Self-processing 2A-polyproteins--a system for co-ordinate expression of multiple proteins in transgenic plants.

Science.gov (United States)

Halpin, C; Cooke, S E; Barakate, A; El Amrani, A; Ryan, M D

1999-02-01

Achieving co-ordinate, high-level and stable expression of multiple transgenes in plants is currently difficult. Expression levels are notoriously variable and influenced by factors that act independently on transgenes at different genetic loci. Instability of expression due to loss, re-arrangement or silencing of transgenes may occur, and is exacerbated by increasing numbers of transgenic loci and repeated use of homologous sequences. Even linking two or more genes within a T-DNA does not necessarily result in co-ordinate expression. Linking proteins in a single open reading frame--a polyprotein--is a strategy for co-ordinate expression used by many viruses. After translation, polyproteins are processed into constituent polypeptides, usually by proteinases encoded within the polyprotein itself. However, in foot-and-mouth disease virus (FMDV), a sequence (2A) of just 16-20 amino acids appears to have the unique capability to mediate cleavage at its own C-terminus by an apparently enzyme-independent, novel type of reaction. This sequence can also mediate cleavage in a heterologous protein context in a range of eukaryotic expression systems. We have constructed a plasmid in which the 2A sequence is inserted between the reporter genes chloramphenicol acetyltransferase (CAT) and beta-glucuronidase (GUS), maintaining a single open reading frame. Here we report that expression of this construct in wheatgerm lysate and transgenic plants results in efficient cleavage of the polyprotein and co-ordinate expression of active CAT and GUS. Self-processing polyproteins using the FMDV 2A sequence could therefore provide a system for ensuring co-ordinated, stable expression of multiple introduced proteins in plant cells.

α-Lipoic acid prevents lipotoxic cardiomyopathy in acyl CoA-synthase transgenic mice

International Nuclear Information System (INIS)

Lee, Young; Naseem, R. Haris; Park, Byung-Hyun; Garry, Daniel J.; Richardson, James A.; Schaffer, Jean E.; Unger, Roger H.

2006-01-01

α-Lipoic acid (α-LA) mimics the hypothalamic actions of leptin on food intake, energy expenditure, and activation of AMP-activated protein kinase (AMPK). To determine if, like leptin, α-LA protects against cardiac lipotoxicity, α-LA was fed to transgenic mice with cardiomyocyte-specific overexpression of the acyl CoA synthase (ACS) gene. Untreated ACS-transgenic mice died prematurely with increased triacylglycerol content and dilated cardiomyopathy, impaired systolic function and myofiber disorganization, apoptosis, and interstitial fibrosis on microscopy. In α-LA-treated ACS-transgenic mice heart size, echocardiogram and TG content were normal. Plasma TG fell 50%, hepatic-activated phospho-AMPK rose 6-fold, sterol regulatory element-binding protein-1c declined 50%, and peroxisome proliferator-activated receptor-γ cofactor-1α mRNA rose 4-fold. Since food restriction did not prevent lipotoxicity, we conclude that α-LA treatment, like hyperleptinemia, protects the heart of ACS-transgenic mice from lipotoxicity

An alternative method for cDNA cloning from surrogate eukaryotic cells transfected with the corresponding genomic DNA.

Science.gov (United States)

Hu, Lin-Yong; Cui, Chen-Chen; Song, Yu-Jie; Wang, Xiang-Guo; Jin, Ya-Ping; Wang, Ai-Hua; Zhang, Yong

2012-07-01

cDNA is widely used in gene function elucidation and/or transgenics research but often suitable tissues or cells from which to isolate mRNA for reverse transcription are unavailable. Here, an alternative method for cDNA cloning is described and tested by cloning the cDNA of human LALBA (human alpha-lactalbumin) from genomic DNA. First, genomic DNA containing all of the coding exons was cloned from human peripheral blood and inserted into a eukaryotic expression vector. Next, by delivering the plasmids into either 293T or fibroblast cells, surrogate cells were constructed. Finally, the total RNA was extracted from the surrogate cells and cDNA was obtained by RT-PCR. The human LALBA cDNA that was obtained was compared with the corresponding mRNA published in GenBank. The comparison showed that the two sequences were identical. The novel method for cDNA cloning from surrogate eukaryotic cells described here uses well-established techniques that are feasible and simple to use. We anticipate that this alternative method will have widespread applications.

Highly efficient generation of knock-in transgenic medaka by CRISPR/Cas9-mediated genome engineering.

Science.gov (United States)

Watakabe, Ikuko; Hashimoto, Hisashi; Kimura, Yukiko; Yokoi, Saori; Naruse, Kiyoshi; Higashijima, Shin-Ichi

2018-01-01

Medaka ( Oryzias latipes ) is a popular animal model used in vertebrate genetic analysis. Recently, an efficient (~ 30%) knock-in system via non-homologous end joining (NHEJ) was established in zebrafish using the CRISPR/Cas9 system. If the same technique were applicable in medaka, it would greatly expand the usefulness of this model organism. The question of the applicability of CRISPR/Cas9 in medaka, however, has yet to be addressed. We report the highly efficient generation of knock-in transgenic medaka via non-homologous end joining (NHEJ). Donor plasmid containing a heat-shock promoter and a reporter gene was co-injected with a short guide RNA (sgRNA) targeted for genome digestion, an sgRNA targeted for donor plasmid digestion, and Cas9 mRNA. Broad transgene expression in the expression domain of a target gene was observed in approximately 25% of injected embryos. By raising these animals, we established stable knock-in transgenic fish with several different constructs for five genetic loci, obtaining transgenic founders at efficiencies of > 50% for all five loci. Further, we show that the method is useful for obtaining mutant alleles. In the experiments where transgene integrations were targeted between the transcription start site and the initiation methionine, the resultant transgenic fish became mutant alleles. With its simplicity, design flexibility, and high efficiency, we propose that CRISPR/Cas9-mediated knock-in via NHEJ will become a standard method for the generation of transgenic and mutant medaka.

Overexpression of TIMP-1 under the MMP-9 promoter interferes with wound healing in transgenic mice

OpenAIRE

Salonurmi, T.; Parikka, M.; Kontusaari, S.; Pirila, E.; Munaut, Carine; Salo, T.; Tryggvason, K.

2004-01-01

We have generated transgenic mice harboring the murine matrix metalloproteinase 9 (MMP-9) promoter cloned in front of human TIMP-1 cDNA. The transgenic mice were viable and fertile and exhibited normal growth and general development. During wound healing the mice were shown to express human TIMP-1 in keratinocytes that normally express MMP-9. However, the healing of skin wounds was significantly retarded with slow migration of keratinocytes over the wound in transgenic mice. In situ zymograph...

Predicting, monitoring and controlling geomechanical effects of CO2 injection

International Nuclear Information System (INIS)

Streit, J.E.; Siggins, A.F.

2005-01-01

A key objective of geological carbon dioxide (CO 2 ) storage in porous rock is long-term subsurface containment of CO 2 . Fault stability and maximum sustainable pore-fluid pressures should be estimated in geomechanical studies in order to avoid damage to reservoir seals and fault seals of storage sites during CO 2 injection. Such analyses rely on predicting the evolution of effective stresses in rocks and faults during CO 2 injection. However, geomechanical analyses frequently do not incorporate poroelastic behaviour of reservoir rock, as relevant poroelastic properties are rarely known. The knowledge of rock poroelastic properties would allow the use of seismic methods for the accurate measurement of the effective stress evolution during CO 2 injection. This paper discussed key geomechanical effects of CO 2 injection into porous rock, and in particular, focused on the effects that the poroelasticity of reservoir rocks and pore pressure/stress coupling have on effective stresses. Relevant geophysical monitoring techniques were also suggested. The paper also outlined how these techniques could be applied to measure stress changes related to poroelastic rock behaviour during CO 2 injection and to test the predictions of sustainable changes in effective stress in CO 2 storage sites. It was concluded that a combination of predictive geomechanical techniques and application of geophysical monitoring techniques is a valid new concept for controlling and monitoring the geomechanical effects of CO 2 storage. 36 refs., 5 figs

DEPLETED HYDROCARBON RESERVOIRS AND CO2 INJECTION WELLS –CO2 LEAKAGE ASSESSMENT

Directory of Open Access Journals (Sweden)

Nediljka Gaurina-Međimurec

2017-03-01

Full Text Available Migration risk assessment of the injected CO2 is one of the fi rst and indispensable steps in determining locations for the implementation of projects for carbon dioxide permanent disposal in depleted hydrocarbon reservoirs. Within the phase of potential storage characterization and assessment, it is necessary to conduct a quantitative risk assessment, based on dynamic reservoir models that predict the behaviour of the injected CO2, which requires good knowledge of the reservoir conditions. A preliminary risk assessment proposed in this paper can be used to identify risks of CO2 leakage from the injection zone and through wells by quantifying hazard probability (likelihood and severity, in order to establish a risk-mitigation plan and to engage prevention programs. Here, the proposed risk assessment for the injection well is based on a quantitative risk matrix. The proposed assessment for the injection zone is based on methodology used to determine a reservoir probability in exploration and development of oil and gas (Probability of Success, abbr. POS, and modifi ed by taking into account hazards that may lead to CO2 leakage through the cap rock in the atmosphere or groundwater. Such an assessment can eliminate locations that do not meet the basic criteria in regard to short-term and long-term safety and the integrity of the site

An Efficient Method for Generation of Transgenic Rats Avoiding Embryo Manipulation

Directory of Open Access Journals (Sweden)

Bhola Shankar Pradhan

2016-01-01

Full Text Available Although rats are preferred over mice as an animal model, transgenic animals are generated predominantly using mouse embryos. There are limitations in the generation of transgenic rat by embryo manipulation. Unlike mouse embryos, most of the rat embryos do not survive after male pronuclear DNA injection which reduces the efficiency of generation of transgenic rat by this method. More importantly, this method requires hundreds of eggs collected by killing several females for insertion of transgene to generate transgenic rat. To this end, we developed a noninvasive and deathless technique for generation of transgenic rats by integrating transgene into the genome of the spermatogonial cells by testicular injection of DNA followed by electroporation. After standardization of this technique using EGFP as a transgene, a transgenic disease model displaying alpha thalassemia was successfully generated using rats. This efficient method will ease the generation of transgenic rats without killing the lives of rats while simultaneously reducing the number of rats used for generation of transgenic animal.

Performance of CO2 enrich CNG in direct injection engine

Science.gov (United States)

Firmansyah, W. B.; Ayandotun, E. Z.; Zainal, A.; Aziz, A. R. A.; Heika, M. R.

2015-12-01

This paper investigates the potential of utilizing the undeveloped natural gas fields in Malaysia with high carbon dioxide (CO2) content ranging from 28% to 87%. For this experiment, various CO2 proportions by volume were added to pure natural gas as a way of simulating raw natural gas compositions in these fields. The experimental tests were carried out using a 4-stroke single cylinder spark ignition (SI) direct injection (DI) compressed natural gas (CNG) engine. The tests were carried out at 180° and 300° before top dead centre (BTDC) injection timing at 3000 rpm, to establish the effects on the engine performance. The results show that CO2 is suppressing the combustion of CNG while on the other hand CNG combustion is causing CO2 dissociation shown by decreasing CO2 emission with the increase in CO2 content. Results for 180° BTDC injection timing shows higher performance compared to 300° BTDC because of two possible reasons, higher volumetric efficiency and higher stratification level. The results also showed the possibility of increasing the CO2 content by injection strategy.

Generation of minipigs with targeted transgene insertion by recombinase-mediated cassette exchange (RMCE) and somatic cell nuclear transfer (SCNT)

DEFF Research Database (Denmark)

Jakobsen, Jannik E.; Johansen, Marianne Gregers; Schmidt, Mette

2012-01-01

minicircles in RMCE in fibroblasts with all four acceptor loci and followed by SCNT, we produced piglets with a single copy of a transgene incorporated into one of the transcriptionally active acceptor loci. The transgene, consisting of a cDNA of the Alzheimer’s disease-causing gene PSEN1M146I driven...

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

International Nuclear Information System (INIS)

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

1989-01-01

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

Repeated intraperitoneal injections of liposomes containing phosphatidic acid and cardiolipin reduce amyloid-β levels in APP/PS1 transgenic mice

DEFF Research Database (Denmark)

Ordóñez-Gutiérrez, Lara; Re, Francesca; Bereczki, Erika

2015-01-01

, it was hypothesized that shifting this equilibrium towards the blood by enhancing peripheral clearance might reduce Aβ levels in the brain: the 'sink effect'. We tested this hypothesis by intraperitoneally injecting APP/PS1 transgenic mice with small unilamellar vesicles containing either phosphatidic acid...... Aβ may be therapeutically relevant in AD. FROM THE CLINICAL EDITOR: Intraperitoneal injection of small unilamellar vesicles containing phosphatidic acid or cardiolipin significantly reduced the amount of amyloid-beta (Aß) peptide in the plasma in a rodent model. Brain levels of Aß were also affected...

Prevalence and Correlates of Heroin–Methamphetamine Co-Injection Among Persons Who Inject Drugs in San Diego, California, and Tijuana, Baja California, Mexico

Science.gov (United States)

Meacham, Meredith C.; Strathdee, Steffanie A.; Rangel, Gudelia; Armenta, Richard F.; Gaines, Tommi L.; Garfein, Richard S.

2016-01-01

Objective: Although persons who inject drugs (PWID) in the western United States–Mexico border region are known to inject both heroin and methamphetamine, little is known about the prevalence and risks associated with co-injection of this depressant–stimulant combination (also known as “goofball” and “Mexican speedball”). Method: Baseline data from parallel cohort studies of PWID conducted concurrently in San Diego, CA, and Tijuana, Mexico, were used to estimate the prevalence and identify correlates of heroin–methamphetamine co-injection. PWID older than 18 years of age who reported injecting illicit drugs in the past month (N = 1,311; 32.7% female) were recruited in San Diego (n = 576) and Tijuana (n = 735) and completed interviewer-administered questionnaires. Bivariate and multivariable logistic regression analyses were used to identify correlates of heroin–methamphetamine co-injection. Results: The prevalence of co-injection in the past 6 months was 39.9% overall and was higher in Tijuana (55.8%) than in San Diego (19.8%). In multivariable analyses adjusting for study cohort, distributive syringe sharing, purchasing syringes prefilled with drugs, finding it hard to get new syringes, reporting great or urgent need for treatment, and younger age were independently associated with co-injection. Past-6-month overdose was significantly associated with higher odds of co-injection in San Diego than in Tijuana. Conclusions: These findings indicate that heroin–methamphetamine co-injection is more common in Tijuana than in San Diego, yet this practice was only associated with overdose in San Diego. Heroin–methamphetamine co-injection was also independently associated with HIV-associated injection risk behaviors. Overdose-prevention interventions should address co-injection of depressants and stimulants. PMID:27588536

Split-Cre complementation restores combination activity on transgene excision in hair roots of transgenic tobacco.

Directory of Open Access Journals (Sweden)

Mengling Wen

Full Text Available The Cre/loxP system is increasingly exploited for genetic manipulation of DNA in vitro and in vivo. It was previously reported that inactive ''split-Cre'' fragments could restore Cre activity in transgenic mice when overlapping co-expression was controlled by two different promoters. In this study, we analyzed recombination activities of split-Cre proteins, and found that no recombinase activity was detected in the in vitro recombination reaction in which only the N-terminal domain (NCre of split-Cre protein was expressed, whereas recombination activity was obtained when the C-terminal (CCre or both NCre and CCre fragments were supplied. We have also determined the recombination efficiency of split-Cre proteins which were co-expressed in hair roots of transgenic tobacco. No Cre recombination event was observed in hair roots of transgenic tobacco when the NCre or CCre genes were expressed alone. In contrast, an efficient recombination event was found in transgenic hairy roots co-expressing both inactive split-Cre genes. Moreover, the restored recombination efficiency of split-Cre proteins fused with the nuclear localization sequence (NLS was higher than that of intact Cre in transgenic lines. Thus, DNA recombination mediated by split-Cre proteins provides an alternative method for spatial and temporal regulation of gene expression in transgenic plants.

scsB, a cDNA encoding the hydrogenosomal beta subunit of succinyl-CoA synthetase from the anaerobic fungus Neocallimastix frontalis

NARCIS (Netherlands)

Brondijk, THC; Durand, R; vanderGiezen, M; Gottschal, JC; Prins, RA; Fevre, M

1996-01-01

A clone containing a Neocallimastix frontalis cDNA assumed to encode the beta subunit of succinyl-CoA synthetase (SCSB) was identified by sequence homology with prokaryotic and eukaryotic counterparts. An open reading frame of 1311 bp was found. The deduced 437 amino acid sequence showed a high

Co-expression of NCED and ALO improves vitamin C level and tolerance to drought and chilling in transgenic tobacco and stylo plants.

Science.gov (United States)

Bao, Gegen; Zhuo, Chunliu; Qian, Chunmei; Xiao, Ting; Guo, Zhenfei; Lu, Shaoyun

2016-01-01

Abscisic acid (ABA) regulates plant adaptive responses to various environmental stresses, while L-ascorbic acid (AsA) that is also named vitamin C is an important antioxidant and involves in plant stress tolerance and the immune system in domestic animals. Transgenic tobacco (Nicotiana tabacum L.) and stylo [Stylosanthes guianensis (Aublet) Swartz], a forage legume, plants co-expressing stylo 9-cis-epoxycarotenoid dioxygenase (SgNCED1) and yeast D-arabinono-1,4-lactone oxidase (ALO) genes were generated in this study, and tolerance to drought and chilling was analysed in comparison with transgenic tobacco overexpressing SgNCED1 or ALO and the wild-type plants. Compared to the SgNCED1 or ALO transgenic plants, in which only ABA or AsA levels were increased, both ABA and AsA levels were increased in transgenic tobacco and stylo plants co-expressing SgNCED1 and ALO genes. Compared to the wild type, an enhanced drought tolerance was observed in SgNCED1 transgenic tobacco plants with induced expression of drought-responsive genes, but not in ALO plants, while an enhanced chilling tolerance was observed in ALO transgenic tobaccos with induced expression of cold-responsive genes, but not in SgNCED1 plants. Co-expression of SgNCED1 and ALO genes resulted in elevated tolerance to both drought and chilling in transgenic tobacco and stylo plants with induced expression of both drought and cold-responsive genes. Our result suggests that co-expression of SgNCED1 and ALO genes is an effective way for use in forage plant improvement for increased tolerance to drought and chilling and nutrition quality. © 2015 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

Prevalence and Correlates of Heroin-Methamphetamine Co-Injection Among Persons Who Inject Drugs in San Diego, California, and Tijuana, Baja California, Mexico.

Science.gov (United States)

Meacham, Meredith C; Strathdee, Steffanie A; Rangel, Gudelia; Armenta, Richard F; Gaines, Tommi L; Garfein, Richard S

2016-09-01

Although persons who inject drugs (PWID) in the western United States-Mexico border region are known to inject both heroin and methamphetamine, little is known about the prevalence and risks associated with co-injection of this depressant-stimulant combination (also known as "goofball" and "Mexican speedball"). Baseline data from parallel cohort studies of PWID conducted concurrently in San Diego, CA, and Tijuana, Mexico, were used to estimate the prevalence and identify correlates of heroin-methamphetamine co-injection. PWID older than 18 years of age who reported injecting illicit drugs in the past month (N = 1,311; 32.7% female) were recruited in San Diego (n = 576) and Tijuana (n = 735) and completed interviewer-administered questionnaires. Bivariate and multivariable logistic regression analyses were used to identify correlates of heroin-meth-amphetamine co-injection. The prevalence of co-injection in the past 6 months was 39.9% overall and was higher in Tijuana (55.8%) than in San Diego (19.8%). In multivariable analyses adjusting for study cohort, distributive syringe sharing, purchasing syringes prefilled with drugs, finding it hard to get new syringes, reporting great or urgent need for treatment, and younger age were independently associated with co-injection. Past-6-month overdose was significantly associated with higher odds of co-injection in San Diego than in Tijuana. These findings indicate that heroin-methamphetamine co-injection is more common in Tijuana than in San Diego, yet this practice was only associated with overdose in San Diego. Heroin-methamphetamine coinjection was also independently associated with HIV-associated injection risk behaviors. Overdose-prevention interventions should address co-injection of depressants and stimulants.

Case history of a successful CO{sub 2} miscible gas WAG injection project

Energy Technology Data Exchange (ETDEWEB)

Harpole, Ken

1998-07-01

A successful fieldwide CO{sub 2} miscible gas injection project has been underway at the East Vacuum Grayburg San Andres Unit (EVGSAU) in eastern New Mexico, USA since 1985. This presentation follows the evolution of CO{sub 2} miscible gas WAG injection operations at EVGSAU and discusses some of the significant changes in reservoir strategy management which have been implemented over the past 13 years. These changes parallel the evolution in the industry's understanding of and experience with CO{sub 2} miscible gs injection processes. The operating problems and reservoir management challenges encountered at East Vacuum - injection performance, sweep efficiency, effective management of WAG operations, rapidly changing requirements for handling produced gas, and maintaining efficient utilization of injected CO{sub 2} - reflect the kinds of challenges typically encountered in managing a large CO{sub 2} injection project. 1 fig., 1 tab.

Evasion of CO2 injected into the ocean in the context of CO2 stabilization

International Nuclear Information System (INIS)

Kheshgi, Haroon S.

2004-01-01

The eventual evasion of injected CO 2 to the atmosphere is one consideration when assessing deep-sea disposal of CO 2 as a potential response option to climate change concerns. Evasion estimated using an ocean carbon cycle model is compared to long-term trajectories for future CO 2 emissions, including illustrative cases leading to stabilization of CO 2 concentration at various levels. Modeled residence time for CO 2 injected into the deep ocean exceeds the 100-year time-scale usually considered in scenarios for future emissions, and the potential impacts of climate change. Illustrative cases leading monotonically to constant CO 2 concentration have been highlighted by the Intergovernmental Panel on Climate Change to give guidance on possible timing of emission reductions that may be required to stabilize greenhouse gas concentrations at various levels. For stabilization cases considered, significant modeled evasion does not occur until long after CO 2 emissions have reached a maximum and begun to decline. Illustrative cases can also lead to a maximum in CO 2 concentration followed by a decline to slowly decreasing concentrations. In such cases, future injection of emissions into the deep ocean leads to lower maximum CO 2 concentration, with less effect on concentration later on in time

cDNA cloning of rat and human medium chain acyl-CoA dehydrogenase (MCAD)

International Nuclear Information System (INIS)

Matsubara, Y.; Kraus, J.P.; Rosenberg, L.E.; Tanaka, K.

1986-01-01

MCAD is one of three mitochondrial flavoenzymes which catalyze the first step in the β-oxidation of straight chain fatty acids. It is a tetramer with a subunit Mr of 45 kDa. MCAD is synthesized in the cytosol as a 49 kDa precursor polypeptide (pMCAD), imported into mitochondria, and cleaved to the mature form. Genetic deficiency of MCAD causes recurrent episodes of hypoglycemic coma accompanied by medium chain dicarboxylic aciduria. Employing a novel approach, the authors now report isolation of partial rat and human cDNA clones encoding pMCAD. mRNA encoding pMCAD was purified to near homogeneity by polysome immunoadsorption using polyclonal monospecific antibody. Single-stranded [ 32 P]labeled cDNA probe was synthesized using the enriched mRNA as template, and was used to screen directly 16,000 colonies from a total rat liver cDNA library constructed in pBR322. One clone (600 bp) was detected by in situ hybridization. Hybrid-selected translation with this cDNA yielded a 49 kDa polypeptide indistinguishable in size from rat pMCAD and immunoprecipitable with anti-MCAD antibody. Using the rat cDNA as probe, 43,000 colonies from a human liver cDNA library were screened. Four identical positive clones (400 bp) were isolated and positively identified by hybrid-selected translation and immunoprecipitation. The sizes of rat and human mRNAs encoding pMCAD were 2.2 kb and 2.4 kb, respectively, as determined by Northern blotting

Optimization of in vitro regeneration and Agrobacterium tumefaciens-mediated transformation with heat-resistant cDNA in Brassica oleracea subsp. italica cv. Green Marvel.

Science.gov (United States)

Ravanfar, Seyed Ali; Aziz, Maheran Abdul; Saud, Halimi Mohd; Abdullah, Janna Ong

2015-11-01

An efficient system for shoot regeneration and Agrobacterium tumefaciens-mediated transformation of Brassica oleracea cv. Green Marvel cultivar is described. This study focuses on developing shoot regeneration from hypocotyl explants of broccoli cv. Green Marvel using thidiazuron (TDZ), zeatin, and kinetin, the optimization of factors affecting Agrobacterium-mediated transformation of the hypocotyl explants with heat-resistant cDNA, followed by the confirmation of transgenicity of the regenerants. High shoot regeneration was observed in 0.05-0.1 mg dm(-3) TDZ. TDZ at 0.1 mg dm(-3) produced among the highest percentage of shoot regeneration (96.67 %) and mean number of shoot formation (6.17). The highest percentage (13.33 %) and mean number (0.17) of putative transformant production were on hypocotyl explants subjected to preculture on shoot regeneration medium (SRM) with 200 µM acetosyringone. On optimization of bacterial density and inoculation time, the highest percentage and mean number of putative transformant production were on hypocotyl explants inoculated with a bacterial dilution of 1:5 for 30 min. Polymerase chain reaction (PCR) assay indicated a transformation efficiency of 8.33 %. The luciferase assay showed stable integration of the Arabidopsis thaliana HSP101 (AtHSP101) cDNA in the transgenic broccoli regenerants. Three out of five transgenic lines confirmed through PCR showed positive hybridization bands of the AtHSP101 cDNA through Southern blot analysis. The presence of AtHSP101 transcripts in the three transgenic broccoli lines indicated by reverse transcription-PCR (RT-PCR) confirmed the expression of the gene. In conclusion, an improved regeneration system has been established from hypocotyl explants of broccoli followed by successful transformation with AtHSP101 for resistance to high temperature.

Simulation of CO2 Injection in Porous Media with Structural Deformation Effect

KAUST Repository

Negara, Ardiansyah

2011-06-18

Carbon dioxide (CO2) sequestration is one of the most attractive methods to reduce the amount of CO2 in the atmosphere by injecting it into the geological formations. Furthermore, it is also an effective mechanism for enhanced oil recovery. Simulation of CO2 injection based on a suitable modeling is very important for explaining the fluid flow behavior of CO2 in a reservoir. Increasing of CO2 injection may cause a structural deformation of the medium. The structural deformation modeling in carbon sequestration is useful to evaluate the medium stability to avoid CO2 leakage to the atmosphere. Therefore, it is important to include such effect into the model. The purpose of this study is to simulate the CO2 injection in a reservoir. The numerical simulations of two-phase flow in homogeneous and heterogeneous porous media are presented. Also, the effects of gravity and capillary pressure are considered. IMplicit Pressure Explicit Saturation (IMPES) and IMplicit Pressure-Displacements and an Explicit Saturation (IMPDES) schemes are used to solve the problems under consideration. Various numerical examples were simulated and divided into two parts of the study. The numerical results demonstrate the effects of buoyancy and capillary pressure as well as the permeability value and its distribution in the domain. Some conclusions that could be derived from the numerical results are the buoyancy of CO2 is driven by the density difference, the CO2 saturation profile (rate and distribution) are affected by the permeability distribution and its value, and the displacements of the porous medium go to constant values at least six to eight months (on average) after injection. Furthermore, the simulation of CO2 injection provides intuitive knowledge and a better understanding of the fluid flow behavior of CO2 in the subsurface with the deformation effect of the porous medium.

Calcium electrotransfer for termination of transgene expression in muscle

DEFF Research Database (Denmark)

Hojman, Pernille; Spanggaard, Iben; Olsen, Caroline Holkman

2011-01-01

Gene electrotransfer is expanding in clinical use, thus we have searched for an emergency procedure to stop transgene expression in case of serious adverse events. Calcium is cytotoxic at high intracellular levels, so we tested effects of calcium electrotransfer on transgene expression in muscle....... A clinical grade calcium solution (20 μl, 168 mM) was injected into transfected mouse or rat tibialis cranialis muscle. Ca(2+) uptake was quantified using calcium 45 ((45)Ca), and voltage and time between injection and pulsation were varied. Extinction of transgene expression was investigated by using both...... voltage pulses of 1000 V/cm. Using these parameters, in vivo imaging showed that transgene expression significantly decreased 4 hr after Ca(2+) electrotransfer and was eliminated within 24 hr. Similarly, serum erythropoietin was reduced by 46% at 4 hr and to control levels at 2 days. Histological analyses...

Restoration of spermatogenesis and male fertility using an androgen receptor transgene.

Directory of Open Access Journals (Sweden)

William H Walker

Full Text Available Androgens signal through the androgen receptor (AR to regulate male secondary sexual characteristics, reproductive tract development, prostate function, sperm production, bone and muscle mass as well as body hair growth among other functions. We developed a transgenic mouse model in which endogenous AR expression was replaced by a functionally modified AR transgene. A bacterial artificial chromosome (BAC was constructed containing all AR exons and introns plus 40 kb each of 5' and 3' regulatory sequence. Insertion of an internal ribosome entry site and the EGFP gene 3' to AR allowed co-expression of AR and EGFP. Pronuclear injection of the BAC resulted in six founder mice that displayed EGFP production in appropriate AR expressing tissues. The six founder mice were mated into a Sertoli cell specific AR knockout (SCARKO background in which spermatogenesis is blocked at the meiosis stage of germ cell development. The AR-EGFP transgene was expressed in a cyclical manner similar to that of endogenous AR in Sertoli cells and fertility was restored as offspring were produced in the absence of Sertoli cell AR. Thus, the AR-EGFP transgene under the control of AR regulatory elements is capable of rescuing AR function in a cell selective, AR-null background. These initial studies provide proof of principle that a strategy employing the AR-EGFP transgene can be used to understand AR functions. Transgenic mice expressing selective modifications of the AR-EGFP transgene may provide crucial information needed to elicit the molecular mechanisms by which AR acts in the testis and other androgen responsive tissues.

Expression of a defence-related intercellular barley peroxidase in transgenic tobacco

DEFF Research Database (Denmark)

Kristensen, B.K.; Brandt, J.; Bojsen, K.

1997-01-01

genetically, phenotypically and biochemically. The T-DNA was steadily inherited through three generations. The barley peroxidase is expressed and sorted to the intercellular space in the transgenic tobacco plants. The peroxidase can be extracted from the intercellular space in two molecular forms from both...... barley and transgenic tobacco. The tobacco expressed forms are indistinguishable from the barley expressed forms as determined by analytical isoelectric focusing (pI 8.5) and Western-blotting. Staining for N-glycosylation showed that one form only was glycosylated. The N-terminus of purified Prx8 from...... transgenic tobacco was blocked by pyroglutamate, after the removal of which, N-terminal sequencing verified the transit signal-peptide cleavage site deduced from the cDNA sequence. Phenotype comparisons show that the constitutive expression of Prx8 lead to growth retardation. However, an infection assay...

The Co-axial Flow of Injectable Solid Hydrogels with Encapsulated Cells

Science.gov (United States)

Stewart, Brandon; Pochan, Darrin; Sathaye, Sameer

2013-03-01

Hydrogels are quickly becoming an important biomaterial that can be used for the safe, localized injection of cancer drugs, the injection of stem cells into areas of interest or other biological applications. Our peptides can be self-assembled in a syringe where they form a gel, sheared by injection and, once in the body, immediately reform a localized pocket of stiff gel. My project has been designed around looking at the possibility of having a co-axial strand, in which one gel can surround another. This co-axial flow can be used to change the physical properties of our gel during injection, such as stiffening our gel using hyaluronic acid or encapsulating cells in the gel and surrounding the gel with growth medium or other biological factors. Rheology on hyaluron stiffened gels and cells encapsulated in gels was performed for comparison to the results from co-axial flow. Confocal microscopy was used to examine the coaxial gels after flow and to determine how the co-axial nature of the gels is affected by the concentration of peptide.

Evasion of CO{sub 2} injected into the ocean in the content of CO{sub 2} stabilization

Energy Technology Data Exchange (ETDEWEB)

Kheshgi, H.S. [ExxonMobil Research and Engineering Co., Annandale, NJ (United States)

2004-08-01

The eventual evasion of injected CO{sub 2} to the atmosphere is one consideration when assessing deep-sea disposal of CO{sub 2} as a potential response option to climate change concerns. Evasion estimated using an ocean carbon cycle model is compared to long-term trajectories for future CO{sub 2} emissions, including illustrative cases leading to stabilization of CO{sub 2} concentration at various levels. Modeled residence time for CO{sub 2} injected into the deep ocean exceeds the 100-year time-scale usually considered in scenarios for future emissions, and the potential impacts of climate change. Illustrative cases leading monotonically to constant CO{sub 2} concentration have been highlighted by the Intergovernmental Panel on Climate Change to give guidance on possible timing of emission reductions that may be required to stabilize greenhouse gas concentrations at various levels. For stabilization cases considered, significant modeled evasion does not occur until long after CO{sub 2} emissions have reached a maximum and begun to decline. Illustrative cases can also lead to a maximum in CO{sub 2} concentration followed by a decline to slowly decreasing concentrations. In such cases, future injection of emissions into the deep ocean leads to lower maximum CO{sub 2} concentration, with less effect on concentration later on in time. (author)

Evasion of CO{sub 2} injected into the ocean in the context of CO{sub 2} stabilization

Energy Technology Data Exchange (ETDEWEB)

Kheshgi, Haroon S

2004-08-01

The eventual evasion of injected CO{sub 2} to the atmosphere is one consideration when assessing deep-sea disposal of CO{sub 2} as a potential response option to climate change concerns. Evasion estimated using an ocean carbon cycle model is compared to long-term trajectories for future CO{sub 2} emissions, including illustrative cases leading to stabilization of CO{sub 2} concentration at various levels. Modeled residence time for CO{sub 2} injected into the deep ocean exceeds the 100-year time-scale usually considered in scenarios for future emissions, and the potential impacts of climate change. Illustrative cases leading monotonically to constant CO{sub 2} concentration have been highlighted by the Intergovernmental Panel on Climate Change to give guidance on possible timing of emission reductions that may be required to stabilize greenhouse gas concentrations at various levels. For stabilization cases considered, significant modeled evasion does not occur until long after CO{sub 2} emissions have reached a maximum and begun to decline. Illustrative cases can also lead to a maximum in CO{sub 2} concentration followed by a decline to slowly decreasing concentrations. In such cases, future injection of emissions into the deep ocean leads to lower maximum CO{sub 2} concentration, with less effect on concentration later on in time.

The U.S. Gas Flooding Experience: CO2 Injection Strategies and Impact on Ultimate Recovery

Energy Technology Data Exchange (ETDEWEB)

Nunez-Lopez, Vanessa [The University of Texas at Austin; Hosseini, Seyyed; Gil-Egui, Ramon

2017-09-29

The Permian Basin in West Texas and southwestern New Mexico has seen 45 years of oil reserve growth through CO2 enhanced oil recovery (CO2 EOR). More than 60 CO2 EOR projects are currently active in the region’s limestone, sandstone and dolomite reservoirs. Water alternating gas (WAG) has been the development strategy of choice in the Permian for several technical and economic reasons. More recently, the technology started to get implemented in the much more porous and permeable clastic depositional systems of the onshore U.S. Gulf Coast. Continued CO2 injection (CGI), as opposed to WAG, was selected as the injection strategy to develop Gulf Coast oil fields, where CO2 injection volumes are significantly larger (up to 6 times larger) than those of the Permian. We conducted a compositional simulation based study with the objective of comparing the CO2 utilization ratios (volume of CO2 injected to produce a barrel of oil) of 4 conventional and novel CO2 injection strategies: (1) continuous gas injection (CGI), (2) water alternating gas (WAG), (3) water curtain injection (WCI), and (4) WAG and WCI combination. These injection scenarios were simulated using the GEM module from the Computer Modeling Group (CMG). GEM is an advanced general equation-of-state compositional simulator, which includes equation of state, CO2 miscible flood, CO2/brine interactions, and complex phase behavior. The simulator is set up to model three fluid phases including water, oil, and gas. Our study demonstrates how the selected field development strategy has a significant impact on the ultimate recovery of CO2-EOR projects, with GCI injection providing maximum oil recovery in absolute volume terms, but with WAG offering a more balanced technical-economical approach.

Connective Tissue Growth Factor Transgenic Mouse Develops Cardiac Hypertrophy, Lean Body Mass and Alopecia.

Science.gov (United States)

Nuglozeh, Edem

2017-07-01

Connective Tissue Growth Factor (CTGF/CCN2) is one of the six members of cysteine-rich, heparin-binding proteins, secreted as modular protein and recognised to play a major function in cell processes such as adhesion, migration, proliferation and differentiation as well as chondrogenesis, skeletogenesis, angiogenesis and wound healing. The capacity of CTGF to interact with different growth factors lends an important role during early and late development, especially in the anterior region of the embryo. CTGF Knockout (KO) mice have several craniofacial defects and bone miss shaped due to an impairment of the vascular system development during chondrogenesis. The aim of the study was to establish an association between multiple modular functions of CTGF and the phenotype and cardiovascular functions in transgenic mouse. Bicistronic cassette was constructed using pIRES expressing vector (Clontech, Palo Alto, CA). The construct harbours mouse cDNA in tandem with LacZ cDNA as a reporter gene under the control of Cytomegalovirus (CMV) promoter. The plasmid was linearised with NotI restriction enzyme, and 50 ng of linearised plasmid was injected into mouse pronucleus for the chimaera production. Immunohistochemical methods were used to assess the colocalisation renin and CTGF as well as morphology and rheology of the cardiovascular system. The chimeric mice were backcrossed against the wild-type C57BL/6 to generate hemizygous (F1) mouse. Most of the offsprings died as a result of respiratory distress and those that survived have low CTGF gene copy number, approximately 40 molecules per mouse genome. The copy number assessment on the dead pups showed 5×10 3 molecules per mouse genome explaining the threshold of the gene in terms of toxicity. Interestingly, the result of this cross showed 85% of the progenies to be positive deviating from Mendelian first law. All F2 progenies died excluding the possibility of establishing the CTGF transgenic mouse line, situation that

Efficient gene knock-out and knock-in with transgenic Cas9 in Drosophila.

Science.gov (United States)

Xue, Zhaoyu; Ren, Mengda; Wu, Menghua; Dai, Junbiao; Rong, Yikang S; Gao, Guanjun

2014-03-21

Bacterial Cas9 nuclease induces site-specific DNA breaks using small gRNA as guides. Cas9 has been successfully introduced into Drosophila for genome editing. Here, we improve the versatility of this method by developing a transgenic system that expresses Cas9 in the Drosophila germline. Using this system, we induced inheritable knock-out mutations by injecting only the gRNA into embryos, achieved highly efficient mutagenesis by expressing gRNA from the promoter of a novel non-coding RNA gene, and recovered homologous recombination-based knock-in of a fluorescent marker at a rate of 4.5% by co-injecting gRNA with a circular DNA donor. Copyright © 2014 Xue et al.

Intratracheal injection of adenovirus containing the human MNSOD transgene protects athymic nude mice from irradiation-induced organizing alveolitis

International Nuclear Information System (INIS)

Epperly, Michael W.; Bray, Jenifer A.; Krager, Stephen; Berry, Luann M.; Gooding, William; Engelhardt, John F.; Zwacka, Ralf; Travis, Elizabeth L.; Greenberger, Joel S.

1999-01-01

Purpose: A dose and volume limiting factor in radiation treatment of thoracic cancer is the development of fibrosis in normal lung. The goal of the present study was to determine whether expression prior to irradiation of a transgene for human manganese superoxide dismutase (MnSOD) or human copper/zinc superoxide dismutase (Cu/ZnSOD) protects against irradiation-induced lung damage in mice. Methods and Materials: Athymic Nude (Nu/J) mice were intratracheally injected with 10 9 plaque-forming units (PFU) of a replication-incompetent mutant adenovirus construct containing the gene for either human MnSOD, human copper/zinc superoxide dismutase (Cu/ZnSOD) or LacZ. Four days later the mice were irradiated to the pulmonary cavity to doses of 850, 900, or 950 cGy. To demonstrate adenoviral infection, nested reverse transcriptase-polymerase chain reaction (RT-PCR) was carried out with primers specific for either human MnSOD or Cu/ZnSOD transgene on freshly explanted lung, trachea, or alveolar type II cells, and immunohistochemistry was used to measure LacZ expression. RNA was extracted on day 0, 1, 4, or 7 after 850 cGy of irradiation from lungs of mice that had previously received adenovirus or had no treatment. Slot blot analysis was performed to quantitate RNA expression for IL-1, tumor necrosis factor (TNF)-α, TGF-β, MnSOD, or Cu/ZnSOD. Lung tissue was explanted and tested for biochemical activity of MnSOD or Cu/ZnSOD after adenovirus injection. Other mice were sacrificed 132 days after irradiation, lungs excised, frozen in OCT, (polyvinyl alcohol, polyethylene glycol mixture) sectioned, H and E stained, and evaluated for percent of the lung demonstrating organizing alveolitis. Results: Mice injected intratracheally with adenovirus containing the gene for human MnSOD had significantly reduced chronic lung irradiation damage following 950 cGy, compared to control mice or mice injected with adenovirus containing the gene for human Cu/ZnSOD or LacZ. Immunohistochemistry

Long-term CO2 injection and its impact on near-surface soil microbiology.

