC were more efficient in removing testosterone (Te) than 17β-estradiol (E2) and showed faster removal rates of oestrogen and androgen than the other materials. Gotthart Maier ZVI was more efficient in removing E2 than Te. OrganoLoc PM-100 OC achieved the lowest final concentration of E2 equivalent (EEQ) and provided maximum removal of both oestrogens and androgens. Copyright © 2013 Elsevier B.V. All rights reserved.

SLS Model Based Design: A Navigation Perspective

NASA Technical Reports Server (NTRS)

Oliver, T. Emerson; Anzalone, Evan; Park, Thomas; Geohagan, Kevin

2018-01-01

The SLS Program has implemented a Model-based Design (MBD) and Model-based Requirements approach for managing component design information and system requirements. This approach differs from previous large-scale design efforts at Marshall Space Flight Center where design documentation alone conveyed information required for vehicle design and analysis and where extensive requirements sets were used to scope and constrain the design. The SLS Navigation Team is responsible for the Program-controlled Design Math Models (DMMs) which describe and represent the performance of the Inertial Navigation System (INS) and the Rate Gyro Assemblies (RGAs) used by Guidance, Navigation, and Controls (GN&C). The SLS Navigation Team is also responsible for navigation algorithms. The navigation algorithms are delivered for implementation on the flight hardware as a DMM. For the SLS Block 1B design, the additional GPS Receiver hardware model is managed as a DMM at the vehicle design level. This paper describes the models, and discusses the processes and methods used to engineer, design, and coordinate engineering trades and performance assessments using SLS practices as applied to the GN&C system, with a particular focus on the navigation components.

Coaxial electrospinning of WO3 nanotubes functionalized with bio-inspired Pd catalysts and their superior hydrogen sensing performance

NASA Astrophysics Data System (ADS)

Choi, Seon-Jin; Chattopadhyay, Saptarshi; Kim, Jae Jin; Kim, Sang-Joon; Tuller, Harry L.; Rutledge, Gregory C.; Kim, Il-Doo

2016-04-01

Macroporous WO3 nanotubes (NTs) functionalized with nanoscale catalysts were fabricated using coaxial electrospinning combined with sacrificial templating and protein-encapsulated catalysts. The macroporous thin-walled nanotubular structures were obtained by introducing colloidal polystyrene (PS) particles to a shell solution of W precursor and poly(vinylpyrrolidone). After coaxial electrospinning with a core liquid of mineral oil and subsequent calcination, open pores with an average diameter of 173 nm were formed on the surface of WO3 NTs due to decomposition of the PS colloids. In addition, catalytic Pd nanoparticles (NPs) were synthesized using bio-inspired protein cages, i.e., apoferritin, and uniformly dispersed within the shell solution and subsequently on the WO3 NTs. The resulting Pd functionalized macroporous WO3 NTs were demonstrated to be high performance hydrogen (H2) sensors. In particular, Pd-functionalized macroporous WO3 NTs exhibited a very high H2 response (Rair/Rgas) of 17.6 at 500 ppm with a short response time. Furthermore, the NTs were shown to be highly selective for H2 compared to other gases such as carbon monoxide (CO), ammonia (NH3), and methane (CH4). The results demonstrate a new synthetic method to prepare highly porous nanotubular structures with well-dispersed nanoscale catalysts, which can provide improved microstructures for chemical sensing.Macroporous WO3 nanotubes (NTs) functionalized with nanoscale catalysts were fabricated using coaxial electrospinning combined with sacrificial templating and protein-encapsulated catalysts. The macroporous thin-walled nanotubular structures were obtained by introducing colloidal polystyrene (PS) particles to a shell solution of W precursor and poly(vinylpyrrolidone). After coaxial electrospinning with a core liquid of mineral oil and subsequent calcination, open pores with an average diameter of 173 nm were formed on the surface of WO3 NTs due to decomposition of the PS colloids. In addition

The role of temperature in the variability and extremes of electricity and gas demand in Great Britain

NASA Astrophysics Data System (ADS)

Thornton, H. E.; Hoskins, B. J.; Scaife, A. A.