Science.gov (United States)

Gwosdz, Simone; West, Julia M; Jones, David; Rakoczy, Jana; Green, Kay; Barlow, Tom; Blöthe, Marco; Smith, Karon; Steven, Michael; Krüger, Martin

2016-12-01

Impacts of long-term CO 2 exposure on environmental processes and microbial populations of near-surface soils are poorly understood. This near-surface long-term CO 2 injection study demonstrated that soil microbiology and geochemistry is influenced more by seasonal parameters than elevated CO 2 Soil samples were taken during a 3-year field experiment including sampling campaigns before, during and after 24 months of continuous CO 2 injection. CO 2 concentrations within CO 2 -injected plots increased up to 23% during the injection period. No CO 2 impacts on geochemistry were detected over time. In addition, CO 2 -exposed samples did not show significant changes in microbial CO 2 and CH 4 turnover rates compared to reference samples. Likewise, no significant CO 2 -induced variations were detected for the abundance of Bacteria, Archaea (16S rDNA) and gene copy numbers of the mcrA gene, Crenarchaeota and amoA gene. The majority (75%-95%) of the bacterial sequences were assigned to five phyla: Firmicutes, Proteobacteria, Actinobacteria, Acidobacteria and Bacteroidetes The majority of the archaeal sequences (85%-100%) were assigned to the thaumarchaeotal cluster I.1b (soil group). Univariate and multivariate statistical as well as principal component analyses showed no significant CO 2 -induced variation. Instead, seasonal impacts especially temperature and precipitation were detected. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Evasion of CO{sub 2} injected into the ocean in the context of CO{sub 2} stabilization

Energy Technology Data Exchange (ETDEWEB)

Haroon S. Kheshgi [ExxonMobil Research and Engineering Company, Annandale, NJ (United States)

2003-07-01

The eventual evasion of injected CO{sub 2} to the atmosphere is one consideration when assessing deep sea disposal of CO{sub 2} as a potential response option to climate change concerns. Evasion estimated using an ocean carbon cycle model is compared to long-term trajectories for future CO{sub 2} emissions, including illustrative cases leading to stabilization of CO{sub 2} concentration at various levels. Modeled residence time for CO{sub 2} injected into the deep ocean exceeds the 100-year time scale usually considered in scenarios for future emissions, and the potential impacts of climate change. Illustrative cases leading monotonically to constant CO{sub 2} concentration have been highlighted by the Intergovernmental Panel on Climate Change to give guidance on possible timing of emission reductions that may be required to stabilize greenhouse gas concentrations at various levels. For stabilization cases considered, significant modeled evasion does not occur until long after CO{sub 2} emissions have reached a maximum and begun to decline. Illustrative cases can also lead to a maximum in CO{sub 2} concentration followed by a decline to slowly decreasing concentrations. In such cases, future injection of emissions into the deep ocean leads to lower maximum CO{sub 2} concentration, with less effect on concentration later on in time. 20 refs., 4 figs.

Comparative study of transgenic Brachypodium distachyon expressing sucrose:fructan 6-fructosyltransferases from wheat and timothy grass with different enzymatic properties.

Science.gov (United States)

Tamura, Ken-Ichi; Sanada, Yasuharu; Tase, Kazuhiro; Kawakami, Akira; Yoshida, Midori; Yamada, Toshihiko

2014-04-01

Fructans can act as cryoprotectants and contribute to freezing tolerance in plant species, such as in members of the grass subfamily Pooideae that includes Triticeae species and forage grasses. To elucidate the relationship of freezing tolerance, carbohydrate composition and degree of polymerization (DP) of fructans, we generated transgenic plants in the model grass species Brachypodium distachyon that expressed cDNAs for sucrose:fructan 6-fructosyltransferases (6-SFTs) with different enzymatic properties: one cDNA encoded PpFT1 from timothy grass (Phleum pratense), an enzyme that produces high-DP levans; a second cDNA encoded wft1 from wheat (Triticum aestivum), an enzyme that produces low-DP levans. Transgenic lines expressing PpFT1 and wft1 showed retarded growth; this effect was particularly notable in the PpFT1 transgenic lines. When grown at 22 °C, both types of transgenic line showed little or no accumulation of fructans. However, after a cold treatment, wft1 transgenic plants accumulated fructans with DP = 3-40, whereas PpFT1 transgenic plants accumulated fructans with higher DPs (20 to the separation limit). The different compositions of the accumulated fructans in the two types of transgenic line were correlated with the differences in the enzymatic properties of the overexpressed 6-SFTs. Transgenic lines expressing PpFT1 accumulated greater amounts of mono- and disaccharides than wild type and wft1 expressing lines. Examination of leaf blades showed that after cold acclimation, PpFT1 overexpression increased tolerance to freezing; by contrast, the freezing tolerance of the wft1 expressing lines was the same as that of wild type plants. These results provide new insights into the relationship of the composition of water-soluble carbohydrates and the DP of fructans to freezing tolerance in plants.

Growth hormone transgenic tilapia (Oreochromis sp.) compensate for increased metabolic rate to preserve exercise performance and hypoxia tolerance

DEFF Research Database (Denmark)

McKenzie, D.J.; Martínez, R.; Morales, A.

2001-01-01

Transgenic tilapia hybrids (Oreochromis mossambicus × O. hornorum) carrying a single copy of a homologous cDNA growth hormone exhibit higher growth rates than their wild-type conspecifics (Martinez et al. 1999). Swimming respirometry was employed to determine whether the increased growth rate...... higher in transgenics, such that aerobic scope was similar in both groups, and there was no difference in maximum sustainable U (5.2 ± 0.5 vs. 4.5 ± 0.7 bl s-1 in transgenics vs. wild-types, respectively). Following 2 h recovery from exercise, tilapia were exposed to progressive hypoxia (stepwise......Pa, respectively). The results indicate that stimulation of growth consequent to ectopic expression of growth hormone in transgenic tilapia (Martinez et al. 1999) is linked to increased rates of maintenance metabolism. The swimming and hypoxia experiments indicate, however, that the transgenics were able...

Modeling of fate and transport of co-injection of H2S with CO2 in deep saline formations

Energy Technology Data Exchange (ETDEWEB)

Zhang, W.; Xu, T.; Li, Y.

2010-12-15

The geological storage of CO{sub 2} in deep saline formations is increasing seen as a viable strategy to reduce the release of greenhouse gases into the atmosphere. However, costs of capture and compression of CO{sub 2} from industrial waste streams containing small quantities of sulfur and nitrogen compounds such as SO{sub 2}, H{sub 2}S and N{sub 2} are very expensive. Therefore, studies on the co-injection of CO{sub 2} containing other acid gases from industrial emissions are very important. In this paper, numerical simulations were performed to study the co-injection of H{sub 2}S with CO{sub 2} in sandstone and carbonate formations. Results indicate that the preferential dissolution of H{sub 2}S gas (compared with CO{sub 2} gas) into formation water results in the delayed breakthrough of H{sub 2}S gas. Co-injection of H{sub 2}S results in the precipitation of pyrite through interactions between the dissolved H{sub 2}S and Fe{sup 2+} from the dissolution of Fe-bearing minerals. Additional injection of H{sub 2}S reduces the capabilities for solubility and mineral trappings of CO{sub 2} compared to the CO{sub 2} only case. In comparison to the sandstone (siliciclastic) formation, the carbonate formation is less favorable to the mineral sequestration of CO{sub 2}. Different from CO{sub 2} mineral trapping, the presence of Fe-bearing siliciclastic and/or carbonate is more favorable to the H{sub 2}S mineral trapping.

The Influence of CO2 Solubility in Brine on Simulation of CO2 Injection into Water Flooded Reservoir and CO2 WAG

DEFF Research Database (Denmark)

Yan, Wei; Stenby, Erling Halfdan

2010-01-01

Injection of CO2 into depleted oil reservoirs is not only a traditional way to enhance oil recovery but also a relatively cheaper way to sequester CO2 underground since the increased oil production can offset some sequestration cost. CO2 injection process is often applied to water flooded...... simulations were made for seven oil samples within a wide range of temperature, pressure and salinity. The results were analyzed in terms of the change in oil recovery due to different phase equilibrium descriptions, the delay in breakthrough and the CO2 lost to the aqueous phase. The influence of different...

Violet Grove CO{sub 2} injection project : monitoring with timelapse VSP surveys

Energy Technology Data Exchange (ETDEWEB)

Coueslan, M.; Lawton, D. [Calgary Univ., Calgary, AB (Canada); Jones, M. [Schlumberger Canada Ltd., Calgary, AB (Canada)

2006-07-01

Several oil and gas fields in western Canada have been depleted through primary production and secondary recovery methods. Injecting carbon dioxide (CO{sub 2}) into a reservoir can enhance oil recovery (EOR) and has the potential benefit of CO{sub 2} sequestration, which reduces greenhouse gas emissions into the atmosphere. It has been estimated that western Canada has a practical CO{sub 2} storage capacity of about 3.3 Gt in its oil and gas reservoirs. In order to claim a reduction in CO{sub 2} emissions, however, the injected CO{sub 2} must be monitored to prove that it is being trapped in these reservoirs. For that reason, the Violet Grove site, near Drayton Valley, Alberta was chosen as a pilot site to study CO{sub 2} injection into a reservoir for enhanced recovery and carbon sequestration purposes. The reservoir is located in the Cardium Formation in the Pembina Field. It was expected that the CO{sub 2} would flow preferentially in the reservoir's dominant fracture orientation, which is northeast-southwest. Simultaneously acquired time-lapse multicomponent surface and borehole seismic surveys were used to monitor changes in the reservoir. Prior to CO{sub 2} injection, a baseline survey was acquired in March 2005. A second survey was acquired 8 months after CO{sub 2} injection. The borehole seismic data displayed higher bandwidth and increased resolution compared to the surface seismic data. The PS-wave borehole seismic data in particular showed much better results. Together, these seismic surveys provide lateral coverage of the area as well as high resolution images near the observation well. Preliminary results from the time-lapse analysis show an increase of 30 to 60 per cent in the reservoir reflectivity amplitudes in the 8 months between the baseline and monitor surveys, suggesting that the CO{sub 2} flood has progressed southwest of the injector, most likely along the dominant fracture trend. 7 refs., 5 figs.

Method and apparatus for efficient injection of CO2 in oceans

Science.gov (United States)

West, Olivia R.; Tsouris, Constantinos; Liang, Liyuan

2003-07-29

A liquid CO.sub.2 injection system produces a negatively buoyant consolidated stream of liquid CO.sub.2, CO.sub.2 hydrate, and water that sinks upon release at ocean depths in the range of 700-1500 m. In this approach, seawater at a predetermined ocean depth is mixed with the liquid CO.sub.2 stream before release into the ocean. Because mixing is conducted at depths where pressures and temperatures are suitable for CO.sub.2 hydrate formation, the consolidated stream issuing from the injector is negatively buoyant, and comprises mixed CO.sub.2 -hydrate/CO.sub.2 -liquid/water phases. The "sinking" characteristic of the produced stream will prolong the metastability of CO.sub.2 ocean sequestration by reducing the CO.sub.2 dissolution rate into water. Furthermore, the deeper the CO.sub.2 hydrate stream sinks after injection, the more stable it becomes internally, the deeper it is dissolved, and the more dispersed is the resulting CO.sub.2 plume. These factors increase efficiency, increase the residence time of CO2 in the ocean, and decrease the cost of CO.sub.2 sequestration while reducing deleterious impacts of free CO.sub.2 gas in ocean water.

CO2 Injectivity in Geological Storages: an Overview of Program and Results of the GeoCarbone-Injectivity Project

International Nuclear Information System (INIS)

Lombard, J.M.; Egermann, P.; Azaroual, M.; Pironon, J.; Broseta, D.; Egermann, P.; Munier, G.; Mouronval, G.

2010-01-01

The objective of the GeoCarbone-Injectivity project was to develop a methodology to study the complex phenomena involved in the near well bore region during CO 2 injection. This paper presents an overview of the program and results of the project, and some further necessary developments. The proposed methodology is based on experiments and simulations at the core scale, in order to understand (physical modelling and definition of constitutive laws) and quantify (calibration of simulation tools) the mechanisms involved in injectivity variations: fluid/rock interactions, transport mechanisms, geomechanical effects. These mechanisms and the associated parameters have then to be integrated in the models at the well bore scale. The methodology has been applied for the study of a potential injection of CO 2 in the Dogger geological formation of the Paris Basin, in collaboration with the other ANR GeoCarbone projects. (authors)

Pyramiding expression of maize genes encoding phosphoenolpyruvate carboxylase (PEPC) and pyruvate orthophosphate dikinase (PPDK) synergistically improve the photosynthetic characteristics of transgenic wheat.

Science.gov (United States)

Zhang, HuiFang; Xu, WeiGang; Wang, HuiWei; Hu, Lin; Li, Yan; Qi, XueLi; Zhang, Lei; Li, ChunXin; Hua, Xia

2014-09-01

Using particle bombardment transformation, we introduced maize pepc cDNA encoding phosphoenolpyruvate carboxylase (PEPC) and ppdk cDNA encoding pyruvate orthophosphate dikinase (PPDK) into the C3 crop wheat to generate transgenic wheat lines carrying cDNA of pepc (PC lines), ppdk (PK lines) or both (PKC lines). The integration, transcription, and expression of the foreign genes were confirmed by Southern blot, Real-time quantitative reverse transcription PCR (Q-RT-PCR), and Western blot analysis. Q-RT-PCR results indicated that the average relative expression levels of pepc and ppdk in the PKC lines reached 10 and 4.6, respectively, compared to their expressions in untransformed plants (set to 1). The enzyme activities of PEPC and PPDK in the PKC lines were 4.3- and 2.1-fold higher, respectively, than in the untransformed control. The maximum daily net photosynthetic rates of the PKC, PC, and PK lines were enhanced by 26.4, 13.3, and 4.5%, respectively, whereas the diurnal accumulations of photosynthesis were 21.3, 13.9, and 6.9%, respectively, higher than in the control. The Fv/Fm of the transgenic plants decreased less than in the control under high temperature and high light conditions (2 weeks after anthesis), suggesting that the transgenic wheat transports more absorbed light energy into a photochemical reaction. The exogenous maize C4-specific pepc gene was more effective than ppdk at improving the photosynthetic performance and yield characteristics of transgenic wheat, while the two genes showed a synergistic effect when they were transformed into the same genetic background, because the PKC lines exhibited improved photosynthetic and physiological traits.

Experiences of PEMEX in CO{sub 2} re-injection; Experiencias de Pemex en reinyeccion de CO{sub 2}

Energy Technology Data Exchange (ETDEWEB)

Sanchez Bujanos, Jose Luis [PEMEX (Mexico)

2008-07-15

Mexico enters the processes of secondary oil recovery the years 50s, by means of water injection. The CO{sub 2} injection is one of the techniques that better results produce in the oil recovery. In this presentation the strategic goals that have been determined within PEMEX Exploracion y Produccion (PEP) are mentioned, and which are the projects of CO{sub 2} injection for oil recovery already existing in Mexico. [Spanish] Mexico entra a los procesos de recuperacion secundaria de petroleo en los anos 50s, mediante inyeccion de agua. La inyeccion de CO{sub 2} es una de las tecnicas que mejores resultados produce en la recuperacion de petroleo. En esta presentacion se mencionan las metas estrategicas que se han fijado dentro de Pemex Exploracion y Produccion (PEP) y cuales son los proyectos de inyeccion de CO{sub 2} para recuperacion de petroleo, existentes actualmente en Mexico.

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

OpenAIRE

Belyavsky, A; Vinogradova, T; Rajewsky, K

1989-01-01

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

Optimizing geologic CO2 sequestration by injection in deep saline formations below oil reservoirs

International Nuclear Information System (INIS)

Han, Weon Shik; McPherson, Brian J.

2009-01-01

The purpose of this research is to present a best-case paradigm for geologic CO 2 storage: CO 2 injection and sequestration in saline formations below oil reservoirs. This includes the saline-only section below the oil-water contact (OWC) in oil reservoirs, a storage target neglected in many current storage capacity assessments. This also includes saline aquifers (high porosity and permeability formations) immediately below oil-bearing formations. While this is a very specific injection target, we contend that most, if not all, oil-bearing basins in the US contain a great volume of such strata, and represent a rather large CO 2 storage capacity option. We hypothesize that these are the best storage targets in those basins. The purpose of this research is to evaluate this hypothesis. We quantitatively compared CO 2 behavior in oil reservoirs and brine formations by examining the thermophysical properties of CO 2 , CO 2 -brine, and CO 2 -oil in various pressure, temperature, and salinity conditions. In addition, we compared the distribution of gravity number (N), which characterizes a tendency towards buoyancy-driven CO 2 migration, and mobility ratio (M), which characterizes the impeded CO 2 migration, in oil reservoirs and brine formations. Our research suggests competing advantages and disadvantages of CO 2 injection in oil reservoirs vs. brine formations: (1) CO 2 solubility in oil is significantly greater than in brine (over 30 times); (2) the tendency of buoyancy-driven CO 2 migration is smaller in oil reservoirs because density contrast between oil and CO 2 is smaller than it between brine and oil (the approximate density contrast between CO 2 and crude oil is ∼100 kg/m 3 and between CO 2 and brine is ∼350 kg/m 3 ); (3) the increased density of oil and brine due to the CO 2 dissolution is not significant (about 7-15 kg/m 3 ); (4) the viscosity reduction of oil due to CO 2 dissolution is significant (from 5790 to 98 mPa s). We compared these competing

Differential induction and spread of tau pathology in young PS19 tau transgenic mice following intracerebral injections of pathological tau from Alzheimer’s disease or corticobasal degeneration brains

Science.gov (United States)

Boluda, Susana; Iba, Michiyo; Zhang, Bin; Raible, Kevin M.; Lee, Virginia M-Y.; Trojanowski, John Q.

2015-01-01

Filamentous tau pathologies are hallmark lesions of several neurodegenerative tauopathies including Alzheimer’s disease (AD) and corticobasal degeneration (CBD) which show cell type-specific and topographically distinct tau inclusions. Growing evidence supports templated transmission of tauopathies through functionally interconnected neuroanatomical pathways suggesting that different self-propagating strains of pathological tau could account for the diverse manifestations of neurodegenerative tauopathies. Here, we describe the rapid and distinct cell type-specific spread of pathological tau following intracerebral injections of CBD or AD brain extracts enriched in pathological tau (designated CBD-Tau and AD-Tau, respectively) in young human mutant P301S tau transgenic (Tg) mice (line PS19) ~6–9 months before they show onset of mutant tau transgene-induced tau pathology. At 1 month post-injection of CBD-Tau, tau inclusions developed predominantly in oligodendrocytes of the fimbria and white matter near the injection sites with infrequent intraneuronal tau aggregates. In contrast, injections of AD-Tau in young PS19 mice induced tau pathology predominantly in neuronal perikarya with little or no oligodendrocyte involvement 1 month post-injection. With longer post-injection survival intervals of up to 6 months, CBD-Tau- and AD-Tau-induced tau pathology spread to different brain regions distant from the injection sites while maintaining the cell type-specific pattern noted above. Finally, CA3 neuron loss was detected 3 months post-injection of AD-Tau but not CBD-Tau. Thus, AD-Tau and CBD-Tau represent specific pathological tau strains that spread differentially and may underlie distinct clinical and pathological features of these two tauopathies. Hence, these strains could become targets to develop disease-modifying therapies for CBD and AD. PMID:25534024

VSP Monitoring of CO2 Injection at the Aneth Oil Field in Utah

Science.gov (United States)

Huang, L.; Rutledge, J.; Zhou, R.; Denli, H.; Cheng, A.; Zhao, M.; Peron, J.

2008-12-01

Remotely tracking the movement of injected CO2 within a geological formation is critically important for ensuring safe and long-term geologic carbon sequestration. To study the capability of vertical seismic profiling (VSP) for remote monitoring of CO2 injection, a geophone string with 60 levels and 96 channels was cemented into a monitoring well at the Aneth oil field in Utah operated by Resolute Natural Resources and Navajo National Oil and Gas Company. The oil field is located in the Paradox Basin of southeastern Utah, and was selected by the Southwest Regional Partnership on Carbon Sequestration, supported by the U.S. Department of Energy, to demonstrate combined enhanced oil recovery (EOR) and CO2 sequestration. The geophones are placed at depths from 805 m to 1704 m, and the oil reservoir is located approximately from 1731 m to 1786 m in depth. A baseline VSP dataset with one zero-offset and seven offset source locations was acquired in October, 2007 before CO2 injection. The offsets/source locations are approximately 1 km away from the monitoring well with buried geophone string. A time-lapse VSP dataset with the same source locations was collected in July, 2008 after five months of CO2/water injection into a horizontal well adjacent to the monitoring well. The total amount of CO2 injected during the time interval between the two VSP surveys was 181,000 MCF (million cubic feet), or 10,500 tons. The time-lapse VSP data are pre-processed to balance the phase and amplitude of seismic events above the oil reservoir. We conduct wave-equation migration imaging and interferometry analysis using the pre-processed time-lapse VSP data. The results demonstrate that time-lapse VSP surveys with high-resolution migration imaging and scattering analysis can provide reliable information about CO2 migration. Both the repeatability of VSP surveys and sophisticated time-lapse data pre-processing are essential to make VSP as an effective tool for monitoring CO2 injection.

Modeling of CO2 migration injected in Weyburn oil reservoir

International Nuclear Information System (INIS)

Zhou Wei; Stenhouse, M.J.; Arthur, R.

2008-01-01

Injecting CO 2 into oil and gas field is a way to enhance oil recovery (EOR) as well as mitigate global warming effect by permanently storing the greenhouse gas into underground. This paper details the models and results of simulating the long-term migration of CO 2 injected into the Weyburn field for both Enhanced Oil Recovery operations and CO 2 sequestration. A System Model was established to define the spatial and temporal extents of the analysis. The Base Scenario was developed to identify key processes, features, and events (FEPs) for the expected evolution of the storage system. A compositional reservoir simulator with equations-of-states (EOS) was used as the modeling tool in order to simulate multiphase, multi-component flow and transport coupled with CO 2 mass partitioning into oil, gas, and water phases. We apply a deterministic treatment to CO 2 migration in the geosphere (natural pathways), whereas the variability of abandoned wells (man-made pathways) necessitates a stochastic treatment. The simulation result was then used to carry out consequence analysis to the local environment. (authors)

Impact of thermal processes on CO2 injectivity into a coal seam

International Nuclear Information System (INIS)

Qu, H Y; Liu, J S; Pan, Z J; Connell, L

2010-01-01

The objective of this study is to investigate how thermal gradients, caused by CO2 injection, expansion and adsorption, affect the permeability and adsorption capacity of coal during CO2 sequestration. A new permeability model is developed in which the concept of elastic modulus reduction ratio is introduced to partition the effective strain between coal matrix and fracture. This model is implemented into a fully coupled mechanical deformation, gas flow and heat transport finite element simulator. To predict the amount of CO2 sequested, the extended Langmuir sorption model is used, with parameters values taken from the literature. The coupled heat and gas flow equations, are solved in COMSOL using the finite element method. The simulation results for a constant volume reservoir demostrate that thermal strain acts to significantly reduce both CO2 injectivity and adsorption capacity. These impacts need to be considered in the calculation of the optimum injection rate and the total sequestration capacity.

Transgenic tobacco plants with improved cyanobacterial Rubisco expression but no extra assembly factors grow at near wild-type rates if provided with elevated CO2.

Science.gov (United States)

Occhialini, Alessandro; Lin, Myat T; Andralojc, P John; Hanson, Maureen R; Parry, Martin A J

2016-01-01

Introducing a carbon-concentrating mechanism and a faster Rubisco enzyme from cyanobacteria into higher plant chloroplasts may improve photosynthetic performance by increasing the rate of CO2 fixation while decreasing losses caused by photorespiration. We previously demonstrated that tobacco plants grow photoautotrophically using Rubisco from Synechococcus elongatus, although the plants exhibited considerably slower growth than wild-type and required supplementary CO2 . Because of concerns that vascular plant assembly factors may not be adequate for assembly of a cyanobacterial Rubisco, prior transgenic plants included the cyanobacterial chaperone RbcX or the carboxysomal protein CcmM35. Here we show that neither RbcX nor CcmM35 is needed for assembly of active cyanobacterial Rubisco. Furthermore, by altering the gene regulatory sequences on the Rubisco transgenes, cyanobacterial Rubisco expression was enhanced and the transgenic plants grew at near wild-type growth rates, although still requiring elevated CO2 . We performed detailed kinetic characterization of the enzymes produced with and without the RbcX and CcmM35 cyanobacterial proteins. These transgenic plants exhibit photosynthetic characteristics that confirm the predicted benefits of introduction of non-native forms of Rubisco with higher carboxylation rate constants in vascular plants and the potential nitrogen-use efficiency that may be achieved provided that adequate CO2 is available near the enzyme. © 2015 The Authors The Plant Journal © 2015 John Wiley & Sons Ltd.

Single-copy insertion of transgenes in Caenorhabditis elegans

DEFF Research Database (Denmark)

Frøkjaer-Jensen, Christian; Davis, M Wayne; Hopkins, Christopher E

2008-01-01

developed a method that inserts a single copy of a transgene into a defined site. Mobilization of a Mos1 transposon generates a double-strand break in noncoding DNA. The break is repaired by copying DNA from an extrachromosomal template into the chromosomal site. Homozygous single-copy insertions can...... be obtained in less than 2 weeks by injecting approximately 20 worms. We have successfully inserted transgenes as long as 9 kb and verified that single copies are inserted at the targeted site. Single-copy transgenes are expressed at endogenous levels and can be expressed in the female and male germlines....

Anxiety-like behavior in transgenic mice with brain expression of neuropeptide Y.

Science.gov (United States)

Inui, A; Okita, M; Nakajima, M; Momose, K; Ueno, N; Teranishi, A; Miura, M; Hirosue, Y; Sano, K; Sato, M; Watanabe, M; Sakai, T; Watanabe, T; Ishida, K; Silver, J; Baba, S; Kasuga, M

1998-01-01

Neuropeptide Y (NPY), one of the most abundant peptide transmitters in the mammalian brain, is assumed to play an important role in behavior and its disorders. To understand the long-term modulation of neuronal functions by NPY, we raised transgenic mice created with a novel central nervous system (CNS) neuron-specific expression vector of human Thy- gene fragment linked to mouse NPY cDNA. In situ hybridization analysis demonstrated transgene-derived NPY expression in neurons (e.g., in the hippocampus, cerebral cortex, and the arcuate nucleus of the hypothalamus) in the transgenic mice. The modest increase of NPY protein in the brain was demonstrated by semiquantitative immunohistochemical analysis and by radioreceptor assay (115% in transgenic mice compared to control littermates). Double-staining experiments indicated colocalization of the transgene-derived NPY message and NPY protein in the same neurons, such as in the arcuate nucleus. The transgenic mice displayed behavioral signs of anxiety and hypertrophy of adrenal zona fasciculata cells, but no change in food intake was observed. The anxiety-like behavior of transgenic mice was reversed, at least in part, by administration of corticotropin-releasing factor (CRF) antagonists, alpha-helical CRF9-41, into the third cerebral ventricle. These results suggest that NPY has a role in anxiety and behavioral responses to stress partly via the CRF neuronal system. This genetic model may provide a unique opportunity to study human anxiety and emotional disorders.

Developing Gene Silencing for the Study and Treatment of Dystonia

Science.gov (United States)

2017-12-01

missense miRNA (MIS). HEK293 cells were co-transfected with human torsinA(WT) or torsinA(ΔE) cDNA and these vectors. Immunofluorecence analysis... cDNA (demonstrating allele-specificity) 18 Figure 7. Supressing striatal expression of human torsinA(ΔE) in vivo. DYT1 transgenic rats

The grapevine VvWRKY2 gene enhances salt and osmotic stress tolerance in transgenic Nicotiana tabacum.

Science.gov (United States)

Mzid, Rim; Zorrig, Walid; Ben Ayed, Rayda; Ben Hamed, Karim; Ayadi, Mariem; Damak, Yosra; Lauvergeat, Virginie; Hanana, Mohsen

2018-06-01

Our study aims to assess the implication of WRKY transcription factor in the molecular mechanisms of grapevine adaptation to salt and water stresses. In this respect, a full-length VvWRKY2 cDNA, isolated from a Vitis vinifera grape berry cDNA library, was constitutively over-expressed in Nicotiana tabacum seedlings. Our results showed that transgenic tobacco plants exhibited higher seed germination rates and better growth, under both salt and osmotic stress treatments, when compared to wild type plants. Furthermore, our analyses demonstrated that, under stress conditions, transgenic plants accumulated more osmolytes, such as soluble sugars and free proline, while no changes were observed regarding electrolyte leakage, H 2 O 2 , and malondialdehyde contents. The improvement of osmotic adjustment may be an important mechanism underlying the role of VvWRKY 2 in promoting tolerance and adaptation to abiotic stresses. Principal component analysis of our results highlighted a clear partition of plant response to stress. On the other hand, we observed a significant adaptation behaviour response for transgenic lines under stress. Taken together, all our findings suggest that over-expression of VvWRKY2 gene has a compelling role in abiotic stress tolerance and, therefore, would provide a useful strategy to promote abiotic stress tolerance in grape via molecular-assisted breeding and/or new biotechnology tools.

From Site Characterization through Safe and Successful CO2 Injection Operation to Post-injection Monitoring and Site Closure - Closing the Full Life Cycle Research at the Ketzin Pilot Site, Germany

Science.gov (United States)

Liebscher, Axel

2017-04-01

Initiated in 2004, the Ketzin pilot site near Berlin, Germany, was the first European onshore storage project for research and development on geological CO2 storage. After comprehensive site characterization the site infrastructure was build comprising three deep wells and the injection facility including pumps and storage tanks. The operational CO2 injection period started in June 2008 and ended in August 2013 when the site entered the post-injection closure period. During these five years, a total amount of 67 kt of CO2 was safely injected into an Upper Triassic saline sandstone aquifer at a depth of 630 m - 650 m. In fall 2013, the first observation well was partially plugged in the reservoir section with CO2 resistant cement; full abandonment of this well finished in 2015 after roughly 2 years of cement plug monitoring. Abandonment of the remaining wells will be finished by summer 2017 and hand-over of liability to the competent authority is scheduled for end of 2017. The CO2 injected was mainly of food grade quality (purity > 99.9%). In addition, 1.5 kt of CO2 from the oxyfuel pilot capture facility "Schwarze Pumpe" (purity > 99.7%) was injected in 2011. The injection period terminated with a CO2-N2 co-injection experiment of 650 t of a 95% CO2/5% N2 mixture in summer 2013 to study the effects of impurities in the CO2 stream on the injection operation. During regular operation, the CO2 was pre-heated on-site to 40°C prior to injection to ensure a single-phase injection process and avoid any phase transition or transient states within the injection facility or the reservoir. Between March and July 2013, just prior to the CO2-N2 co-injection experiment, the injection temperature was stepwise decreased down to 10°C within a "cold-injection" experiment to study the effects of two-phase injection conditions. During injection operation, the combination of different geochemical and geophysical monitoring methods enabled detection and mapping of the spatial and

Immunoglobulin gene expression and regulation of rearrangement in kappa transgenic mice

International Nuclear Information System (INIS)

Ritchie, K.A.

1986-01-01

Transgenic mice were produced by microinjection of the functionally rearranged immunoglobulin kappa gene from the myeloma MOPC-21 into the male pronucleus of fertilized mouse eggs, and implantation of the microinjected embryos into foster mothers. Mice that integrated the injected gene were detected by hybridizing tail DNA dots with radioactively labelled pBR322 plasmid DNA, which detects pBR322 sequences left as a tag on the microinjected DNA. Mice that integrated the injected gene (six males) were mated and the DNA, RNA and serum kappa chains of their offspring were analyzed. A rabbit anti-mouse kappa chain antiserum was also produced for use in detection of mouse kappa chains on protein blots. Hybridomas were produced from the spleen cells of these kappa transgenic mice to immortalize representative B cells and to investigate expression of the transgenic kappa gene, its effect on allelic exclusion, and its effect on the control of light chain gene rearrangement and expression. The results show that the microinjected DNA is integrated as concatamers in unique single or, rarely, two separate sites in the genome. The concatamers are composed of several copies (16 to 64) of injected DNA arranged in a head to tail fashion. The transgene is expressed into protein normally and in a tissue specific fashion. For the first time in these transgenic mice, all tissues contain a functionally rearranged and potentially expressible immunoglobulin gene. The transgene is expressed only in B cells and not in hepatocytes, for example. This indicates that rearrangement of immunoglobulin genes is necessary but not sufficient for the tissue specific expression of these genes by B cells

Secretion of a recombinant protein without a signal peptide by the exocrine glands of transgenic rabbits.

Directory of Open Access Journals (Sweden)

Andrea Kerekes

Full Text Available Transgenic rabbits carrying mammary gland specific gene constructs are extensively used for excreting recombinant proteins into the milk. Here, we report refined phenotyping of previously generated Venus transposon-carrying transgenic rabbits with particular emphasis on the secretion of the reporter protein by exocrine glands, such as mammary, salivary, tear and seminal glands. The Sleeping Beauty (SB transposon transgenic construct contains the Venus fluorophore cDNA, but without a signal peptide for the secretory pathway, driven by the ubiquitous CAGGS (CAG promoter. Despite the absence of a signal peptide, the fluorophore protein was readily detected in milk, tear, saliva and seminal fluids. The expression pattern was verified by Western blot analysis. Mammary gland epithelial cells of SB-CAG-Venus transgenic lactating does also showed Venus-specific expression by tissue histology and fluorescence microscopy. In summary, the SB-CAG-Venus transgenic rabbits secrete the recombinant protein by different glands. This finding has relevance not only for the understanding of the biological function of exocrine glands, but also for the design of constructs for expression of recombinant proteins in dairy animals.

Secretion of a recombinant protein without a signal peptide by the exocrine glands of transgenic rabbits.

Science.gov (United States)

Kerekes, Andrea; Hoffmann, Orsolya Ivett; Iski, Gergely; Lipták, Nándor; Gócza, Elen; Kues, Wilfried A; Bősze, Zsuzsanna; Hiripi, László

2017-01-01

Transgenic rabbits carrying mammary gland specific gene constructs are extensively used for excreting recombinant proteins into the milk. Here, we report refined phenotyping of previously generated Venus transposon-carrying transgenic rabbits with particular emphasis on the secretion of the reporter protein by exocrine glands, such as mammary, salivary, tear and seminal glands. The Sleeping Beauty (SB) transposon transgenic construct contains the Venus fluorophore cDNA, but without a signal peptide for the secretory pathway, driven by the ubiquitous CAGGS (CAG) promoter. Despite the absence of a signal peptide, the fluorophore protein was readily detected in milk, tear, saliva and seminal fluids. The expression pattern was verified by Western blot analysis. Mammary gland epithelial cells of SB-CAG-Venus transgenic lactating does also showed Venus-specific expression by tissue histology and fluorescence microscopy. In summary, the SB-CAG-Venus transgenic rabbits secrete the recombinant protein by different glands. This finding has relevance not only for the understanding of the biological function of exocrine glands, but also for the design of constructs for expression of recombinant proteins in dairy animals.

Cloning and characterization of human very-long-chain acyl-CoA dehydrogenase cDNA, chromosomal assignment of the gene and identification in four patients of nine different mutations within the VLCAD gene

DEFF Research Database (Denmark)

Andresen, B S; Bross, P; Vianey-Saban, C

1996-01-01

Very-long-chain acyl-CoA dehydrogenase (VLCAD) is one of four straight-chain acyl-CoA dehydrogenase (ACD) enzymes, which are all nuclear encoded mitochondrial flavoproteins catalyzing the initial step in fatty acid beta-oxidation. We have used the very fast, Rapid Amplification of cDNA Ends (RACE...

Site of pheromone biosynthesis and isolation of HMG-CoA reductase cDNA in the cotton boll weevil, Anthonomus grandis.

Science.gov (United States)

Taban, A Huma; Fu, Jessica; Blake, Jacob; Awano, Ami; Tittiger, Claus; Blomquist, Gary J

2006-08-01

Isolated gut tissue from male cotton boll weevil, Anthonomus grandis (Coleoptera: Curculionidae), incorporated radiolabeled acetate into components that co-eluted with monoterpenoid pheromone components on HPLC. This demonstrates that pheromone components of male A. grandis are produced de novo and strongly suggests that pheromone biosynthesis occurs in gut tissue. A central enzyme in isoprenoid biosynthesis is 3-hydroxy-3-methylglutaryl-CoA reductase (HMG-R), and a full-length HMG-R cDNA was isolated from A. grandis. The predicted translation product was 54 and 45% identical to HMG-R from Ips paraconfusus and Drosophila melanogaster, respectively. HMG-R gene expression gradually increased with age in male A. grandis, which correlates with pheromone production. However, topical application of JH III did not significantly increase HMG-R mRNA levels.

Attenuation of 6-hydroxydopamine-induced dopaminergic nigrostriatal lesions in superoxide dismutase transgenic mice

International Nuclear Information System (INIS)

Cadet, J.L.; Hirata, H.; Asanuma, M.

1998-01-01

6-Hydroxydopamine is a neurotoxin that produces degeneration of the nigrostriatal dopaminergic pathway in rodents. Its toxicity is thought to involve the generation of superoxide anion secondary to its autoxidation. To examine the effects of the overexpression of Cu,Zn-superoxide dismutase activity on 6-hydroxydopamine-induced dopaminergic neuronal damage, we have measured the effects of 6-hydroxydopamine on striatal and nigral dopamine transporters and nigral tyrosine hydroxylase-immunoreactive neurons in Cu,Zn-superoxide dismutase transgenic mice. Intracerebroventricular injection of 6-hydroxydopamine (50 μg) in non-transgenic mice produced reductions in the size of striatal area and an enlargement of the cerebral ventricle on both sides of the brains of mice killed two weeks after the injection. In addition, 6-hydroxydopamine caused marked decreases in striatal and nigral [ 125 I]RTI-121-labelled dopamine transporters not only on the injected side but also on the non-injected side of non-transgenic mice; this was associated with decreased cell number and size of tyrosine hydroxylase-immunoreactive dopamine neurons in the substantia nigra pars compacta on both sides in these mice. In contrast, superoxide dismutase transgenic mice were protected against these neurotoxic effects of 6-hydroxydopamine, with the homozygous transgenic mice showing almost complete protection.These results provide further support for a role of superoxide anion in the toxic effects of 6-hydroxydopamine. They also provide further evidence that reactive oxygen species may be the main determining factors in the neurodegenerative effects of catecholamines. (Copyright (c) 1998 Elsevier Science B.V., Amsterdam. All rights reserved.)

[Steam and air co-injection in removing TCE in 2D-sand box].

Science.gov (United States)

Wang, Ning; Peng, Sheng; Chen, Jia-Jun

2014-07-01

Steam and air co-injection is a newly developed and promising soil remediation technique for non-aqueous phase liquids (NAPLs) in vadose zone. In this study, in order to investigate the mechanism of the remediation process, trichloroethylene (TCE) removal using steam and air co-injection was carried out in a 2-dimensional sandbox with different layered sand structures. The results showed that co-injection perfectly improved the "tailing" effect compared to soil vapor extraction (SVE), and the remediation process of steam and air co-injection could be divided into SVE stage, steam strengthening stage and heat penetration stage. Removal ratio of the experiment with scattered contaminant area was higher and removal speed was faster. The removal ratios from the two experiments were 93.5% and 88.2%, and the removal periods were 83.9 min and 90.6 min, respectively. Steam strengthened the heat penetration stage. The temperature transition region was wider in the scattered NAPLs distribution experiment, which reduced the accumulation of TCE. Slight downward movement of TCE was observed in the experiment with TCE initially distributed in a fine sand zone. And such downward movement of TCE reduced the TCE removal ratio.

Podocyte changes upon induction of albuminuria in Thy-1.1 transgenic mice.

NARCIS (Netherlands)

Smeets, B.; Dijkman, H.B.P.M.; Loeke, N. te; Son, J.P.H.F. van; Steenbergen, E.; Assmann, K.J.M.; Wetzels, J.F.M.; Groenen, P.J.T.A.

2003-01-01

BACKGROUND: Thy-1.1 transgenic mice, characterized by ectopic expression of the Thy-1.1 protein on podocytes, spontaneously develop proteinuria and focal glomerulosclerosis (FGS). Injection of a monoclonal antibody (mAb) directed against the Thy-1.1 protein in young transgenic mice induces a massive

Case study on ground surface deformation induced by CO2 injection into coal seam

International Nuclear Information System (INIS)

Li Hong; Tang Chun'an

2010-01-01

To monitor a geomechanical response of injecting CO 2 into relatively shallow coal seams, tiltmeters were set as an array to cover the ground surface area surrounding the injection well, and to measure the ground deformation during a well fracturing stimulation and a short-term CO 2 injection test. In this paper, an attempt to establish a quantitative relationship between the in-situ coal swelling and the corresponding ground deformation was made by means of numerical simulation study. (authors)

[Double-ambient CO2 concentration affects the growth, development and sucking behavior of non-target brown plant hopper Nilaparvata lugens fed on transgenic Bt rice.

Science.gov (United States)

Lu, Yong Qing; Dai, Yang; Yu, Xiu Ying; Yu, Fu-Lan; Jiang, Shou Lin; Zhou, Zong Yuan; Chen, Fa Jun

2018-02-01

In recent years, the two issues of climate change including elevated CO 2 etc., and resistance of transgenic Bt crops against non-target insect pests have received widespread attention. Elevated CO 2 can affect the herbivorous insects. To date, there is no consensus about the effect of elevated CO 2 on the suck-feeding insect pests (non-target insect pests of transgenic Bt crops). Its effects on the suck-feeding behavior have rarely been reported. In this study, CO 2 levels were set up in artificial climate chamber to examined the effects of ambient (400 μL·L -1 ) and double-ambient (800 μL·L -1 ) CO 2 levels on the suck-feeding behavior, growth, development, and reproduction of the non-target insect pest of transgenic Bt rice, brown planthopper, Nilaparvata lugens. The results showed that CO 2 level significantly affected the egg and nymph duration, longevity and body mass of adults, and feeding behavior of the 4th and 5th instar nymphs, while had no effect on the fecundity of N. lugens. The duration of eggs and nymphs, and the longevity of female adults were significantly shortened by 4.0%, 4.2% and 6.6% respectively, the proportion of the macropterous adults was significantly increased by 11.6%, and the body mass of newly hatched female adults was significantly decreased by 2.2% by elevated CO 2 . In addition, elevated CO 2 significantly enhanced the stylet puncturing efficiency of the 4th and 5th instar nymphs of N. lugens. The duration ofphloem ingestion of the N4b waveform was significantly prolonged by 60.0% and 50.1%, and the frequency significantly was increased by 230.0% and 155.9% for the 4th and 5th instar nymphs of N. lugens by elevated CO 2 , respectively. It was concluded that double-ambient CO 2 could promote the growth and development of N. lugens through enhancing its suck-feeding, shorten the generation life-span and increase the macropertous adults' proportion of N. lugens. Thus, it could result in the occurrence of non-target rice

Normalized cDNA libraries

Science.gov (United States)

Soares, Marcelo B.; Efstratiadis, Argiris

1997-01-01

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

Antisense repression of sucrose phosphate synthase in transgenic muskmelon alters plant growth and fruit development

International Nuclear Information System (INIS)

Tian, Hongmei; Ma, Leyuan; Zhao, Cong; Hao, Hui; Gong, Biao; Yu, Xiyan; Wang, Xiufeng

2010-01-01

To unravel the roles of sucrose phosphate synthase (SPS) in muskmelon (Cucumis melo L.), we reduced its activity in transgenic muskmelon plants by an antisense approach. For this purpose, an 830 bp cDNA fragment of muskmelon sucrose phosphate synthase was expressed in antisense orientation behind the 35S promoter of the cauliflower mosaic virus. The phenotype of the antisense plants clearly differed from that of control plants. The transgenic plant leaves were markedly smaller, and the plant height and stem diameter were obviously shorter and thinner. Transmission electron microscope observation revealed that the membrane degradation of chloroplast happened in transgenic leaves and the numbers of grana and grana lamella in the chloroplast were significantly less, suggesting that the slow growth and weaker phenotype of transgenic plants may be due to the damage of the chloroplast ultrastructure, which in turn results in the decrease of the net photosynthetic rate. The sucrose concentration and levels of sucrose phosphate synthase decreased in transgenic mature fruit, and the fruit size was smaller than the control fruit. Together, our results suggest that sucrose phosphate synthase may play an important role in regulating the muskmelon plant growth and fruit development.

Antisense repression of sucrose phosphate synthase in transgenic muskmelon alters plant growth and fruit development

Energy Technology Data Exchange (ETDEWEB)

Tian, Hongmei; Ma, Leyuan; Zhao, Cong; Hao, Hui; Gong, Biao [College of Horticulture Science and Engineering, State Key Laboratory of Crop Biology, Shandong Agricultural University, Tai' an, Shandong 271018 (China); Yu, Xiyan, E-mail: yuxiyan@sdau.edu.cn [College of Horticulture Science and Engineering, State Key Laboratory of Crop Biology, Shandong Agricultural University, Tai' an, Shandong 271018 (China); Wang, Xiufeng, E-mail: xfwang@sdau.edu.cn [College of Horticulture Science and Engineering, State Key Laboratory of Crop Biology, Shandong Agricultural University, Tai' an, Shandong 271018 (China)

2010-03-12

To unravel the roles of sucrose phosphate synthase (SPS) in muskmelon (Cucumis melo L.), we reduced its activity in transgenic muskmelon plants by an antisense approach. For this purpose, an 830 bp cDNA fragment of muskmelon sucrose phosphate synthase was expressed in antisense orientation behind the 35S promoter of the cauliflower mosaic virus. The phenotype of the antisense plants clearly differed from that of control plants. The transgenic plant leaves were markedly smaller, and the plant height and stem diameter were obviously shorter and thinner. Transmission electron microscope observation revealed that the membrane degradation of chloroplast happened in transgenic leaves and the numbers of grana and grana lamella in the chloroplast were significantly less, suggesting that the slow growth and weaker phenotype of transgenic plants may be due to the damage of the chloroplast ultrastructure, which in turn results in the decrease of the net photosynthetic rate. The sucrose concentration and levels of sucrose phosphate synthase decreased in transgenic mature fruit, and the fruit size was smaller than the control fruit. Together, our results suggest that sucrose phosphate synthase may play an important role in regulating the muskmelon plant growth and fruit development.

Expression of porcine myostatin prodomain genomic sequence leads to a decrease in muscle growth, but significant intramuscular fat accretion in transgenic pigs.

Science.gov (United States)

Myostatin, a member of TGF-beta superfamily, is a dominant inhibitor of skeletal muscle development and growth. Previously, skeletal muscle-specific over-expression of myostatin prodomain cDNA (5’-region 886 nucleotide) dramatically increased growth performance and muscle mass in transgenic mice. I...

Insect resistance to Nilaparvata lugens and Cnaphalocrocis medinalis in transgenic indica rice and the inheritance of gna+sbti transgenes.

Science.gov (United States)

Li, Guiying; Xu, Xinping; Xing, Hengtai; Zhu, Huachen; Fan, Qin

2005-04-01

Molecular genetic analysis and insect bioassay of transgenic indica rice 'Zhuxian B' plants carrying snowdrop lectin gene (gna) and soybean trypsin inhibitor gene (sbti) were investigated in detail. PCR, 'dot' blot and PCR-Southern blot analysis showed that both transgenes had been incorporated into the rice genome and transmitted up to R3 progeny in most lines tested. Some transgenic lines exhibited Mendelian segregation, but the other showed either 1:1 (positive: negative for the transgenes) or other aberrant segregation patterns. The segregation patterns of gna gene crossed between R2 and R3 progeny. In half of transgenic R3 lines, gna and sbti transgenes co-segregated. Two independent homozygous lines expressing double transgenes were identified in R3 progeny. Southern blot analysis demonstrated that the copy numbers of integrated gna and sbti transgenes varied from one to ten in different lines. Insect bioassay data showed that most transgenic plants had better resistance to both Nilaparvata lugens (Stahl) and Cnaphalocrocis medinalis (Guenee) than wild-type plants. The insect resistance of transgenic lines increased with the increase in transgene positive ratio in most of the transgenic lines. In all, we obtained nine lines of R3 transgenic plants, including one pure line, which had better resistance to both N lugens and C medinalis than wild-type plants. Copyright 2005 Society of Chemical Industry.

Fatty acyl-CoA reductases of birds

Directory of Open Access Journals (Sweden)

Hellenbrand Janine

2011-12-01

Full Text Available Abstract Background Birds clean and lubricate their feathers with waxes that are produced in the uropygial gland, a holocrine gland located on their back above the tail. The type and the composition of the secreted wax esters are dependent on the bird species, for instance the wax ester secretion of goose contains branched-chain fatty acids and unbranched fatty alcohols, whereas that of barn owl contains fatty acids and alcohols both of which are branched. Alcohol-forming fatty acyl-CoA reductases (FAR catalyze the reduction of activated acyl groups to fatty alcohols that can be esterified with acyl-CoA thioesters forming wax esters. Results cDNA sequences encoding fatty acyl-CoA reductases were cloned from the uropygial glands of barn owl (Tyto alba, domestic chicken (Gallus gallus domesticus and domestic goose (Anser anser domesticus. Heterologous expression in Saccharomyces cerevisiae showed that they encode membrane associated enzymes which catalyze a NADPH dependent reduction of acyl-CoA thioesters to fatty alcohols. By feeding studies of transgenic yeast cultures and in vitro enzyme assays with membrane fractions of transgenic yeast cells two groups of isozymes with different properties were identified, termed FAR1 and FAR2. The FAR1 group mainly synthesized 1-hexadecanol and accepted substrates in the range between 14 and 18 carbon atoms, whereas the FAR2 group preferred stearoyl-CoA and accepted substrates between 16 and 20 carbon atoms. Expression studies with tissues of domestic chicken indicated that FAR transcripts were not restricted to the uropygial gland. Conclusion The data of our study suggest that the identified and characterized avian FAR isozymes, FAR1 and FAR2, can be involved in wax ester biosynthesis and in other pathways like ether lipid synthesis.

Fatty acyl-CoA reductases of birds

Science.gov (United States)

2011-01-01

Background Birds clean and lubricate their feathers with waxes that are produced in the uropygial gland, a holocrine gland located on their back above the tail. The type and the composition of the secreted wax esters are dependent on the bird species, for instance the wax ester secretion of goose contains branched-chain fatty acids and unbranched fatty alcohols, whereas that of barn owl contains fatty acids and alcohols both of which are branched. Alcohol-forming fatty acyl-CoA reductases (FAR) catalyze the reduction of activated acyl groups to fatty alcohols that can be esterified with acyl-CoA thioesters forming wax esters. Results cDNA sequences encoding fatty acyl-CoA reductases were cloned from the uropygial glands of barn owl (Tyto alba), domestic chicken (Gallus gallus domesticus) and domestic goose (Anser anser domesticus). Heterologous expression in Saccharomyces cerevisiae showed that they encode membrane associated enzymes which catalyze a NADPH dependent reduction of acyl-CoA thioesters to fatty alcohols. By feeding studies of transgenic yeast cultures and in vitro enzyme assays with membrane fractions of transgenic yeast cells two groups of isozymes with different properties were identified, termed FAR1 and FAR2. The FAR1 group mainly synthesized 1-hexadecanol and accepted substrates in the range between 14 and 18 carbon atoms, whereas the FAR2 group preferred stearoyl-CoA and accepted substrates between 16 and 20 carbon atoms. Expression studies with tissues of domestic chicken indicated that FAR transcripts were not restricted to the uropygial gland. Conclusion The data of our study suggest that the identified and characterized avian FAR isozymes, FAR1 and FAR2, can be involved in wax ester biosynthesis and in other pathways like ether lipid synthesis. PMID:22151413

Efficient generation of knock-in transgenic zebrafish carrying reporter/driver genes by CRISPR/Cas9-mediated genome engineering.

Science.gov (United States)

Kimura, Yukiko; Hisano, Yu; Kawahara, Atsuo; Higashijima, Shin-ichi

2014-10-08

The type II bacterial CRISPR/Cas9 system is rapidly becoming popular for genome-engineering due to its simplicity, flexibility, and high efficiency. Recently, targeted knock-in of a long DNA fragment via homology-independent DNA repair has been achieved in zebrafish using CRISPR/Cas9 system. This raised the possibility that knock-in transgenic zebrafish could be efficiently generated using CRISPR/Cas9. However, how widely this method can be applied for the targeting integration of foreign genes into endogenous genomic loci is unclear. Here, we report efficient generation of knock-in transgenic zebrafish that have cell-type specific Gal4 or reporter gene expression. A donor plasmid containing a heat-shock promoter was co-injected with a short guide RNA (sgRNA) targeted for genome digestion, a sgRNA targeted for donor plasmid digestion, and Cas9 mRNA. We have succeeded in establishing stable knock-in transgenic fish with several different constructs for 4 genetic loci at a frequency being exceeding 25%. Due to its simplicity, design flexibility, and high efficiency, we propose that CRISPR/Cas9-mediated knock-in will become a standard method for the generation transgenic zebrafish.

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

Lifescience Database Archive (English)

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

Purification of a jojoba embryo fatty acyl-coenzyme A reductase and expression of its cDNA in high erucic acid rapeseed.

Science.gov (United States)

Metz, J G; Pollard, M R; Anderson, L; Hayes, T R; Lassner, M W

2000-03-01

The jojoba (Simmondsia chinensis) plant produces esters of long-chain alcohols and fatty acids (waxes) as a seed lipid energy reserve. This is in contrast to the triglycerides found in seeds of other plants. We purified an alcohol-forming fatty acyl-coenzyme A reductase (FAR) from developing embryos and cloned the cDNA encoding the enzyme. Expression of a cDNA in Escherichia coli confers FAR activity upon those cells and results in the accumulation of fatty alcohols. The FAR sequence shows significant homology to an Arabidopsis protein of unknown function that is essential for pollen development. When the jojoba FAR cDNA is expressed in embryos of Brassica napus, long-chain alcohols can be detected in transmethylated seed oils. Resynthesis of the gene to reduce its A plus T content resulted in increased levels of alcohol production. In addition to free alcohols, novel wax esters were detected in the transgenic seed oils. In vitro assays revealed that B. napus embryos have an endogenous fatty acyl-coenzyme A: fatty alcohol acyl-transferase activity that could account for this wax synthesis. Thus, introduction of a single cDNA into B. napus results in a redirection of a portion of seed oil synthesis from triglycerides to waxes.

Downhole fluid injection systems, CO2 sequestration methods, and hydrocarbon material recovery methods

Science.gov (United States)

Schaef, Herbert T.; McGrail, B. Peter

2015-07-28

Downhole fluid injection systems are provided that can include a first well extending into a geological formation, and a fluid injector assembly located within the well. The fluid injector assembly can be configured to inject a liquid CO2/H2O-emulsion into the surrounding geological formation. CO2 sequestration methods are provided that can include exposing a geological formation to a liquid CO2/H2O-emulsion to sequester at least a portion of the CO2 from the emulsion within the formation. Hydrocarbon material recovery methods are provided that can include exposing a liquid CO2/H2O-emulsion to a geological formation having the hydrocarbon material therein. The methods can include recovering at least a portion of the hydrocarbon material from the formation.

Controlled CO2 injection into a shallow aquifer and leakage detection monitoring practices at the K-COSEM site, Korea

Science.gov (United States)

Lee, S. S.; Joun, W.; Ju, Y. J.; Ha, S. W.; Jun, S. C.; Lee, K. K.

2017-12-01

Artificial carbon dioxide injection into a shallow aquifer system was performed with two injection types imitating short- and long-term CO2 leakage events into a shallow aquifer. One is pulse type leakage of CO2 (6 hours) under a natural hydraulic gradient (0.02) and the other is long-term continuous injection (30 days) under a forced hydraulic gradient (0.2). Injection and monitoring tests were performed at the K-COSEM site in Eumseong, Korea where a specially designed well field had been installed for artificial CO2 release tests. CO2-infused and tracer gases dissolved groundwater was injected through a well below groundwater table and monitoring were conducted in both saturated and unsaturated zones. Real-time monitoring data on CO2 concentration and hydrochemical parameters, and periodical measurements of several gas tracers (He, Ar, Kr, SF6) were obtained. The pulse type short-term injection test was carried out prior to the long-term injection test. Results of the short-term injection test, under natural hydraulic gradient, showed that CO2 plume migrated along the preferential pathway identified through hydraulic interference tests. On the other hand, results of the long-term injection test indicated the CO2 plume migration path was aligned to the forced hydraulic gradient. Compared to the short-term test, the long-term injection formed detectable CO2 concentration change in unsaturated wellbores. Recovery data of tracer gases made breakthrough curves compatible to numerical simulation results. The monitoring results indicated that detection of CO2 leakage into groundwater was more effectively performed by using a pumping and monitoring method in order to capture by-passing plume. With this concept, an effective real-time monitoring method was proposed. Acknowledgement: Financial support was provided by the "R&D Project on Environmental Management of Geologic CO2storage" from the KEITI (Project number : 2014001810003)

An active role for endogenous beta-1,3-glucanase genes in transgene-mediated co-suppression in tobacco.

Science.gov (United States)

Sanders, Matthew; Maddelein, Wendy; Depicker, Anna; Van Montagu, Marc; Cornelissen, Marc; Jacobs, John

2002-11-01

Post-transcriptional gene silencing (PTGS) is characterized by the accumulation of short interfering RNAs that are proposed to mediate sequence-specific degradation of cognate and secondary target mRNAs. In plants, it is unclear to what extent endogenous genes contribute to this process. Here, we address the role of the endogenous target genes in transgene-mediated PTGS of beta-1,3-glucanases in tobacco. We found that mRNA sequences of the endogenous glucanase glb gene with varying degrees of homology to the Nicotiana plumbaginifolia gn1 transgene are targeted by the silencing machinery, although less efficiently than corresponding transgene regions. Importantly, we show that endogene-specific nucleotides in the glb sequence provide specificity to the silencing process. Consistent with this finding, small sense and antisense 21- to 23-nucleotide RNAs homologous to the endogenous glb gene were detected. Combined, these data demonstrate that a co-suppressed endogenous glucan ase gene is involved in signal amplification and selection of homologous targets, and show that endogenous genes can actively participate in PTGS in plants. The findings are introduced as a further sophistication of the post-transciptional silencing model.

Numerical Investigation into the Impact of CO2-Water-Rock Interactions on CO2 Injectivity at the Shenhua CCS Demonstration Project, China

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Guodong Yang

2017-01-01

Full Text Available A 100,000 t/year demonstration project for carbon dioxide (CO2 capture and storage in the deep saline formations of the Ordos Basin, China, has been successfully completed. Field observations suggested that the injectivity increased nearly tenfold after CO2 injection commenced without substantial pressure build-up. In order to evaluate whether this unique phenomenon could be attributed to geochemical changes, reactive transport modeling was conducted to investigate CO2-water-rock interactions and changes in porosity and permeability induced by CO2 injection. The results indicated that using porosity-permeability relationships that include tortuosity, grain size, and percolation porosity, other than typical Kozeny-Carman porosity-permeability relationship, it is possible to explain the considerable injectivity increase as a consequence of mineral dissolution. These models might be justified in terms of selective dissolution along flow paths and by dissolution or migration of plugging fines. In terms of geochemical changes, dolomite dissolution is the largest source of porosity increase. Formation physical properties such as temperature, pressure, and brine salinity were found to have modest effects on mineral dissolution and precipitation. Results from this study could have practical implications for a successful CO2 injection and enhanced oil/gas/geothermal production in low-permeability formations, potentially providing a new basis for screening of storage sites and reservoirs.

Large-scale CO2 injection demos for the development of monitoring and verification technology and guidelines (CO2ReMoVe)

Energy Technology Data Exchange (ETDEWEB)

Wildenborg, T.; David, P. [TNO Built Environment and Geosciences, Princetonlaan 6, 3584 CB Utrecht (Netherlands); Bentham, M.; Chadwick, A.; Kirk, K. [British Geological Survey, Kingsley Dunham Centre, Keyworth, Nottingham NG12 5GG (United Kingdom); Dillen, M. [SINTEF Petroleum Research, Trondheim (Norway); Groenenberg, H. [Unit Policy Studies, Energy Research Centre of the Netherlands ECN, Amsterdam (Netherlands); Deflandre, J.P.; Le Gallo, J. [Institut Francais du Petrole, Rueil-Malmaison (France)

2009-04-15

The objectives of the EU project CO2ReMoVe are to undertake the research and development necessary to establish scientifically based standards for monitoring future CCS operations and to develop the performance assessment methodologies necessary to demonstrate the long-term reliability of geological storage of CO2. This could in turn lead to guidelines for the certification of sites suitable for CCS on a wide scale. Crucial to the project portfolio are the continuing large-scale CO2 injection operation at Sleipner, the injection operation at In Salah (Algeria) and the recently started injection project at Snoehvit (Norway). Two pilot sites are also currently in the project portfolio, Ketzin in Germany and K12-B in the offshore continental shelf of the Netherlands.

Simulation of CO2 Injection in Porous Media with Structural Deformation Effect

KAUST Repository

Negara, Ardiansyah

2011-01-01

Carbon dioxide (CO2) sequestration is one of the most attractive methods to reduce the amount of CO2 in the atmosphere by injecting it into the geological formations. Furthermore, it is also an effective mechanism for enhanced oil recovery

Surface-downhole and crosshole geoelectrics for monitoring of brine injection at the Ketzin CO2 storage site

Science.gov (United States)

Rippe, Dennis; Bergmann, Peter; Labitzke, Tim; Wagner, Florian; Schmidt-Hattenberger, Cornelia

2016-04-01

The Ketzin pilot site in Germany is the longest operating on-shore CO2 storage site in Europe. From June 2008 till August 2013, a total of ˜67,000 tonnes of CO2 were safely stored in a saline aquifer at depths of 630 m to 650 m. The storage site has now entered the abandonment phase, and continuation of the multi-disciplinary monitoring as part of the national project "CO2 post-injection monitoring and post-closure phase at the Ketzin pilot site" (COMPLETE) provides the unique chance to participate in the conclusion of the complete life cycle of a CO2 storage site. As part of the continuous evaluation of the functionality and integrity of the CO2 storage in Ketzin, from October 12, 2015 till January 6, 2015 a total of ˜2,900 tonnes of brine were successfully injected into the CO2 reservoir, hereby simulating in time-lapse the natural backflow of brine and the associated displacement of CO2. The main objectives of this brine injection experiment include investigation of how much of the CO2 in the pore space can be displaced by brine and if this displacement of CO2 during the brine injection differs from the displacement of formation fluid during the initial CO2 injection. Geophysical monitoring of the brine injection included continuous geoelectric measurements accompanied by monitoring of pressure and temperature conditions in the injection well and two adjacent observation wells. During the previous CO2 injection, the geoelectrical monitoring concept at the Ketzin pilot site consisted of permanent crosshole measurements and non-permanent large-scale surveys (Kiessling et al., 2010). Time-lapse geoelectrical tomographies derived from the weekly crosshole data at near-wellbore scale complemented by six surface-downhole surveys at a scale of 1.5 km showed a noticeable resistivity signature within the target storage zone, which was attributed to the CO2 plume (Schmidt-Hattenberger et al., 2011) and interpreted in terms of relative CO2 and brine saturations (Bergmann

Cloning of genes and developing transgenic crops with enhanced tolerance to salinity and drought (abstract)

International Nuclear Information System (INIS)

Bansal, K.C.; Chinnusamy, V.; Tayal, D.; Das, A.; Goel, D.; Yadav, V.; Singh, A.K.; Lakhshmi, K.

2005-01-01

Abiotic stresses represent the most limiting factors affecting agricultural productivity. In India more than 60% of total cultivated land is still rainfed and crops experience frequent droughts. Thus, we need to develop transgenic crops tolerant to drought, and other related abiotic stress factors such as salinity, low and high temperature stresses. At the National Research Centre on Plant Biotechnology, Indian Agricultural Research Institute (ICAR), we have initiated a programme on developing transgenic crops tolerant to a range of abiotic stresses. The major emphasis is on developing transgenic potato, tomato, mustard, rice and wheat. While, transgenic plants of potato. tomato and mustard have already been generated with osmotin gene and are at different stages of testing, other key genes imparting tolerance to abiotic stresses are being isolated from different species for producing transgenic rice and wheat cultivars tolerant to multiple stresses. Genes that have been isolated in our laboratory include ascorbate peroxidase gene (TaApx) and genes encoding transcription factor, CBFs (TaCBF2 and TaCBP3) from a drought tolerant wheat cultivar (C306), Lea1 cDNA from Brassica species, codA from Arthrobacter globiformis, and otsBA operon from E. coli. Apart from these stress-related genes, we have isolated a few stress-inducible promoters for deploying them in gene stacking in developing transgenic crops with enhanced tolerance to multiple abiotic stresses. The results will be presented. (author)

Experimental Investigations into CO2 Interactions with Injection Well Infrastructure for CO2 Storage

Science.gov (United States)

Syed, Amer; Shi, Ji-Quan; Durucan, Sevket; Nash, Graham; Korre, Anna

2013-04-01

Wellbore integrity is an essential requirement to ensure the success of a CO2 Storage project as leakage of CO2 from the injection or any other abandoned well in the storage complex, could not only severely impede the efficiency of CO2 injection and storage but also may result in potential adverse impact on the surrounding environment. Early research has revealed that in case of improper well completions and/or significant changes in operating bottomhole pressure and temperature could lead to the creation of microannulus at cement-casing interface which may constitute a preferential pathway for potential CO2 leakage during and post injection period. As a part of a European Commission funded CO2CARE project, the current research investigates the sealing behaviour of such microannulus at the cement-casing interface under simulated subsurface reservoir pressure and temperature conditions and uses the findings to develop a methodology to assess the overall integrity of CO2 storage. A full scale wellbore experimental test set up was constructed for use under elevated pressure and temperature conditions as encountered in typical CO2 storage sites. The wellbore cell consists of an assembly of concentric elements of full scale casing (Diameter= 0.1524m), cement sheath and an outer casing. The stainless steel outer ring is intended to simulate the stiffness offered by the reservoir rock to the displacement applied at the wellbore. The Central Loading Mechanism (CLM) consists of four case hardened shoes that can impart radial load onto the well casing. The radial movement of the shoes is powered through the synchronised movement of four precision jacks controlled hydraulically which could impart radial pressures up to 15 MPa. The cell body is a gas tight enclosure that houses the wellbore and the central loading mechanism. The setup is enclosed in a laboratory oven which acts both as temperature and safety enclosure. Prior to a test, cement mix is set between the casing and

Recent progress on technologies and applications of transgenic ...

African Journals Online (AJOL)

USER

2010-06-14

Jun 14, 2010 ... embryo with DNA or viral vectors is possible, but this approach does not have the ... re-injecting them to cock testis is the most efficient and cost-effective strategy to .... transgenic manipulation, but also allows the selection of.

Pore network quantification of sandstones under experimental CO2 injection using image analysis

Science.gov (United States)

Berrezueta, Edgar; González-Menéndez, Luís; Ordóñez-Casado, Berta; Olaya, Peter

2015-04-01

Automated-image identification and quantification of minerals, pores and textures together with petrographic analysis can be applied to improve pore system characterization in sedimentary rocks. Our case study is focused on the application of these techniques to study the evolution of rock pore network subjected to super critical CO2-injection. We have proposed a Digital Image Analysis (DIA) protocol that guarantees measurement reproducibility and reliability. This can be summarized in the following stages: (i) detailed description of mineralogy and texture (before and after CO2-injection) by optical and scanning electron microscopy (SEM) techniques using thin sections; (ii) adjustment and calibration of DIA tools; (iii) data acquisition protocol based on image capture with different polarization conditions (synchronized movement of polarizers); (iv) study and quantification by DIA that allow (a) identification and isolation of pixels that belong to the same category: minerals vs. pores in each sample and (b) measurement of changes in pore network, after the samples have been exposed to new conditions (in our case: SC-CO2-injection). Finally, interpretation of the petrography and the measured data by an automated approach were done. In our applied study, the DIA results highlight the changes observed by SEM and microscopic techniques, which consisted in a porosity increase when CO2 treatment occurs. Other additional changes were minor: variations in the roughness and roundness of pore edges, and pore aspect ratio, shown in the bigger pore population. Additionally, statistic tests of pore parameters measured were applied to verify that the differences observed between samples before and after CO2-injection were significant.

Co-overexpressing a Plasma Membrane and a Vacuolar Membrane Sodium/Proton Antiporter Significantly Improves Salt Tolerance in Transgenic Arabidopsis Plants

Science.gov (United States)

Pehlivan, Necla; Sun, Li; Jarrett, Philip; Yang, Xiaojie; Mishra, Neelam; Chen, Lin; Kadioglu, Asim; Shen, Guoxin; Zhang, Hong

2016-01-01

The Arabidopsis gene AtNHX1 encodes a vacuolar membrane-bound sodium/proton (Na+/H+) antiporter that transports Na+ into the vacuole and exports H+ into the cytoplasm. The Arabidopsis gene SOS1 encodes a plasma membrane-bound Na+/H+ antiporter that exports Na+ to the extracellular space and imports H+ into the plant cell. Plants rely on these enzymes either to keep Na+ out of the cell or to sequester Na+ into vacuoles to avoid the toxic level of Na+ in the cytoplasm. Overexpression of AtNHX1 or SOS1 could improve salt tolerance in transgenic plants, but the improved salt tolerance is limited. NaCl at concentration >200 mM would kill AtNHX1-overexpressing or SOS1-overexpressing plants. Here it is shown that co-overexpressing AtNHX1 and SOS1 could further improve salt tolerance in transgenic Arabidopsis plants, making transgenic Arabidopsis able to tolerate up to 250 mM NaCl treatment. Furthermore, co-overexpression of AtNHX1 and SOS1 could significantly reduce yield loss caused by the combined stresses of heat and salt, confirming the hypothesis that stacked overexpression of two genes could substantially improve tolerance against multiple stresses. This research serves as a proof of concept for improving salt tolerance in other plants including crops. PMID:26985021

Generation and characterization of a transgenic pig carrying a DsRed-monomer reporter gene.

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Chih-Jen Chou

Full Text Available Pigs are an optimal animal for conducting biomedical research because of their anatomical and physiological resemblance to humans. In contrast to the abundant resources available in the study of mice, few fluorescent protein-harboring porcine models are available for preclinical studies. In this paper, we report the successful generation and characterization of a transgenic DsRed-Monomer porcine model.The transgene comprised a CMV enhancer/chicken-beta actin promoter and DsRed monomeric cDNA. Transgenic pigs were produced by using pronuclear microinjection. PCR and Southern blot analyses were applied for identification of the transgene. Histology, blood examinations and computed tomography were performed to study the health conditions. The pig amniotic fluid progenitor/stem cells were also isolated to examine the existence of red fluorescence and differentiation ability.Transgenic pigs were successfully generated and transmitted to offspring at a germ-line transmission rate of 43.59% (17/39. Ubiquitous expression of red fluorescence was detected in the brain, eye, tongue, heart, lung, liver, pancreas, spleen, stomach, small intestine, large intestine, kidney, testis, and muscle; this was confirmed by histology and western blot analyses. In addition, we confirmed the differentiation potential of amniotic fluid progenitor stem cells isolated from the transgenic pig.This red fluorescent pig can serve as a host for other fluorescent-labeled cells in order to study cell-microenvironment interactions, and can provide optimal red-fluorescent-labeled cells and tissues for research in developmental biology, regenerative medicine, and xenotransplantation.

Podocyte changes upon induction of albuminuria in Thy-1.1 transgenic mice.

Science.gov (United States)

Smeets, Bart; Dijkman, Henry B P M; te Loeke, Nathalie A J M; van Son, Jacco P H F; Steenbergen, Eric J; Assmann, Karel J M; Wetzels, Jack F M; Groenen, Patricia J T A

2003-12-01

Thy-1.1 transgenic mice, characterized by ectopic expression of the Thy-1.1 protein on podocytes, spontaneously develop proteinuria and focal glomerulosclerosis (FGS). Injection of a monoclonal antibody (mAb) directed against the Thy-1.1 protein in young transgenic mice induces a massive albuminuria that is followed by an accelerated FGS within 3 weeks. This albuminuria is complement and leukocyte independent. The time course of proteinuria, the pathogenesis of the acute proteinuria and the dose dependency of FGS are unknown. Albuminuria was measured in Thy-1.1 transgenic mice after injection of different doses of anti-Thy-1.1 mAb and at different time points within the first 24 h after injection. Podocytic foot processes and slit pore diameter were quantitated by electron microscopy. Changes in expression of slit pore constituents (podocin, CD2AP, nephrin and ZO-1), cytoskeleton-associated proteins (actin, alpha-actinin, ezrin and synaptopodin), the GDH-podocyte adhesion molecules alpha(3)-integrin, and heparan sulfate were studied by immunofluorescence. FGS was scored by light microscopy at 3 weeks after induction of albuminuria. Albuminuria in Thy-1.1 transgenic mice was observed within 10 min after anti-Thy-1.1 mAb injection. This rapid development of albuminuria was accompanied by a reduction in number of podocytic foot processes from 20.0 +/- 0.7/10 microm glomerular basement membrane (GBM) in saline-treated transgenic mice to 8.0 +/- 0.5 and 2.2 +/- 0.2 in anti-Thy-1.1-treated mice, at 10 min and 8 h after treatment, respectively. In addition, we observed a significant decrease in width of remaining slit pores, from 32.7 +/- 1.1 to 26.8 +/- 1.4 nm at 10 min after mAb injection. By immunofluorescence, we did not observe major changes in the expression pattern of any of the proteins studied. There was no correlation between the injected dose of the anti-Thy-1.1 mAb and the acute albuminuria. In contrast, the percentage of FGS at 3 weeks correlated with the

Mode of transgene expression after fusion to early or late viral genes of a conditionally replicating adenovirus via an optimized internal ribosome entry site in vitro and in vivo

International Nuclear Information System (INIS)

Rivera, Angel A.; Wang Minghui; Suzuki, Kaori; Uil, Taco G.; Krasnykh, Victor; Curiel, David T.; Nettelbeck, Dirk M.

2004-01-01

The expression of therapeutic genes by oncolytic viruses is a promising strategy to improve viral oncolysis, to augment gene transfer compared with a nonreplicating adenoviral vector, or to combine virotherapy and gene therapy. Both the mode of transgene expression and the locale of transgene insertion into the virus genome critically determine the efficacy of this approach. We report here on the properties of oncolytic adenoviruses which contain the luciferase cDNA fused via an optimized internal ribosome entry site (IRES) to the immediate early adenoviral gene E1A (AdΔE1AIL), the early gene E2B (AdΔE2BIL), or the late fiber gene (AdΔfiberIL). These viruses showed distinct kinetics of transgene expression and luciferase activity. Early after infection, luciferase activities were lower for these viruses, especially for AdΔE2BIL, compared with nonreplicating AdTL, which contained the luciferase gene expressed from the strong CMV promoter. However, 6 days after infection, luciferase activities were approximately four (AdΔE1AIL) to six (AdΔfiberIL) orders of magnitude higher than for AdTL, reflecting virus replication and efficient transgene expression. Similar results were obtained in vivo after intratumoral injection of AdΔE2BIL, AdΔfiberIL, and AdTL. AdΔfiberIL and the parental virus, Ad5-Δ24, resulted in similar cytotoxicity, but AdΔE2BIL and AdΔE1AIL were slightly attenuated. Disruption of the expression of neighboring viral genes by insertion of the transgene was minimal for AdΔE2BIL and AdΔfiberIL, but substantial for AdΔE1AIL. Our observations suggest that insertion of IRES-transgene cassettes into viral transcription units is an attractive strategy for the development of armed oncolytic adenoviruses with defined kinetics and strength of transgene expression

The Effect of CO2 Injection on Macroalgae Gelidium latifolium Biomass Growth Rate and Carbohydrate Content

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Mujizat Kawaroe

2016-06-01

Full Text Available There are many species of macroalga grow in marine ecosystem and potentially as raw material for bioethanol resource. Bioethanol is a conversion result of carbohydrate, one of macroalgae biomass content. The exploration of macroalgae require information about  growth rate ability to determine availability in the nature. This research analyze growth rate and carbohydrate content of marine macroalga Gelidium latifolium on cultivation using varied injection of carbon dioxide and aeration. The treatments were control (K, 2000 cc CO2 injection and aeration (P1, 3000 cc CO2 injection and aeration (P2, 2000 cc CO2 injection without aeration (P3, and 3000 cc CO2 injection without aeration (P4. Samples weight were 3 gram in early cultivation on laboratorium scale for 42 days observation. The results showed that the daily growth rate Gelidium latifolium during the study ranged from 0.02-1.06%. The highest daily growth rate was 1.06±0.14% (P2. Carbohydrate yield was 18.23% in early cultivation then 19.40% (K and P2, 20.40% (P1, 16.87% (K3, and 16.40% (P4 after cultivation. The high of carbohydrates value may not guarantee the sustainable Gelidium latifolium biomass utilization as raw material for bioethanol production because of the low growth rate, thus it is necessary to modified and encourage cultivation method effectively. Keywords: CO2 injection, growth rate, carbohydrate, macroalgae, Gelidium latifolium

RFLP for Duchenne muscular dystrophy cDNA clone 30-2

Energy Technology Data Exchange (ETDEWEB)

Walker, A P; Bartlett, R J; Laing, N G; Siddique, T; Yamaoka, L H; Chen, J C; Hung, W Y; Roses, A D [Duke Univ. Medical Center, Durham, NC (USA)

1988-09-26

30-2 is one of 6 cDNA clones which comprise the cDNA for the Duchenne muscular dystrophy gene. It is a 1.15 kb fragment in the EcoRI site of Bluescribe. TaqI (T{down arrow}CGA) identifies two bands with alleles at 3.7 and 3.5 kb, as well as eight constant bands at 9.0, 7.5, 4.6, 3.6, 3.4, 2.5, 1.7 and 1.4 kb. The allele frequency was studied in 47 unrelated DMD males: 3.7 kb allele 0.45; and 3.5 kb allele 0.55. Co-dominant X-linked segregation was demonstrated in two 2-generation families. 1.1% agarose gels required to resolve the bands. The polymorphism is also recognized by PERT 87-15.

Sensitivity of CO2 storage performance to varying rates and dynamic injectivity in the Bunter Sandstone, UK

Science.gov (United States)

Kolster, C.; Mac Dowell, N.; Krevor, S. C.; Agada, S.

2016-12-01

Carbon capture and storage (CCS) is needed for meeting legally binding greenhouse gas emissions targets in the UK (ECCC 2016). Energy systems models have been key to identifying the importance of CCS but they tend to impose few constraints on the availability and use of geologic CO2 storage reservoirs. Our aim is to develop simple models that use dynamic representations of limits on CO2 storage resources. This will allow for a first order representation of the storage reservoir for use in systems models with CCS. We use the ECLIPSE reservoir simulator and a model of the Southern North Sea Bunter Sandstone saline aquifer. We analyse reservoir performance sensitivities to scenarios of varying CO2 injection demand for a future UK low carbon energy market. With 12 injection sites, we compare the impact of injecting at a constant 2MtCO2/year per site and varying this rate by a factor of 1.8 and 0.2 cyclically every 5 and 2.5 years over 50 years of injection. The results show a maximum difference in average reservoir pressure of 3% amongst each case and a similar variation in plume migration extent. This suggests that simplified models can maintain accuracy by using average rates of injection over similar time periods. Meanwhile, by initiating injection at rates limited by pressurization at the wellhead we find that injectivity steadily increases. As a result, dynamic capacity increases. We find that instead of injecting into sites on a need basis, we can strategically inject the CO2 into 6 of the deepest sites increasing injectivity for the first 15 years by 13%. Our results show injectivity as highly dependent on reservoir heterogeneity near the injection site. Injecting 1MTCO2/year into a shallow, low permeability and porosity site instead of into a deep injection site with high permeability and porosity reduces injectivity in the first 5 years by 52%. ECCC. 2016. Future of Carbon Capture and Storage in the UK. UK Parliament House of Commons, Energy and Climate Change

Generation of marker-free transgenic hexaploid wheat via an Agrobacterium-mediated co-transformation strategy in commercial Chinese wheat varieties.

Science.gov (United States)

Wang, Ke; Liu, Huiyun; Du, Lipu; Ye, Xingguo

2017-05-01

Genotype specificity is a big problem lagging the development of efficient hexaploid wheat transformation system. Increasingly, the biosecurity of genetically modified organisms is garnering public attention, so the generation of marker-free transgenic plants is very important to the eventual potential commercial release of transgenic wheat. In this study, 15 commercial Chinese hexaploid wheat varieties were successfully transformed via an Agrobacterium-mediated method, with efficiency of up to 37.7%, as confirmed by the use of Quickstix strips, histochemical staining, PCR analysis and Southern blotting. Of particular interest, marker-free transgenic wheat plants from various commercial Chinese varieties and their F 1 hybrids were successfully obtained for the first time, with a frequency of 4.3%, using a plasmid harbouring two independent T-DNA regions. The average co-integration frequency of the gus and the bar genes located on the two independent T-DNA regions was 49.0% in T 0 plants. We further found that the efficiency of generating marker-free plants was related to the number of bar gene copies integrated in the genome. Marker-free transgenic wheat plants were identified in the progeny of three transgenic lines that had only one or two bar gene copies. Moreover, silencing of the bar gene was detected in 30.7% of T 1 positive plants, but the gus gene was never found to be silenced in T 1 plants. Bisulphite genomic sequencing suggested that DNA methylation in the 35S promoter of the bar gene regulatory region might be the main reason for bar gene silencing in the transgenic plants. © 2016 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

Numerical modeling of injection and mineral trapping of CO2 withH2S and SO2 in a Sandstone Formation

Energy Technology Data Exchange (ETDEWEB)

Xu, Tianfu; Apps, John A.; Pruess, Karsten; Yamamoto, Hajime

2004-09-07

Carbon dioxide (CO{sub 2}) injection into deep geologic formations could decrease the atmospheric accumulation of this gas from anthropogenic sources. Furthermore, by co-injecting H{sub 2}S or SO{sub 2}, the products respectively of coal gasification or combustion, with captured CO{sub 2}, problems associated with surface disposal would be mitigated. We developed models that simulate the co-injection of H{sub 2}S or SO{sub 2} with CO{sub 2} into an arkose formation at a depth of about 2 km and 75 C. The hydrogeology and mineralogy of the injected formation are typical of those encountered in Gulf Coast aquifers of the United States. Six numerical simulations of a simplified 1-D radial region surrounding the injection well were performed. The injection of CO{sub 2} alone or co-injection with SO{sub 2} or H{sub 2}S results in a concentrically zoned distribution of secondary minerals surrounding a leached and acidified region adjacent to the injection well. Co-injection of SO{sub 2} with CO{sub 2} results in a larger and more strongly acidified zone, and alteration differs substantially from that caused by the co-injection of H{sub 2}S or injection of CO{sub 2} alone. Precipitation of carbonates occurs within a higher pH (pH > 5) peripheral zone. Significant quantities of CO{sub 2} are sequestered by ankerite, dawsonite, and lesser siderite. The CO{sub 2} mineral-trapping capacity of the formation can attain 40-50 kg/m{sup 3} medium for the selected arkose. In contrast, secondary sulfates precipitate at lower pH (pH < 5) within the acidified zone. Most of the injected SO{sub 2} is transformed and immobilized through alunite precipitation with lesser amounts of anhydrite and minor quantities of pyrite. The dissolved CO{sub 2} increases with time (enhanced solubility trapping). The mineral alteration induced by injection of CO{sub 2} with either SO{sub 2} or H{sub 2}S leads to corresponding changes in porosity. Significant increases in porosity occur in the acidified

Expression of a maize Myb transcription factor driven by a putative silk-specific promoter significantly enhances resistance to Helicoverpa zea in transgenic maize.

Science.gov (United States)

Johnson, Eric T; Berhow, Mark A; Dowd, Patrick F

2007-04-18

Hi II maize (Zea mays) plants were engineered to express maize p1 cDNA, a Myb transcription factor, controlled by a putative silk specific promoter, for secondary metabolite production and corn earworm resistance. Transgene expression did not enhance silk color, but about half of the transformed plant silks displayed browning when cut, which indicated the presence of p1-produced secondary metabolites. Levels of maysin, a secondary metabolite with insect toxicity, were highest in newly emerged browning silks. The insect resistance of transgenic silks was also highest at emergence, regardless of maysin levels, which suggests that other unidentified p1-induced molecules likely contributed to larval mortality. Mean survivor weights of corn earworm larvae fed mature browning transgenic silks were significantly lower than weights of those fed mature nonbrowning transgenic silks. Some transgenic pericarps browned with drying and contained similar molecules found in pericarps expressing a dominant p1 allele, suggesting that the promoter may not be silk-specific.

High-efficiency production of human serum albumin in the posterior silk glands of transgenic silkworms, Bombyx mori L.

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Qiujie Qian

Full Text Available Human serum albumin (HSA is an important biological preparation with a variety of biological functions in clinical applications. In this study, the mRNA of a fusion transposase derived from the pESNT-PBase plasmid and a pBHSA plasmid containing the HSA gene under the control of a fibroin light chain (FL promoter were co-injected into fertilized eggs. Fifty-six transgenic silkworm pedigrees expressing theexogenous recombinant HSA (rHSA in the posterior silk glands (PSGs with stable inheritance were successfully obtained. The SDS-PAGE and Western blot results confirmed that the rHSA was secreted into the transgenic silkworm cocoon, and the rHSA could be easily extracted with phosphate-buffered saline (PBS. In our research, the isolated highest amount rHSA constituted up to 29.1% of the total soluble protein of the cocoon shell, indicating that the transgenic silkworm produced an average of 17.4 μg/mg of rHSA in the cocoon shell. The production of soluble rHSA in the PSGs by means of generating transgenic silkworms is a novel approach, whereby a large amount of virus-free and functional HSA can be produced through the simple rearing of silkworms.