2016-11-01

The daily relationship of electricity and gas demand with temperature in Great Britain is analysed from 1975 to 2013 and 1996 to 2013 respectively. The annual mean and annual cycle amplitude of electricity demand exhibit low frequency variability. This low frequency variability is thought to be predominantly driven by socio-economic changes rather than temperature variation. Once this variability is removed, both daily electricity and gas demand have a strong anti-correlation with temperature (r elec = -0.90 , r gas = -0.94). However these correlations are inflated by the changing demand-temperature relationship during spring and autumn. Once the annual cycles of temperature and demand are removed, the correlations are {r}{{elec}}=-0.60 and {r}{{gas}}=-0.83. Winter then has the strongest demand-temperature relationship, during which a 1 °C reduction in daily temperature typically gives a ˜1% increase in daily electricity demand and a 3%-4% increase in gas demand. Extreme demand periods are assessed using detrended daily temperature observations from 1772. The 1 in 20 year peak day electricity and gas demand estimates are, respectively, 15% (range 14%-16%) and 46% (range 44%-49%) above their average winter day demand during the last decade. The risk of demand exceeding recent extreme events, such as during the winter of 2009/2010, is also quantified.

[Separation and determination of eight plant hormones by reversed-phase high performance liquid chromatography].

PubMed

Fang, N; Hou, S; Shao, X; He, Y; Zhao, G

1998-09-01

In this paper, reversed-phase high performance liquid chromatographic technique was used for the separation and determination of eight plant hormones. Methanol-water-acetic acid system was chosen as the mobile phase. The effects of different separation conditions, such as the methanol and acetic acid concentrations in mobile phase, on the retention behaviours of eight plant hormones in this system were studied. The general trends in retention behaviours could be correlated to the methanol concentration in mobile phase. The experimental results showed that the optimum separation was achieved with following gradient elution condition: 0-3 minutes, 70% (water percentage in mobile phase), 3-13 minutes, 70%-20%, 13-48 minutes, 20%. Benzene was added to be as the internal standard. Under this experimental condition, the eight plant hormones could be separated completely and detected quantitatively at 260 nm within 16 minutes. The calibration curves for the eight compounds gave linearity over a wide range. The correlation coefficients of each components were r(ZT) = 0.9971, r(GAs) = 0.9999, r(K) = 0.9997, r(BA) = 0.9995, r(IAA) = 0.9998, r(IPA) = 0.9982, r(IBA) = 0.9995 and r(NAA) = 0.9995. The method is rapid, simple and efficient. It is a suitable method for the accurate determination of gibberellic acid (GA) and alpha-naphthaleneacetic acid (alpha-NAA) in products for agricultural use.

User's guide to the Residual Gas Analyzer (RGA)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Artman, S.A.

1988-08-04

The Residual Gas Analyzer (RGA), a Model 100C UTI quadrupole mass spectrometer, measures the concentrations of selected masses in the Fusion Energy Division's (FED) Advanced Toroidal Facility (ATF). The RGA software is a VAX FORTRAN computer program which controls the experimental apparatus, records the raw data, performs data reduction, and plots the data. The RGA program allows data to be collected from an RGA on ATF or from either of two RGAs in the laboratory. In the laboratory, the RGA diagnostic plays an important role in outgassing studied on various candidate materials for fusion experiments. One such material, graphite, ismore » being used more often in fusion experiments due to its ability to withstand high power loads. One of the functions of the RGA diagnostic is aid in the determination of the best grade of graphite to be used in these experiments and to study the procedures used to condition it. A procedure of particular interest involves baking the graphite sample in order to remove impurities that may be present in it. These impurities can be studied while in the ATF plasma or while being baked and outgassed in the laboratory. The Residual Gas Analyzer is a quadrupole mass spectrometer capable of scanning masses ranging in size from 1 atomic mass unit (amu) to 300 amu while under computer control. The procedure for collecting data for a particular mass is outlined.« less

Identification and characterization of a NBS–LRR class resistance gene analog in Pistacia atlantica subsp. Kurdica