Effects of Elevated Carbon Dioxide on the Growth and Foliar Chemistry of Transgenic Bt Cotton

Institute of Scientific and Technical Information of China (English)

Gang Wu; Fa-Jun Chen; Feng Ge; Yu-Cheng Sun

2007-01-01

A field study was carried out to quantify plant growth and the foliar chemistry of transgenic Bacillus thuringiensis (Bt)cotton (cv. GK-12) exposed to ambient CO2 and elevated (double-ambient) CO2 for different lengths of time (1, 2 and 3 months) in 2004 and 2005. The results indicated that CO2 levels significantly affected plant height, leaf area per plant and leaf chemistry of transgenic Bt cotton. Significantly, higher plant height and leaf area per plant were observed after cotton plants that were grown in elevated CO2 were compared with plants grown in ambient CO2 for 1, 2 and 3 months in the investigation. Simultaneously, significant interaction between CO2 level x investigating year was observed in leaf area per plant. Moreover, foliar total amino acids were increased by 14%, 13%, 11% and 12%, 14%, 10% in transgenic Bt cotton after exposed to elevated CO2 for 1, 2 or 3 months compared with ambient CO2 in 2004 and 2005, respectively. Condensed tannin occurrence increased by 17%, 11%, 9% in 2004 and 12%, 11%, 9% in 2005 in transgenic Bt cotton after being exposed to elevated CO2 for 1, 2 or 3 months compared with ambient CO2 for the same time. However, Bt toxin decreased by 3.0%,2.9%, 3.1% and 2.4%, 2.5%, 2.9% in transgenic Bt cotton after exposed to elevated CO2 for 1, 2 or 3months compared with ambient CO2 for same time in 2004 and 2005, respectively. Furthermore, there was prominent interaction on the foliar total amino acids between the CO2 level and the time of cotton plant being exposed to elevated CO2. It is presumed that elevated CO2 can alter the plant growth and hence ultimately the phenotype allocation to foliar chemistical components of transgenic Bt cotton, which may in turn, affect the plant-herbivore interactions.

Improved forage digestibility of tall fescue (Festuca arundinacea) by transgenic down-regulation of cinnamyl alcohol dehydrogenase.

Science.gov (United States)

Chen, Lei; Auh, Chung-Kyoon; Dowling, Paul; Bell, Jeremey; Chen, Fang; Hopkins, Andrew; Dixon, Richard A; Wang, Zeng-Yu

2003-11-01

Lignification of cell walls during plant development has been identified as the major factor limiting forage digestibility and concomitantly animal productivity. cDNA sequences encoding a key lignin biosynthetic enzyme, cinnamyl alcohol dehydrogenase (CAD), were cloned from the widely grown monocotyledonous forage species tall fescue (Festuca arundinacea Schreb.). Recombinant tall fescue CAD expressed in E. coli exhibited the highest V(max)/K(m) values when coniferaldehyde and sinapaldehyde were used as substrates. Transgenic tall fescue plants carrying either sense or antisense CAD gene constructs were obtained by microprojectile bombardment of single genotype-derived embryogenic suspension cells. Severely reduced levels of mRNA transcripts and significantly reduced CAD enzymatic activities were found in two transgenic plants carrying sense and antisense CAD transgenes, respectively. These CAD down-regulated transgenic lines had significantly decreased lignin content and altered ratios of syringyl (S) to guaiacyl (G), G to p-hydroxyphenyl (H) and S to H units. No significant changes in cellulose, hemicellulose, neutral sugar composition, p-coumaric acid and ferulic acid levels were observed in the transgenic plants. Increases of in vitro dry matter digestibility of 7.2-9.5% were achieved in the CAD down-regulated lines, thus providing a novel germplasm to be used for the development of grass cultivars with improved forage quality.

Assessment of brine migration risks along vertical pathways due to CO2 injection

Science.gov (United States)

Kissinger, Alexander; Class, Holger

2015-04-01

Global climate change, shortage of resources and the growing usage of renewable energy sources has lead to a growing demand for the utilization of subsurface systems. Among these competing uses are Carbon Capture and Storage (CCS), geothermal energy, nuclear waste disposal, 'renewable' methane or hydrogen storage as well as the ongoing production of fossil resources like oil, gas and coal. Additionally, these technologies may also create conflicts with essential public interests such as water supply. For example, the injection of CO2 into the subsurface causes an increase in pressure reaching far beyond the actual radius of influence of the CO2 plume, potentially leading to large amounts of displaced salt water. In this work we focus on the large scale impacts of CO2 storage on brine migration but the methodology and the obtained results may also apply to other fields like waste water disposal, where large amounts of fluid are injected into the subsurface. In contrast to modeling on the reservoir scale the spatial scale required for this work is much larger in both vertical and lateral direction, as the regional hydrogeology has to be considered. Structures such as fault zones, hydrogeological windows in the Rupelian clay or salt domes are considered as potential pathways for displaced fluids into shallow systems and their influence has to be taken into account. We put the focus of our investigations on the latter type of scenario, since there is still a poor understanding of the role that salt diapirs would play in CO2 storage projects. As there is hardly any field data available on this scale, we compare different levels of model complexity in order to identify the relevant processes for brine displacement and simplify the modeling process wherever possible, for example brine injection vs. CO2 injection, simplified geometries vs. the complex formation geometry and the role of salt induced density differences on flow. Further we investigate the impact of the

Caprock Integrity during Hydrocarbon Production and CO2 Injection in the Goldeneye Reservoir

Science.gov (United States)

Salimzadeh, Saeed; Paluszny, Adriana; Zimmerman, Robert

2016-04-01

Carbon Capture and Storage (CCS) is a key technology for addressing climate change and maintaining security of energy supplies, while potentially offering important economic benefits. UK offshore, depleted hydrocarbon reservoirs have the potential capacity to store significant quantities of carbon dioxide, produced during power generation from fossil fuels. The Goldeneye depleted gas condensate field, located offshore in the UK North Sea at a depth of ~ 2600 m, is a candidate for the storage of at least 10 million tons of CO2. In this research, a fully coupled, full-scale model (50×20×8 km), based on the Goldeneye reservoir, is built and used for hydro-carbon production and CO2 injection simulations. The model accounts for fluid flow, heat transfer, and deformation of the fractured reservoir. Flow through fractures is defined as two-dimensional laminar flow within the three-dimensional poroelastic medium. The local thermal non-equilibrium between injected CO2 and host reservoir has been considered with convective (conduction and advection) heat transfer. The numerical model has been developed using standard finite element method with Galerkin spatial discretisation, and finite difference temporal discretisation. The geomechanical model has been implemented into the object-oriented Imperial College Geomechanics Toolkit, in close interaction with the Complex Systems Modelling Platform (CSMP), and validated with several benchmark examples. Fifteen major faults are mapped from the Goldeneye field into the model. Modal stress intensity factors, for the three modes of fracture opening during hydrocarbon production and CO2 injection phases, are computed at the tips of the faults by computing the I-Integral over a virtual disk. Contact stresses -normal and shear- on the fault surfaces are iteratively computed using a gap-based augmented Lagrangian-Uzawa method. Results show fault activation during the production phase that may affect the fault's hydraulic conductivity

Performance Analysis of Cold Energy Recovery from CO2 Injection in Ship-Based Carbon Capture and Storage (CCS

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Hwalong You

2014-11-01

Full Text Available Carbon capture and storage (CCS technology is one of the practical solutions for mitigating the effects of global warming. When captured CO2 is injected into storage sites, the CO2 is subjected to a heating process. In a conventional CO2 injection system, CO2 cold energy is wasted during this heating process. This study proposes a new CO2 injection system that takes advantage of the cold energy using the Rankine cycle. The study compared the conventional system with the new CO2 injection system in terms of specific net power consumption, exergy efficiency, and life-cycle cost (LCC to estimate the economic effects. The results showed that the new system reduced specific net power consumption and yielded higher exergy efficiency. The LCC of the new system was more economical. Several cases were examined corresponding to different conditions, specifically, discharge pressure and seawater temperature. This information may affect decision-making when CCS projects are implemented.

Methane Production from Gas Hydrate Deposits through Injection of Supercritical CO₂

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Matthias Haeckel

2012-06-01

Full Text Available The recovery of natural gas from CH₄-hydrate deposits in sub-marine and sub-permafrost environments through injection of CO₂ is considered a suitable strategy towards emission-neutral energy production. This study shows that the injection of hot, supercritical CO₂ is particularly promising. The addition of heat triggers the dissociation of CH₄-hydrate while the CO₂, once thermally equilibrated, reacts with the pore water and is retained in the reservoir as immobile CO₂-hydrate. Furthermore, optimal reservoir conditions of pressure and temperature are constrained. Experiments were conducted in a high-pressure flow-through reactor at different sediment temperatures (2 °C, 8 °C, 10 °C and hydrostatic pressures (8 MPa, 13 MPa. The efficiency of both, CH₄ production and CO₂ retention is best at 8 °C, 13 MPa. Here, both CO₂- and CH₄-hydrate as well as mixed hydrates can form. At 2 °C, the production process was less effective due to congestion of transport pathways through the sediment by rapidly forming CO₂-hydrate. In contrast, at 10 °C CH₄ production suffered from local increases in permeability and fast breakthrough of the injection fluid, thereby confining the accessibility to the CH₄ pool to only the most prominent fluid channels. Mass and volume balancing of the collected gas and fluid stream identified gas mobilization as equally important process parameter in addition to the rates of methane hydrate dissociation and hydrate conversion. Thus, the combination of heat supply and CO₂ injection in one supercritical phase helps to overcome the mass transfer limitations usually observed in experiments with cold liquid or gaseous CO₂.

Generation of antiviral transgenic chicken using spermatogonial ...

African Journals Online (AJOL)

This study was conducted in order to generate anti-viral transgenic chickens through transfected spermatogonial stem cell with fusion gene EGFP-MMx. After injecting fusion gene EGFP-MMx into testes, tissues frozen section, polymerase chain reaction (PCR) and dot blot of testes was performed at 30, 40, 50, 60, 70 and 80 ...

DNA Nanoparticles: Detection of Long-Term Transgene Activity in Brain using Bioluminescence Imaging

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David M. Yurek

2011-09-01

Full Text Available In this study, we used bioluminescence imaging (BLI to track long-term transgene activity following the transfection of brain cells using a nonviral gene therapy technique. Formulations of deoxyribonucleic acid (DNA combined with 30-mer lysine polymers (substituted with 10 kDa polyethylene glycol form nanoparticles that transfect brain cells in vivo and produce transgene activity. Here we show that a single intracerebral injection of these DNA nanoparticles (DNPs into the rat cortex, striatum, or substantia nigra results in long-term and persistent luciferase transgene activity over an 8- to 11-week period as evaluated by in vivo BLI analysis, and single injections of DNPs into the mouse striatum showed stable luciferase transgene activity for 1 year. Compacted DNPs produced in vivo signals 7- to 34-fold higher than DNA alone. In contrast, ex vivo BLI analysis, which is subject to less signal quenching from surrounding tissues, demonstrated a DNP to DNA alone ratio of 76- to 280-fold. Moreover, the ex vivo BLI analysis confirmed that signals originated from the targeted brain structures. In summary, BLI permits serial analysis of luciferase transgene activity at multiple brain locations following gene transfer with DNPs. Ex vivo analysis may permit more accurate determination of relative activities of gene transfer vectors.

Fate of injected CO2 in the Wilcox Group, Louisiana, Gulf Coast Basin: Chemical and isotopic tracers of microbial–brine–rock–CO2 interactions

Science.gov (United States)

Shelton, Jenna L.; McIntosh, Jennifer C.; Warwick, Peter D.; Lee Zhi Yi, Amelia

2014-01-01

The “2800’ sandstone” of the Olla oil field is an oil and gas-producing reservoir in a coal-bearing interval of the Paleocene–Eocene Wilcox Group in north-central Louisiana, USA. In the 1980s, this producing unit was flooded with CO2 in an enhanced oil recovery (EOR) project, leaving ∼30% of the injected CO2 in the 2800’ sandstone post-injection. This study utilizes isotopic and geochemical tracers from co-produced natural gas, oil and brine to determine the fate of the injected CO2, including the possibility of enhanced microbial conversion of CO2 to CH4 via methanogenesis. Stable carbon isotopes of CO2, CH4 and DIC, together with mol% CO2 show that 4 out of 17 wells sampled in the 2800’ sandstone are still producing injected CO2. The dominant fate of the injected CO2appears to be dissolution in formation fluids and gas-phase trapping. There is some isotopic and geochemical evidence for enhanced microbial methanogenesis in 2 samples; however, the CO2 spread unevenly throughout the reservoir, and thus cannot explain the elevated indicators for methanogenesis observed across the entire field. Vertical migration out of the target 2800’ sandstone reservoir is also apparent in 3 samples located stratigraphically above the target sand. Reservoirs comparable to the 2800’ sandstone, located along a 90-km transect, were also sampled to investigate regional trends in gas composition, brine chemistry and microbial activity. Microbial methane, likely sourced from biodegradation of organic substrates within the formation, was found in all oil fields sampled, while indicators of methanogenesis (e.g. high alkalinity, δ13C-CO2 and δ13C-DIC values) and oxidation of propane were greatest in the Olla Field, likely due to its more ideal environmental conditions (i.e. suitable range of pH, temperature, salinity, sulfate and iron concentrations).

Co-overexpressing a Plasma Membrane and a Vacuolar Membrane Sodium/Proton Antiporter Significantly Improves Salt Tolerance in Transgenic Arabidopsis Plants.

Science.gov (United States)

Pehlivan, Necla; Sun, Li; Jarrett, Philip; Yang, Xiaojie; Mishra, Neelam; Chen, Lin; Kadioglu, Asim; Shen, Guoxin; Zhang, Hong

2016-05-01

The Arabidopsis gene AtNHX1 encodes a vacuolar membrane-bound sodium/proton (Na(+)/H(+)) antiporter that transports Na(+) into the vacuole and exports H(+) into the cytoplasm. The Arabidopsis gene SOS1 encodes a plasma membrane-bound Na(+)/H(+) antiporter that exports Na(+) to the extracellular space and imports H(+) into the plant cell. Plants rely on these enzymes either to keep Na(+) out of the cell or to sequester Na(+) into vacuoles to avoid the toxic level of Na(+) in the cytoplasm. Overexpression of AtNHX1 or SOS1 could improve salt tolerance in transgenic plants, but the improved salt tolerance is limited. NaCl at concentration >200 mM would kill AtNHX1-overexpressing or SOS1-overexpressing plants. Here it is shown that co-overexpressing AtNHX1 and SOS1 could further improve salt tolerance in transgenic Arabidopsis plants, making transgenic Arabidopsis able to tolerate up to 250 mM NaCl treatment. Furthermore, co-overexpression of AtNHX1 and SOS1 could significantly reduce yield loss caused by the combined stresses of heat and salt, confirming the hypothesis that stacked overexpression of two genes could substantially improve tolerance against multiple stresses. This research serves as a proof of concept for improving salt tolerance in other plants including crops. © The Author 2016. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists.

Recovery of avian metapneumovirus subgroup C from cDNA: cross-recognition of avian and human metapneumovirus support proteins.

Science.gov (United States)

Govindarajan, Dhanasekaran; Buchholz, Ursula J; Samal, Siba K

2006-06-01

Avian metapneumovirus (AMPV) causes an acute respiratory disease in turkeys and is associated with "swollen head syndrome" in chickens, contributing to significant economic losses for the U.S. poultry industry. With a long-term goal of developing a better vaccine for controlling AMPV in the United States, we established a reverse genetics system to produce infectious AMPV of subgroup C entirely from cDNA. A cDNA clone encoding the entire 14,150-nucleotide genome of AMPV subgroup C strain Colorado (AMPV/CO) was generated by assembling five cDNA fragments between the T7 RNA polymerase promoter and the autocatalytic hepatitis delta virus ribozyme of a transcription plasmid, pBR 322. Transfection of this plasmid, along with the expression plasmids encoding the N, P, M2-1, and L proteins of AMPV/CO, into cells stably expressing T7 RNA polymerase resulted in the recovery of infectious AMPV/CO. Characterization of the recombinant AMPV/CO showed that its growth properties in tissue culture were similar to those of the parental virus. The potential of AMPV/CO to serve as a viral vector was also assessed by generating another recombinant virus, rAMPV/CO-GFP, that expressed the enhanced green fluorescent protein (GFP) as a foreign protein. Interestingly, GFP-expressing AMPV and GFP-expressing human metapneumovirus (HMPV) could be recovered using the support plasmids of either virus, denoting that the genome promoters are conserved between the two metapneumoviruses and can be cross-recognized by the polymerase complex proteins of either virus. These results indicate a close functional relationship between AMPV/CO and HMPV.

Identification and suppression of the p-coumaroyl CoA:hydroxycinnamyl alcohol transferase in Zea mays L.

Science.gov (United States)

Marita, Jane M; Hatfield, Ronald D; Rancour, David M; Frost, Kenneth E

2014-06-01

Grasses, such as Zea mays L. (maize), contain relatively high levels of p-coumarates (pCA) within their cell walls. Incorporation of pCA into cell walls is believed to be due to a hydroxycinnamyl transferase that couples pCA to monolignols. To understand the role of pCA in maize development, the p-coumaroyl CoA:hydroxycinnamyl alcohol transferase (pCAT) was isolated and purified from maize stems. Purified pCAT was subjected to partial trypsin digestion, and peptides were sequenced by tandem mass spectrometry. TBLASTN analysis of the acquired peptide sequences identified a single full-length maize cDNA clone encoding all the peptide sequences obtained from the purified enzyme. The cDNA clone was obtained and used to generate an RNAi construct for suppressing pCAT expression in maize. Here we describe the effects of suppression of pCAT in maize. Primary screening of transgenic maize seedling leaves using a new rapid analytical platform was used to identify plants with decreased amounts of pCA. Using this screening method, mature leaves from fully developed plants were analyzed, confirming reduced pCA levels throughout plant development. Complete analysis of isolated cell walls from mature transgenic stems and leaves revealed that lignin levels did not change, but pCA levels decreased and the lignin composition was altered. Transgenic plants with the lowest levels of pCA had decreased levels of syringyl units in the lignin. Thus, altering the levels of pCAT expression in maize leads to altered lignin composition, but does not appear to alter the total amount of lignin present in the cell walls. © 2014 The Authors The Plant Journal © 2014 John Wiley & Sons Ltd.

Effect of oxygen co-injected with carbon dioxide on Gothic shale caprock–CO₂–brine interaction during geologic carbon sequestration

Energy Technology Data Exchange (ETDEWEB)

Jung, Hun Bok; Um, Wooyong; Cantrell, Kirk J.

2013-09-01

Co-injection of oxygen, a significant component in CO₂ streams produced by the oxyfuel combustion process, can cause a significant alteration of the redox state in deep geologic formations during geologic carbon sequestration. The potential impact of co-injected oxygen on the interaction between synthetic CO₂–brine (0.1 M NaCl) and shale caprock (Gothic shale from the Aneth Unit in Utah) and mobilization of trace metals was investigated at ~ 10 MPa and ~ 75 °C. A range of relative volume percentages of O₂ to CO₂ (0, 1, 4 and 8%) were used in these experiments to address the effect of oxygen on shale–CO₂–brine interaction under various conditions. Major mineral phases in Gothic shale are quartz, calcite, dolomite, montmorillonite, and pyrite. During Gothic shale–CO₂–brine interaction in the presence of oxygen, pyrite oxidation occurred extensively and caused enhanced dissolution of calcite and dolomite. Pyrite oxidation and calcite dissolution subsequently resulted in the precipitation of Fe(III) oxides and gypsum (CaSO₄·2H₂O). In the presence of oxygen, dissolved Mn and Ni were elevated because of oxidative dissolution of pyrite. The mobility of dissolved Ba was controlled by barite (BaSO₄) precipitation in the presence of oxygen. Dissolved U in the experimental brines increased to ~ 8–14 μg/L, with concentrations being slightly higher in the absence of oxygen than in the presence of oxygen. Experimental and modeling results indicate the interaction between shale caprock and oxygen co-injected with CO₂ during geologic carbon sequestration can exert significant impacts on brine pH, solubility of carbonate minerals, stability of sulfide minerals, and mobility of trace metals. The major impact of oxygen is most likely to occur in the zone near CO₂ injection wells where impurity gases can accumulate. Finally, oxygen in CO₂

Overexpression of CsANR increased flavan-3-ols and decreased anthocyanins in transgenic tobacco.

Science.gov (United States)

Kumar, Vinay; Yadav, Sudesh Kumar

2013-06-01

Anthocyanins and flavan-3-ols are distributed widely in plants and synthesized by a common biosynthetic pathway. Anthocyanin reductase (ANR) represents branching-point enzyme of this pathway converting anthocyanidins to flavan-3-ols. Since tea contains highest amount of flavonoids, a cDNA encoding anthocyanin reductase from tea (CsANR) was overexpressed in transgenic tobacco to check the influence on anthocyanin and flavan-3-ols. The transgenic tobacco was confirmed by genomic PCR and expression of transgene was analyzed through semiquantitative PCR. Interestingly flowers of transgenic tobacco were light pink/white in color instead of dark pink in wild tobacco, documenting the decrease in anthocyanins content. Upon measurement, flower anthocyanin content was found to be lesser. While flavan-3-ols (epicatechin and epigallocatechin) contents were increased in leaf tissue of transgenic lines. The expressions of other endogenous flavonoid biosynthetic pathway genes in different floral parts (sepal, petal, stamen, and carpel) of CsANR overexpressing tobacco as well as wild tobacco were analyzed. The transcript levels of PAL and CHI genes were downregulated, while transcript levels of F3H, FLS, CHS, ANR1, and ANR2 genes were upregulated in all floral parts of CsANR transgenic plants compared to wild tobacco. The expressions of DFR and ANS genes were also spatially modulated in different floral parts due to overexpression of CsANR. Thus, CsANR overexpression increased flavan-3-ols and decreased anthocyanin content by modulating the expressions of various flavonoid biosynthetic pathway genes in flower of tobacco. These changes might be responsible for the observed pollen tube in the pollens of CsANR overexpressing transgenic tobacco when they were still in the anther before pollination.

Enhanced phytoremediation of mixed heavy metal (mercury)-organic pollutants (trichloroethylene) with transgenic alfalfa co-expressing glutathione S-transferase and human P450 2E1.

Science.gov (United States)

Zhang, Yuanyuan; Liu, Junhong; Zhou, Yuanming; Gong, Tingyun; Wang, Jing; Ge, Yinlin

2013-09-15

Soil contamination is a global environmental problem and many efforts have been made to find efficient remediation methods over the last decade. Moreover, remediation of mixed contaminated soils are more difficult. In the present study, transgenic alfalfa plants pKHCG co-expressing glutathione S-transferase (GST) and human P450 2E1 (CYP2E1) genes were used for phytoremediation of mixed mercury (Hg)-trichloroethylene (TCE) contaminants. Simultaneous expression of GST and CYP2E1 may produce a significant synergistic effect, and leads to improved resistance and accumulation to heavy metal-organic complex contaminants. Based on the tolerance and accumulation assays, pKHCG transgenic plants were more resistant to Hg/TCE complex pollutants and many folds higher in Hg/TCE-accumulation than the non-transgenic control plants in mixed contaminated soil. It is confirmed that GST and CYP2E1 co-expression may be a useful strategy to help achieve mixed heavy metal-organic pollutants phytoremediation. Copyright © 2013 Elsevier B.V. All rights reserved.

Transgenic animals and their application in medicine

OpenAIRE

Bagle TR, Kunkulol RR, Baig MS, More SY

2013-01-01

Transgenic animals are animals that are genetically altered to have traits that mimic symptoms of specific human pathologies. They provide genetic models of various human diseases which are important in understanding disease and developing new targets. In early 1980 Gordon and co-workers described the first gene addition experiment using the microinjection technology and since then the impact of transgenic technology on basic research has been significant. Within 20 years of its inception, AT...

Assessment of stress tolerance, productivity, and forage quality in T1 transgenic alfalfa co-overexpressing ZxNHX and ZxVP1-1 from Zygophyllum xanthoxylum

Directory of Open Access Journals (Sweden)

Peng Kang

2016-10-01

Full Text Available Salinization, desertification, and soil nutrient deprivation are threatening the production of alfalfa (Medicago sativa L. in northern China. We have previously generated T0 transgenic alfalfa co-overexpressing Zygophyllum xanthoxylum ZxNHX and ZxVP1-1 genes with enhanced salt and drought tolerance. To further develop this excellent breeding material into the new forage cultivar, stress tolerance, productivity, and forage quality of T1 transgenic alfalfa (GM were assessed in this study. The GM inherited the traits of salt and drought tolerance from T0 generation. Most importantly, co-overexpression of ZxNHX and ZxVP1-1 enhanced the tolerance to Pi deficiency in GM, which was associated with more Pi accumulation in plants. Meanwhile, T1 transgenic alfalfa developed a larger root system with increased root size, root dry weight and root/shoot ratio, which may be one important reason for the improvement of phosphorus nutrition and high biomass accumulation in GM under various conditions. GM also accumulated more crude protein, crude fibre, crude fat, and crude ash than wild-type (WT plants, especially under stress conditions and in the field. More interestingly, the crude fat contents sharply dropped in WT (by 66%-74%, whereas showed no change or decreased less in GM, when subjected to salinity, drought or low-Pi. Our results indicate that T1 transgenic alfalfa co-overexpressing ZxNHX and ZxVP1-1 shows stronger stress tolerance, higher productivity and better forage quality. This study provides a solid foundation for creating the alfalfa cultivars with high yield, good quality and wide adaptability on saline, dry and nutrient-deprived marginal lands of northern China.

Transgenic alfalfa plants co-expressing glutathione S-transferase (GST) and human CYP2E1 show enhanced resistance to mixed contaminates of heavy metals and organic pollutants

International Nuclear Information System (INIS)

Zhang, Yuanyuan; Liu, Junhong

2011-01-01

Transgenic alfalfa plants simultaneously expressing human CYP2E1 and glutathione S-transferase (GST) were generated from hypocotyl segments by the use of an Agrobacterium transformation system for the phytoremediation of the mixed contaminated soil with heavy metals and organic pollutants. The transgenic alfalfa plants were screened by a combination of kanamycin resistance, PCR, GST and CYP2E1 activity and Western blot analysis. The capabilities of mixed contaminants (heavy metals-organic compounds) resistance of pKHCG transgenic alfalfa plants became markedly increased compared with the transgenic alfalfa plants expressing single gene (GST or CYP2E1) and the non-transgenic control plants. The pKHCG alfalfa plants exhibited strong resistance towards the mixtures of cadmium (Cd) and trichloroethylene (TCE) that were metabolized by the introduced GST and CYP2E1 in combination. Our results show that the pKHCG transgenic alfalfa plants have good potential for phytoremediation because they have cross-tolerance towards the complex contaminants of heavy metals and organic pollutants. Therefore, these transgenic alfalfa plants co-expressing GST and human P450 CDNAs may have a great potential for phytoremediation of mixed environmental contaminants.

Transgenic alfalfa plants co-expressing glutathione S-transferase (GST) and human CYP2E1 show enhanced resistance to mixed contaminates of heavy metals and organic pollutants

Energy Technology Data Exchange (ETDEWEB)

Zhang, Yuanyuan [Department of Pharmaceutics, Qingdao University of Science and Technology, 53 Zhengzhou Road, P.O. Box 70, Qingdao 266042 (China); Liu, Junhong, E-mail: liujh@qust.edu.cn [Department of Pharmaceutics, Qingdao University of Science and Technology, 53 Zhengzhou Road, P.O. Box 70, Qingdao 266042 (China)

2011-05-15

Transgenic alfalfa plants simultaneously expressing human CYP2E1 and glutathione S-transferase (GST) were generated from hypocotyl segments by the use of an Agrobacterium transformation system for the phytoremediation of the mixed contaminated soil with heavy metals and organic pollutants. The transgenic alfalfa plants were screened by a combination of kanamycin resistance, PCR, GST and CYP2E1 activity and Western blot analysis. The capabilities of mixed contaminants (heavy metals-organic compounds) resistance of pKHCG transgenic alfalfa plants became markedly increased compared with the transgenic alfalfa plants expressing single gene (GST or CYP2E1) and the non-transgenic control plants. The pKHCG alfalfa plants exhibited strong resistance towards the mixtures of cadmium (Cd) and trichloroethylene (TCE) that were metabolized by the introduced GST and CYP2E1 in combination. Our results show that the pKHCG transgenic alfalfa plants have good potential for phytoremediation because they have cross-tolerance towards the complex contaminants of heavy metals and organic pollutants. Therefore, these transgenic alfalfa plants co-expressing GST and human P450 CDNAs may have a great potential for phytoremediation of mixed environmental contaminants.

D2O laser pumped by an injection-locked CO2 laser for ion-temperature measurements

International Nuclear Information System (INIS)

Okada, Tatsuo; Ohga, Tetsuaki; Yokoo, Masakazu; Muraoka, Katsunori; Akazaki, Masanori.

1986-01-01

The cooperative Thomson scattering method is one of the various new techniques proposed for measuring the temperature of ions in nuclear fusion critical plasma, for which a high-performance FIR laser pumped by an injection-locked CO 2 laser is required. This report deals with D 2 O laser with a wavelength of 385 μm which is pumped by injection-locked single-mole TEA CO 2 laser composed of a driver laser and an output-stage laser. A small-sized automatic pre-ionization type laser is employed for the driver. The resonator of the driver laser consists of a plane grating of littrow arrangement and ZnSe plane output mirrors with reflection factor of 50 %. An aperture and ZnSe etalon are inserted in the resonator to produce single transverse- and longitudinal-mode oscillation, respectively. The output-stage laser is also of the automatic pre-ionization type. Theoretically, an injection power of 0.1 pW/mm 3 is required for a CO 2 laser. Single-mode oscillation of several hundred nW/mm 3 can be produced by the CO 2 laser used in this study. Tuning of the output-stage laser is easily controlled by the driver laser. High stability of the injection-locked operation is demonstrated. CO 2 laser beam is introduced into the D 2 O laser through a KCl window to excite D 2 O laser beam in the axial direction. Input and output characteristics of the D 2 O laser are shown. Also presented are typical pulse shapes from the D 2 O laser pumped by a free-running CO 2 laser pulse or by an injection-locked single-mode CO 2 laser pulse. (Nogami, K.)

Microaspiration of esophageal gland cells and cDNA library construction for identifying parasitism genes of plant-parasitic nematodes.

Science.gov (United States)

Hussey, Richard S; Huang, Guozhong; Allen, Rex

2011-01-01

Identifying parasitism genes encoding proteins secreted from a plant-parasitic nematode's esophageal gland cells and injected through its stylet into plant tissue is the key to understanding the molecular basis of nematode parasitism of plants. Parasitism genes have been cloned by directly microaspirating the cytoplasm from the esophageal gland cells of different parasitic stages of cyst or root-knot nematodes to provide mRNA to create a gland cell-specific cDNA library by long-distance reverse-transcriptase polymerase chain reaction. cDNA clones are sequenced and deduced protein sequences with a signal peptide for secretion are identified for high-throughput in situ hybridization to confirm gland-specific expression.

Assessment of Factors Influencing Effective CO₂ Storage Capacity and Injectivity in Eastern Gas Shales

Energy Technology Data Exchange (ETDEWEB)

Godec, Michael [Advanced Resources International, Inc., Arlington, VA (United States)

2013-06-30

Building upon advances in technology, production of natural gas from organic-rich shales is rapidly developing as a major hydrocarbon supply option in North America and around the world. The same technology advances that have facilitated this revolution - dense well spacing, horizontal drilling, and hydraulic fracturing - may help to facilitate enhanced gas recovery (EGR) and carbon dioxide (CO₂) storage in these formations. The potential storage of CO₂ in shales is attracting increasing interest, especially in Appalachian Basin states that have extensive shale deposits, but limited CO₂ storage capacity in conventional reservoirs. The goal of this cooperative research project was to build upon previous and on-going work to assess key factors that could influence effective EGR, CO₂ storage capacity, and injectivity in selected Eastern gas shales, including the Devonian Marcellus Shale, the Devonian Ohio Shale, the Ordovician Utica and Point Pleasant shale and equivalent formations, and the late Devonian-age Antrim Shale. The project had the following objectives: (1) Analyze and synthesize geologic information and reservoir data through collaboration with selected State geological surveys, universities, and oil and gas operators; (2) improve reservoir models to perform reservoir simulations to better understand the shale characteristics that impact EGR, storage capacity and CO₂ injectivity in the targeted shales; (3) Analyze results of a targeted, highly monitored, small-scale CO₂ injection test and incorporate into ongoing characterization and simulation work; (4) Test and model a smart particle early warning concept that can potentially be used to inject water with uniquely labeled particles before the start of CO₂ injection; (5) Identify and evaluate potential constraints to economic CO₂ storage in gas shales, and propose development approaches that overcome these constraints

Optical modulation of transgene expression in retinal pigment epithelium

Science.gov (United States)

Palanker, D.; Lavinsky, D.; Chalberg, T.; Mandel, Y.; Huie, P.; Dalal, R.; Marmor, M.

2013-03-01

Over a million people in US alone are visually impaired due to the neovascular form of age-related macular degeneration (AMD). The current treatment is monthly intravitreal injections of a protein which inhibits Vascular Endothelial Growth Factor, thereby slowing progression of the disease. The immense financial and logistical burden of millions of intravitreal injections signifies an urgent need to develop more long-lasting and cost-effective treatments for this and other retinal diseases. Viral transfection of ocular cells allows creation of a "biofactory" that secretes therapeutic proteins. This technique has been proven successful in non-human primates, and is now being evaluated in clinical trials for wet AMD. However, there is a critical need to down-regulate gene expression in the case of total resolution of retinal condition, or if patient has adverse reaction to the trans-gene products. The site for genetic therapy of AMD and many other retinal diseases is the retinal pigment epithelium (RPE). We developed and tested in pigmented rabbits, an optical method to down-regulate transgene expression in RPE following vector delivery, without retinal damage. Microsecond exposures produced by a rapidly scanning laser vaporize melanosomes and destroy a predetermined fraction of the RPE cells selectively. RPE continuity is restored within days by migration and proliferation of adjacent RPE, but since the transgene is not integrated into the nucleus it is not replicated. Thus, the decrease in transgene expression can be precisely determined by the laser pattern density and further reduced by repeated treatment without affecting retinal structure and function.

Simulation of CO2 Sequestration at Rock Spring Uplift, Wyoming: Heterogeneity and Uncertainties in Storage Capacity, Injectivity and Leakage

Energy Technology Data Exchange (ETDEWEB)

Deng, Hailin [Los Alamos National Laboratory; Dai, Zhenxue [Los Alamos National Laboratory; Jiao, Zunsheng [Wyoming State Geological Survey; Stauffer, Philip H. [Los Alamos National Laboratory; Surdam, Ronald C. [Wyoming State Geological Survey

2011-01-01

Many geological, geochemical, geomechanical and hydrogeological factors control CO{sub 2} storage in subsurface. Among them heterogeneity in saline aquifer can seriously influence design of injection wells, CO{sub 2} injection rate, CO{sub 2} plume migration, storage capacity, and potential leakage and risk assessment. This study applies indicator geostatistics, transition probability and Markov chain model at the Rock Springs Uplift, Wyoming generating facies-based heterogeneous fields for porosity and permeability in target saline aquifer (Pennsylvanian Weber sandstone) and surrounding rocks (Phosphoria, Madison and cap-rock Chugwater). A multiphase flow simulator FEHM is then used to model injection of CO{sub 2} into the target saline aquifer involving field-scale heterogeneity. The results reveal that (1) CO{sub 2} injection rates in different injection wells significantly change with local permeability distributions; (2) brine production rates in different pumping wells are also significantly impacted by the spatial heterogeneity in permeability; (3) liquid pressure evolution during and after CO{sub 2} injection in saline aquifer varies greatly for different realizations of random permeability fields, and this has potential important effects on hydraulic fracturing of the reservoir rock, reactivation of pre-existing faults and the integrity of the cap-rock; (4) CO{sub 2} storage capacity estimate for Rock Springs Uplift is 6614 {+-} 256 Mt at 95% confidence interval, which is about 36% of previous estimate based on homogeneous and isotropic storage formation; (5) density profiles show that the density of injected CO{sub 2} below 3 km is close to that of the ambient brine with given geothermal gradient and brine concentration, which indicates CO{sub 2} plume can sink to the deep before reaching thermal equilibrium with brine. Finally, we present uncertainty analysis of CO{sub 2} leakage into overlying formations due to heterogeneity in both the target saline

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

International Nuclear Information System (INIS)

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

2013-01-01

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

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

Science.gov (United States)

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

2014-01-01

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

Molecular cloning of lupin leghemoglobin cDNA

DEFF Research Database (Denmark)

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

1987-01-01

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

Quantification of a maximum injection volume of CO2 to avert geomechanical perturbations using a compositional fluid flow reservoir simulator

Science.gov (United States)

Jung, Hojung; Singh, Gurpreet; Espinoza, D. Nicolas; Wheeler, Mary F.

2018-02-01

Subsurface CO2 injection and storage alters formation pressure. Changes of pore pressure may result in fault reactivation and hydraulic fracturing if the pressure exceeds the corresponding thresholds. Most simulation models predict such thresholds utilizing relatively homogeneous reservoir rock models and do not account for CO2 dissolution in the brine phase to calculate pore pressure evolution. This study presents an estimation of reservoir capacity in terms of allowable injection volume and rate utilizing the Frio CO2 injection site in the coast of the Gulf of Mexico as a case study. The work includes laboratory core testing, well-logging data analyses, and reservoir numerical simulation. We built a fine-scale reservoir model of the Frio pilot test in our in-house reservoir simulator IPARS (Integrated Parallel Accurate Reservoir Simulator). We first performed history matching of the pressure transient data of the Frio pilot test, and then used this history-matched reservoir model to investigate the effect of the CO2 dissolution into brine and predict the implications of larger CO2 injection volumes. Our simulation results -including CO2 dissolution- exhibited 33% lower pressure build-up relative to the simulation excluding dissolution. Capillary heterogeneity helps spread the CO2 plume and facilitate early breakthrough. Formation expansivity helps alleviate pore pressure build-up. Simulation results suggest that the injection schedule adopted during the actual pilot test very likely did not affect the mechanical integrity of the storage complex. Fault reactivation requires injection volumes of at least about sixty times larger than the actual injected volume at the same injection rate. Hydraulic fracturing necessitates much larger injection rates than the ones used in the Frio pilot test. Tested rock samples exhibit ductile deformation at in-situ effective stresses. Hence, we do not expect an increase of fault permeability in the Frio sand even in the presence of

Development of transgenic rats producing human β-amyloid precursor protein as a model for Alzheimer's disease: Transgene and endogenous APP genes are regulated tissue-specifically

Directory of Open Access Journals (Sweden)

Chan Anthony WS

2008-02-01

Full Text Available Abstract Background Alzheimer's disease (AD is a devastating neurodegenerative disorder that affects a large and growing number of elderly individuals. In addition to idiopathic disease, AD is also associated with autosomal dominant inheritance, which causes a familial form of AD (FAD. Some instances of FAD have been linked to mutations in the β-amyloid protein precursor (APP. Although there are numerous mouse AD models available, few rat AD models, which have several advantages over mice, have been generated. Results Fischer 344 rats expressing human APP driven by the ubiquitin-C promoter were generated via lentiviral vector infection of Fischer 344 zygotes. We generated two separate APP-transgenic rat lines, APP21 and APP31. Serum levels of human amyloid-beta (Aβ40 were 298 pg/ml for hemizygous and 486 pg/ml for homozygous APP21 animals. Serum Aβ42 levels in APP21 homozygous rats were 135 pg/ml. Immunohistochemistry in brain showed that the human APP transgene was expressed in neurons, but not in glial cells. These findings were consistent with independent examination of enhanced green fluorescent protein (eGFP in the brains of eGFP-transgenic rats. APP21 and APP31 rats expressed 7.5- and 3-times more APP mRNA, respectively, than did wild-type rats. Northern blots showed that the human APP transgene, driven by the ubiquitin-C promoter, is expressed significantly more in brain, kidney and lung compared to heart and liver. A similar expression pattern was also seen for the endogenous rat APP. The unexpected similarity in the tissue-specific expression patterns of endogenous rat APP and transgenic human APP mRNAs suggests regulatory elements within the cDNA sequence of APP. Conclusion This manuscript describes the generation of APP-transgenic inbred Fischer 344 rats. These are the first human AD model rat lines generated by lentiviral infection. The APP21 rat line expresses high levels of human APP and could be a useful model for AD. Tissue

Virally and physically transgenized equine adipose-derived stromal cells as a cargo for paracrine secreted factors

Directory of Open Access Journals (Sweden)

Cavirani Sandro

2010-09-01

Full Text Available Abstract Background Adipose-Derived Stromal Cells have been shown to have multiple lineage differentiation properties and to be suitable for tissues regeneration in many degenerative processes. Their use has been proposed for the therapy of joint diseases and tendon injuries in the horse. In the present report the genetic manipulation of Equine Adipose-Derived Stromal Cells has been investigated. Results Equine Adipose-Derived Stromal Cells were successfully virally transduced as well as transiently and stably transfected with appropriate parameters, without detrimental effect on their differentiation properties. Moreover, green fluorescent protein alone, fused to neo gene, or co-expressed as bi-cistronic reporter constructs, driven by viral and house-keeping gene promoters, were tested. The better expressed cassette was employed to stably transfect Adipose-Derived Stromal Cells for cell therapy purposes. Stably transfected Equine Adipose-Derived Stromal Cells with a heterologous secreted viral antigen were able to immunize horses upon injection into the lateral wall of the neck. Conclusion This study provides the methods to successfully transgenize Adipose-Derived Stromal Cells both by lentiviral vector and by transfection using optimized constructs with suitable promoters and reporter genes. In conclusion these findings provide a working platform for the delivery of potentially therapeutic proteins to the site of cells injection via transgenized Equine Adipose-Derived Stromal Cells.

Mineral storage of CO2/H2S gas mixture injection in basaltic rocks

Science.gov (United States)

Clark, D. E.; Gunnarsson, I.; Aradottir, E. S.; Oelkers, E. H.; Sigfússon, B.; Snæbjörnsdottír, S. Ó.; Matter, J. M.; Stute, M.; Júlíusson, B. M.; Gíslason, S. R.

2017-12-01

Carbon capture and storage is one solution to reducing CO2 emissions in the atmosphere. The long-term geological storage of buoyant supercritical CO2 requires high integrity cap rock. Some of the risk associated with CO2 buoyancy can be overcome by dissolving CO2 into water during its injection, thus eliminating its buoyancy. This enables injection into fractured rocks, such as basaltic rocks along oceanic ridges and on continents. Basaltic rocks are rich in divalent cations, Ca2+, Mg2+ and Fe2+, which react with CO2 dissolved in water to form stable carbonate minerals. This possibility has been successfully tested as a part of the CarbFix CO2storage pilot project at the Hellisheiði geothermal power plant in Iceland, where they have shown mineralization occurs in less than two years [1, 2]. Reykjavik Energy and the CarbFix group has been injecting a mixture of CO2 and H2S at 750 m depth and 240-250°C since June 2014; by 1 January 2016, 6290 tons of CO2 and 3530 tons of H2S had been injected. Once in the geothermal reservoir, the heat exchange and sufficient dissolution of the host rock neutralizes the gas-charged water and saturates the formation water respecting carbonate and sulfur minerals. A thermally stable inert tracer was also mixed into the stream to monitor the subsurface transport and to assess the degree of subsurface carbonation and sulfide precipitation [3]. Water and gas samples have been continuously collected from three monitoring wells and geochemically analyzed. Based on the results, mineral saturation stages have been defined. These results and tracer mass balance calculations are used to evaluate the rate and magnitude of CO2 and H2S mineralization in the subsurface, with indications that mineralization of carbon and sulfur occurs within months. [1] Gunnsarsson, I., et al. (2017). Rapid and cost-effective capture and subsurface mineral storage of carbon and sulfur. Manuscript submitted for publication. [2] Matter, J., et al. (2016). Rapid

Overexpression of an alfalfa GDP-mannose 3, 5-epimerase gene enhances acid, drought and salt tolerance in transgenic Arabidopsis by increasing ascorbate accumulation.

Science.gov (United States)

Ma, Lichao; Wang, Yanrong; Liu, Wenxian; Liu, Zhipeng

2014-11-01

GDP-mannose 3', 5'-epimerase (GME) catalyses the conversion of GDP-D-mannose to GDP-L-galactose, an important step in the ascorbic acid (ascorbic acid) biosynthetic pathway in higher plants. In this study, a novel cDNA fragment (MsGME) encoding a GME protein was isolated and characterised from alfalfa (Medicago sativa). An expression analysis confirmed that MsGME expression was induced by salinity, PEG and acidity stresses. MsGME overexpression in Arabidopsis enhanced tolerance of the transgenic plants to salt, drought and acid. Real-time PCR analysis revealed that the transcript levels of GDP-D-mannose pyrophosphorylase (GMP), L-galactose-phosphate 1-P phosphatase (GP) and GDP-L-galactose phosphorylase (GGP) were increased in transgenic Arabidopsis (T3 generation). Moreover, the ascorbate content was increased in transgenic Arabidopsis. Our results suggest that MsGME can effectively enhance tolerance of transgenic Arabidopsis to acid, drought and salt by increasing ascorbate accumulation.

Cloning of the human androgen receptor cDNA

International Nuclear Information System (INIS)

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

1988-01-01

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

Co-transforming bar and CsALDH Genes Enhanced Resistance to Herbicide and Drought and Salt Stress in Transgenic Alfalfa (Medicago sativa L.)

Science.gov (United States)

Duan, Zhen; Zhang, Daiyu; Zhang, Jianquan; Di, Hongyan; Wu, Fan; Hu, Xiaowen; Meng, Xuanchen; Luo, Kai; Zhang, Jiyu; Wang, Yanrong

2015-01-01

Drought and high salinity are two major abiotic factors that restrict the productivity of alfalfa. By application of the Agrobacterium-mediated transformation method, an oxidative responsive gene, CsALDH12A1, from the desert grass Cleistogenes songorica together with the bar gene associated with herbicide resistance, were co-transformed into alfalfa (Medicago sativa L.). From the all 90 transformants, 16 were positive as screened by spraying 1 mL L-1 10% Basta solution and molecularly diagnosis using PCR. Real-time PCR analysis indicated that drought and salt stress induced high CsALDH expression in the leaves of the transgenic plants. The CsALDH expression levels under drought (15 d) and salt stress (200 mM NaCl) were 6.11 and 6.87 times higher than in the control plants, respectively. In comparison to the WT plants, no abnormal phenotypes were observed among the transgenic plants, which showed significant enhancement of tolerance to 15 d of drought and 10 d of salinity treatment. Evaluation of the physiological and biochemical indices during drought and salt stress of the transgenic plants revealed relatively lower Na+ content and higher K+ content in the leaves relative to the WT plants, a reduction of toxic on effects and maintenance of osmotic adjustment. In addition, the transgenic plants could maintain a higher relative water content level, higher shoot biomass, fewer changes in the photosystem, decreased membrane injury, and a lower level of osmotic stress. These results indicate that the co-expression of the introduced bar and CsALDH genes enhanced the herbicide, drought and salt tolerance of alfalfa and therefore can potentially be used as a novel genetic resource for the future breeding programs to develop new cultivars. PMID:26734025

Stable isotope reactive transport modeling in water-rock interactions during CO2 injection

Science.gov (United States)

Hidalgo, Juan J.; Lagneau, Vincent; Agrinier, Pierre

2010-05-01

Stable isotopes can be of great usefulness in the characterization and monitoring of CO2 sequestration sites. Stable isotopes can be used to track the migration of the CO2 plume and identify leakage sources. Moreover, they provide unique information about the chemical reactions that take place on the CO2-water-rock system. However, there is a lack of appropriate tools that help modelers to incorporate stable isotope information into the flow and transport models used in CO2 sequestration problems. In this work, we present a numerical tool for modeling the transport of stable isotopes in groundwater reactive systems. The code is an extension of the groundwater single-phase flow and reactive transport code HYTEC [2]. HYTEC's transport module was modified to include element isotopes as separate species. This way, it is able to track isotope composition of the system by computing the mixing between the background water and the injected solution accounting for the dependency of diffusion on the isotope mass. The chemical module and database have been expanded to included isotopic exchange with minerals and the isotope fractionation associated with chemical reactions and mineral dissolution or precipitation. The performance of the code is illustrated through a series of column synthetic models. The code is also used to model the aqueous phase CO2 injection test carried out at the Lamont-Doherty Earth Observatory site (Palisades, New York, USA) [1]. References [1] N. Assayag, J. Matter, M. Ader, D. Goldberg, and P. Agrinier. Water-rock interactions during a CO2 injection field-test: Implications on host rock dissolution and alteration effects. Chemical Geology, 265(1-2):227-235, July 2009. [2] Jan van der Lee, Laurent De Windt, Vincent Lagneau, and Patrick Goblet. Module-oriented modeling of reactive transport with HYTEC. Computers & Geosciences, 29(3):265-275, April 2003.

Co2 injection into oil reservoir associated with structural deformation

KAUST Repository

El-Amin, Mohamed

2012-01-01

In this work, the problem of structural deformation with two-phase flow of carbon sequestration is presented. A model to simulate miscible CO2 injection with structural deformation in the aqueous phase is established. In the first part of this paper, we developed analytical solution for the problem under consideration with certain types of boundary conditions, namely, Dirichlet and Neumann boundary conditions. The second part concerns to numerical simulation using IMPDES scheme. A simulator based on cell-centered finite difference method is used to solve this equations system. Distributions of CO2 saturation, and horizontal and vertical displacements have been introduced.

Impact of CO2 injection protocol on fluid-solid reactivity: high-pressure and temperature microfluidic experiments in limestone

Science.gov (United States)

Jimenez-Martinez, Joaquin; Porter, Mark; Carey, James; Guthrie, George; Viswanathan, Hari

2017-04-01

Geological sequestration of CO2 has been proposed in the last decades as a technology to reduce greenhouse gas emissions to the atmosphere and mitigate the global climate change. However, some questions such as the impact of the protocol of CO2 injection on the fluid-solid reactivity remain open. In our experiments, two different protocols of injection are compared at the same conditions (8.4 MPa and 45 C, and constant flow rate 0.06 ml/min): i) single phase injection, i.e., CO2-saturated brine; and ii) simultaneous injection of CO2-saturated brine and scCO2. For that purpose, we combine a unique high-pressure/temperature microfluidics experimental system, which allows reproducing geological reservoir conditions in geo-material substrates (i.e., limestone, Cisco Formation, Texas, US) and high resolution optical profilometry. Single and multiphase flow through etched fracture networks were optically recorded with a microscope, while processes of dissolution-precipitation in the etched channels were quantified by comparison of the initial and final topology of the limestone micromodels. Changes in hydraulic conductivity were quantified from pressure difference along the micromodel. The simultaneous injection of CO2-saturated brine and scCO2, reduced the brine-limestone contact area and also created a highly heterogeneous velocity field (i.e., low velocities regions or stagnation zones, and high velocity regions or preferential paths), reducing rock dissolution and enhancing calcite precipitation. The results illustrate the contrasting effects of single and multiphase flow on chemical reactivity and suggest that multiphase flow by isolating parts of the flow system can enhance CO2 mineralization.

Testing and modelling of a novel oil-free co-rotating scroll machine with water injection

International Nuclear Information System (INIS)

Mendoza, Luis Carlos; Lemofouet, Sylvain; Schiffmann, Jürg

2017-01-01

Highlights: • Performance of novel oil-free co-rotating scroll expander presented. • Water injection allows reaching quasi-isothermal expansion process. • Comparison between experimental data, semi-empirical and deterministic models. • Flank leakage, water injection and rotor speed effects have been analyzed experimentally. • Design guidelines for co-rotating scroll improvement proposed. - Abstract: Efficient compressed air energy storage requires reversible isothermal compression and expansion devices. The isothermal compression and expansion processes can either be approached by several stages with intercooling or by the more convenient injection of a liquid, often water. While volumetric machines are readily available for dry processes the compression and expansion of a gas with the presence of liquid is still problematic. The concept of a co-rotating scroll has been identified as a promising technology to cope with the presence of liquid. The current paper discusses the first experimental results of an oil-free co-rotating scroll prototype tested in expansion mode on a wide range of rotational speeds, varying water injection flow rates and with different nominal flank clearances. A maximal overall isothermal efficiency of 34% and a maximum output power of 1.74 kW_e_l were measured with this first prototype, providing the proof of the technical feasibility of the oil-free co-rotating scroll expander concept. The experimental data indicate a positive effect of water injection suggesting good heat transfer behaviour between the water and the air in the individual chambers, which is a result of the relatively long residence time compared to other volumetric concepts. The experimental sensitivity analysis yields a strong dependency of the machine performance on both the nominal flank clearance and on the injected water rate. The analysis through a semi-empirical model suggests the inversion of a classical trend, i.e. the increase in total leakage area with

Transgenic chickens expressing human urokinase-type plasminogen activator.

Science.gov (United States)

Lee, Sung Ho; Gupta, Mukesh Kumar; Ho, Young Tae; Kim, Teoan; Lee, Hoon Taek

2013-09-01

Urokinase-type plasminogen activator is a serine protease that is clinically used in humans for the treatment of thrombolytic disorders and vascular diseases such as acute ischemic stroke and acute peripheral arterial occlusion. This study explored the feasibility of using chickens as a bioreactor for producing human urokinase-type plasminogen activator (huPA). Recombinant huPA gene, under the control of a ubiquitous Rous sarcoma virus promoter, was injected into the subgerminal cavity of freshly laid chicken eggs at stage X using the replication-defective Moloney murine leukemia virus (MoMLV)-based retrovirus vectors encapsidated with VSV-G (vesicular stomatitis virus G) glycoprotein. A total of 38 chicks, out of 573 virus-injected eggs, hatched and contained the huPA gene in their various body parts. The mRNA transcript of the huPA gene was present in various organs, including blood and egg, and was germ-line transmitted to the next generation. The level of active huPA protein was 16-fold higher in the blood of the transgenic chicken than in the nontransgenic chicken (P huPA protein in eggs increased from 7.82 IU/egg in the G0 generation to 17.02 IU/egg in the G1 generation. However, huPA-expressing embryos had reduced survival and hatchability at d 18 and 21 of incubation, respectively, and the blood clotting time was significantly higher in transgenic chickens than their nontransgenic counterparts (P huPA transgenic chickens could be successfully produced by the retroviral vector system. Transgenic chickens, expressing the huPA under the control of a ubiquitous promoter, may not only be used as a bioreactor for pharming of the huPA drug but also be useful for studying huPA-induced bleeding and other disorders.

Assessment of CO2 Mineralization and Dynamic Rock Properties at the Kemper Pilot CO2 Injection Site

Science.gov (United States)

Qin, F.; Kirkland, B. L.; Beckingham, L. E.

2017-12-01

CO2-brine-mineral reactions following CO2 injection may impact rock properties including porosity, permeability, and pore connectivity. The rate and extent of alteration largely depends on the nature and evolution of reactive mineral interfaces. In this work, the potential for geochemical reactions and the nature of the reactive mineral interface and corresponding hydrologic properties are evaluated for samples from the Lower Tuscaloosa, Washita-Fredericksburg, and Paluxy formations. These formations have been identified as future regionally extensive and attractive CO2 storage reservoirs at the CO2 Storage Complex in Kemper County, Mississippi, USA (Project ECO2S). Samples from these formations were obtained from the Geological Survey of Alabama and evaluated using a suite of complementary analyses. The mineral composition of these samples will be determined using petrography and powder X-ray Diffraction (XRD). Using these compositions, continuum-scale reactive transport simulations will be developed and the potential CO2-brine-mineral interactions will be examined. Simulations will focus on identifying potential reactive minerals as well as the corresponding rate and extent of reactions. The spatial distribution and accessibility of minerals to reactive fluids is critical to understanding mineral reaction rates and corresponding changes in the pore structure, including pore connectivity, porosity and permeability. The nature of the pore-mineral interface, and distribution of reactive minerals, will be determined through imaging analysis. Multiple 2D scanning electron microscopy (SEM) backscattered electron (BSE) images and energy dispersive x-ray spectroscopy (EDS) images will be used to create spatial maps of mineral distributions. These maps will be processed to evaluate the accessibility of reactive minerals and the potential for flow-path modifications following CO2 injection. The "Establishing an Early CO2 Storage Complex in Kemper, MS" project is funded by

Injection, flow, and mixing of CO2 in porous media with residual gas.

Energy Technology Data Exchange (ETDEWEB)

Oldenburg, C.M.; Doughty, C.A.

2010-09-01

Geologic structures associated with depleted natural gas reservoirs are desirable targets for geologic carbon sequestration (GCS) as evidenced by numerous pilot and industrial-scale GCS projects in these environments world-wide. One feature of these GCS targets that may affect injection is the presence of residual CH{sub 4}. It is well known that CH{sub 4} drastically alters supercritical CO{sub 2} density and viscosity. Furthermore, residual gas of any kind affects the relative permeability of the liquid and gas phases, with relative permeability of the gas phase strongly dependent on the time-history of imbibition or drainage, i.e., dependent on hysteretic relative permeability. In this study, the effects of residual CH{sub 4} on supercritical CO{sub 2} injection were investigated by numerical simulation in an idealized one-dimensional system under three scenarios: (1) with no residual gas; (2) with residual supercritical CO{sub 2}; and (3) with residual CH{sub 4}. We further compare results of simulations that use non-hysteretic and hysteretic relative permeability functions. The primary effect of residual gas is to decrease injectivity by decreasing liquid-phase relative permeability. Secondary effects arise from injected gas effectively incorporating residual gas and thereby extending the mobile gas plume relative to cases with no residual gas. Third-order effects arise from gas mixing and associated compositional effects on density that effectively create a larger plume per unit mass. Non-hysteretic models of relative permeability can be used to approximate some parts of the behavior of the system, but fully hysteretic formulations are needed to accurately model the entire system.

GFP-Mutant Human Tau Transgenic Mice Develop Tauopathy Following CNS Injections of Alzheimer's Brain-Derived Pathological Tau or Synthetic Mutant Human Tau Fibrils.

Science.gov (United States)

Gibbons, Garrett S; Banks, Rachel A; Kim, Bumjin; Xu, Hong; Changolkar, Lakshmi; Leight, Susan N; Riddle, Dawn M; Li, Chi; Gathagan, Ronald J; Brown, Hannah J; Zhang, Bin; Trojanowski, John Q; Lee, Virginia M-Y

2017-11-22

Neurodegenerative proteinopathies characterized by intracellular aggregates of tau proteins, termed tauopathies, include Alzheimer's disease (AD), frontotemporal lobar degeneration (FTLD) with tau pathology (FTLD-tau), and related disorders. Pathological tau proteins derived from human AD brains (AD-tau) act as proteopathic seeds that initiate the templated aggregation of soluble tau upon intracerebral injection into tau transgenic (Tg) and wild-type mice, thereby modeling human tau pathology. In this study, we found that aged Tg mice of both sexes expressing human tau proteins harboring a pathogenic P301L MAPT mutation labeled with green fluorescent protein (T40PL-GFP Tg mouse line) exhibited hyperphosphorylated tau mislocalized to the somatodentritic domain of neurons, but these mice did not develop de novo insoluble tau aggregates, which are characteristic of human AD and related tauopathies. However, intracerebral injections of either T40PL preformed fibrils (PFFs) or AD-tau seeds into T40PL-GFP mice induced abundant intraneuronal pathological inclusions of hyperphosphorylated T40PL-GFP. These injections of pathological tau resulted in the propagation of tau pathology from the injection site to neuroanatomically connected brain regions, and these tau inclusions consisted of both T40PL-GFP and WT endogenous mouse tau. Primary neurons cultured from the brains of neonatal T40PL-GFP mice provided an informative in vitro model for examining the uptake and localization of tau PFFs. These findings demonstrate the seeded aggregation of T40PL-GFP in vivo by synthetic PFFs and human AD-tau and the utility of this system to study the neuropathological spread of tau aggregates. SIGNIFICANCE STATEMENT The stereotypical spread of pathological tau protein aggregates have recently been attributed to the transmission of proteopathic seeds. Despite the extensive use of transgenic mouse models to investigate the propagation of tau pathology in vivo , details of the aggregation

Studying the Specific Activity of the Amide Form of HLDF-6 Peptide using the Transgenic Model of Alzheimer's Disease.

Science.gov (United States)

Bogachouk, A P; Storozheva, Z I; Telegin, G B; Chernov, A S; Proshin, A T; Sherstnev, V V; Zolotarev, Yu A; Lipkin, V M

2017-01-01

The neuroprotective and nootropic activities of the amide form (AF) of the HLDF-6 peptide (TGENHR-NH 2 ) were studied in transgenic mice of the B6C3-Tg(APPswe,PSEN1de9)85Dbo (Tg+) line (the animal model of familial Alzheimer's disease (AD)). The study was performed in 4 mouse groups: group 1 (study group): Tg+ mice intranasally injected with the peptide at a dose of 250 μg/kg; group 2 (active control): Tg+ mice intranasally injected with normal saline; group 3 (control 1): Tg- mice; and group 4 (control 2): C57Bl/6 mice. The cognitive functions were evaluated using three tests: the novel object recognition test, the conditioned passive avoidance task, and the Morris water maze. The results testify to the fact that the pharmaceutical substance (PhS) based on the AF of HLDF-6 peptide at a dose of 250 μg/kg administered intranasally efficiently restores the disturbed cognitive functions in transgenic mice. These results are fully consistent with the data obtained in animal models of Alzheimer's disease induced by the injection of the beta-amyloid (βA) fragment 25-35 into the giant-cell nucleus basalis of Meynert or by co-injection of the βA fragment 25-35 and ibotenic acid into the hippocampus, and the model of ischemia stroke (chronic bilateral occlusion of carotids, 2VO). According to the overall results, PhS based on AF HLDF-6 was chosen as an object for further investigation; the dose of 250 μg/kg was used as an effective therapeutic dose. Intranasal administration was the route for delivery.

Second-strand cDNA synthesis: classical method

International Nuclear Information System (INIS)

Gubler, U.

1987-01-01

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

Characterization and Ectopic Expression of CoWRI1, an AP2/EREBP Domain-Containing Transcription Factor from Coconut (Cocos nucifera L.) Endosperm, Changes the Seeds Oil Content in Transgenic Arabidopsis thaliana and Rice (Oryza sativa L.).

Science.gov (United States)

Sun, RuHao; Ye, Rongjian; Gao, Lingchao; Zhang, Lin; Wang, Rui; Mao, Ting; Zheng, Yusheng; Li, Dongdong; Lin, Yongjun

2017-01-01

Coconut ( Cocos nucifera L.) is a key tropical crop and a member of the monocotyledonous family Arecaceae ( Palmaceae ). Few genes and related metabolic processes involved in coconut endosperm development have been investigated. In this study, a new member of the WRI1 gene family was isolated from coconut endosperm and was named CoWRI1 . Its transcriptional activities and interactions with the acetyl-CoA carboxylase ( BCCP2 ) promoter of CoWRI1 were confirmed by the yeast two-hybrid and yeast one-hybrid approaches, respectively. Functional characterization was carried out through seed-specific expression in Arabidopsis and endosperm-specific expression in rice. In transgenic Arabidopsis , high over-expressions of CoWRI1 in seven independent T2 lines were detected by quantitative real-time PCR. The relative mRNA accumulation of genes encoding enzymes involved in either fatty acid biosynthesis or triacylglycerols assembly (BCCP2, KASI, MAT, ENR, FATA, and GPDH) were also assayed in mature seeds. Furthermore, lipid and fatty acids C16:0 and C18:0 significantly increased. In two homozygous T2 transgenic rice lines (G5 and G2), different CoWRI1 expression levels were detected, but no CoWRI1 transcripts were detected in the wild type. Analyses of the seed oil content, starch content, and total protein content indicated that the two T2 transgenic lines showed a significant increase ( P oil content. The transgenic lines also showed a significant increase in starch content, whereas total protein content decreased significantly. Further analysis of the fatty acid composition revealed that palmitic acid (C16:0) and linolenic acid (C18:3) increased significantly in the seeds of the transgenic rice lines, but oleic acid (C18:1) levels significantly declined.

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

NARCIS (Netherlands)

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

1996-01-01

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

Application of intracytoplasmic sperm injection (ICSI) for fertilization and development in birds.

Science.gov (United States)

Shimada, Kiyoshi; Ono, Tamao; Mizushima, Shusei

2014-01-15

Intracytoplasmic sperm injection (ICSI) technology in birds has been hampered due to opacity of oocyte. We developed ICSI-assisted fertilization and gene transfer in quail. This paper reviews recent advances of our ICSI experiments. The oocyte retrieved from the oviduct and a quail sperm was injected into the oocyte under a stereomicroscope. The oocyte was cultured for 24h at 41°C under 5% CO2 in air. The fertilization and development was assessed by microscopic observation. The fertility rate ranged 12-18% and development varied from stage II to V in trials. To improve the fertility rate, phospholipase C (PLC) zeta was injected with a sperm. It was increased to 37-50%. Furthermore, injection of inositol trisphosphate increased to over 85%. Quail oocyte can be fertilized with chicken sperm and so can testicular elongated spermatid. To extend embryonic development, chicken eggshell was used as a surrogate culture at 37°C after the 24h incubation at 41°C under 5% CO2 in air. It survived up to 2days thereafter. Finally, gene transfer was attempted in quail egg. The sperm membrane was disrupted with Triton X-100 (TX-100) and was injected with PLCzeta cRNA and enhanced green fluorescent protein (EGFP) gene in oocyte. The GFP expression was evaluated at 24h incubation at 41°C under 5% CO2 in air in the embryos. While the expression was not detected in the control oocytes, the experimental treatment induced blastoderm development (44%) of the oocytes and 86% of blastoderm showed fluorescent emission. In addition, PCR analysis detected EGFP fragments in 50% of GFP-expressing blastoderm. Our ICSI method may be the first step toward the production of transgenic birds. Copyright © 2013 Elsevier Inc. All rights reserved.

Evaluation of miscible and immiscible CO2 injection in one of the Iranian oil fields

Directory of Open Access Journals (Sweden)

Aref Hashemi Fath

2014-09-01

Full Text Available Carbon dioxide (CO2 flooding is one of the most important methods for enhanced oil recovery (EOR because it not only increases oil recovery efficiency but also causes a reduction of greenhouse gas emissions. It is a very complex system, involving phase behavior that could increase the recovery of oil by means of swelling, evaporation and decreasing viscosity of the oil. In this study, a reservoir modeling approach was used to evaluate immiscible and miscible CO2 flooding in a fractured oil field. To reduce simulation time, we grouped fluid components into 10 pseudo-components. The 3-parameter, Peng–Robinson Equation of State (EOS was used to match PVT experimental data by using the PVTi software. A one-dimensional slim-tube model was defined using ECLIPSE 300 software to determine the minimum miscibility pressure (MMP for injection of CO2. We used FloGrid software for making a reservoir static model and the reservoir model was calibrated using manual and assisted history matching methods. Then various scenarios of natural depletion, immiscible and miscible CO2 injection have been simulated by ECLIPSE 300 software and then the simulation results of scenarios have been compared. Investigation of simulation results shows that the oil recovery factor in miscible CO2 injection scenario is more than other methods.

Expression of Finger Millet EcDehydrin7 in Transgenic Tobacco Confers Tolerance to Drought Stress.

Science.gov (United States)

Singh, Rajiv Kumar; Singh, Vivek Kumar; Raghavendrarao, Sanagala; Phanindra, Mullapudi Lakshmi Venkata; Venkat Raman, K; Solanke, Amolkumar U; Kumar, Polumetla Ananda; Sharma, Tilak Raj

2015-09-01

One of the critical alarming constraints for agriculture is water scarcity. In the current scenario, global warming due to climate change and unpredictable rainfall, drought is going to be a master player and possess a big threat to stagnating gene pool of staple food crops. So it is necessary to understand the mechanisms that enable the plants to cope with drought stress. In this study, effort was made to prospect the role of EcDehydrin7 protein from normalized cDNA library of drought tolerance finger millet in transgenic tobacco. Biochemical and molecular analyses of T0 transgenic plants were done for stress tolerance. Leaf disc assay, seed germination test, dehydration assay, and chlorophyll estimation showed EcDehydrin7 protein directly link to drought tolerance. Northern and qRT PCR analyses shows relatively high expression of EcDehydrin7 protein compare to wild type. T0 transgenic lines EcDehydrin7(11) and EcDehydrin7(15) shows superior expression among all lines under study. In summary, all results suggest that EcDehydrin7 protein has a remarkable role in drought tolerance and may be used for sustainable crop breeding program in other food crops.

Persistent interferon transgene expression by RNA interference-mediated silencing of interferon receptors.

Science.gov (United States)

Takahashi, Yuki; Vikman, Elin; Nishikawa, Makiya; Ando, Mitsuru; Watanabe, Yoshihiko; Takakura, Yoshinobu

2010-09-01

The in vivo half-life of interferons (IFNs) is very short, and its extension would produce a better therapeutic outcome in IFN-based therapy. Delivery of IFN genes is one solution for providing a sustained supply. IFNs have a variety of functions, including the suppression of transgene expression, through interaction with IFN receptors (IFNRs). This suppression could prevent IFNs from being expressed from vectors delivered. Silencing the expression of IFNAR and IFNGR, the receptors for type I and II IFNs, respectively, in cells expressing IFNs may prolong transgene expression of IFNs. Mouse melanoma B16-BL6 cells or mouse liver were selected as a site expressing IFNs (not a target for IFN gene therapy) and IFN-expressing plasmid DNA was delivered with or without small interfering RNA (siRNA) targeting IFNRs. Transfection of B16-BL6 cells with siRNA targeting IFNAR1 subunit (IFNAR1) resulted in the reduced expression of IFNAR on the cell surface. This silencing significantly increased the IFN-beta production in cells that were transfected with IFN-beta-expressing plasmid DNA. Similar results were obtained with the combination of IFN-gamma and IFNGR. Co-injection of IFN-beta-expressing plasmid DNA with siRNA targeting IFNAR1 into mice resulted in sustained plasma concentration of IFN-beta. These results provide experimental evidence that the RNAi-mediated silencing of IFNRs in cells expressing IFN, such as hepatocytes, is an effective approach for improving transgene expression of IFNs when their therapeutic target comprises cells other than those expressing IFNs.

Monoterpene engineering in a woody plant Eucalyptus camaldulensis using a limonene synthase cDNA.

Science.gov (United States)

Ohara, Kazuaki; Matsunaga, Etsuko; Nanto, Kazuya; Yamamoto, Kyoko; Sasaki, Kanako; Ebinuma, Hiroyasu; Yazaki, Kazufumi

2010-01-01

Metabolic engineering aimed at monoterpene production has become an intensive research topic in recent years, although most studies have been limited to herbal plants including model plants such as Arabidopsis. The genus Eucalyptus includes commercially important woody plants in terms of essential oil production and the pulp industry. This study attempted to modify the production of monoterpenes, which are major components of Eucalyptus essential oil, by introducing two expression constructs containing Perilla frutescens limonene synthase (PFLS) cDNA, whose gene products were designed to be localized in either the plastid or cytosol, into Eucalyptus camaldulensis. The expression of the plastid-type and cytosol-type PFLS cDNA in transgenic E. camaldulensis was confirmed by real-time polymerase chain reaction (PCR). Gas chromatography with a flame ionization detector analyses of leaf extracts revealed that the plastidic and cytosolic expression of PFLS yielded 2.6- and 4.5-times more limonene than that accumulated in wild-type E. camaldulensis, respectively, while the ectopic expression of PFLS had only a small effect on the emission of limonene from the leaves of E. camaldulensis. Surprisingly, the high level of PFLS in Eucalyptus was accompanied by a synergistic increase in the production of 1,8-cineole and alpha-pinene, two major components of Eucalyptus monoterpenes. This genetic engineering of monoterpenes demonstrated a new potential for molecular breeding in woody plants.

Triazole-linked DNA as a primer surrogate in the synthesis of first-strand cDNA.

Science.gov (United States)

Fujino, Tomoko; Yasumoto, Ken-ichi; Yamazaki, Naomi; Hasome, Ai; Sogawa, Kazuhiro; Isobe, Hiroyuki

2011-11-04

A phosphate-eliminated nonnatural oligonucleotide serves as a primer surrogate in reverse transcription reaction of mRNA. Despite of the nonnatural triazole linkages in the surrogate, the reverse transcriptase effectively elongated cDNA sequences on the 3'-downstream of the primer by transcription of the complementary sequence of mRNA. A structure-activity comparison with the reference natural oligonucleotides shows the superior priming activity of the surrogate containing triazole-linkages. The nonnatural linkages also protect the transcribed cDNA from digestion reactions with 5'-exonuclease and enable us to remove noise transcripts of unknown origins. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Characterizing near-surface CO2 conditions before injection - Perspectives from a CCS project in the Illinois Basin, USA

Science.gov (United States)

Locke, R.A.; Krapac, I.G.; Lewicki, J.L.; Curtis-Robinson, E.

2011-01-01

The Midwest Geological Sequestration Consortium is conducting a large-scale carbon capture and storage (CCS) project in Decatur, Illinois, USA to demonstrate the ability of a deep saline formation to store one million tonnes of carbon dioxide (CO2) from an ethanol facility. Beginning in early 2011, CO2 will be injected at a rate of 1,000 tonnes/day for three years into the Mount Simon Sandstone at a depth of approximately 2,100 meters. An extensive Monitoring, Verification, and Accounting (MVA) program has been undertaken for the Illinois Basin Decatur Project (IBDP) and is focused on the 0.65 km2 project site. Goals include establishing baseline conditions to evaluate potential impacts from CO2 injection, demonstrating that project activities are protective of human health and the environment, and providing an accurate accounting of stored CO2. MVA efforts are being conducted pre-, during, and post- CO2 injection. Soil and net CO2 flux monitoring has been conducted for more than one year to characterize near-surface CO2 conditions. More than 2,200 soil CO2 flux measurements have been manually collected from a network of 118 soil rings since June 2009. Three ring types have been evaluated to determine which type may be the most effective in detecting potential CO 2 leakage. Bare soil, shallow-depth rings were driven 8 cm into the ground and were prepared to minimize surface vegetation in and near the rings. Bare soil, deep-depth rings were prepared similarly, but were driven 46 cm. Natural-vegetation, shallow-depth rings were driven 8 cm and are most representative of typical vegetation conditions. Bare-soil, shallow-depth rings had the smallest observed mean flux (1.78 ??mol m-2 s-1) versus natural-vegetation, shallow-depth rings (3.38 ??mol m-2 s-1). Current data suggest bare ring types would be more sensitive to small CO2 leak signatures than natural ring types because of higher signal to noise ratios. An eddy covariance (EC) system has been in use since June

Sequence of human protamine 2 cDNA

Energy Technology Data Exchange (ETDEWEB)

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

1988-08-11

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

Helper-dependent adenovirus achieve more efficient and persistent liver transgene expression in non-human primates under immunosuppression.

Science.gov (United States)

Unzu, C; Melero, I; Hervás-Stubbs, S; Sampedro, A; Mancheño, U; Morales-Kastresana, A; Serrano-Mendioroz, I; de Salamanca, R E; Benito, A; Fontanellas, A

2015-11-01

Helper-dependent adenoviral (HDA) vectors constitute excellent gene therapy tools for metabolic liver diseases. We have previously shown that an HDA vector encoding human porphobilinogen deaminase (PBGD) corrects acute intermittent porphyria mice. Now, six non-human primates were injected in the left hepatic lobe with the PBGD-encoding HDA vector to study levels and persistence of transgene expression. Intrahepatic administration of 5 × 10(12) viral particles kg(-1) (10(10) infective units kg(-1)) of HDA only resulted in transient (≈14 weeks) transgene expression in one out of three individuals. In contrast, a more prolonged 90-day immunosuppressive regimen (tacrolimus, mycophenolate, rituximab and steroids) extended meaningful transgene expression for over 76 weeks in two out of two cases. Transgene expression under immunosuppression (IS) reached maximum levels 6 weeks after HDA administration and gradually declined reaching a stable plateau within the therapeutic range for acute porphyria. The non-injected liver lobes also expressed the transgene because of vector circulation. IS controlled anticapsid T-cell responses and decreased the induction of neutralizing antibodies. Re-administration of HDA-hPBGD at week +78 achieved therapeutically meaningful transgene expression only in those animals receiving IS again at the time of this second vector exposure. Overall, immunity against adenoviral capsids poses serious hurdles for long-term HDA-mediated liver transduction, which can be partially circumvented by pharmacological IS.

Monitoring a pilot CO2 injection experiment in a shallow aquifer using 3D cross-well electrical resistance tomography

Science.gov (United States)

Yang, X.; Lassen, R. N.; Looms, M. C.; Jensen, K. H.

2014-12-01

Three dimensional electrical resistance tomography (ERT) was used to monitor a pilot CO2 injection experiment at Vrøgum, Denmark. The purpose was to evaluate the effectiveness of the ERT method for monitoring the two opposing effects from gas-phase and dissolved CO2 in a shallow unconfined siliciclastic aquifer. Dissolved CO2 increases water electrical conductivity (EC) while gas phase CO2 reduce EC. We injected 45kg of CO2 into a shallow aquifer for 48 hours. ERT data were collected for 50 hours following CO2 injection. Four ERT monitoring boreholes were installed on a 5m by 5m square grid and each borehole had 24 electrodes at 0.5 m electrode spacing at depths from 1.5 m to 13 m. ERT data were inverted using a difference inversion algorithm for bulk EC. 3D ERT successfully detected the CO2 plume distribution and growth in the shallow aquifer. We found that the changes of bulk EC were dominantly positive following CO2 injection, indicating that the effect of dissolved CO2 overwhelmed that of gas phase CO2. The pre-injection baseline resistivity model clearly showed a three-layer structure of the site. The electrically more conductive glacial sand layer in the northeast region are likely more permeable than the overburden and underburden and CO2 plumes were actually confined in this layer. Temporal bulk EC increase from ERT agreed well with water EC and cross-borehole ground penetrating radar data. ERT monitoring offers a competitive advantage over water sampling and GPR methods because it provides 3D high-resolution temporal tomographic images of CO2 distribution and it can also be automated for unattended operation. This work was performed under the auspices of the U.S. Department of Energy by Lawrence Livermore National Laboratory under contract DE-AC52-07NA27344. Lawrence Livermore National Security, LLC. LLNL IM release#: LLNL-PROC-657944.

Studying the Specific Activity of the Amide Form of HLDF-6 Peptide using the Transgenic Model of Alzheimer’s Disease

Science.gov (United States)

Bogachouk, A. P.; Storozheva, Z. I.; Telegin, G. B.; Chernov, A. S.; Proshin, A. T.; Sherstnev, V. V.; Zolotarev, Yu. A.; Lipkin, V. M.

2017-01-01

The neuroprotective and nootropic activities of the amide form (AF) of the HLDF-6 peptide (TGENHR-NH2) were studied in transgenic mice of the B6C3-Tg(APPswe,PSEN1de9)85Dbo (Tg+) line (the animal model of familial Alzheimer’s disease (AD)). The study was performed in 4 mouse groups: group 1 (study group): Tg+ mice intranasally injected with the peptide at a dose of 250 μg/kg; group 2 (active control): Tg+ mice intranasally injected with normal saline; group 3 (control 1): Tg- mice; and group 4 (control 2): C57Bl/6 mice. The cognitive functions were evaluated using three tests: the novel object recognition test, the conditioned passive avoidance task, and the Morris water maze. The results testify to the fact that the pharmaceutical substance (PhS) based on the AF of HLDF-6 peptide at a dose of 250 μg/kg administered intranasally efficiently restores the disturbed cognitive functions in transgenic mice. These results are fully consistent with the data obtained in animal models of Alzheimer’s disease induced by the injection of the beta-amyloid (βA) fragment 25-35 into the giant-cell nucleus basalis of Meynert or by co-injection of the βA fragment 25-35 and ibotenic acid into the hippocampus, and the model of ischemia stroke (chronic bilateral occlusion of carotids, 2VO). According to the overall results, PhS based on AF HLDF-6 was chosen as an object for further investigation; the dose of 250 μg/kg was used as an effective therapeutic dose. Intranasal administration was the route for delivery. PMID:29104777

Method for construction of normalized cDNA libraries

Science.gov (United States)

Soares, Marcelo B.; Efstratiadis, Argiris

1998-01-01

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

Salt Sensitive Tet-Off-Like Systems to Knockdown Primordial Germ Cell Genes for Repressible Transgenic Sterilization in Channel Catfish, Ictalurus punctatus.

Science.gov (United States)

Li, Hanbo; Su, Baofeng; Qin, Guyu; Ye, Zhi; Alsaqufi, Ahmed; Perera, Dayan A; Shang, Mei; Odin, Ramjie; Vo, Khoi; Drescher, David; Robinson, Dalton; Zhang, Dan; Abass, Nermeen; Dunham, Rex A

2017-05-31

Repressible knockdown approaches were investigated for transgenic sterilization in channel catfish, Ictalurus punctatus . Two primordial germ cell (PGC) marker genes, nanos and dead end , were targeted for knockdown, and an off-target gene, vasa , was monitored. Two potentially salt sensitive repressible promoters, zebrafish adenylosuccinate synthase 2 (ADSS) and zebrafish racemase (Rm), were each coupled with four knockdown strategies: ds-sh RNA targeting the 5' end (N1) or 3' end (N2) of channel catfish nanos , full-length cDNA sequence of channel catfish nanos for overexpression (cDNA) and ds-sh RNA targeting channel catfish dead end (DND). Each construct had an untreated group and treated group with sodium chloride as the repressor compound. Spawning rates of full-sibling P₁ fish exposed or not exposed to the constructs as treated and untreated embryos were 93% and 59%, respectively, indicating potential sterilization of fish and repression of the constructs. Although the mRNA expression data of PGC marker genes were inconsistent in P₁ fish, most F₁ individuals were able to downregulate the target genes in untreated groups and repress the knockdown process in treated groups. The results indicate that repressible transgenic sterilization is feasible for reproductive control of fish, but more data from F₂ or F₃ are needed for evaluation.

Effects of GC temperature and carrier gas flow rate on on-line oxygen isotope measurement as studied by on-column CO injection.

Science.gov (United States)

Chen, Zhi-Gang; Yin, Xi-Jie; Zhou, Youping

2015-08-01

Although deemed important to δ 18 O measurement by on-line high-temperature conversion techniques, how the GC conditions affect δ 18 O measurement is rarely examined adequately. We therefore directly injected different volumes of CO or CO-N 2 mix onto the GC column by a six-port valve and examined the CO yield, CO peak shape, CO-N 2 separation, and δ 18 O value under different GC temperatures and carrier gas flow rates. The results show the CO peak area decreases when the carrier gas flow rate increases. The GC temperature has no effect on peak area. The peak width increases with the increase of CO injection volume but decreases with the increase of GC temperature and carrier gas flow rate. The peak intensity increases with the increase of GC temperature and CO injection volume but decreases with the increase of carrier gas flow rate. The peak separation time between N 2 and CO decreases with an increase of GC temperature and carrier gas flow rate. δ 18 O value decreases with the increase of CO injection volume (when half m/z 28 intensity is rate. On average, the δ 18 O value of the injected CO is about 1‰ higher than that of identical reference CO. The δ 18 O distribution pattern of the injected CO is probably a combined result of ion source nonlinearity and preferential loss of C 16 O or oxygen isotopic exchange between zeolite and CO. For practical application, a lower carrier gas flow rate is therefore recommended as it has the combined advantages of higher CO yield, better N 2 -CO separation, lower He consumption, and insignificant effect on δ 18 O value, while a higher-than-60 °C GC temperature and a larger-than-100 µl CO volume is also recommended. When no N 2 peak is expected, a higher GC temperature is recommended, and vice versa. Copyright © 2015 John Wiley & Sons, Ltd. Copyright © 2015 John Wiley & Sons, Ltd.