PubMed Central

Bahramnejad, Bahman

2014-01-01

P. atlantica subsp. Kurdica, with the local name of Baneh, is a wild medicinal plant which grows in Kurdistan, Iran. The identification of resistance gene analogs holds great promise for the development of resistant cultivars. A PCR approach with degenerate primers designed according to conserved NBS-LRR (nucleotide binding site-leucine rich repeat) regions of known disease-resistance (R) genes was used to amplify and clone homologous sequences from P. atlantica subsp. Kurdica. A DNA fragment of the expected 500-bp size was amplified. The nucleotide sequence of this amplicon was obtained through sequencing and the predicted amino acid sequence compared to the amino acid sequences of known R-genes revealed significant sequence similarity. Alignment of the deduced amino acid sequence of P. atlantica subsp. Kurdica resistance gene analog (RGA) showed strong identity, ranging from 68% to 77%, to the non-toll interleukin receptor (non-TIR) R-gene subfamily from other plants. A P-loop motif (GMMGGEGKTT), a conserved and hydrophobic motif GLPLAL, a kinase-2a motif (LLVLDDV), when replaced by IAVFDDI in PAKRGA1 and a kinase-3a (FGPGSRIII) were presented in all RGA. A phylogenetic tree, based on the deduced amino-acid sequences of PAKRGA1 and RGAs from different species indicated that they were separated in two clusters, PAKRGA1 being on cluster II. The isolated NBS analogs can be eventually used as guidelines to isolate numerous R-genes in Pistachio. PMID:27843981

In vitro bioassay investigations of the endocrine disrupting potential of steviol glycosides and their metabolite steviol, components of the natural sweetener Stevia.

PubMed

Shannon, Maeve; Rehfeld, Anders; Frizzell, Caroline; Livingstone, Christina; McGonagle, Caoimhe; Skakkebaek, Niels E; Wielogórska, Ewa; Connolly, Lisa

2016-05-15

The food industry is moving towards the use of natural sweeteners such as those produced by Stevia rebaudiana due to the number of health and safety concerns surrounding artificial sweeteners. Despite the fact that these sweeteners are natural; they cannot be assumed safe. Steviol glycosides have a steroidal structure and therefore may have the potential to act as an endocrine disruptor in the body. Reporter gene assays (RGAs), H295R steroidogenesis assay and Ca(2+) fluorimetry based assays using human sperm cells have been used to assess the endocrine disrupting potential of two steviol glycosides: stevioside and rebaudioside A, and their metabolite steviol. A decrease in transcriptional activity of the progestagen receptor was seen following treatment with 25,000 ng/ml steviol in the presence of progesterone (157 ng/ml) resulting in a 31% decrease in progestagen response (p=<0.01). At the level of steroidogenesis, the metabolite steviol (500-25,000 ng/ml) increased progesterone production significantly by 2.3 fold when exposed to 10,000 ng/ml (p=<0.05) and 5 fold when exposed to 25,000 ng/ml (p=<0.001). Additionally, steviol was found to induce an agonistic response on CatSper, a progesterone receptor of sperm, causing a rapid influx of Ca(2+). The response was fully inhibited using a specific CatSper inhibitor. These findings highlight the potential for steviol to act as a potential endocrine disruptor. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Spatio-temporal diversification of the cell wall matrix materials in the developing stomatal complexes of Zea mays.

PubMed

Giannoutsou, E; Apostolakos, P; Galatis, B

2016-11-01

The matrix cell wall materials, in developing Zea mays stomatal complexes are asymmetrically distributed, a phenomenon appearing related to the local cell wall expansion and deformation, the establishment of cell polarity, and determination of the cell division plane. In cells of developing Zea mays stomatal complexes, definite cell wall regions expand determinately and become locally deformed. This differential cell wall behavior is obvious in the guard cell mother cells (GMCs) and the subsidiary cell mother cells (SMCs) that locally protrude towards the adjacent GMCs. The latter, emitting a morphogenetic stimulus, induce polarization/asymmetrical division in SMCs. Examination of immunolabeled specimens revealed that homogalacturonans (HGAs) with a high degree of de-esterification (2F4- and JIM5-HGA epitopes) and arabinogalactan proteins are selectively distributed in the extending and deformed cell wall regions, while their margins are enriched with rhamnogalacturonans (RGAs) containing highly branched arabinans (LM6-RGA epitope). In SMCs, the local cell wall matrix differentiation constitutes the first structural event, indicating the establishment of cell polarity. Moreover, in the premitotic GMCs and SMCs, non-esterified HGAs (2F4-HGA epitope) are preferentially localized in the cell wall areas outlining the cytoplasm where the preprophase band is formed. In these areas, the forthcoming cell plate fuses with the parent cell walls. These data suggest that the described heterogeneity in matrix cell wall materials is probably involved in: (a) local cell wall expansion and deformation, (b) the transduction of the inductive GMC stimulus, and (c) the determination of the division plane in GMCs and SMCs.