Reduction of birefringence in a skin-layer of injection molded polymer strips using CO{sub 2} laser irradiation

Energy Technology Data Exchange (ETDEWEB)

Kurosaki, Yasuo; Satoh, Isao; Saito, Takushi [Tokyo Inst. of Tech. (Japan). Dept. of Mechanical Intelligent Systems Engineering

1995-12-31

Injection molding of polymers is currently utilized for numerous industrial applications. Because of high productivity and stable quality of molded products, the injection-molding process makes the production costs lower, and therefore, is expected to spread more widely in the future. This paper deals with a technique for improving the optical quality of injection molded polymer products using radiative heating. The birefringence frozen in a skin-layer of the molded part was reduced by CO{sub 2} laser heating, and the efficiency of this technique was investigated experimentally. Namely, a simple numerical calculation was performed to estimate the heating efficiency of CO{sub 2} laser in the polymer, effects of radiation heating on the skin-layer of the molded polymer were observed by using a mold with transparent windows, and the residual birefringence frozen in the final molded specimen was measured. The results clearly showed that the birefringence in the skin-layer of injection molded polymer strips was reduced with CO{sub 2} laser heating. The authors believe that the proposed method for reducing the birefringence frozen in injection-molded polymer products is suitable for practical molding, because process time required for the injection-molding is only slightly increased with this method.

Pre injection characterisation and evaluation of CO{sub 2} sequestration potential in the Haizume formation, Niigata basin, Japan; Caracterisation avant injection et evaluation du potentiel de sequestration de CO{sub 2} dans la formation de Haizume, bassin de Niigata, Japon. Modelisation geochimique des interactions eau-mineraux-CO{sub 2}

Energy Technology Data Exchange (ETDEWEB)

Zwingmann, N. [CSIRO Petroleum, ARRC, Bentley, WA (Australia); Mito, S.; Sorai, M.; Ohsumi, T. [RITE, Kyoto (Japan); Zwingmann, N. [Western, Univ. (Australia); Sorai, M. [Mitsubishi Research Institute, Inc. (Japan)

2005-03-15

The Research Institute of Innovative Technology for the Earth (RITE) is carrying out a small-scale CO{sub 2} injection field experiment to investigate the feasibility of geological sequestration of CO{sub 2} greenhouse gas in the south-west of Nagaoka City, Niigata Prefecture, Japan. Prior to the injection geochemical reactions caused by CO{sub 2} injections were investigated using the geochemical modelling code (EQ3/6). The injection formation is the sedimentary marine Haizume Formation (Pleistocene) in the Uonuma Group, which is covered by a mud-stone seal. The formation is mainly composed of quartz, plagioclase, feldspar, pyroxene, and clays (smectite, chlorite). The sandstone shows minor consolidation and grain size is medium to coarse sand. The total dissolved solid (TDS) of the formation water is approximately 6100 mg/l and the water contains a high Ca{sup 2+} ({>=} 20% of Na{sup +} concentration). The geochemical model was used for an initial adjustment of the formation water chemistry to the formation conditions and a modelling of the formation water-mineral-CO{sub 2} reactions. The modelling results showed a high reactivity of the minerals in the CO{sub 2} rich environment and high mineral conversion rate within the formation. At the final state, approximately 23 mol of CO{sub 2} were taken into 1 kg of formation water and more than 90% of this was stored within carbonate minerals. In this simulation, some uncertainty is associated with the time scale and a more detailed investigation is planned and will address accurate evaluation. (authors)

Suppression of inflammation by dexamethasone prolongs adenoviral vector-mediated transgene expression in the facial nucleus of the rat

NARCIS (Netherlands)

Hermens, W.T.J.M.C.; Verhaagen, J

1998-01-01

Adenoviral vector directed gene transfer to rat facial motoneurons occurs efficiently following intra-parenchymal injection of relatively high dosages (> or =10(7) pfu per injection) of a prototype first generation adenoviral vector. However, high level of transgene expression, as observed during

Transgenic expression of lactoferrin imparts enhanced resistance to head blight of wheat caused by Fusarium graminearum.

Science.gov (United States)

Han, Jigang; Lakshman, Dilip K; Galvez, Leny C; Mitra, Sharmila; Baenziger, Peter Stephen; Mitra, Amitava

2012-03-09

The development of plant gene transfer systems has allowed for the introgression of alien genes into plant genomes for novel disease control strategies, thus providing a mechanism for broadening the genetic resources available to plant breeders. Using the tools of plant genetic engineering, a broad-spectrum antimicrobial gene was tested for resistance against head blight caused by Fusarium graminearum Schwabe, a devastating disease of wheat (Triticum aestivum L.) and barley (Hordeum vulgare L.) that reduces both grain yield and quality. A construct containing a bovine lactoferrin cDNA was used to transform wheat using an Agrobacterium-mediated DNA transfer system to express this antimicrobial protein in transgenic wheat. Transformants were analyzed by Northern and Western blots to determine lactoferrin gene expression levels and were inoculated with the head blight disease fungus F. graminearum. Transgenic wheat showed a significant reduction of disease incidence caused by F. graminearum compared to control wheat plants. The level of resistance in the highly susceptible wheat cultivar Bobwhite was significantly higher in transgenic plants compared to control Bobwhite and two untransformed commercial wheat cultivars, susceptible Wheaton and tolerant ND 2710. Quantification of the expressed lactoferrin protein by ELISA in transgenic wheat indicated a positive correlation between the lactoferrin gene expression levels and the levels of disease resistance. Introgression of the lactoferrin gene into elite commercial wheat, barley and other susceptible cereals may enhance resistance to F. graminearum.

Intrathecal injection of naked plasmid DNA provides long-term expression of secreted proteins.

Science.gov (United States)

Hughes, Travis S; Langer, Stephen J; Johnson, Kirk W; Chavez, Raymond A; Watkins, Linda R; Milligan, Erin D; Leinwand, Leslie A

2009-01-01

Therapeutic benefit has been reported to result from intrathecal (i.t.) injection of transgene vectors, including naked DNA. However, most studies using naked DNA have measured only the transgene expression of intracellular proteins. Here we demonstrate that i.t. injection of naked DNA can result in long-term expression of secreted proteins. Plasmids expressing either secreted alkaline phosphatase (SEAP) or human interleukin-10 (hIL-10) were injected into the i.t. space in rats, and transgene products were repeatedly measured in the cerebrospinal fluid (CSF). Both SEAP and hIL-10 were maximal at 1 and 2 days after the injection and still detectable at 4 months. The utilization of a plasmid having two features that are hypothesized to increase gene expression (matrix attachment regions (MARs) and lack of CpG dinucleotides) resulted in a significant increase in gene expression. Reinjection of SEAP or hIL-10 plasmids after 4 months significantly increased protein levels at 1 and 14 days after the reinjection. SEAP was uniformly distributed between the DNA delivery site (approximately vertebral level T13) and the lumbar puncture site (L5/L6 inter-vertebral space), was reduced at the cisterna magna, and was detectable, though at much lower levels, in serum. These data suggest that naked DNA has the potential to be used as a therapeutic tool for applications that require long-term release of transgenes into the CSF.

Generation of a Homozygous Transgenic Rat Strain Stably Expressing a Calcium Sensor Protein for Direct Examination of Calcium Signaling.

Science.gov (United States)

Szebényi, Kornélia; Füredi, András; Kolacsek, Orsolya; Pergel, Enikő; Bősze, Zsuzsanna; Bender, Balázs; Vajdovich, Péter; Tóvári, József; Homolya, László; Szakács, Gergely; Héja, László; Enyedi, Ágnes; Sarkadi, Balázs; Apáti, Ágota; Orbán, Tamás I

2015-08-03

In drug discovery, prediction of selectivity and toxicity require the evaluation of cellular calcium homeostasis. The rat is a preferred laboratory animal for pharmacology and toxicology studies, while currently no calcium indicator protein expressing rat model is available. We established a transgenic rat strain stably expressing the GCaMP2 fluorescent calcium sensor by a transposon-based methodology. Zygotes were co-injected with mRNA of transposase and a CAG-GCaMP2 expressing construct, and animals with one transgene copy were pre-selected by measuring fluorescence in blood cells. A homozygous rat strain was generated with high sensor protein expression in the heart, kidney, liver, and blood cells. No pathological alterations were found in these animals, and fluorescence measurements in cardiac tissue slices and primary cultures demonstrated the applicability of this system for studying calcium signaling. We show here that the GCaMP2 expressing rat cardiomyocytes allow the prediction of cardiotoxic drug side-effects, and provide evidence for the role of Na(+)/Ca(2+) exchanger and its beneficial pharmacological modulation in cardiac reperfusion. Our data indicate that drug-induced alterations and pathological processes can be followed by using this rat model, suggesting that transgenic rats expressing a calcium-sensitive protein provide a valuable system for pharmacological and toxicological studies.

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

Lifescience Database Archive (English)

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

Creating Transgenic shRNA Mice by Recombinase-Mediated Cassette Exchange

Science.gov (United States)

Premsrirut, Prem K.; Dow, Lukas E.; Park, Youngkyu; Hannon, Gregory J.; Lowe, Scott W.

2014-01-01

RNA interference (RNAi) enables sequence-specific, experimentally induced silencing of virtually any gene by tapping into innate regulatory mechanisms that are conserved among most eukaryotes. The principles that enable transgenic RNAi in cell lines can also be used to create transgenic animals, which express short-hairpin RNAs (shRNAs) in a regulated or tissue-specific fashion. However, RNAi in transgenic animals is somewhat more challenging than RNAi in cultured cells. The activities of promoters that are commonly used for shRNA expression in cell culture can vary enormously in different tissues, and founder lines also typically vary in transgene expression due to the effects of their single integration sites. There are many ways to produce mice carrying shRNA transgenes and the method described here uses recombinase-mediated cassette exchange (RMCE). RMCE permits insertion of the shRNA transgene into a well-characterized locus that gives reproducible and predictable expression in each founder and enhances the probability of potent expression in many cell types. This procedure is more involved and complex than simple pronuclear injection, but if even a few shRNA mice are envisioned, for example, to probe the functions of several genes, the effort of setting up the processes outlined below are well worthwhile. Note that when creating a transgenic mouse, one should take care to use the most potent shRNA possible. As a rule of thumb, the sequence chosen should provide >90% knockdown when introduced into cultured cells at single copy (e.g., on retroviral infection at a multiplicity of ≤0.3). PMID:24003198

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

International Nuclear Information System (INIS)

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

1987-01-01

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

Composite potato plants with transgenic roots on non-transgenic shoots: a model system for studying gene silencing in roots.

Science.gov (United States)

Horn, Patricia; Santala, Johanna; Nielsen, Steen Lykke; Hühns, Maja; Broer, Inge; Valkonen, Jari P T

2014-12-01

Composite potato plants offer an extremely fast, effective and reliable system for studies on gene functions in roots using antisense or inverted-repeat but not sense constructs for gene inactivation. Composite plants, with transgenic roots on a non-transgenic shoot, can be obtained by shoot explant transformation with Agrobacterium rhizogenes. The aim of this study was to generate composite potato plants (Solanum tuberosum) to be used as a model system in future studies on root-pathogen interactions and gene silencing in the roots. The proportion of transgenic roots among the roots induced was high (80-100%) in the four potato cultivars tested (Albatros, Desirée, Sabina and Saturna). No wild-type adventitious roots were formed at mock inoculation site. All strains of A. rhizogenes tested induced phenotypically normal roots which, however, showed a reduced response to cytokinin as compared with non-transgenic roots. Nevertheless, both types of roots were infected to a similar high rate with the zoospores of Spongospora subterranea, a soilborne potato pathogen. The transgenic roots of composite potato plants expressed significantly higher amounts of β-glucuronidase (GUS) than the roots of a GUS-transgenic potato line event. Silencing of the uidA transgene (GUS) was tested by inducing roots on the GUS-transgenic cv. Albatros event with strains of A. rhizogenes over-expressing either the uidA sense or antisense transcripts, or inverted-repeat or hairpin uidA RNA. The three last mentioned constructs caused 2.5-4.0 fold reduction in the uidA mRNA expression. In contrast, over-expression of uidA resulted in over 3-fold increase in the uidA mRNA and GUS expression, indicating that sense-mediated silencing (co-suppression) was not functional in roots. The results suggest that composite plants offer a useful experimental system for potato research, which has gained little previous attention.

Bacterial magnetic particles improve testes-mediated transgene efficiency in mice.

Science.gov (United States)

Wang, Chao; Sun, Guanghong; Wang, Ye; Kong, Nana; Chi, Yafei; Yang, Leilei; Xin, Qiliang; Teng, Zhen; Wang, Xu; Wen, Yujun; Li, Ying; Xia, Guoliang

2017-11-01

Nano-scaled materials have been proved to be ideal DNA carriers for transgene. Bacterial magnetic particles (BMPs) help to reduce the toxicity of polyethylenimine (PEI), an efficient gene-transferring agent, and assist tissue transgene ex vivo. Here, the effectiveness of the BMP-PEI complex-conjugated foreign DNAs (BPDs) in promoting testes-mediated gene transfer (TMGT) in mouse was compared with that of liposome-conjugated foreign DNAs. The results proved that through testes injection, the clusters of BPDs successfully reached the cytoplasm and the nuclear of spermatogenesis cell, and expressed in testes of transgene founder mice. Additionally, the ratio of founder mice obtained from BPDs (88%) is about 3 times higher than the control (25%) (p mice from BPD group were significantly improved, as compared with the control (p mice within the first filial was significantly higher in BPDs compared with the control (73.8% versus 11.6%, p mice in vivo.

Primary structure of bovine pituitary secretory protein I (chromogranin A) deduced from the cDNA sequence

International Nuclear Information System (INIS)

Ahn, T.G.; Cohn, D.V.; Gorr, S.U.; Ornstein, D.L.; Kashdan, M.A.; Levine, M.A.

1987-01-01

Secretory protein I (SP-I), also referred to as chromogranin A, is an acidic glycoprotein that has been found in every tissue of endocrine and neuroendocrine origin examined but never in exocrine or epithelial cells. Its co-storage and co-secretion with peptide hormones and neurotransmitters suggest that it has an important endocrine or secretory function. The authors have isolated cDNA clones from a bovine pituitary λgt11 expression library using an antiserum to parathyroid SP-I. The largest clone (SP4B) hybridized to a transcript of 2.1 kilobases in RNA from parathyroid, pituitary, and adrenal medulla. Immunoblots of bacterial lysates derived from SP4B lysognes demonstrated specific antibody binding to an SP4B/β-galactosidase fusion protein (160 kDa) with a cDNA-derived component of 46 kDa. Radioimmunoassay of the bacterial lystates with SP-I antiserum yielded parallel displacement curves of 125 I-labeled SP-I by the SP4B lysate and authentic SP-I. SP4B contains a cDNA of 1614 nucleotides that encodes a 449-amino acid protein (calculated mass, 50 kDa). The nucleotide sequences of the pituitary SP-I cDNA and adrenal medullary SP-I cDNAs are nearly identical. Analysis of genomic DNA suggests that pituitary, adrenal, and parathyroid SP-I are products of the same gene

Primary structure of bovine pituitary secretory protein I (chromogranin A) deduced from the cDNA sequence

Energy Technology Data Exchange (ETDEWEB)

Ahn, T.G.; Cohn, D.V.; Gorr, S.U.; Ornstein, D.L.; Kashdan, M.A.; Levine, M.A.

1987-07-01

Secretory protein I (SP-I), also referred to as chromogranin A, is an acidic glycoprotein that has been found in every tissue of endocrine and neuroendocrine origin examined but never in exocrine or epithelial cells. Its co-storage and co-secretion with peptide hormones and neurotransmitters suggest that it has an important endocrine or secretory function. The authors have isolated cDNA clones from a bovine pituitary lambdagt11 expression library using an antiserum to parathyroid SP-I. The largest clone (SP4B) hybridized to a transcript of 2.1 kilobases in RNA from parathyroid, pituitary, and adrenal medulla. Immunoblots of bacterial lysates derived from SP4B lysognes demonstrated specific antibody binding to an SP4B/..beta..-galactosidase fusion protein (160 kDa) with a cDNA-derived component of 46 kDa. Radioimmunoassay of the bacterial lystates with SP-I antiserum yielded parallel displacement curves of /sup 125/I-labeled SP-I by the SP4B lysate and authentic SP-I. SP4B contains a cDNA of 1614 nucleotides that encodes a 449-amino acid protein (calculated mass, 50 kDa). The nucleotide sequences of the pituitary SP-I cDNA and adrenal medullary SP-I cDNAs are nearly identical. Analysis of genomic DNA suggests that pituitary, adrenal, and parathyroid SP-I are products of the same gene.

Ectopic expression of the agouti gene in transgenic mice causes obesity, features of type II diabetes, and yellow fur

Energy Technology Data Exchange (ETDEWEB)

Klebig, M.L.; Woychik, R.P. [Oak Ridge National Laboratory, Oak Ridge, TN (United States); Wilkinson, J.E. [Univ. of Tennessee, Knoxville, TN (United States); Geisler, J.G. [Oak Ridge National Laboratory, Oak Ridge, TN (United States)]|[Univ. of Tennessee, Knoxville, TN (United States)

1995-05-23

Mice that carry the lethal yellow (A{sup y}) or viable yellow (A{sup vy}) mutation, two dominant mutations of the agouti (a) gene in mouse chromosome 2, exhibit a phenotype that includes yellow fur, marked obesity, a form of type II diabetes associated with insulin resistance, and an increased susceptibility to tumor development. Molecular analyses of these and several other dominant {open_quotes}obese yellow{close_quotes} a-locus mutations suggested that ectopic expression of the normal agouti protein gives rise to this complex pleiotropic phenotype. We have now tested this hypothesis directly by generating transgenic mice that ectopically express an agouti cDNA clone encoding the normal agouti protein in all tissues examined. Transgenic mice of both sexes have yellow fur, become obese, and develop hyperinsulinemia. In addition, male transgenic mice develop hyperglycemia by 12-20 weeks of age. These results demonstrate conclusively that the ectopic agouti expression is responsible for most, if not all, of the phenotypic traits of the dominant, obese yellow mutants. 42 refs., 5 figs.

High blood pressure in transgenic mice carrying the rat angiotensinogen gene.

Science.gov (United States)

Kimura, S; Mullins, J J; Bunnemann, B; Metzger, R; Hilgenfeldt, U; Zimmermann, F; Jacob, H; Fuxe, K; Ganten, D; Kaling, M

1992-01-01

Transgenic mice were generated by injecting the entire rat angiotensinogen gene into the germline of NMRI mice. The resulting transgenic animals were characterized with respect to hemodynamics, parameters of the renin angiotension system, and expression of the transgene. The transgenic line TGM(rAOGEN)123 developed hypertension with a mean arterial blood pressure of 158 mmHg in males and 132 mmHg in females. In contrast, the transgenic line TGM(rAOGEN)92 was not hypertensive. Rat angiotensinogen was detectable only in plasma of animals of line 123. Total plasma angiotensinogen and plasma angiotensin II concentrations were about three times as high as those of negative control mice. In TGM(rAOGEN)123 the transgene was highly expressed in liver and brain. Transcripts were also detected in heart, kidney and testis. In TGM(rAOGEN)92 the brain was the main expressing organ. In situ hybridization revealed an mRNA distribution in the brain of TGM(rAOGEN)123 similar to the one in rat. In TGM(rAOGEN)92 the expression pattern in the brain was aberrant. These data indicate that overexpression of the angiotensinogen gene in liver and brain leads to the development of hypertension in transgenic mice. The TGM(rAOGEN)123 constitutes a high angiotensin II type of hypertension and may provide a new experimental animal model to study the kinetics and function of the renin angiotensin system. Images PMID:1547785

Subsurface impact of CO2: Response of carbonate rocks and wellbore cement to supercritical CO2 injection and long-term storage. Geologica Ultraiectina (310)

NARCIS (Netherlands)

Liteanu, E.

2009-01-01

Capture of CO2 at fossil fuel power station coupled with geological storage in empty oil and gas reservoirs is widely viewed as the most promising option for reducing CO2 emissions to the atmosphere, i.e. for climate change mitigation. Injection of CO2 into such reservoirs will change their chemical

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

Science.gov (United States)

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

2006-05-01

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

Cancer-specific transgene expression mediated by systemic injection of nanoparticles.

Science.gov (United States)

Chisholm, Edward J; Vassaux, Georges; Martin-Duque, Pilar; Chevre, Raphael; Lambert, Olivier; Pitard, Bruno; Merron, Andrew; Weeks, Mark; Burnet, Jerome; Peerlinck, Inge; Dai, Ming-Shen; Alusi, Ghassan; Mather, Stephen J; Bolton, Katherine; Uchegbu, Ijeoma F; Schatzlein, Andreas G; Baril, Patrick

2009-03-15

The lack of safe and efficient systemic gene delivery vectors has largely reduced the potential of gene therapy in the clinic. Previously, we have reported that polypropylenimine dendrimer PPIG3/DNA nanoparticles are capable of tumor transfection upon systemic administration in tumor-bearing mice. To be safely applicable in the clinic, it is crucial to investigate the colloidal stability of nanoparticles and to monitor the exact biodistribution of gene transfer in the whole body of the live subject. Our biophysical characterization shows that dendrimers, when complexed with DNA, are capable of forming spontaneously in solution a supramolecular assembly that possesses all the features required to diffuse in experimental tumors through the enhanced permeability and retention effect. We show that these nanoparticles are of sizes ranging from 33 to 286 nm depending on the DNA concentration, with a colloidal stable and well-organized fingerprint-like structure in which DNA molecules are condensed with an even periodicity of 2.8 nm. Whole-body nuclear imaging using small-animal nano-single-photon emission computed tomography/computer tomography scanner and the human Na/I symporter (NIS) as reporter gene shows unique and highly specific tumor targeting with no detection of gene transfer in any of the other tissues of tumor-bearing mice. Tumor-selective transgene expression was confirmed by quantitative reverse transcription-PCR at autopsy of scanned animals, whereas genomic PCR showed that the tumor sites are the predominant sites of nanoparticle accumulation. Considering that NIS imaging of transgene expression has been recently validated in humans, our data highlight the potential of these nanoparticles as a new formulation for cancer gene therapy.

Procedure for normalization of cDNA libraries

Science.gov (United States)

Bonaldo, Maria DeFatima; Soares, Marcelo Bento

1997-01-01

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

Geomechanical Modeling of Fault Responses and the Potential for Notable Seismic Events during Underground CO2 Injection

Science.gov (United States)

Rutqvist, J.; Cappa, F.; Mazzoldi, A.; Rinaldi, A.

2012-12-01

The importance of geomechanics associated with large-scale geologic carbon storage (GCS) operations is now widely recognized. There are concerns related to the potential for triggering notable (felt) seismic events and how such events could impact the long-term integrity of a CO2 repository (as well as how it could impact the public perception of GCS). In this context, we review a number of modeling studies and field observations related to the potential for injection-induced fault reactivations and seismic events. We present recent model simulations of CO2 injection and fault reactivation, including both aseismic and seismic fault responses. The model simulations were conducted using a slip weakening fault model enabling sudden (seismic) fault rupture, and some of the numerical analyses were extended to fully dynamic modeling of seismic source, wave propagation, and ground motion. The model simulations illustrated what it will take to create a magnitude 3 or 4 earthquake that would not result in any significant damage at the groundsurface, but could raise concerns in the local community and could also affect the deep containment of the stored CO2. The analyses show that the local in situ stress field, fault orientation, fault strength, and injection induced overpressure are critical factors in determining the likelihood and magnitude of such an event. We like to clarify though that in our modeling we had to apply very high injection pressure to be able to intentionally induce any fault reactivation. Consequently, our model simulations represent extreme cases, which in a real GCS operation could be avoided by estimating maximum sustainable injection pressure and carefully controlling the injection pressure. In fact, no notable seismic event has been reported from any of the current CO2 storage projects, although some unfelt microseismic activities have been detected by geophones. On the other hand, potential future commercial GCS operations from large power plants

Cloning of the γ-aminobutyric acid (GABA) ρ1 cDNA: A GABA receptor subunit highly expressed in the retina

International Nuclear Information System (INIS)

Cutting, G.R.; Lu, Luo; Kasch, L.M.; Montrose-Rafizadeh, C.; Antonarakis, S.E.; Guggino, W.B.; Kazazian, H.H. Jr.; O'Hara, B.F.; Donovan, D.M.; Shimada, Shoichi; Uhl, G.R.

1991-01-01

Type A γ-aminobutyric acid (GABA A ) receptors are a family of ligand-gated chloride channels that are the major inhibitory neurotransmitter receptors in the nervous system. Molecular cloning has revealed diversity in the subunits that compose this heterooligomeric receptor, but each previously elucidated subunit displays amino acid similarity in conserved structural elements. The authors have used these highly conserved regions to identify additional members of this family by using the polymerase chain reaction (PCR). One PCR product was used to isolate a full-length cDNA from a human retina cDNA library. The mature protein predicted from this cDNA sequence is 458 amino acids long and displays between 30 and 38% amino acid similarity to the previously identified GABA A subunits. This gene is expressed primarily in the retina but transcripts are also detected in the brain, lung, and thymus. Injection of Xenopus oocytes with RNA transcribed in vitro produces a GABA-responsive chloride conductance and expression of the cDNA in COS cells yields GABA-displaceable muscimol binding. These features are consistent with our identification of a GABA subunit, GABA ρ 1 , with prominent retinal expression that increases the diversity and tissue specificity of this ligand-gated ion-channel receptor family

Rock physical aspects of CO{sub 2} injection in chalk

Energy Technology Data Exchange (ETDEWEB)

Alam, M.M.

2011-04-15

Impact of supercritical CO{sub 2} on the petrophysical and rock-mechanics properties of Ekofisk Formation and Tor Formation chalk from South Arne field, Danish North Sea, chalk was investigated. A series of laboratory experiments was performed on core material collected from the reservoir zone of the South Arne field in order to reveal the changes with respect to porosity, specific surface, pore stiffness, wettability, mineralogy and mechanical failure. In addition, a theoretical rock physical background was also established in order to be able to make sensible interpretation of laboratory data. Sound wave velocity was used as the central tool to study any change in petrophysical and rock mechanical properties. The main focus was to achieve a better understanding of effective stress coefficient (also known as Biot's coefficient); by means of which effective stress can be predicted more accurately. Independent theoretical studies were made on diagenesis, surface properties and stiffness of chalk and their relation with sonic velocity (or Biot's coefficient calculated from sonic velocity). The knowledge and experience from these studies was combined to achieve the main research objective of monitoring changes in hydrocarbon reservoirs in chalk due to CO{sub 2} injection. In order to understand the development of chalk from calcareous ooze and achieving pore stiffness, the diagenesis process of a sedimentary sequence from Kerguelen Plateau in the Indian Ocean was studied. The principal objective of the study was to explore how different porosity reduction mechanisms change the strength of these deep sea carbonate-rich sediments and how these mechanisms can be traced from the change in Biot's coefficient, alpha. In calcareous ooze, alpha was found close to one. Mechanical compaction reduces porosity, but only leads to a minor decrease in alpha. Recrystallization process renders particles smoother, but do not lead to reduction in alpha unless it gives

CO2 injection into submarine, CH4-hydrate bearing sediments: Parameter studies towards the development of a hydrate conversion technology

Science.gov (United States)

Deusner, Christian; Bigalke, Nikolaus; Kossel, Elke; Haeckel, Matthias

2013-04-01

In the recent past, international research efforts towards exploitation of submarine and permafrost hydrate reservoirs have increased substantially. Until now, findings indicate that a combination of different technical means such as depressurization, thermal stimulation and chemical activation is the most promising approach for producing gas from natural hydrates. Moreover, emission neutral exploitation of CH4-hydrates could potentially be achieved in a combined process with CO2 injection and storage as CO2-hydrate. In the German gas hydrate initiative SUGAR, a combination of experimental and numerical studies is used to elucidate the process mechanisms and technical parameters on different scales. Experiments were carried out in the novel high-pressure flow-through system NESSI (Natural Environment Simulator for sub-Seafloor Interactions). Recent findings suggest that the injection of heated, supercritical CO2 is beneficial for both CH4 production and CO2 retention. Among the parameters tested so far are the CO2 injection regime (alternating vs. continuous injection) and the reservoir pressure / temperature conditions. Currently, the influence of CO2 injection temperature is investigated. It was shown that CH4 production is optimal at intermediate reservoir temperatures (8 ° C) compared to lower (2 ° C) and higher temperatures (10 ° C). The reservoir pressure, however, was of minor importance for the production efficiency. At 8 ° C, where CH4- and CO2-hydrates are thermodynamically stable, CO2-hydrate formation appears to be slow. Eventual clogging of fluid conduits due to CO2-rich hydrate formation force open new conduits, thereby tapping different regions inside the CH4-hydrate sample volume for CH4gas. In contrast, at 2 ° C immediate formation of CO2-hydrate results in rapid and irreversible obstruction of the entire pore space. At 10 ° C pure CO2-hydrates can no longer be formed. Consequently the injected CO2 flows through quickly and interaction with

Hydro-geophysical responses to the injection of CO2 in core plugs of Berea sandstone

Science.gov (United States)

Song, I.; Park, K. G.

2017-12-01

We have built a laboratory-scale core flooding system to measure the relative permeability of a core sample and the acoustic response to the CO2 saturation degree at in situ condition of pressure and temperature down to a few kilometer depths. The system consisted of an acoustic velocity core holder (AVC model from the Core Laboratories) between upstream where CO2 and H2O were injected separately and downstream where the mixed fluids came out of a core sample. Core samples with 4 cm in diameter and 5 cm in length of Berea sandstone were in turn placed in the core holder for confining and axial pressures. The flooding operations of the multiphase fluids were conducted through the sample at 40ºC in temperature and 8 MPa in backpressure. CO2 and H2O in the physical condition were injected separately into a sample at constant rate with various ratios. The two phases were mixed during flowing through the sample. The mixed fluids out of the sample were separated again by their different densities in a chamber equipped with a level gauge of the interface. From the level change of the water in the separator, we measured the volume of water coming out of the sample for each test with a constant ratio of the injection rates. Then it was possible to calculate the saturation degree of CO2 from the difference between input volume and output volume of water. The differential pressure between upstream and downstream was directly measured to calculate the relative permeability as a function of the CO2 saturation degree. We also conducted ultrasonic measurements using piezoelectric sensors on the end plugs. An electric pulse was given to a sensor on one end of sample, and then ultrasonic waves were recorded from the other end. The various ratios of injection rate of CO2 and H2O into Berea sandstone yielded a range of 0.1-0.7 in CO2 saturation degree. The relative permeability was obtained at the condition of steady-state flow for given stages from the velocity of each phase and

Coal char combustion under a CO{sub 2}-rich atmosphere: Implications for pulverized coal injection in a blast furnace

Energy Technology Data Exchange (ETDEWEB)

Borrego, A.G.; Casal, M.D. [Instituto Nacional del Carbon, CSIC. P.O. Box 73, 33080 Oviedo (Spain); Osorio, E.; Vilela, A.C.F. [Laboratorio de Siderurgia, DEMET/PPGEM - Universidade Federal do Rio Grande do Sul. P.O. Box 15021, 91501-970 Porto Alegre (Brazil)

2008-11-15

Pulverized coal injection (PCI) is employed in blast furnace tuyeres attempting to maximize the injection rate without increasing the amount of unburned char inside the stack of the blast furnace. When coal is injected with air through the injection lance, the resolidified char will burn in an atmosphere with a progressively lower oxygen content and higher CO{sub 2} concentration. In this study an experimental approach was followed to separate the combustion process into two distinct devolatilization and combustion steps. Initially coal was injected into a drop tube furnace (DTF) operating at 1300 C in an atmosphere with a low oxygen concentration to ensure the combustion of volatiles and prevent the formation of soot. Then the char was refired into the DTF at the same temperature under two different atmospheres O{sub 2}/N{sub 2} (typical combustion) and O{sub 2}/CO{sub 2} (oxy-combustion) with the same oxygen concentration. Coal injection was also performed under a higher oxygen concentration in atmospheres typical for both combustion and oxy-combustion. The fuels tested comprised a petroleum coke and coals currently used for PCI injection ranging from high volatile to low volatile bituminous rank. Thermogravimetric analyses and microscopy techniques were used to establish the reactivity and appearance of the chars. Overall similar burnouts were achieved with N{sub 2} and CO{sub 2} for similar oxygen concentrations and therefore no loss in burnout should be expected as a result of enrichment in CO{sub 2} in the blast furnace gas. The advantage of increasing the amount of oxygen in a reacting atmosphere during burnout was found to be greater, the higher the rank of the coal. (author)

Rapid Integration of Multi-copy Transgenes Using Optogenetic Mutagenesis in Caenorhabditis elegans

Directory of Open Access Journals (Sweden)

Kentaro Noma

2018-06-01

Full Text Available Stably transmitted transgenes are indispensable for labeling cellular components and manipulating cellular functions. In Caenorhabditis elegans, transgenes are generally generated as inheritable multi-copy extrachromosomal arrays, which can be stabilized in the genome through a mutagenesis-mediated integration process. Standard methods to integrate extrachromosomal arrays primarily use protocols involving ultraviolet light plus trimethylpsoralen or gamma- or X-ray irradiation, which are laborious and time-consuming. Here, we describe a one-step integration method, following germline-mutagenesis induced by mini Singlet Oxygen Generator (miniSOG. Upon blue light treatment, miniSOG tagged to histone (Histone-miniSOG generates reactive oxygen species (ROS and induces heritable mutations, including DNA double-stranded breaks. We demonstrate that we can bypass the need to first establish extrachromosomal transgenic lines by coupling microinjection of desired plasmids with blue light illumination on Histone-miniSOG worms to obtain integrants in the F3 progeny. We consistently obtained more than one integrant from 12 injected animals in two weeks. This optogenetic approach significantly reduces the amount of time and labor for transgene integration. Moreover, it enables to generate stably expressed transgenes that cause toxicity in animal growth.

Quantitative monitoring of CO2 injection at Sleipner using seismic full waveform inversion in the time lapse mode and rock physics modeling

International Nuclear Information System (INIS)

Queisser, M.P.

2012-01-01

Carbon capture and sequestration is a technology to achieve a considerable deceleration of CO 2 emission promptly. Since 1996 one of the largest CO 2 storage projects is taking place at Sleipner in the Norwegian North Sea. In order to monitor injected CO 2 , time lapse seismic monitoring surveys have been carried out. Estimating subsurface parameters from the Sleipner seismic data is a challenging problem due to the specific geology of the storage reservoir, which is further complicated by injected CO 2 . Most seismic imaging methods enable only qualitative insights into the subsurface. Motivated by the need for a quantitative seismic monitoring of the injected CO 2 , I have applied 2D seismic full waveform inversion to seismic data sets from Sleipner from 1994 (baseline), 1999 and 2006 along three seismic lines to infer subsurface parameters and parameter changes in the storage reservoir. The P-wave velocity is the major parameter, as it is the most sensitive to CO 2 injection. An energy preconditioning of the gradient has been implemented. The usual source wavelet calibration did not prove to be reliable. An alternative source calibration has been successfully applied. By comparing seismic images with inversion results, I found that using seismic images to locate CO 2 accumulations in the subsurface may be misleading. The quantitative imaging approach using full waveform inversion resulted in a consistent evolution of the model parameter with time. Major reductions in P-wave velocity and hence the CO 2 accumulations could be quantitatively imaged down to a resolution of 10 m. Observed travel time shifts due to CO 2 injection are comparable to those derived from the inversion result. In order to estimate CO 2 saturations, rock physical concepts have been combined and extended to arrive at a rock physical formulation of the subsurface at Sleipner. I used pseudo Monte Carlo rock physics modeling to assess the influence of lithologic heterogeneity on the CO 2

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

Lifescience Database Archive (English)

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

Consequences of thermal fracture developments due to injection cold CO2 into depleted gas fields

NARCIS (Netherlands)

Peters, E.; Pizzocolo, F.; Loeve, D.; Fokker, P.A.; Hofstee, C.; Orlic, B.; Maas, J.G.

2013-01-01

CO2 storage is planned in a depleted gas field called P18, which is located offshore in the vicinity of the Dutch coast. This project is also known as the ROAD project, which is the Rotterdam capture and storage demonstration project. In the P18-4 compartment, cold CO2 will be injected into a

Motor Function and Dopamine Release Measurements in Transgenic Huntington’s Disease Model Rats

Science.gov (United States)

Ortiz, Andrea N.; Osterhaus, Gregory L.; Lauderdale, Kelli; Mahoney, Luke; Fowler, Stephen C.; von Hörsten, Stephan; Riess, Olaf; Johnson, Michael A.

2013-01-01

Huntington’s disease (HD) is a fatal, genetic, neurodegenerative disorder characterized by deficits in motor and cognitive function. Here, we have quantitatively characterized motor deficiencies and dopamine release dynamics in transgenic HD model rats. Behavioral analyses were conducted using a newly-developed force-sensing runway and a previously-developed force-plate actometer. Gait disturbances were readily observed in transgenic HD rats at 12 to 15 months of age. Additionally, dopamine system challenge by ip injection of amphetamine also revealed that these rats were resistant to the expression of focused stereotypy compared to wild-type controls. Moreover, dopamine release, evoked by the application of single and multiple electrical stimulus pulses applied at different frequencies, and measured using fast-scan cyclic voltammetry at carbon-fiber microelectrodes, was diminished in transgenic HD rats compared to age-matched wild-type control rats. Collectively, these results underscore the potential contribution of dopamine release alterations to the expression of motor impairments in transgenic HD rats. PMID:22418060

Impact of hydrogeological and geomechanical properties on surface uplift at a CO2 injection site: Parameter estimation and uncertainty quantification

Science.gov (United States)

Newell, P.; Yoon, H.; Martinez, M. J.; Bishop, J. E.; Arnold, B. W.; Bryant, S.

2013-12-01

It is essential to couple multiphase flow and geomechanical response in order to predict a consequence of geological storage of CO2. In this study, we estimate key hydrogeologic features to govern the geomechanical response (i.e., surface uplift) at a large-scale CO2 injection project at In Salah, Algeria using the Sierra Toolkit - a multi-physics simulation code developed at Sandia National Laboratories. Importantly, a jointed rock model is used to study the effect of postulated fractures in the injection zone on the surface uplift. The In Salah Gas Project includes an industrial-scale demonstration of CO2 storage in an active gas field where CO2 from natural gas production is being re-injected into a brine-filled portion of the structure downdip of the gas accumulation. The observed data include millimeter scale surface deformations (e.g., uplift) reported in the literature and injection well locations and rate histories provided by the operators. Our preliminary results show that the intrinsic permeability and Biot coefficient of the injection zone are important. Moreover pre-existing fractures within the injection zone affect the uplift significantly. Estimation of additional (i.e., anisotropy ratio) and coupled parameters will help us to develop models, which account for the complex relationship between mechanical integrity and CO2 injection-induced pressure changes. Uncertainty quantification of model predictions will be also performed using various algorithms including null-space Monte Carlo and polynomial-chaos expansion methods. This work will highlight that our coupled reservoir and geomechanical simulations associated with parameter estimation can provide a practical solution for designing operating conditions and understanding subsurface processes associated with the CO2 injection. This work is supported as part of the Center for Frontiers of Subsurface Energy Security, an Energy Frontier Research Center funded by the U.S. Department of Energy, Office

Transgenic expression of lactoferrin imparts enhanced resistance to head blight of wheat caused by Fusarium graminearum

Directory of Open Access Journals (Sweden)

Han Jigang

2012-03-01

Full Text Available Abstract Background The development of plant gene transfer systems has allowed for the introgression of alien genes into plant genomes for novel disease control strategies, thus providing a mechanism for broadening the genetic resources available to plant breeders. Using the tools of plant genetic engineering, a broad-spectrum antimicrobial gene was tested for resistance against head blight caused by Fusarium graminearum Schwabe, a devastating disease of wheat (Triticum aestivum L. and barley (Hordeum vulgare L. that reduces both grain yield and quality. Results A construct containing a bovine lactoferrin cDNA was used to transform wheat using an Agrobacterium-mediated DNA transfer system to express this antimicrobial protein in transgenic wheat. Transformants were analyzed by Northern and Western blots to determine lactoferrin gene expression levels and were inoculated with the head blight disease fungus F. graminearum. Transgenic wheat showed a significant reduction of disease incidence caused by F. graminearum compared to control wheat plants. The level of resistance in the highly susceptible wheat cultivar Bobwhite was significantly higher in transgenic plants compared to control Bobwhite and two untransformed commercial wheat cultivars, susceptible Wheaton and tolerant ND 2710. Quantification of the expressed lactoferrin protein by ELISA in transgenic wheat indicated a positive correlation between the lactoferrin gene expression levels and the levels of disease resistance. Conclusions Introgression of the lactoferrin gene into elite commercial wheat, barley and other susceptible cereals may enhance resistance to F. graminearum.

Salt Sensitive Tet-Off-Like Systems to Knockdown Primordial Germ Cell Genes for Repressible Transgenic Sterilization in Channel Catfish, Ictalurus punctatus

Directory of Open Access Journals (Sweden)

Hanbo Li

2017-05-01

Full Text Available Repressible knockdown approaches were investigated for transgenic sterilization in channel catfish, Ictalurus punctatus. Two primordial germ cell (PGC marker genes, nanos and dead end, were targeted for knockdown, and an off-target gene, vasa, was monitored. Two potentially salt sensitive repressible promoters, zebrafish adenylosuccinate synthase 2 (ADSS and zebrafish racemase (Rm, were each coupled with four knockdown strategies: ds-sh RNA targeting the 5′ end (N1 or 3′ end (N2 of channel catfish nanos, full-length cDNA sequence of channel catfish nanos for overexpression (cDNA and ds-sh RNA targeting channel catfish dead end (DND. Each construct had an untreated group and treated group with sodium chloride as the repressor compound. Spawning rates of full-sibling P1 fish exposed or not exposed to the constructs as treated and untreated embryos were 93% and 59%, respectively, indicating potential sterilization of fish and repression of the constructs. Although the mRNA expression data of PGC marker genes were inconsistent in P1 fish, most F1 individuals were able to downregulate the target genes in untreated groups and repress the knockdown process in treated groups. The results indicate that repressible transgenic sterilization is feasible for reproductive control of fish, but more data from F2 or F3 are needed for evaluation.

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

Science.gov (United States)

Ares, Manuel

2014-01-01

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

Constructing and detecting a cDNA library for mites.

Science.gov (United States)

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

2015-10-01

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

Molecular cloning of growth hormone encoding cDNA of Indian

Indian Academy of Sciences (India)

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

A cold-induced myo-inositol transporter-like gene confers tolerance to multiple abiotic stresses in transgenic tobacco plants.