Linkage maps of grapevine displaying the chromosomal locations of 420 microsatellite markers and 82 markers for R-gene candidates.

PubMed

Di Gaspero, G; Cipriani, G; Adam-Blondon, A-F; Testolin, R

2007-05-01

Genetic maps functionally oriented towards disease resistance have been constructed in grapevine by analysing with a simultaneous maximum-likelihood estimation of linkage 502 markers including microsatellites and resistance gene analogs (RGAs). Mapping material consisted of two pseudo-testcrosses, 'Chardonnay' x 'Bianca' and 'Cabernet Sauvignon' x '20/3' where the seed parents were Vitis vinifera genotypes and the male parents were Vitis hybrids carrying resistance to mildew diseases. Individual maps included 320-364 markers each. The simultaneous use of two mapping crosses made with two pairs of distantly related parents allowed mapping as much as 91% of the markers tested. The integrated map included 420 Simple Sequence Repeat (SSR) markers that identified 536 SSR loci and 82 RGA markers that identified 173 RGA loci. This map consisted of 19 linkage groups (LGs) corresponding to the grape haploid chromosome number, had a total length of 1,676 cM and a mean distance between adjacent loci of 3.6 cM. Single-locus SSR markers were randomly distributed over the map (CD = 1.12). RGA markers were found in 18 of the 19 LGs but most of them (83%) were clustered on seven LGs, namely groups 3, 7, 9, 12, 13, 18 and 19. Several RGA clusters mapped to chromosomal regions where phenotypic traits of resistance to fungal diseases such as downy mildew and powdery mildew, bacterial diseases such as Pierce's disease, and pests such as dagger and root-knot nematode, were previously mapped in different segregating populations. The high number of RGA markers integrated into this new map will help find markers linked to genetic determinants of different pest and disease resistances in grape.

The heat capacity of hydrous cordierite above 295 K

NASA Astrophysics Data System (ADS)

Carey, J. William

1993-04-01

The heat capacity of synthetic hydrous cordierite (Mg2Al4Si5O18·nH2O) has been determined by differential scanning calorimetry (DSC) from 295 to 425 K as a function of H2O content. Six samples with H2O contents ranging from 0 to 0.82 per formula unit were examined. The partial molar heat capacity of H2O in cordierite over the measured temperature interval is independent of composition and temperature within experimental uncertainty and is equal to 43.3 ±0.8 J/mol/ K. This value exceeds the molar heat capacity of gaseous H2O by 9.7 J/mol/K, but is significantly smaller than the heat capacity of H2O in several zeolites and liquid H2O. A statistical-mechanical model of the heat capacity of adsorbed gas species (Barrer 1978) is used to extrapolate the heat capacity of hydrous cordierite to temperatures greater than 425 K. In this model, the heat capacity of hydrous cordierite (Crd·nH2O) is represented as follows: Cp(Crd · nH2O) = Cp(Crd)+ n{Cp(H2O, gas)+ R(gas constant)} (1) An examination of calorimetric data for hydrous beryl, analcime, mordenite, and clinoptilolite (Hemingway et al. 1986; Johnson et al. 1982, 1991, 1992) demonstrates the general applicability of the statistical-mechanical model for the extrapolation of heat capacity data of zeolitic minerals. The heat capacity data for cordierite are combined with the data of Carey and Navrotsky (1992) to obtain the molar enthalpy of formation and enthalpy of hydration of hydrous cordierite as a function of temperature.