Science.gov (United States)

Sambe, Mame Abdou Nahr; He, Xueying; Tu, Qinghua; Guo, Zhenfei

2015-03-01

A full length cDNA encoding a myo-inositol transporter-like protein, named as MfINT-like, was cloned from Medicago sativa subsp. falcata (herein falcata), a species with greater cold tolerance than alfalfa (M. sativa subsp. sativa). MfINT-like is located on plasma membranes. MfINT-like transcript was induced 2-4 h after exogenous myo-inositol treatment, 24-96 h with cold, and 96 h by salinity. Given that myo-inositol accumulates higher in falcata after 24 h of cold treatment, myo-inositol is proposed to be involved in cold-induced expression of MfINT-like. Higher levels of myo-inositol was observed in leaves of transgenic tobacco plants overexpressing MfINT-like than the wild-type but not in the roots of plants grown on myo-inositol containing medium, suggesting that transgenic plants had higher myo-inositol transport activity than the wild-type. Transgenic plants survived better to freezing temperature, and had lower ion leakage and higher maximal photochemical efficiency of photosystem II (Fv /Fm ) after chilling treatment. In addition, greater plant fresh weight was observed in transgenic plants as compared with the wild-type when plants were grown under drought or salinity stress. The results suggest that MfINT-like mediated transport of myo-inositol is associated with plant tolerance to abiotic stresses. © 2014 Scandinavian Plant Physiology Society.

Reactive transport modeling to study changes in water chemistry induced by CO2 injection at the Frio-I brine pilot

Energy Technology Data Exchange (ETDEWEB)

Kharaka, Y.K; Doughty, C.; Freifeld, B.M.; Daley, T.M.; Xu, T.

2009-11-01

To demonstrate the potential for geologic storage of CO{sub 2} in saline aquifers, the Frio-I Brine Pilot was conducted, during which 1600 tons of CO{sub 2} were injected into a high-permeability sandstone and the resulting subsurface plume of CO{sub 2} was monitored using a variety of hydrogeological, geophysical, and geochemical techniques. Fluid samples were obtained before CO{sub 2} injection for baseline geochemical characterization, during the CO{sub 2} injection to track its breakthrough at a nearby observation well, and after injection to investigate changes in fluid composition and potential leakage into an overlying zone. Following CO{sub 2} breakthrough at the observation well, brine samples showed sharp drops in pH, pronounced increases in HCO{sub 3}{sup -} and aqueous Fe, and significant shifts in the isotopic compositions of H{sub 2}O and dissolved inorganic carbon. Based on a calibrated 1-D radial flow model, reactive transport modeling was performed for the Frio-I Brine Pilot. A simple kinetic model of Fe release from the solid to aqueous phase was developed, which can reproduce the observed increases in aqueous Fe concentration. Brine samples collected after half a year had lower Fe concentrations due to carbonate precipitation, and this trend can be also captured by our modeling. The paper provides a method for estimating potential mobile Fe inventory, and its bounding concentration in the storage formation from limited observation data. Long-term simulations show that the CO{sub 2} plume gradually spreads outward due to capillary forces, and the gas saturation gradually decreases due to its dissolution and precipitation of carbonates. The gas phase is predicted to disappear after 500 years. Elevated aqueous CO{sub 2} concentrations remain for a longer time, but eventually decrease due to carbonate precipitation. For the Frio-I Brine Pilot, all injected CO{sub 2} could ultimately be sequestered as carbonate minerals.

Cloning of the cDNA for human 12-lipoxygenase

International Nuclear Information System (INIS)

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

1990-01-01

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

Vertically-Integrated Dual-Continuum Models for CO2 Injection in Fractured Aquifers

Science.gov (United States)

Tao, Y.; Guo, B.; Bandilla, K.; Celia, M. A.

2017-12-01

Injection of CO2 into a saline aquifer leads to a two-phase flow system, with supercritical CO2 and brine being the two fluid phases. Various modeling approaches, including fully three-dimensional (3D) models and vertical-equilibrium (VE) models, have been used to study the system. Almost all of that work has focused on unfractured formations. 3D models solve the governing equations in three dimensions and are applicable to generic geological formations. VE models assume rapid and complete buoyant segregation of the two fluid phases, resulting in vertical pressure equilibrium and allowing integration of the governing equations in the vertical dimension. This reduction in dimensionality makes VE models computationally more efficient, but the associated assumptions restrict the applicability of VE model to formations with moderate to high permeability. In this presentation, we extend the VE and 3D models for CO2 injection in fractured aquifers. This is done in the context of dual-continuum modeling, where the fractured formation is modeled as an overlap of two continuous domains, one representing the fractures and the other representing the rock matrix. Both domains are treated as porous media continua and can be modeled by either a VE or a 3D formulation. The transfer of fluid mass between rock matrix and fractures is represented by a mass transfer function connecting the two domains. We have developed a computational model that combines the VE and 3D models, where we use the VE model in the fractures, which typically have high permeability, and the 3D model in the less permeable rock matrix. A new mass transfer function is derived, which couples the VE and 3D models. The coupled VE-3D model can simulate CO2 injection and migration in fractured aquifers. Results from this model compare well with a full-3D model in which both the fractures and rock matrix are modeled with 3D models, with the hybrid VE-3D model having significantly reduced computational cost. In

Functional cloning using pFB retroviral cDNA expression libraries.

Science.gov (United States)

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

2002-09-01

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

Monitoring a large volume CO2 injection: Year two results from SECARB project at Denbury’s Cranfield, Mississippi, USA

Science.gov (United States)

Hovorka, Susan D.; Meckel, Timothy A.; Trevino, Ramon H.; Lu, Jiemin; Nicot, Jean-Philippe; Choi, Jong-Won; Freeman, David; Cook, Paul; Daley, Thomas M.; Ajo-Franklin, Jonathan B.; Freifeild, Barry M.; Doughty, Christine; Carrigan, Charles R.; La Brecque, Doug; Kharaka, Yousif K.; Thordsen, James J.; Phelps, Tommy J.; Yang, Changbing; Romanak, Katherine D.; Zhang, Tongwei; Holt, Robert M.; Lindler, Jeffery S.; Butsch, Robert J.

2011-01-01

The Southeast Regional Carbon Sequestration Partnership (SECARB) early project in western Mississippi has been testing monitoring tools and approaches to document storage efficiency and storage permanence under conditions of CO2 EOR as well as downdip injection into brine. Denbury Onshore LLC is host for the study and has brought a depleted oil and gas reservoir, Cranfield Field, under CO2 flood. Injection was started in July 2008 and has now achieved injection rates greater than 1.2 million tons/year though 23 wells, with cumulative mass injected as of August, 2010 of 2.2 million metric tons. Injection is into coarse grained fluvial deposits of the Cretaceous lower Tuscaloosa Formation in a gentle anticline at depths of 3300 m. A team of researchers from 10 institutions has collected data from five study areas, each with a different goal and different spatial and temporal scale.

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

NARCIS (Netherlands)

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

2000-01-01

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

cDNA library construction of two human Demodexspecies.

Science.gov (United States)

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

2017-06-01

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

Magneto-plasmonic Au-Coated Co nanoparticles synthesized via hot-injection method

Science.gov (United States)

Souza, João B., Jr.; Varanda, Laudemir C.

2018-02-01

A synthetic procedure is described for the obtaining of superparamagnetic Co nanoparticles (NPs) via hot-injection method in the presence of sodium borohydride. The Co NPs obtained have an average diameter of 5.3 nm and saturation magnetization of 115 emu g-1. A modified Langevin equation is fitted to the magnetization curves using a log-normal distribution for the particle diameter and an effective field to account for dipolar interactions. The calculated magnetic diameter of the Co NPs is 0.6 nm smaller than TEM-derived values, implying a magnetic dead layer of 0.3 nm. The magnetic core is coated with Au to prevent oxidation, resulting in water-stable magneto-plasmonic Co/Au core/shell NPs with saturation of 71.6 emu g-1. The coating adds a localized surface plasmon resonance property with absorbance in the so-called ‘therapeutic window’ (690-900 nm), suitable for biomedical applications. It is suggested that these multifunctional NPs are distinguished as a potential platform for applied and fundamental research.

Pollen Competition as a Reproductive Isolation Barrier Represses Transgene Flow between Compatible and Co-Flowering Citrus Genotypes

Science.gov (United States)

Pons, Elsa; Navarro, Antonio; Ollitrault, Patrick; Peña, Leandro

2011-01-01

Background/Objective Despite potential benefits granted by genetically modified (GM) fruit trees, their release and commercialization raises concerns about their potential environmental impact, and the transfer via pollen of transgenes to cross-compatible cultivars is deemed to be the greatest source for environmental exposure. Information compiled from field trials on GM trees is essential to propose measures to minimize the transgene dispersal. We have conducted a field trial of seven consecutive years to investigate the maximum frequency of pollen-mediated crop-to-crop transgene flow in a citrus orchard, and its relation to the genetic, phenological and environmental factors involved. Methodology/Principal Findings Three different citrus genotypes carrying the uidA (GUS) tracer marker gene (pollen donors) and a non-GM self-incompatible contiguous citrus genotype (recipient) were used in conditions allowing natural entomophilous pollination to occur. The examination of 603 to 2990 seeds per year showed unexpectedly low frequencies (0.17–2.86%) of transgene flow. Paternity analyses of the progeny of subsets of recipient plants using 10 microsatellite (SSR) loci demonstrated a higher mating competence of trees from another non-GM pollen source population that greatly limited the mating chance of the contiguous cross-compatible and flowering-synchronized transgenic pollen source. This mating superiority could be explained by a much higher pollen competition capacity of the non-GM genotypes, as was confirmed through mixed-hand pollinations. Conclusions/Significance Pollen competition strongly contributed to transgene confinement. Based on this finding, suitable isolation measures are proposed for the first time to prevent transgene outflow between contiguous plantings of citrus types that may be extendible to other entomophilous transgenic fruit tree species. PMID:21991359

Changes in Oleic Acid Content of Transgenic Soybeans by Antisense RNA Mediated Posttranscriptional Gene Silencing

Directory of Open Access Journals (Sweden)

Ling Zhang

2014-01-01

Full Text Available The Delta-12 oleate desaturase gene (FAD2-1, which converts oleic acid into linoleic acid, is the key enzyme determining the fatty acid composition of seed oil. In this study, we inhibited the expression of endogenous Delta-12 oleate desaturase GmFad2-1b gene by using antisense RNA in soybean Williams 82. By employing the soybean cotyledonary-node method, a part of the cDNA of soybean GmFad2-1b 801 bp was cloned for the construction of a pCAMBIA3300 vector under the soybean seed promoter BCSP. Leaf painting, LibertyLink strip, PCR, Southern blot, qRT-PCR, and fatty acid analysis were used to detect the insertion and expression of GmFad2-1b in the transgenic soybean lines. The results indicate that the metabolically engineered plants exhibited a significant increase in oleic acid (up to 51.71% and a reduction in palmitic acid (to <3% in their seed oil content. No structural differences were observed between the fatty acids of the transgenic and the nontransgenic oil extracts.

Flow regime analysis for fluid injection into a confined aquifer: implications for CO2 sequestration

Science.gov (United States)

Guo, B.; Zheng, Z.; Celia, M. A.; Stone, H.

2015-12-01

Carbon dioxide injection into a confined saline aquifer may be modeled as an axisymmetric two-phase flow problem. Assuming the two fluids segregate in the vertical direction due to strong buoyancy, and neglecting capillary pressure and miscibility, the lubrication approximation leads to a nonlinear advection-diffusion equation that describes the evolution of the sharp fluid-fluid interface. The flow behaviors in the system are controlled by two dimensionless groups: M, the viscosity ratio of the displaced fluid relative to injected fluid, and Γ , the gravity number, which represents the relative importance of buoyancy and fluid injection. Four different analytical solutions can be derived as the asymptotic approximations, representing specific values of the parameter pairs. The four solutions correspond to: (1) Γ 1; and (4) Γ >> 1, any M values. The first two of these solutions are new, while the third corresponds to the solution of Nordbotten and Celia (2006) for confined injections and the fourth corresponds to the solution of (Lyle et al., 2005) for gravity currents in an unconfined aquifer. Overall, the various axisymmetric flows can be summarized in a Γ-M regime diagram with five distinct dynamic behaviors including the four asymptotic regimes and an intermediate regime (Fig. 1). Data from a number of CO2 injection sites around the world can be used to compute the two dimensionless groups Γ and M associated with each injection. When plotted on the regime diagram, these values show the flow behavior for each injection and how the values vary from site to site. For all the CO2 injections, M is always larger than 1, while Γ can range from 0.01 up to 100. The pairs of (Γ, M) with lower Γ values correspond to solution (3), while the ones with higher Γ values can move up to the intermediate regime and the flow regime for solution (4). The higher values of Γ correspond to pilot-scale injections with low injection rates; most industrial-scale injection

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

Lifescience Database Archive (English)

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

Final report of a scalable, automated, semipermanent seismic array (SASSA) method for detecting CO₂ extent during geologic CO₂ injection

Energy Technology Data Exchange (ETDEWEB)

Burnison, Shaughn [University of North Dakota Energy & Environmental Research Center, Grand Forks, ND (United States); Livers-Douglas, Amanda [University of North Dakota Energy & Environmental Research Center, Grand Forks, ND (United States); Barajas-Olalde, Cesar [University of North Dakota Energy & Environmental Research Center, Grand Forks, ND (United States); Jin, Lu [University of North Dakota Energy & Environmental Research Center, Grand Forks, ND (United States); Vettleson, Heidi [University of North Dakota Energy & Environmental Research Center, Grand Forks, ND (United States); Hamiling, John [University of North Dakota Energy & Environmental Research Center, Grand Forks, ND (United States); Gorecki, Charles [University of North Dakota Energy & Environmental Research Center, Grand Forks, ND (United States)

2017-12-15

The scalable, automated, semipermanent seismic array (SASSA) project led and managed by the Energy & Environmental Research Center (EERC) was designed as a 3-year proof-of-concept study to evaluate and demonstrate an innovative application of the seismic method. The concept was to use a sparse surface array of 96 nodal seismic sensors paired with a single, remotely operated active seismic source at a fixed location to monitor for CO₂ saturation changes in a subsurface reservoir by processing the data for time-lapse changes at individual, strategically chosen reservoir reflection points. The combination of autonomous equipment and modern processing algorithms was used to apply the seismic method in a manner different from the normal paradigm of collecting a spatially dense data set to produce an image. It was used instead to monitor individual, strategically chosen reservoir reflection points for detectable signal character changes that could be attributed to the passing of a CO₂ saturation front or, possibly, changes in reservoir pressure. Data collection occurred over the course of 1 year at an oil field undergoing CO₂ injection for enhanced oil recovery (EOR) and focused on four overlapping “five-spot” EOR injector–producer patterns. Selection, procurement, configuration, installation, and testing of project equipment and collection of five baseline data sets were completed in advance of CO₂ injection within the study area. Weekly remote data collection produced 41 incremental time-lapse records for each of the 96 nodes. Validation was provided by two methods: 1) a conventional 2-D seismic line acquired through the center of the study area before injection started and again after the project ended and processed in a time-lapse manner and 2) by CO₂ saturation maps created from reservoir simulations based on injection and production history matching. Interpreted results were encouraging but mixed, with

Muscle-directed gene therapy for phenylketonuria (PKU): Development of transgenic mice with muscle-specific phenylalanine hydroxylase expression

Energy Technology Data Exchange (ETDEWEB)

Harding, C.O.; Messing, A.; Wolff, J.A. [Univ. of Wisconsin, Madison, WI (United States)

1994-09-01

Phenylketonuria (PKU) is an attractive target for gene therapy because of shortcomings in current therapy including lifelong commitment to a difficult and expensive diet, persistent mild cognitive deficits in some children despite adequate dietary therapy, and maternal PKU syndrome. Phenylalanine hydroxylase (PAH) is normally expressed only in liver, but we propose to treat PKU by introducing the gene for PAH into muscle. In order to evaluate both the safety and efficacy of this approach, we have a developed a trangenic mouse which expresses PAH in both cardiac and skeletal muscle. The transgene includes promoter and enhancer sequences from the mouse muscle creatine kinase (MCK) gene fused to the mouse liver PAH cDNA. Mice which have inherited the transgene are healthy, active, and do not exhibit any signs of muscle weakness or wasting. Ectopic PAH expression in muscle is not detrimental to the health, neurologic function, or reproduction of the mice. Pah{sup enu2} hyperphenylalaninemic mice, a model of human PAH deficiency, bred to carry the transgene have substantial PAH expression in cardiac and skeletal muscle but none in liver. Muscle PAH expression alone does not complement the hyperphenylalaninemic phenotype of Pah{sup enu2} mice. However, administration of reduced tetrahydrobiopterin to transgenic Pah{sup enu2} mice is associated with a 25% mean decrease in serum phenylalanine levels. We predict that ectopic expression of PAH in muscle along with adequate muscle supplies of reduced biopterin cofactor will decrease hyperphenylalaninemia in PKU.

Decreased TK activity alters growth, yield and tolerance to low temperature and low light intensity in transgenic cucumber plants.

Science.gov (United States)

Bi, Huangai; Dong, Xubing; Wu, Guoxiu; Wang, Meiling; Ai, Xizhen

2015-02-01

Four CsTK antisense transgenic cucumber plants were obtained. Decreased TK activity decreased the photosynthetic rate, seed germination rate, growth yield, and the tolerance to low temperature and weak light stress. Transketolase (TK, EC 2.2.1.1) is a key enzyme in the photosynthetic carbon reduction cycle (Calvin cycle). A cDNA fragment (526 bp) encoding transketolase was cloned from cucumber plants (Cucumis sativa L. cv 'Jinyou 3') by RT-PCR. The antisense expression [(PBI-CsTK(-)] vector containing the CsTK gene fragment was constructed. The resulting plasmid was introduced into the cucumber inbred lines '08-1' using the agrobacterium-mediated method, and four antisense transgenic cucumber plants were obtained. Decreased CsTK expression either unaltered or slightly increased the mRNA abundance and activities of the other main enzymes in the Calvin cycle, however, it decreased the TK activity and net photosynthetic rate (Pn) in antisense transgenic cucumber leaves. Antisense plants showed decreases in the growth, ratio of female flowers and yield compared with the wild-type (WT) plants. The decrease in Pn, stomatal conductance (Gs), transpiration rate (Tr), photochemical efficiency (Fv/Fm) and actual photochemical efficiency of PSII (ΦPSII) and the increase in electrolyte leakage (EL) were greater in antisense transgenic plants than in WT plants under low temperature (5 °C) and low light intensity (100 μmol m(-2) s(-1)).

Transient changes in shallow groundwater chemistry during the MSU ZERT CO2 injection experiment

Science.gov (United States)

Apps, J.A.; Zheng, Lingyun; Spycher, N.; Birkholzer, J.T.; Kharaka, Y.; Thordsen, J.; Kakouros, E.; Trautz, R.

2011-01-01

Food-grade CO2 was injected into a shallow aquifer through a perforated pipe placed horizontally 1-2 m below the water table at the Montana State University Zero Emission Research and Technology (MSU-ZERT) field site at Bozeman, Montana. The possible impact of elevated CO2 levels on groundwater quality was investigated by analyzing 80 water samples taken before, during, and following CO2 injection. Field determinations and laboratory analyses showed rapid and systematic changes in pH, alkalinity, and conductance, as well as increases in the aqueous concentrations of trace element species. The geochemical data were first evaluated using principal component analysis (PCA) in order to identify correlations between aqueous species. The PCA findings were then used in formulating a geochemical model to simulate the processes likely to be responsible for the observed increases in the concentrations of dissolved constituents. Modeling was conducted taking into account aqueous and surface complexation, cation exchange, and mineral precipitation and dissolution. Reasonable matches between measured data and model results suggest that: (1) CO2 dissolution in the groundwater causes calcite to dissolve. (2) Observed increases in the concentration of dissolved trace metals result likely from Ca+2-driven ion exchange with clays (smectites) and sorption/desorption reactions likely involving Fe (hydr)oxides. (3) Bicarbonate from CO2 dissolution appears to compete for sorption with anionic species such as HAsO4-2, potentially increasing dissolved As levels in groundwater. ?? 2011 Published by Elsevier Ltd.

Physical Property Changes During CO2 Injection into Sandstone from Pukpyeong Formation, South Korea: Pore-scale Approach

Science.gov (United States)

Han, J.; Keehm, Y.

2010-12-01

Carbon dioxide is believed to be responsible for global warming and climate change, and Korea government puts a great effort in CCS (Carbon Capture and Storage). The geological sequestration is regarded as one viable option and we are looking for prospecting formations for carbon storage. In this paper, we present a new approach to determine physical property changes during CO2 injection and preliminary results from applying the method to one of prospective Tertiary formation in South Korea. The so-called computational rock physics method is composed of three steps: 1) acquisition of high-resolution pore microstructures by X-ray micro-tomography; 2) CO2 injection simulation using lattice-Boltzmann (LB) two-phase flow simulation; and 3) FEM property simulations (electrical and elastic) at different CO2 saturations during the injection. We have been shown the viability of the method last year. This year we applied this method to one of CS (carbon storage) target area, Pukpyeong formation located in north-eastern part of South Korea. From thin section analysis, we found that the formation is composed of mudstone, sandstone and conglomerate, and most of them are poorly consolidated. The mudstone and poorly-sorted conglomerate are believed to have very low permeability, and the effect of CO2 injection would be significant. Thus we focus on sandstone units and get pore microstructure of those units. We then performed the computational rock physics analysis, and present the relations of Vp - CO2 saturation, and electrical conductivity - CO2 saturation for a few sand units. We also present the preliminary upscaling results by putting combined sandstone and mudstone units into FEM modeling. The modeling results implies that the new computational approach can be very useful to characterizing the CS sites especially in early stage. Acknowledgement: This work was supported by the Energy R&D program of the Korea Institute of Energy Technology Evaluation and Planning (KETEP

Optimal scheduling for enhanced coal bed methane production through CO2 injection

International Nuclear Information System (INIS)

Huang, Yuping; Zheng, Qipeng P.; Fan, Neng; Aminian, Kashy

2014-01-01

Highlights: • A novel deterministic optimization model for CO 2 -ECBM production scheduling. • Maximize the total profit from both sales of natural gas and CO 2 credits trading in the carbon market. • A stochastic model incorporating uncertainties and dynamics of NG price and CO 2 credit. - Abstract: Enhanced coal bed methane production with CO 2 injection (CO 2 -ECBM) is an effective technology for accessing the natural gas embedded in the traditionally unmineable coal seams. The revenue via this production process is generated not only by the sales of coal bed methane, but also by trading CO 2 credits in the carbon market. As the technology of CO 2 -ECBM becomes mature, its commercialization opportunities are also springing up. This paper proposes applicable mathematical models for CO 2 -ECBM production and compares the impacts of their production schedules on the total profit. A novel basic deterministic model for CO 2 -ECBM production including the technical and chemical details is proposed and then a multistage stochastic programming model is formulated in order to address uncertainties of natural gas price and CO 2 credit. Both models are nonlinear programming problems, which are solved by commercial nonlinear programming software BARON via GAMS. Numerical experiments show the benefits (e.g., expected profit gain) of using stochastic models versus deterministic models

Oviduct-Specific Expression of Human Neutrophil Defensin 4 in Lentivirally Generated Transgenic Chickens

Science.gov (United States)

Liu, Tongxin; Wu, Hanyu; Cao, Dainan; Li, Qingyuan; Zhang, Yaqiong; Li, Ning; Hu, Xiaoxiang

2015-01-01

The expression of oviduct-specific recombinant proteins in transgenic chickens is a promising technology for the production of therapeutic biologics in eggs. In this study, we constructed a lentiviral vector encoding an expression cassette for human neutrophil defensin 4 (HNP4), a compound that displays high activity against Escherichia coli, and produced transgenic chickens that expressed the recombinant HNP4 protein in egg whites. After the antimicrobial activity of the recombinant HNP4 protein was tested at the cellular level, a 2.8-kb ovalbumin promoter was used to drive HNP4 expression specifically in oviduct tissues. From 669 injected eggs, 218 chickens were successfully hatched. Ten G0 roosters, with semens identified as positive for the transgene, were mated with wild-type hens to generate G1 chickens. From 1,274 total offspring, fifteen G1 transgenic chickens were positive for the transgene, which was confirmed by PCR and Southern blotting. The results of the Southern blotting and genome walking indicated that a single copy of the HNP4 gene was integrated into chromosomes 1, 2, 3, 4, 6 and 24 of the chickens. As expected, HNP4 expression was restricted to the oviduct tissues, and the levels of both transcriptional and translational HNP4 expression varied greatly in transgenic chickens with different transgene insertion sites. The amount of HNP4 protein expressed in the eggs of G1 and G2 heterozygous transgenic chickens ranged from 1.65 μg/ml to 10.18 μg/ml. These results indicated that the production of transgenic chickens that expressed HNP4 protein in egg whites was successful. PMID:26020529

Analysis of transgenic wheat (Triticum aestivum L.) harboring a maize (Zea mays L.) gene for plastid EF-Tu: segregation pattern, expression and effects of the transgene.

Science.gov (United States)

Fu, Jianming; Ristic, Zoran

2010-06-01

We previously reported that transgenic wheat (Triticum aestivum L.) carrying a maize (Zea mays L.) gene (Zmeftu1) for chloroplast protein synthesis elongation factor, EF-Tu, displays reduced thermal aggregation of leaf proteins, reduced injury to photosynthetic membranes (thylakoids), and enhanced rate of CO(2) fixation following exposure to heat stress (18 h at 45 degrees C) [Fu et al. in Plant Mol Biol 68:277-288, 2008]. In the current study, we investigated the segregation pattern and expression of the transgene Zmeftu1 and determined the grain yield of transgenic plants after exposure to a brief heat stress (18 h at 45 degrees C). We also assessed thermal aggregation of soluble leaf proteins in transgenic plants, testing the hypothesis that increased levels of EF-Tu will lead to a non-specific protection of leaf proteins against thermal aggregation. The transgenic wheat displayed a single-gene pattern of segregation of Zmeftu1. Zmeftu1 was expressed, and the transgenic plants synthesized and accumulated three anti-EF-Tu cross-reacting polypeptides of similar molecular mass but different pI, suggesting the possibility of posttranslational modification of this protein. The transgenic plants also showed better grain yield after exposure to heat stress compared with their non-transgenic counterparts. Soluble leaf proteins of various molecular masses displayed lower thermal aggregation in transgenic than in non-transgenic wheat. The results suggest that overexpression of chloroplast EF-Tu can be beneficial to wheat tolerance to heat stress. Moreover, the results also support the hypothesis that EF-Tu contributes to heat tolerance by acting as a molecular chaperone and protecting heat-labile proteins from thermal aggregation in a non-specific manner.

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

Lifescience Database Archive (English)

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

Stable large-scale CO2 storage in defiance of an energy system based on renewable energy - Modelling the impact of varying CO2 injection rates on reservoir behavior

Science.gov (United States)

Bannach, Andreas; Hauer, Rene; Martin, Streibel; Stienstra, Gerard; Kühn, Michael

2015-04-01

The IPCC Report 2014 strengthens the need for CO2 storage as part of CCS or BECCS to reach ambitious climate goals despite growing energy demand in the future. The further expansion of renewable energy sources is a second major pillar. As it is today in Germany the weather becomes the controlling factor for electricity production by fossil fuelled power plants which lead to significant fluctuations of CO2-emissions which can be traced in injection rates if the CO2 were captured and stored. To analyse the impact of such changing injection rates on a CO2 storage reservoir. two reservoir simulation models are applied: a. An (smaller) reservoir model approved by gas storage activities for decades, to investigate the dynamic effects in the early stage of storage filling (initial aquifer displacement). b. An anticline structure big enough to accommodate a total amount of ≥ 100 Mega tons CO2 to investigate the dynamic effects for the entire operational life time of the storage under particular consideration of very high filling levels (highest aquifer compression). Therefore a reservoir model was generated. The defined yearly injection rate schedule is based on a study performed on behalf of IZ Klima (DNV GL, 2014). According to this study the exclusive consideration of a pool of coal-fired power plants causes the most intensive dynamically changing CO2 emissions and hence accounts for variations of a system which includes industry driven CO2 production. Besides short-term changes (daily & weekly cycles) seasonal influences are also taken into account. Simulation runs cover a variation of injection points (well locations at the top vs. locations at the flank of the structure) and some other largely unknown reservoir parameters as aquifer size and aquifer mobility. Simulation of a 20 year storage operation is followed by a post-operational shut-in phase which covers approximately 500 years to assess possible effects of changing injection rates on the long-term reservoir

Construction and characterization of cDNA library for IRM-2 mice

International Nuclear Information System (INIS)

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

2010-01-01

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

Regional-scale brine migration along vertical pathways due to CO2 injection - Part 1: The participatory modeling approach

Science.gov (United States)

Scheer, Dirk; Konrad, Wilfried; Class, Holger; Kissinger, Alexander; Knopf, Stefan; Noack, Vera

2017-06-01

Saltwater intrusion into potential drinking water aquifers due to the injection of CO2 into deep saline aquifers is one of the potential hazards associated with the geological storage of CO2. Thus, in a site selection process, models for predicting the fate of the displaced brine are required, for example, for a risk assessment or the optimization of pressure management concepts. From the very beginning, this research on brine migration aimed at involving expert and stakeholder knowledge and assessment in simulating the impacts of injecting CO2 into deep saline aquifers by means of a participatory modeling process. The involvement exercise made use of two approaches. First, guideline-based interviews were carried out, aiming at eliciting expert and stakeholder knowledge and assessments of geological structures and mechanisms affecting CO2-induced brine migration. Second, a stakeholder workshop including the World Café format yielded evaluations and judgments of the numerical modeling approach, scenario selection, and preliminary simulation results. The participatory modeling approach gained several results covering brine migration in general, the geological model sketch, scenario development, and the review of the preliminary simulation results. These results were included in revised versions of both the geological model and the numerical model, helping to improve the analysis of regional-scale brine migration along vertical pathways due to CO2 injection.

RNA Sequencing Identifies Upregulated Kyphoscoliosis Peptidase and Phosphatidic Acid Signaling Pathways in Muscle Hypertrophy Generated by Transgenic Expression of Myostatin Propeptide

Directory of Open Access Journals (Sweden)

Yuanxin Miao

2015-04-01

Full Text Available Myostatin (MSTN, a member of the transforming growth factor-β superfamily, plays a crucial negative role in muscle growth. MSTN mutations or inhibitions can dramatically increase muscle mass in most mammal species. Previously, we generated a transgenic mouse model of muscle hypertrophy via the transgenic expression of the MSTN N-terminal propeptide cDNA under the control of the skeletal muscle-specific MLC1 promoter. Here, we compare the mRNA profiles between transgenic mice and wild-type littermate controls with a high-throughput RNA sequencing method. The results show that 132 genes were significantly differentially expressed between transgenic mice and wild-type control mice; 97 of these genes were up-regulated, and 35 genes were down-regulated in the skeletal muscle. Several genes that had not been reported to be involved in muscle hypertrophy were identified, including up-regulated myosin binding protein H (mybph, and zinc metallopeptidase STE24 (Zmpste24. In addition, kyphoscoliosis peptidase (Ky, which plays a vital role in muscle growth, was also up-regulated in the transgenic mice. Interestingly, a pathway analysis based on grouping the differentially expressed genes uncovered that cardiomyopathy-related pathways and phosphatidic acid (PA pathways (Dgki, Dgkz, Plcd4 were up-regulated. Increased PA signaling may increase mTOR signaling, resulting in skeletal muscle growth. The findings of the RNA sequencing analysis help to understand the molecular mechanisms of muscle hypertrophy caused by MSTN inhibition.

RNA sequencing identifies upregulated kyphoscoliosis peptidase and phosphatidic acid signaling pathways in muscle hypertrophy generated by transgenic expression of myostatin propeptide.

Science.gov (United States)

Miao, Yuanxin; Yang, Jinzeng; Xu, Zhong; Jing, Lu; Zhao, Shuhong; Li, Xinyun

2015-04-09

Myostatin (MSTN), a member of the transforming growth factor-β superfamily, plays a crucial negative role in muscle growth. MSTN mutations or inhibitions can dramatically increase muscle mass in most mammal species. Previously, we generated a transgenic mouse model of muscle hypertrophy via the transgenic expression of the MSTN N-terminal propeptide cDNA under the control of the skeletal muscle-specific MLC1 promoter. Here, we compare the mRNA profiles between transgenic mice and wild-type littermate controls with a high-throughput RNA sequencing method. The results show that 132 genes were significantly differentially expressed between transgenic mice and wild-type control mice; 97 of these genes were up-regulated, and 35 genes were down-regulated in the skeletal muscle. Several genes that had not been reported to be involved in muscle hypertrophy were identified, including up-regulated myosin binding protein H (mybph), and zinc metallopeptidase STE24 (Zmpste24). In addition, kyphoscoliosis peptidase (Ky), which plays a vital role in muscle growth, was also up-regulated in the transgenic mice. Interestingly, a pathway analysis based on grouping the differentially expressed genes uncovered that cardiomyopathy-related pathways and phosphatidic acid (PA) pathways (Dgki, Dgkz, Plcd4) were up-regulated. Increased PA signaling may increase mTOR signaling, resulting in skeletal muscle growth. The findings of the RNA sequencing analysis help to understand the molecular mechanisms of muscle hypertrophy caused by MSTN inhibition.

76 FR 56982 - Announcement of Federal Underground Injection Control (UIC) Class VI Program for Carbon Dioxide (CO2

Science.gov (United States)

2011-09-15

...-9465-1] Announcement of Federal Underground Injection Control (UIC) Class VI Program for Carbon Dioxide... Injection Control (UIC) Class VI Program for Carbon Dioxide (CO 2 ) Geologic Sequestration (GS) Wells under... highlighted in the ``Report of the Interagency Task Force on Carbon Capture and Storage'' (August 2010), it is...

Comparison of intravenous and intraperitoneal [{sup 123}I]IBZM injection for dopamine D2 receptor imaging in mice

Energy Technology Data Exchange (ETDEWEB)

Meyer, Philipp T. [Department of Neurology, University Hospital Aachen, 52074 Aachen (Germany); Department of Nuclear Medicine, University Hospital Aachen, 52074 Aachen (Germany)], E-mail: pmeyer@ukaachen.de; Salber, Dagmar [C. and O. Vogt Institute of Brain Research, University Hospital Duesseldorf, 40225 Duesseldorf (Germany); Schiefer, Johannes [Department of Neurology, University Hospital Aachen, 52074 Aachen (Germany); Cremer, Markus [Institute of Neurosciences and Biophysics - Medicine, Research Center Juelich, 52425 Juelich (Germany); Schaefer, Wolfgang M. [Department of Nuclear Medicine, University Hospital Aachen, 52074 Aachen (Germany); Kosinski, Christoph M. [Department of Neurology, University Hospital Aachen, 52074 Aachen (Germany); Langen, Karl-Josef [Institute of Neurosciences and Biophysics - Medicine, Research Center Juelich, 52425 Juelich (Germany)

2008-07-15

Introduction: Intraperitoneal (IP) injection represents an attractive alternative route of radiotracer administration for small animal imaging, e.g., for longitudinal studies in transgenic mouse models. We explored the cerebral kinetics of the reversible dopamine D2 receptor ligand [{sup 123}I]IBZM after IP injection in mice. Methods: Cerebral [{sup 123}I]IBZM kinetics were assessed by ex vivo autoradiography in mice sacrificed between 30 and 200 min after IP or intravenous (IV) injection. The striatum-to-cerebellum (S/C) uptake ratio at 140 min was evaluated in wild-type mice and R6/2 transgenic mice (a Huntington's disease model) in comparison with in vitro autoradiography using [{sup 3}H]raclopride. Results: [{sup 123}I]IBZM uptake was slower and lower after IP injection [maximum uptake in striatum 5.6% injected dose per gram (ID/g) at 60 min] than IV injection (10.5%ID/g at 30 min). Between 60 and 120 min, striatal (cerebellar) uptake after IP injection reached 63% (91%) of the uptake after IV injection. The S/C uptake ratio increased to 15.5 at 200 min after IP injection, which corresponds to 87% of the IV injection value (17.8). Consistent with in vitro [{sup 3}H]raclopride autoradiography, the S/C ratio given by ex vivo [{sup 123}I]IBZM autoradiography (140 min after IP injection) was significantly reduced in R6/2 mice. Conclusions: Although IP injection resulted in slower kinetics, relevant measures of dopamine D2 receptor availability were comparable. Thus, IP injection represents a promising route of tracer administration for small animal [{sup 123}I]IBZM SPECT. This should considerably simplify the implementation of longitudinal small animal neuroimaging studies, e.g., in transgenic mouse models.

Physicochemical effects of discrete CO2-SO2 mixtures on injection and storage in a sandstone aquifer

NARCIS (Netherlands)

Waldmann, S.; Hofstee, C.; Koenen, M.; Loeve, D.

2016-01-01

Geological storage of captured CO2, which typically will contain certain amounts of impurities, in salineaquifers is of potential to reduce greenhouse gas emissions into the atmosphere. The co-injection of theimpurity SO2has an effect on the chemical reactivity of the fluid and solid phases as well

An AANAT/ASMT transgenic animal model constructed with CRISPR/Cas9 system serving as the mammary gland bioreactor to produce melatonin-enriched milk in sheep.

Science.gov (United States)

Ma, Teng; Tao, Jingli; Yang, Minghui; He, Changjiu; Tian, Xiuzhi; Zhang, Xiaosheng; Zhang, Jinlong; Deng, Shoulong; Feng, Jianzhong; Zhang, Zhenzhen; Wang, Jing; Ji, Pengyun; Song, Yukun; He, Pingli; Han, Hongbing; Fu, Juncai; Lian, Zhengxing; Liu, Guoshi

2017-08-01

Melatonin as a potent antioxidant exhibits important nutritional and medicinal values. To produce melatonin-enriched milk will benefit the consumers. In this study, a sheep bioreactor which generates melatonin-enriched milk has been successfully developed by the technology that combined CRISPR/Cas9 system and microinjection. The AANAT and ASMT were cloned from pineal gland of Dorper sheep (Ovis aries). The in vitro studies found that AANAT and ASMT were successfully transferred to the mammary epithelial cell lines and significantly increased melatonin production in the culture medium compared to the nontransgenic cell lines. In addition, the Cas9 mRNA, sgRNA, and the linearized vectors pBC1-AANAT and pBC1-ASMT were co-injected into the cytoplasm of pronuclear embryos which were implanted into ewes by oviducts transferring. Thirty-four transgenic sheep were generated with the transgenic positive rate being roughly 35% which were identified by Southern blot and sequencing. Seven carried transgenic AANAT, two carried ASMT, and 25 carried both of AANAT and ASMT genes. RT-PCR and Western blot demonstrated that the lambs expressed these genes in their mammary epithelial cells and these animals produced melatonin-enriched milk. This is the first report to show a functional AANAT and ASMT transgenic animal model which produce significantly high levels of melatonin milk compared to their wild-type counterparts. The advanced technologies used in the study laid a foundation for generating large transgenic livestock, for example, the cows, which can produce high level of melatonin milk. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Present-day stress analysis in the St. Lawrence Lowlands from borehole breakouts and implications for Co2 injection

Energy Technology Data Exchange (ETDEWEB)

Konstantinovskaya, E.; Malo, M. [Institut national de la recherche scientifique, Centre Eau, Terre et Environnement (INRS-ETE), 490, de la Couronne, Quebec, G1K 9A9 (Canada)], email: Elena.Konstantinovskaya@ete.inrs.ca; Castillo, D.A.; Hughes, Baker [Reservoir Development Services, GeoMechanics, 5373 West Alabama Street, Suite 300, Houston, Texas, 77056 (United States)], email: David.Castillo@BakerHughes.com

2011-07-01

An essential part of reservoir-geomechanical study in the St. Lawrence Lowlands, Quebec, consists of evaluating the geomechanical response of the rock and caprock of the reservoir to different injection scenarios and the long-term storage of CO2. The first phase of this study consists of determining current stress direction. The next step consists of assessing the potential shear failure or reactivation of pre-existing faults and fracture sets caused by changes in the reservoir pressure because of CO2 injection. According to study results, the orientation of the average maximum horizontal stress direction SHmax obtained from stress-induced wellbore borehole breakouts in this area is N59 degree E plus or minus 20 degree. The wellbore breakouts in 17 wells go from 250 m to 4 km depth. A major purpose of this geomechanical study is to estimate to what extent sustainable fluid pressures for CO2 injection can be increased without causing fracturing, faulting, or reactivation of pre-existing faults. To this end, one must determine prevailing stresses (directions and magnitudes), fault geometries and rock strengths.

Plant breeding by using radiation mutation - Development of radiation indicator plants by molecular breeding

Energy Technology Data Exchange (ETDEWEB)

Liu, Jang Ryol; Kwak, Sang Soo; Kwon, Seok Yoon [Korea Research Institute of Bioscience and Biotechnology, Taejon (Korea)

2000-04-01

- tSOD1, cytosolic CuZnSOD cDNA was cloned from tobacco cDNA library by PCR. To develop the under-producing the transgenic plants, the vectors were constructed using by antisense and co-supressing technology. The transgenic tobacco plants were confirmed that over 60% of kanamycin-resistant plants were introduced the foreign gene by PCR and transformed one copy through Southern blot analysis. - In an attempt to identify marker genes for gamma irradiation of plants, expression patterns of diverse genes upon gamma irradiation of young tobacco plants were investigated. With the knowledge of distinctive expression patterns of diverse genes, irradiation-indicating marker plants could be developed by engineering and monitoring multiple radiation-responsive genes. Additionally, a gamma irradiation-responsive NtTMK1 receptor-like kinase gene was molecular biologically characterized. -Uranium reductase gene (Cytochrome C3) and radiation resistance gene (recA) have been cloned from Desulfovibrio and Deinococcus radiodurans. -Two plant transformation vectors (pCYC3 and pDrecA) have been constructed. - Tobacco transgenic plants of have been obtained. 52 refs., 5 figs. (Author)

Characterization of the porcine carboxypeptidase E cDNA

DEFF Research Database (Denmark)

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

2007-01-01

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

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

Energy Technology Data Exchange (ETDEWEB)

NONE

2001-03-01

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

Generation and characterization of human heme oxygenase-1 transgenic pigs.