Reporter enzyme inhibitor study to aid assembly of orthogonal reporter gene assays.

PubMed

Ho, Pei-i; Yue, Kimberley; Pandey, Pramod; Breault, Lyne; Harbinski, Fred; McBride, Aaron J; Webb, Brian; Narahari, Janaki; Karassina, Natasha; Wood, Keith V; Hill, Adam; Auld, Douglas S

2013-05-17

Reporter gene assays (RGAs) are commonly used to measure biological pathway modulation by small molecules. Understanding how such compounds interact with the reporter enzyme is critical to accurately interpret RGA results. To improve our understanding of reporter enzymes and to develop optimal RGA systems, we investigated eight reporter enzymes differing in brightness, emission spectrum, stability, and substrate requirements. These included common reporter enzymes such as firefly luciferase (Photinus pyralis), Renilla reniformis luciferase, and β-lactamase, as well as mutated forms of R. reniformis luciferase emitting either blue- or green-shifted luminescence, a red-light emitting form of Luciola cruciata firefly luciferase, a mutated form of Gaussia princeps luciferase, and a proprietary luciferase termed "NanoLuc" derived from the luminescent sea shrimp Oplophorus gracilirostris. To determine hit rates and structure-activity relationships, we screened a collection of 42,460 PubChem compounds at 10 μM using purified enzyme preparations. We then compared hit rates and chemotypes of actives for each enzyme. The hit rates ranged from <0.1% for β-lactamase to as high as 10% for mutated forms of Renilla luciferase. Related luciferases such as Renilla luciferase mutants showed high degrees of inhibitor overlap (40-70%), while unrelated luciferases such as firefly luciferases, Gaussia luciferase, and NanoLuc showed <10% overlap. Examination of representative inhibitors in cell-based assays revealed that inhibitor-based enzyme stabilization can lead to increases in bioluminescent signal for firefly luciferase, Renilla luciferase, and NanoLuc, with shorter half-life reporters showing increased activation responses. From this study we suggest strategies to improve the construction and interpretation of assays employing these reporter enzymes.

Genome Mapping and Molecular Breeding of Tomato

PubMed Central

Foolad, Majid R.

2007-01-01

The cultivated tomato, Lycopersicon esculentum, is the second most consumed vegetable worldwide and a well-studied crop species in terms of genetics, genomics, and breeding. It is one of the earliest crop plants for which a genetic linkage map was constructed, and currently there are several molecular maps based on crosses between the cultivated and various wild species of tomato. The high-density molecular map, developed based on an L. esculentum × L. pennellii cross, includes more than 2200 markers with an average marker distance of less than 1 cM and an average of 750 kbp per cM. Different types of molecular markers such as RFLPs, AFLPs, SSRs, CAPS, RGAs, ESTs, and COSs have been developed and mapped onto the 12 tomato chromosomes. Markers have been used extensively for identification and mapping of genes and QTLs for many biologically and agriculturally important traits and occasionally for germplasm screening, fingerprinting, and marker-assisted breeding. The utility of MAS in tomato breeding has been restricted largely due to limited marker polymorphism within the cultivated species and economical reasons. Also, when used, MAS has been employed mainly for improving simply-inherited traits and not much for improving complex traits. The latter has been due to unavailability of reliable PCR-based markers and problems with linkage drag. Efforts are being made to develop high-throughput markers with greater resolution, including SNPs. The expanding tomato EST database, which currently includes ∼214 000 sequences, the new microarray DNA chips, and the ongoing sequencing project are expected to aid development of more practical markers. Several BAC libraries have been developed that facilitate map-based cloning of genes and QTLs. Sequencing of the euchromatic portions of the tomato genome is paving the way for comparative and functional analysis of important genes and QTLs. PMID:18364989

An in vitro investigation of endocrine disrupting effects of the mycotoxin alternariol

DOE Office of Scientific and Technical Information (OSTI.GOV)

Frizzell, Caroline; Ndossi, Doreen; Sokoine University of Agriculture, Morogoro

2013-08-15

Alternariol (AOH) is a mycotoxin commonly produced by Alternaria alternata on a wide range of foods. Few studies to date have been performed to evaluate the effects of AOH on endocrine activity. The present study makes use of in vitro mammalian cellular based assays and gene expression to investigate the ability of AOH to act as an endocrine disruptor by various modes of action. Reporter gene assays (RGAs), incorporating natural steroid hormone receptors for oestrogens, androgens, progestagens and glucocorticoids were used to identify endocrine disruption at the level of nuclear receptor transcriptional activity, and the H295R steroidogenesis assay was usedmore » to assess endocrine disruption at the level of gene expression and steroid hormone production. AOH exhibited a weak oestrogenic response when tested in the oestrogen responsive RGA and binding of progesterone to the progestagen receptor was shown to be synergistically increased in the presence of AOH. H295R cells when exposed to 0.1–1000 ng/ml AOH, did not cause a significant change in testosterone and cortisol hormones but exposure to 1000 ng/ml (3.87 μM) AOH resulted in a significant increase in estradiol and progesterone production. In the gene expression study following exposure to 1000 ng/ml (3.87 μM) AOH, only one gene NR0B1 was down-regulated, whereas expression of mRNA for CYP1A1, MC2R, HSD3B2, CYP17, CYP21, CYP11B2 and CYP19 was up-regulated. Expression of the other genes investigated did not change significantly. In conclusion AOH is a weak oestrogenic mycotoxin that also has the ability to interfere with the steroidogenesis pathway. - Highlights: • Alternariol was investigated for endocrine disrupting activity. • Reporter gene assays and the H295R steroidogenesis assay have been used. • An oestrogenic effect of alternariol was observed. • This can lead to an increase in expression of the progesterone receptor. • Alternariol is capable of modulating hormone production and gene

Identification and characterization of potential NBS-encoding resistance genes and induction kinetics of a putative candidate gene associated with downy mildew resistance in Cucumis

PubMed Central

2010-01-01

Background Due to the variation and mutation of the races of Pseudoperonospora cubensis, downy mildew has in recent years become the most devastating leaf disease of cucumber worldwide. Novel resistance to downy mildew has been identified in the wild Cucumis species, C. hystrix Chakr. After the successful hybridization between C. hystrix and cultivated cucumber (C. sativus L.), an introgression line (IL5211S) was identified as highly resistant to downy mildew. Nucleotide-binding site and leucine-rich repeat (NBS-LRR) genes are the largest class of disease resistance genes cloned from plant with highly conserved domains, which can be used to facilitate the isolation of candidate genes associated with downy mildew resistance in IL5211S. Results Degenerate primers that were designed based on the conserved motifs in the NBS domain of resistance (R) proteins were used to isolate NBS-type sequences from IL5211S. A total of 28 sequences were identified and named as cucumber (C. sativus = CS) resistance gene analogs as CSRGAs. Polygenetic analyses separated these sequences into four different classes. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis showed that these CSRGAs expressed at different levels in leaves, roots, and stems. In addition, introgression from C. hystrix induced expression of the partial CSRGAs in cultivated cucumber, especially CSRGA23, increased four-fold when compared to the backcross parent CC3. Furthermore, the expression of CSRGA23 under P. cubensis infection and abiotic stresses was also analyzed at different time points. Results showed that the P. cubensis treatment and four tested abiotic stimuli, MeJA, SA, ABA, and H2O2, triggered a significant induction of CSRGA23 within 72 h of inoculation. The results indicate that CSRGA23 may play a critical role in protecting cucumber against P. cubensis through a signaling the pathway triggered by these molecules. Conclusions Four classes of NBS-type RGAs were successfully isolated

Multi-Objective Optimization of Transmission Lines / Elektropārvades Līnijas Daudzkriteriālā Optimizācija

NASA Astrophysics Data System (ADS)

Berjozkina, S.; Sauhats, A.; Neimane, V.

2013-10-01

Introduction of new advanced electrical connections into a transmission grid reduces the capacity of existing overhead lines (OHLs). At the same time, designing & building of new OHLs and substations involves considerable technical, environmental and economical problems. The authors propose a concept of the multi-objective optimization for selection of transmission line routes, towers (their type, placement and geometry), of conductors, insulators, dampers, earthing and lightning protection systems, span lengths, etc.. The optimization is organized in five stages. At the first and second stages a search for optimum solutions is performed along with determination of the main impacting factors. The next two stages present a two-objective optimization based on Pareto's approach. At the last stage (exemplified by a case study), the probability of the restriction removal conditions is assessed, and preventive measures are identified. The presented approach uses a real line design and is intended for minimizing the total invested capital and maximizing the net present value. In the framework of this approach 20 alternatives have been elaborated, which can successfully be applied in the cases described in the paper. Elektropārvades tīklam rodas nepieciešamība pēc jauniem elektriskajiem pieslēgumiem, kas noved pie esošo gaisvadu līniju jaudas nepietiekamības. Viens no iespējamajiem pastāvošās problēmas risinājumiem ir jaunu gaisvadu līniju un apakšstacijas būvniecība. Gaisvadu līniju projektēšana ir saistīta ar ievērojamām tehniskām, vides un ekonomiskām problēmām. Darbā aprakstīta elektropārvades līnijas optimālās trases izvēles daudzkritēriju optimizācijas koncepcija, ieskaitot balstu tipa, balstu izvietojuma koordināšu, balstu ģeometrijas, vadu tipu un parametru, izolatoru tipu, vibroslāpētāju tipu, zibensaizsardzības un zemēšanas sistēmu, kā arī laidumu garumu izvēles optimizāciju. Optimizācijas uzdevums tiek organiz

Two alternative recessive quantitative trait loci influence resistance to spring black stem and leaf spot in Medicago truncatula.

PubMed

Kamphuis, Lars G; Lichtenzveig, Judith; Oliver, Richard P; Ellwood, Simon R

2008-03-26

cluster of Toll/Interleukin1 receptor-nucleotide binding site-leucine-rich repeat (TIR-NBS-LRR) genes and disease resistance protein-like genes, while no resistance gene analogues (RGAs) are apparent in the genomic sequence of the reference accession A17 at the rnpm2 locus. The induction of defence responses and cell death in the susceptible interaction following infection by P. medicaginis suggested this pathogen is not negatively affected by these responses and may promote them. A QTL for resistance was revealed in each of two populations derived from crosses between a resistant accession and two different susceptible accessions. Both loci are recessive in nature, and the simplest explanation for the existence of two separate QTLs is the occurrence of host genotype-specific susceptibility loci that may interact with undetermined P. medicaginis virulence factors.

SLS Navigation Model-Based Design Approach

NASA Technical Reports Server (NTRS)

Oliver, T. Emerson; Anzalone, Evan; Geohagan, Kevin; Bernard, Bill; Park, Thomas

2018-01-01

The SLS Program chose to implement a Model-based Design and Model-based Requirements approach for managing component design information and system requirements. This approach differs from previous large-scale design efforts at Marshall Space Flight Center where design documentation alone conveyed information required for vehicle design and analysis and where extensive requirements sets were used to scope and constrain the design. The SLS Navigation Team has been responsible for the Program-controlled Design Math Models (DMMs) which describe and represent the performance of the Inertial Navigation System (INS) and the Rate Gyro Assemblies (RGAs) used by Guidance, Navigation, and Controls (GN&C). The SLS Navigation Team is also responsible for the navigation algorithms. The navigation algorithms are delivered for implementation on the flight hardware as a DMM. For the SLS Block 1-B design, the additional GPS Receiver hardware is managed as a DMM at the vehicle design level. This paper provides a discussion of the processes and methods used to engineer, design, and coordinate engineering trades and performance assessments using SLS practices as applied to the GN&C system, with a particular focus on the Navigation components. These include composing system requirements, requirements verification, model development, model verification and validation, and modeling and analysis approaches. The Model-based Design and Requirements approach does not reduce the effort associated with the design process versus previous processes used at Marshall Space Flight Center. Instead, the approach takes advantage of overlap between the requirements development and management process, and the design and analysis process by efficiently combining the control (i.e. the requirement) and the design mechanisms. The design mechanism is the representation of the component behavior and performance in design and analysis tools. The focus in the early design process shifts from the development and