Directory of Open Access Journals (Sweden)

Hye-Jung Yeom

Full Text Available Xenotransplantation using transgenic pigs as an organ source is a promising strategy to overcome shortage of human organ for transplantation. Various genetic modifications have been tried to ameliorate xenograft rejection. In the present study we assessed effect of transgenic expression of human heme oxygenase-1 (hHO-1, an inducible protein capable of cytoprotection by scavenging reactive oxygen species and preventing apoptosis caused by cellular stress during inflammatory processes, in neonatal porcine islet-like cluster cells (NPCCs. Transduction of NPCCs with adenovirus containing hHO-1 gene significantly reduced apoptosis compared with the GFP-expressing adenovirus control after treatment with either hydrogen peroxide or hTNF-α and cycloheximide. These protective effects were diminished by co-treatment of hHO-1 antagonist, Zinc protoporphyrin IX. We also generated transgenic pigs expressing hHO-1 and analyzed expression and function of the transgene. Human HO-1 was expressed in most tissues, including the heart, kidney, lung, pancreas, spleen and skin, however, expression levels and patterns of the hHO-1 gene are not consistent in each organ. We isolate fibroblast from transgenic pigs to analyze protective effect of the hHO-1. As expected, fibroblasts derived from the hHO-1 transgenic pigs were significantly resistant to both hydrogen peroxide damage and hTNF-α and cycloheximide-mediated apoptosis when compared with wild-type fibroblasts. Furthermore, induction of RANTES in response to hTNF-α or LPS was significantly decreased in fibroblasts obtained from the hHO-1 transgenic pigs. These findings suggest that transgenic expression of hHO-1 can protect xenografts when exposed to oxidative stresses, especially from ischemia/reperfusion injury, and/or acute rejection mediated by cytokines. Accordingly, hHO-1 could be an important candidate molecule in a multi-transgenic pig strategy for xenotransplantation.

Generation and characterization of human heme oxygenase-1 transgenic pigs.

Science.gov (United States)

Yeom, Hye-Jung; Koo, Ok Jae; Yang, Jaeseok; Cho, Bumrae; Hwang, Jong-Ik; Park, Sol Ji; Hurh, Sunghoon; Kim, Hwajung; Lee, Eun Mi; Ro, Han; Kang, Jung Taek; Kim, Su Jin; Won, Jae-Kyung; O'Connell, Philip J; Kim, Hyunil; Surh, Charles D; Lee, Byeong-Chun; Ahn, Curie

2012-01-01

Xenotransplantation using transgenic pigs as an organ source is a promising strategy to overcome shortage of human organ for transplantation. Various genetic modifications have been tried to ameliorate xenograft rejection. In the present study we assessed effect of transgenic expression of human heme oxygenase-1 (hHO-1), an inducible protein capable of cytoprotection by scavenging reactive oxygen species and preventing apoptosis caused by cellular stress during inflammatory processes, in neonatal porcine islet-like cluster cells (NPCCs). Transduction of NPCCs with adenovirus containing hHO-1 gene significantly reduced apoptosis compared with the GFP-expressing adenovirus control after treatment with either hydrogen peroxide or hTNF-α and cycloheximide. These protective effects were diminished by co-treatment of hHO-1 antagonist, Zinc protoporphyrin IX. We also generated transgenic pigs expressing hHO-1 and analyzed expression and function of the transgene. Human HO-1 was expressed in most tissues, including the heart, kidney, lung, pancreas, spleen and skin, however, expression levels and patterns of the hHO-1 gene are not consistent in each organ. We isolate fibroblast from transgenic pigs to analyze protective effect of the hHO-1. As expected, fibroblasts derived from the hHO-1 transgenic pigs were significantly resistant to both hydrogen peroxide damage and hTNF-α and cycloheximide-mediated apoptosis when compared with wild-type fibroblasts. Furthermore, induction of RANTES in response to hTNF-α or LPS was significantly decreased in fibroblasts obtained from the hHO-1 transgenic pigs. These findings suggest that transgenic expression of hHO-1 can protect xenografts when exposed to oxidative stresses, especially from ischemia/reperfusion injury, and/or acute rejection mediated by cytokines. Accordingly, hHO-1 could be an important candidate molecule in a multi-transgenic pig strategy for xenotransplantation.

Enhancer-promoter interference and its prevention in transgenic plants

Science.gov (United States)

Transcriptional enhancer elements have been shown to override the specificity of nearby promoters in a position- and orientation-independent manner. This is problematic when multiple enhancers/promoters co-exist within a single transgenic construct as it has the potential to cause the mis-expressio...

Simulation of a Potential CO2 Storage in the West Paris Basin: Site Characterization and Assessment of the Long-Term Hydrodynamical and Geochemical Impacts Induced by the CO2 Injection

Directory of Open Access Journals (Sweden)

Estublier Audrey

2017-07-01

Full Text Available This article presents the preliminary results of a study carried out as part of a demonstration project of CO2 storage in the Paris Basin. This project funded by ADEME (French Environment and Energy Management Agency and several industrial partners (TOTAL, ENGIE, EDF, Lafarge, Air Liquide, Vallourec aimed to study the possibility to set up an experimental infrastructure of CO2 transport and storage. Regarding the storage, the objectives were: (1 to characterize the selected site by optimizing the number of wells in a CO2 injection case of 200 Mt over 50 years in the Trias, (2 to simulate over time the CO2 migration and the induced pressure field, and (3 to analyze the geochemical behavior of the rock over the long term (1,000 years. The preliminary site characterization study revealed that only the southern area of Keuper succeeds to satisfy this injection criterion using only four injectors. However, a complementary study based on a refined fluid flow model with additional secondary faults concluded that this zone presents the highest potential of CO2 injection but without reaching the objective of 200 Mt with a reasonable number of wells. The simulation of the base scenario, carried out before the model refinement, showed that the overpressure above 0.1 MPa covers an area of 51,869 km2 in the Chaunoy formation, 1,000 years after the end of the injection, which corresponds to the whole West Paris Basin, whereas the CO2 plume extension remains small (524 km2. This overpressure causes brine flows at the domain boundaries and a local overpressure in the studied oil fields. Regarding the preliminary risk analysis of this project, the geochemical effects induced by the CO2 injection were studied by simulating the fluid-rock interactions with a coupled geochemical and fluid flow model in a domain limited to the storage complex. A one-way coupling of two models based on two domains fitting into each other was developed using dynamic boundary

Study of the reactive processes during CO2 injection into sedimentary reservoirs: Experimental quantification of the processes at meso-scale

International Nuclear Information System (INIS)

Luquot, L.

2008-11-01

In order to minimize CO 2 atmospheric concentration, a solution consists in sequestrating CO 2 in geological reservoirs. To estimate long term risks, it is necessary to quantify the couplings between reaction processes as well as structural and hydrodynamical modifications. We realised two experimental benches enabling injecting CO 2 -enriched-brine in conditions corresponding to in situ storage (T ≤ 200 C and P ≤ 200 bar) and developed an experimental protocol using X-Ray microtomography and fluid and rock analyses in order to measure the variations of physical and chemical parameters. The study of carbonated reservoirs near the injection well, allows quantifying different k-phi relationships depending on the dissolution processes and triggered by the local fluid chemical composition and initials conditions. Away from the injection well, we observe carbonate precipitation decreasing the permeability. The study of fractured cap-rock samples shows that alternative percolation of CO 2 -enriched-brine and CO 2 gas increases the fracture permeability. The study of silicated rocks indicates carbonate precipitation in zeolite sandstone and sintered dunite grains. Nevertheless, in zeolite sandstone we also observe the precipitation of clay particles located in the fluid pathways which decrease strongly the permeability. (author)

Discussion of the influence of CO and CH4 in CO2 transport, injection, and storage for CCS technology.

Science.gov (United States)

Blanco, Sofía T; Rivas, Clara; Bravo, Ramón; Fernández, Javier; Artal, Manuela; Velasco, Inmaculada

2014-09-16

This paper discusses the influence of the noncondensable impurities CO and CH4 on Carbon Capture and Storage (CCS) technology. We calculated and drew conclusions about the impact of both impurities in the CO2 on selected transport, injection, and storage parameters (pipeline pressure drop, storage capacity, etc.), whose analysis is necessary for the safe construction and operation of CO2 pipelines and for the secure long-term geological storage of anthropogenic CO2. To calculate these parameters, it is necessary to acquire data on the volumetric properties and the vapor-liquid equilibrium of the fluid being subjected to CCS. In addition to literature data, we used new experimental data, which are presented here and were obtained for five mixtures of CO2+CO with compositions characteristic of the typical emissions of the E.U. and the U.S.A. Temperatures and pressures are based on relevant CO2 pipeline and geological storage site values. From our experimental results, Peng-Robinson, PC-SAFT, and GERG Equations of State for were validated CO2+CO under the conditions of CCS. We conclude that the concentration of both impurities strongly affects the studied parameters, with CO being the most influential and problematic. The overall result of these negative effects is an increase in the difficulties, risks, and overall costs of CCS.

Geologic Carbon Sequestration: Mitigating Climate Change by Injecting CO2 Underground (LBNL Summer Lecture Series)

Energy Technology Data Exchange (ETDEWEB)

Oldenburg, Curtis M. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Earth Sciences Division

2009-07-21

Summer Lecture Series 2009: Climate change provides strong motivation to reduce CO2 emissions from the burning of fossil fuels. Carbon dioxide capture and storage involves the capture, compression, and transport of CO2 to geologically favorable areas, where its injected into porous rock more than one kilometer underground for permanent storage. Oldenburg, who heads Berkeley Labs Geologic Carbon Sequestration Program, will focus on the challenges, opportunities, and research needs of this innovative technology.

Impact of supercritical CO2 injection on petrophysical and rock mechanics properties of chalk: an experimental study on chalk from South Arne field, North Sea

DEFF Research Database (Denmark)

Alam, Mohammad Monzurul; Hjuler, Morten Leth; Christensen, Helle Foged

2011-01-01

Changes in chalk due to EOR by injecting supercritical CO2 (CO2-EOR) can ideally be predicted by applying geophysical methods designed from laboratory-determined petrophysical and rock mechanics properties. A series of petrophysical and rock mechanics tests were performed on Ekofisk Formation...... and Tor Formation chalk of the South Arne field to reveal the changes in petrophysical and rock mechanics properties of chalk due to the injection of CO2 at supercritical state. An increase in porosity and decrease in specific surface was observed due to injection of supercritical CO2. This indicates...... as indicated by NMR T2 relaxation time was observed. Rock mechanics testing indicates that in 30% porosity chalk from the South Arne field, injection of supercritical CO2 has no significant effect on shear strength and compaction properties, while there is probably a slight decrease in stiffness properties...

Experimental and numerical study on the fracture of rocks during injection of CO2-saturated water

Science.gov (United States)

Li, Qi; Wu, Zhishen; Lei, Xing-Lin; Murakami, Yutaka; Satoh, Takashi

2007-02-01

Geological sequestration of CO2 into depleted hydrocarbon reserviors or saline aquifers presents the enormous potential to reduce greenhouse gas emission from fossil fuels. However, it may give rise to a complicated coupling physical and chemical process. One of the processes is the hydro-mechanical impact of CO2 injection. During the injection project, the increase of pore pressures of storing formations can induce the instability, which finally results in a catastrophic failure of disposal sites. This paper focuses mainly on the role of CO2-saturated water in the fracturing behavior of rocks. To investigate how much the dissolved CO2 can influence the pore pressure change of rocks, acoustic emission (AE) experiments were performed on sandstone and granite samples under triaxial conditions. The main innovation of this paper is to propose a time dependent porosity method to simulate the abrupt failure process, which is observed in the laboratory and induced by the pore pressure change due to the volume dilatancy of rocks, using a finite element scheme associated with two-phase characteristics. The results successfully explained the phenomena obtained in the physical experiments.

Flexible CO2 laser and submucosal gel injection for safe endoluminal resection in the intestines.

Science.gov (United States)

Au, Joyce T; Mittra, Arjun; Wong, Joyce; Carpenter, Susanne; Carson, Joshua; Haddad, Dana; Monette, Sebastien; Ezell, Paula; Patel, Snehal; Fong, Yuman

2012-01-01

The CO(2) laser's unique wavelength of 10.6 μm has the advantage of being readily absorbed by water but historically limited it to line-of-sight procedures. Through recent technological advances, a flexible CO(2) laser fiber has been developed and holds promise for endoluminal surgery. We examined whether this laser, along with injection of a water-based gel in the submucosal space, will allow safe dissection of the intestines and enhance the potential of this tool for minimally invasive surgery. Using an ex vivo model with porcine intestines, spot ablation was performed with the flexible CO(2) laser at different power settings until transmural perforation. Additionally, excisions of mucosal patches were performed by submucosal dissection with and without submucosal injection of a water-based gel. With spot ablation at 5 W, none of the specimens was perforated by 5 min, which was the maximum recorded time. The time to perforation was significantly shorter with increased laser power, and gel pretreatment protected the intestines against spot ablation, increasing the time to perforation from 6 to 37 s at 10 W and from 1 to 7 s at 15 W. During excision of mucosal patches, 56 and 83% of untreated intestines perforated at 5 and 10 W, respectively. Gel pretreatment prior to excision protected all intestines against perforation. These specimens were verified to be intact by inflation with air to over 100 mmHg. Furthermore, excision of the mucosal patch was complete in gel-pretreated specimens, whereas 22% of untreated specimens had residual islands of mucosa after excision. The flexible CO(2) laser holds promise as a precise dissection and cutting tool for endoluminal surgery of the intestines. Pretreatment with a submucosal injection of a water-based gel protects the intestines from perforation during ablation and mucosal dissection.

cDNA microarray screening in food safety

International Nuclear Information System (INIS)

Roy, Sashwati; Sen, Chandan K.

2006-01-01

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

[TSA improve transgenic porcine cloned embryo development and transgene expression].

Science.gov (United States)

Kong, Qing-Ran; Zhu, Jiang; Huang, Bo; Huan, Yan-Jun; Wang, Feng; Shi, Yong-Qian; Liu, Zhong-Feng; Wu, Mei-Ling; Liu, Zhong-Hua

2011-07-01

Uncompleted epigenetic reprogramming is attributed to the low efficiency of producing transgenic cloned animals. Histone modification associated with epigenetics can directly influence the embryo development and transgene expression. Trichostatin A (TSA), as an inhibitor of histone deacetylase, can change the status of histone acetylation, improve somatic cell reprogramming, and enhance cloning efficiency. TSA prevents the chromatin structure from being condensed, so that transcription factor could binds to DNA sequence easily and enhance transgene expression. Our study established the optimal TSA treatment on porcine donor cells and cloned embryos, 250 nmol/L, 24 h and 40 nmol/L, 24 h, respectively. Furthermore, we found that both the cloned embryo and the donor cell treated by TSA resulted in the highest development efficiency. Meanwhile, TSA can improve transgene expression in donor cell and cloned embryo. In summary, TSA can significantly improve porcine reconstructed embryo development and transgene expression.

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

African Journals Online (AJOL)

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

Spin injection from Co2MnGa into an InGaAs quantum well

DEFF Research Database (Denmark)

Hickey, M. C.; Damsgaard, Christian Danvad; Holmes, S. N.

2008-01-01

We have demonstrated spin injection from a full Heusler alloy Co2MnGa thin film into a (100) InGaAs quantum well in a semiconductor light-emitting diode structure at a temperature of 5 K. The detection is performed in the oblique Hanle geometry, allowing quantification of the effective spin lifet...

An overexpression of chalcone reductase of Pueraria montana var. lobata alters biosynthesis of anthocyanin and 5'-deoxyflavonoids in transgenic tobacco.

Science.gov (United States)

Joung, Jae-youl; Kasthuri, G Mangai; Park, Ji-young; Kang, Won-jin; Kim, Hyun-soon; Yoon, Bong-sik; Joung, Hyouk; Jeon, Jae-heung

2003-03-28

We isolated the chalcone reductase (pl-chr) gene of Pueraria montana var. lobata by using a PCR strategy from cDNA pools of storage roots. A high level of expression of RNA was found in both stems and roots. The genomic Southern blot result suggests that pl-chr exists as a member of a small gene family. By introducing a pl-chr gene under the control of the 35S CaMV promoter into the pink-flowering Xanthi line of Nicotiana tabacum, the flower color was changed from pink to white-to-pink. The contents of anthocyanin in the flowers of the transgenic lines were dramatically decreased by 40%, but the total UV absorption compounds remained unchanged. The production of liquiritigenin in pl-chr overexpressed transgenic tobacco lines was confirmed by HPLC and MS analysis. The introduction of pl-chr gene provides a method to redirect the flavonoid pathway into 5'-deoxyflavonoid production in non-legume crops, in order to manipulate the phenylpropanoid pathway for isoflavonoid production.

Overexpression of CaDSR6 increases tolerance to drought and salt stresses in transgenic Arabidopsis plants.

Science.gov (United States)

Kim, Eun Yu; Seo, Young Sam; Park, Ki Youl; Kim, Soo Jin; Kim, Woo Taek

2014-11-15

The partial CaDSR6 (Capsicum annuum Drought Stress Responsive 6) cDNA was previously identified as a drought-induced gene in hot pepper root tissues. However, the cellular role of CaDSR6 with regard to drought stress tolerance was unknown. In this report, full-length CaDSR6 cDNA was isolated. The deduced CaDSR6 protein was composed of 234 amino acids and contained an approximately 30 amino acid-long Asp-rich domain in its central region. This Asp-rich domain was highly conserved in all plant DSR6 homologs identified and shared a sequence identity with the N-terminal regions of yeast p23(fyp) and human hTCTP, which contain Rab protein binding sites. Transgenic Arabidopsis plants overexpressing CaDSR6 (35S:CaDSR6-sGFP) were tolerant to high salinity, as identified by more vigorous root growth and higher levels of total chlorophyll than wild type plants. CaDSR6-overexpressors were also more tolerant to drought stress compared to wild type plants. The 35S:CaDSR6-sGFP leaves retained their water content and chlorophyll more efficiently than wild type leaves in response to dehydration stress. The expression of drought-induced marker genes, such as RD20, RD22, RD26, RD29A, RD29B, RAB18, KIN2, ABF3, and ABI5, was markedly increased in CaDSR6-overexpressing plants relative to wild type plants under both normal and drought conditions. These results suggest that overexpression of CaDSR6 is associated with increased levels of stress-induced genes, which, in turn, conferred a drought tolerant phenotype in transgenic Arabidopsis plants. Overall, our data suggest that CaDSR6 plays a positive role in the response to drought and salt stresses. Copyright © 2014 Elsevier B.V. All rights reserved.

Infectious Maize rayado fino virus from cloned cDNA

Science.gov (United States)

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

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

DEFF Research Database (Denmark)

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

1995-01-01

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

Transformation of pecan and regeneration of transgenic plants.

Science.gov (United States)

McGranahan, G H; Leslie, C A; Dandekar, A M; Uratsu, S L; Yates, I E

1993-09-01

A gene transfer system developed for walnut (Juglans regia L.) was successfully applied to pecan (Carya illinoensis [Wang] K. Koch). Repetitively embryogenic somatic embryos derived from open-pollinated seed of 'Elliott', 'Wichita', and 'Schley' were co-cultivated with Agrobacterium strain EHA 101/pCGN 7001, which contains marker genes for beta-glucuronidase activity and resistance to kanamycin. Several modifications of the standard walnut transformation techniques were tested, including a lower concentration of kanamycin and a modified induction medium, but these treatments had no measurable effect on efficiency of transformation. Nineteen of the 764 viable inoculated embryos produced transgenic subclones; 13 of these were from the line 'Elliott'6, 3 from 'Schley'5/3, and 3 from 'Wichita'9. Transgenic embryos of 'Wichita'9 germinated most readily and three subclones were successfully micropropagated. Three transgenic plants of one of these subclones were obtained by grafting the tissue cultured shoots to seedling pecan rootstock in the greenhouse. Gene insertion, initially detected by GUS activity, was confirmed by detection of integrated T-DNA sequences using Southern analysis.

TaSK5, an abiotic stress-inducible GSK3/shaggy-like kinase from wheat, confers salt and drought tolerance in transgenic Arabidopsis.

Science.gov (United States)

Christov, Nikolai Kirilov; Christova, Petya Koeva; Kato, Hideki; Liu, Yuelin; Sasaki, Kentaro; Imai, Ryozo

2014-11-01

A novel cold-inducible GSK3/shaggy-like kinase, TaSK5, was isolated from winter wheat using a macroarray-based differential screening approach. TaSK5 showed high similarity to Arabidopsis subgroup I GSK3/shaggy-like kinases ASK-alpha, AtSK-gamma and ASK-epsilon. RNA gel blot analyses revealed TaSK5 induction by cold and NaCl treatments and to a lesser extent by drought treatment. TaSK5 functionally complemented the cold- and salt-sensitive phenotypes of a yeast GSK3/shaggy-like kinase mutant, △mck1. Transgenic Arabidopsis plants overexpressing TaSK5 cDNA showed enhanced tolerance to salt and drought stresses. By contrast, the tolerance of the transgenic plants to freezing stress was not altered. Microarray analysis revealed that a number of abiotic stress-inducible genes were constitutively induced in the transgenic Arabidopsis plants, suggesting that TaSK5 may function in a novel signal transduction pathway that appears to be unrelated to DREB1/CBF regulon and may involve crosstalk between abiotic and hormonal signals. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

A rapid method for screening arrayed plasmid cDNA library by PCR

International Nuclear Information System (INIS)

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

1999-01-01

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

Rapid characterization of transgenic and non-transgenic soybean oils by chemometric methods using NIR spectroscopy

Science.gov (United States)

Luna, Aderval S.; da Silva, Arnaldo P.; Pinho, Jéssica S. A.; Ferré, Joan; Boqué, Ricard

Near infrared (NIR) spectroscopy and multivariate classification were applied to discriminate soybean oil samples into non-transgenic and transgenic. Principal Component Analysis (PCA) was applied to extract relevant features from the spectral data and to remove the anomalous samples. The best results were obtained when with Support Vectors Machine-Discriminant Analysis (SVM-DA) and Partial Least Squares-Discriminant Analysis (PLS-DA) after mean centering plus multiplicative scatter correction. For SVM-DA the percentage of successful classification was 100% for the training group and 100% and 90% in validation group for non transgenic and transgenic soybean oil samples respectively. For PLS-DA the percentage of successful classification was 95% and 100% in training group for non transgenic and transgenic soybean oil samples respectively and 100% and 80% in validation group for non transgenic and transgenic respectively. The results demonstrate that NIR spectroscopy can provide a rapid, nondestructive and reliable method to distinguish non-transgenic and transgenic soybean oils.

Effectiveness of hCMV, mEF1a and mAct promoters on driving of foreign gene expression in transgenic zebrafish

Directory of Open Access Journals (Sweden)

. Alimuddin

2007-01-01

Full Text Available Highly unsaturated fatty acids (HUFA, especially eicosapentaenoic acid (EPA and docosahexaenoic acid (DHA have long been recognized for its beneficial effect for human health and development.   The D6 fatty acid desaturase is generally considered to be the rate-limiting factor in HUFA biosynthesis.  Here, as the first step of study, we conducted experiment to select an appropriate construct that allows higher expression levels of masu salmon (Oncorhynchus masou D6-desaturase gene in zebrafish (Danio rerio in order to increase its activity for synthesizing EPA/DHA.  Salmon D6-desaturase cDNA (sD6 was separately ligated with human cytomegalovirus (hCMV, medaka elongation factor 1a (mEF1a and medaka b-actin (mAct promoters.  The resulted construct was designated as hCMV-sD6, mEF1a-sD6 and mAct-sD6, respectively.  Each of the constructs in circular DNA form was microinjected into 1-cell stage embryos at a concentration of 30mg/ml. Transgenic individuals were identified by polymerase chain reaction (PCR and their expression levels were analyzed by reverse transcription PCR.  The first (F1 and second (F2 generation was produced by crossing the transgenic founder F0 and F1, respectively, with wild-type fish.  The results showed that the highest transient gene expression level was obtained from the mAct-D6 construct, followed respectively by EF1a-D6 and hCMV-D6 construct. The transmission rate of transgene into F1 generation was 4.2%-44.1%, and into F2 was followed the Mendellian segregation pattern.   Expression of transgene in F2 generation was varied between strains regarding as the mosaics of F0 fish.  Now, a transgenic system to study the modification of fatty acid biosynthesis pathways in fish was established.  Further investigations are to produce fish containing higher levels of EPA and DHA. Keywords: desaturase, nutraceutical fatty acid, transgenic, zebrafish, masu salmon   Abstrak Promoter merupakan regulator yang menentukan

cDNA encoding a polypeptide including a hevein sequence

Energy Technology Data Exchange (ETDEWEB)

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

1993-02-16

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

Reactivity of hydrocarbons in response to injection of a CO2/O2 mixture under depleted reservoir conditions: experimental and numerical modeling

International Nuclear Information System (INIS)

Pacini-Petitjean, Claire

2015-01-01

The geological storage of CO 2 (CO 2 Capture-Storage - CCS) and the Enhanced Oil Recovery (EOR) by CO 2 injection into petroleum reservoirs could limit CO 2 atmospheric accumulation. However, CO 2 can be associated with oxygen. To predict the hydrocarbon evolution under these conditions involves the study of oxidation mechanisms. Oxidation experiment and kinetic detailed modeling were carried out with pure compounds. The comparison between experimental and modeling results led to the construction of a hydrocarbon oxidation kinetic model and emphasized the parameters leading to auto ignition. The good agreement between our experiments and modeling are promising for the development of a tool predicting the critical temperature leading to auto-ignition and the evolution of hydrocarbon composition, to estimate the stability of a petroleum system in CO 2 injection context. (author) [fr

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

Science.gov (United States)

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

2000-08-01

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

An improved method for RNA isolation and cDNA library construction from immature seeds of Jatropha curcas L

Directory of Open Access Journals (Sweden)

Kaur Jatinder

2010-05-01

Full Text Available Abstract Background RNA quality and quantity is sometimes unsuitable for cDNA library construction, from plant seeds rich in oil, polysaccharides and other secondary metabolites. Seeds of jatropha (Jatropha curcas L. are rich in fatty acids/lipids, storage proteins, polysaccharides, and a number of other secondary metabolites that could either bind and/or co-precipitate with RNA, making it unsuitable for downstream applications. Existing RNA isolation methods and commercial kits often fail to deliver high-quality total RNA from immature jatropha seeds for poly(A+ RNA purification and cDNA synthesis. Findings A protocol has been developed for isolating good quality total RNA from immature jatropha seeds, whereby a combination of the CTAB based RNA extraction method and a silica column of a commercial plant RNA extraction kit is used. The extraction time was reduced from two days to about 3 hours and the RNA was suitable for poly(A+ RNA purification, cDNA synthesis, cDNA library construction, RT-PCR, and Northern hybridization. Based on sequence information from selected clones and amplified PCR product, the cDNA library seems to be a good source of full-length jatropha genes. The method was equally effective for isolating RNA from mustard and rice seeds. Conclusions This is a simple CTAB + silica column method to extract high quality RNA from oil rich immature jatropha seeds that is suitable for several downstream applications. This method takes less time for RNA extraction and is equally effective for other tissues where the quality and quantity of RNA is highly interfered by the presence of fatty acids, polysaccharides and polyphenols.

Vulnerability to drug-related infections and co-infections among injecting drug users in Budapest, Hungary

Science.gov (United States)

Neaigus, Alan; Ujhelyi, Eszter

2009-01-01

Background: Drug-related infectious diseases are among the major health consequences of drug use, and any existing drug-related infection may predispose injecting drug users (IDUs) to other infections. Methods: We assessed among IDUs in Budapest, Hungary the prevalence of and vulnerability to selected drug-related infections and co-infections. The sample consisted of 186 participants recruited between October 2005 and December 2006. Results: We found 0% HIV, 37% HCV, 24% HAV, and 14% past HBV infection. Infections with Herpes 1 or 2, tuberculosis, Chlamydia, syphilis, and gonorrhoea were 79%, 12%, 7%, 4%, and 0%, respectively. Co-infection with HAV/HCV was 12%, HBV/HCV 9%, HAV/HBV 7%, and HAV/HBV/HCV 4%. Those over age 30, the ethnic Roma, and the homeless were more likely to have any hepatitis and a higher number of drug-related infections. Amphetamine injectors were more likely to have a higher number of drug-related infections and those who travelled within Hungary were more likely to have any STI. However, those who worked at least part time and those who were in treatment were less likely to have drug-related infections. Conclusions: These results highlight the need of interventions in Hungary to reach and focus on marginalized (Roma or homeless) IDUs and address not only injecting and sex risk, but also hygienic living and injecting conditions. Furthermore, structural interventions to increase social integration (working or being in treatment) may improve welfare and decrease drug use and infection risk tied to drug use/injection among disadvantaged, marginalized, mostly minority populations. PMID:19224936

Expression of the double-stranded RNA of the soybean pod borer Leguminivora glycinivorella (Lepidoptera: Tortricidae) ribosomal protein P0 gene enhances the resistance of transgenic soybean plants.

Science.gov (United States)

Meng, Fanli; Li, Yang; Zang, Zhenyuan; Li, Na; Ran, Ruixue; Cao, Yingxue; Li, Tianyu; Zhou, Quan; Li, Wenbin

2017-12-01

The soybean pod borer [SPB; Leguminivora glycinivorella (Matsumura) (Lepidoptera: Tortricidae)] is the most important soybean pest in northeastern Asia. Silencing genes using plant-mediated RNA-interference is a promising strategy for controlling SPB infestations. The ribosomal protein P0 is important for protein translation and DNA repair in the SPB. Thus, transferring P0 double-stranded RNA (dsRNA) into plants may help prevent SPB-induced damage. We investigated the effects of SpbP0 dsRNA injections and SpbP0 dsRNA-expressing transgenic soybean plants on the SPB. Larval mortality rates were greater for SpbP0 dsRNA-injected larvae (96%) than for the control larvae (31%) at 14 days after injections. Transgenic T 2 soybean plants expressing SpbP0 dsRNA sustained less damage from SPB larvae than control plants. In addition, the expression level of the SpbP0 gene decreased and the mortality rate increased when SPB larvae were fed on T 3 transgenic soybean pods. Moreover, the surviving larvae were deformed and exhibited inhibited growth. Silencing SpbP0 expression is lethal to the SPB. Transgenic soybean plants expressing SpbP0 dsRNA are more resistant to the SPB than wild-type plants. Thus, SpbP0 dsRNA-expressing transgenic plants may be useful for controlling insect pests. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

Brain cDNA clone for human cholinesterase

International Nuclear Information System (INIS)

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

1987-01-01

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

Production of the first transgenic cassava in Africa via direct shoot ...

African Journals Online (AJOL)

Administrator

2006-10-02

Oct 2, 2006 ... organogenesis systems for recovering transgenic cassava plants is described. The contributions of ... acetic acid), SSA (sub-Saharan Africa), picloram (3, 5, 6- ..... ed between Agrobacterium strains. ..... the fundamental requirements for the co-cultivated .... FAOSTAT Statistical Database: Agriculture Data.

Effect of gas field production and CO2 injection on brine flow and salt precipitation

NARCIS (Netherlands)

Loeve, D.; Tambach, T.J.; Hofstee, C.; Plug, W.J.; Maas, J.

2012-01-01

This paper reports modeling of gas field produc-tion and CO2 injection from a theoretical reser-voir based on characteristics of the P18 gas field in the Dutch offshore, which consists of four geological deposits with different petrophysical properties. We especially focus on the brine flow during

Cloning and characterization of the human colipase cDNA

International Nuclear Information System (INIS)

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

1990-01-01

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

Characterization of pancreatic lesions from MT-tgf alpha, Ela-myc and MT-tgf alpha/Ela-myc single and double transgenic mice.

Science.gov (United States)

Liao, Dezhong Joshua; Wang, Yong; Wu, Jiusheng; Adsay, Nazmi Volkan; Grignon, David; Khanani, Fayyaz; Sarkar, Fazlul H

2006-07-05

In order to identify good animal models for investigating therapeutic and preventive strategies for pancreatic cancer, we analyzed pancreatic lesions from several transgenic models and made a series of novel findings. Female MT-tgf alpha mice of the MT100 line developed pancreatic proliferation, acinar-ductal metaplasia, multilocular cystic neoplasms, ductal adenocarcinomas and prominent fibrosis, while the lesions in males were less severe. MT-tgf alpha-ES transgenic lines of both sexes developed slowly progressing lesions that were similar to what was seen in MT100 males. In both MT100 and MT-tgf alpha-ES lines, TGF alpha transgene was expressed mainly in proliferating ductal cells. Ela-myc transgenic mice with a mixed C57BL/6, SJL and FVB genetic background developed pancreatic tumors at 2-7 months of age, and half of the tumors were ductal adenocarcinomas, similar to what was reported originally by Sandgren et al 1. However, in 20% of the mice, the tumors metastasized to the liver. MT100/Ela-myc and MT-tgf alpha-ES/Ela-myc double transgenic mice developed not only acinar carcinomas and mixed carcinomas as previously reported but also various ductal-originated lesions, including multilocular cystic neoplasms and ductal adenocarcinomas. The double transgenic tumors were more malignant and metastasized to the liver at a higher frequency (33%) compared with the Ela-myc tumors. Sequencing of the coding region of p16ink4, k-ras and Rb cDNA in small numbers of pancreatic tumors did not identify mutations. The short latency for tumor development, the variety of tumor morphology and the liver metastases seen in Ela-myc and MT-tgf alpha/Ela-myc mice make these animals good models for investigating new therapeutic and preventive strategies for pancreatic cancer.

Co-transforming bar and CsLEA enhanced tolerance to drought and salt stress in transgenic alfalfa (Medicago sativa L.).

Science.gov (United States)

Zhang, Jiyu; Duan, Zhen; Zhang, Daiyu; Zhang, Jianquan; Di, Hongyan; Wu, Fan; Wang, Yanrong

2016-03-25

Drought and high salinity are two major abiotic factors that restrict alfalfa productivity. A dehydrin protein, CsLEA, from the desert grass Cleistogenes songorica was transformed into alfalfa (Medicago sativa L.) via Agrobacterium-mediated transformation using the bar gene as a selectable marker, and the drought and salt stress tolerances of the transgenic plants were assessed. Thirty-nine of 119 transformants were positive, as screened by Basta, and further molecularly authenticated using PCR and RT-PCR. Phenotype observations revealed that the transgenic plants grew better than the wild-type (WT) plants after 15d of drought stress and 10d of salt stress: the leaves of WT alfalfa turned yellow, whereas the transgenic alfalfa leaves only wilted; after rewatering, the transgenic plants returned to a normal state, though the WT plants could not be restored. Evaluation of physiologic and biochemical indices during drought and salt stresses showed a relatively lower Na(+) content in the leaves of the transgenic plants, which would reduce toxic ion effects. In addition, the transgenic plants were able to maintain a higher relative water content (RWC), higher shoot biomass, fewer photosystem changes, decreased membrane injury, and a lower level of osmotic stress injury. These results demonstrate that overexpression of the CsLEA gene can enhance the drought and salt tolerance of transgenic alfalfa; in addition, carrying the bar gene in the genome may increase herbicide resistance. Copyright © 2016 Elsevier Inc. All rights reserved.

Measurement of electrical impedance of a Berea sandstone core during the displacement of saturated brine by oil and CO2 injections

Science.gov (United States)

Liu, Yu; Xue, Ziqiu; Park, Hyuck; Kiyama, Tamotsu; Zhang, Yi; Nishizawa, Osamu; Chae, Kwang-seok

2015-12-01

Complex electrical impedance measurements were performed on a brine-saturated Berea sandstone core while oil and CO2 were injected at different pressures and temperatures. The saturations of brine, oil, and CO2 in the core were simultaneously estimated using an X-ray computed tomography scanner. The formation factor of this Berea core and the resistivity indexes versus the brine saturations were calculated using Archie's law. The experimental results found different flow patterns of oil under different pressures and temperatures. Fingers were observed for the first experiment at 10 MPa and 40 °C. The fingers were restrained as the viscosity ratio of oil and water changed in the second (10 MPa and 25 °C) and third (5 MPa and 25 °C) experiments. The resistivity index showed an exponential increase with a decrease in brine saturation. The saturation exponent varied from 1.4 to 4.0 at different pressure and temperature conditions. During the oil injection procedure, the electrical impedance increased with oil saturation and was significantly affected by different oil distributions; therefore, the impedance varied whether the finger was remarkable or not, even if the oil saturation remained constant. During the CO2 injection steps, the impedance showed almost no change with CO2 saturation because the brine in the pores became immobile after the oil injection.

Geomechanical effects on CO₂ leakage through fault zones during large-scale underground injection

Energy Technology Data Exchange (ETDEWEB)

Rinaldi, Antonio P. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Rutqvist, Jonny [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Cappa, Frédéric [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Univ. of Nice Sophia-Antipolis, Nice (France). Cote d' Azur Observatory. GeoAzur

2013-12-01

The importance of geomechanics—including the potential for faults to reactivate during large-scale geologic carbon sequestration operations—has recently become more widely recognized. However, notwithstanding the potential for triggering notable (felt) seismic events, the potential for buoyancy-driven CO₂ to reach potable groundwater and the ground surface is actually more important from public safety and storage-efficiency perspectives. In this context, this paper extends the previous studies on the geomechanical modeling of fault responses during underground carbon dioxide injection, focusing on the short-term integrity of the sealing caprock, and hence on the potential for leakage of either brine or CO₂ to reach the shallow groundwater aquifers during active injection. We consider stress/strain-dependent permeability and study the leakage through the fault zone as its permeability changes during a reactivation, also causing seismicity. We analyze several scenarios related to the volume of CO₂ injected (and hence as a function of the overpressure), involving both minor and major faults, and analyze the profile risks of leakage for different stress/strain-permeability coupling functions. We conclude that whereas it is very difficult to predict how much fault permeability could change upon reactivation, this process can have a significant impact on the leakage rate. Moreover, our analysis shows that induced seismicity associated with fault reactivation may not necessarily open up a new flow path for leakage. Results show a poor correlation between magnitude and amount of fluid leakage, meaning that a single event is generally not enough to substantially change the permeability along the entire fault length. Finally, and consequently, even if some changes in permeability occur, this does not mean that the CO₂ will migrate up along the entire fault, breaking through the caprock to enter the overlying aquifer.

Transgene detection by digital droplet PCR.

Directory of Open Access Journals (Sweden)

Dirk A Moser

Full Text Available Somatic gene therapy is a promising tool for the treatment of severe diseases. Because of its abuse potential for performance enhancement in sports, the World Anti-Doping Agency (WADA included the term 'gene doping' in the official list of banned substances and methods in 2004. Several nested PCR or qPCR-based strategies have been proposed that aim at detecting long-term presence of transgene in blood, but these strategies are hampered by technical limitations. We developed a digital droplet PCR (ddPCR protocol for Insulin-Like Growth Factor 1 (IGF1 detection and demonstrated its applicability monitoring 6 mice injected into skeletal muscle with AAV9-IGF1 elements and 2 controls over a 33-day period. A duplex ddPCR protocol for simultaneous detection of Insulin-Like Growth Factor 1 (IGF1 and Erythropoietin (EPO transgenic elements was created. A new DNA extraction procedure with target-orientated usage of restriction enzymes including on-column DNA-digestion was established. In vivo data revealed that IGF1 transgenic elements could be reliably detected for a 33-day period in DNA extracted from whole blood. In vitro data indicated feasibility of IGF1 and EPO detection by duplex ddPCR with high reliability and sensitivity. On-column DNA-digestion allowed for significantly improved target detection in downstream PCR-based approaches. As ddPCR provides absolute quantification, it ensures excellent day-to-day reproducibility. Therefore, we expect this technique to be used in diagnosing and monitoring of viral and bacterial infection, in detecting mutated DNA sequences as well as profiling for the presence of foreign genetic material in elite athletes in the future.

Dehydrins Impart Protection against Oxidative Stress in Transgenic Tobacco Plants.

Science.gov (United States)

Halder, Tanmoy; Upadhyaya, Gouranga; Basak, Chandra; Das, Arup; Chakraborty, Chandrima; Ray, Sudipta

2018-01-01

Environmental stresses generate reactive oxygen species (ROS) which might be detrimental to the plants when produced in an uncontrolled way. However, the plants ameliorate such stresses by synthesizing antioxidants and enzymes responsible for the dismutation of ROS. Additionally, the dehydrins were also able to protect the inactivation of the enzyme lactate dehydrogenase against hydroxyl radicals (OH ⋅ ) generated during Fenton's reaction. SbDhn1 and SbDhn2 overexpressing transgenic tobacco plants were able to protect against oxidative damage. Transgenic tobacco lines showed better photosynthetic efficiency along with high chlorophyll content, soluble sugar and proline. However, the malonyl dialdehyde (MDA) content was significantly lower in transgenic lines. Experimental evidence demonstrates the protective effect of dehydrins on electron transport chain in isolated chloroplast upon methyl viologen (MV) treatment. The transgenic tobacco plants showed significantly lower superoxide radical generation () upon MV treatment. The accumulation of the H 2 O 2 was also lower in the transgenic plants. Furthermore, in the transgenic plants the expression of ROS scavenging enzymes was higher compared to non-transformed (NT) or vector transformed (VT) plants. Taken together these data, during oxidative stress dehydrins function by scavenging the () directly and also by rendering protection to the enzymes responsible for the dismutation of () thereby significantly reducing the amount of hydrogen peroxides formed. Increase in proline content along with other antioxidants might also play a significant role in stress amelioration. Dehydrins thus function co-operatively with other protective mechanisms under oxidative stress conditions rendering protection in stress environment.

Use of the viral 2A peptide for bicistronic expression in transgenic mice

Directory of Open Access Journals (Sweden)

Trichas Georgios

2008-09-01

Full Text Available Abstract Background Transgenic animals are widely used in biomedical research and biotechnology. Multicistronic constructs, in which several proteins are encoded by a single messenger RNA, are commonly used in genetically engineered animals. This is currently done by using an internal ribosomal entry site to separate the different coding regions. 2A peptides result in the co-translational 'cleavage' of proteins and are an attractive alternative to the internal ribosomal entry site. They are more reliable than the internal ribosomal entry site and lead to expression of multiple cistrons at equimolar levels. They work in a wide variety of eukaryotic cells, but to date have not been demonstrated to function in transgenic mice in an inheritable manner. Results To test 2A function in transgenic mice and uncover any possible toxicity of widespread expression of the 2A peptide, we made a bicistronic reporter construct containing the coding sequence for a membrane localised red fluorescent protein (Myr-TdTomato and a nuclear localised green fluorescent protein (H2B-GFP, separated by a 2A sequence. When this reporter is transfected into HeLa cells, the two fluorescent proteins correctly localise to mutually exclusive cellular compartments, demonstrating that the bicistronic construct is a reliable readout of 2A function. The two fluorescent proteins also correctly localise when the reporter is electroporated into chick neural tube cells. We made two independent transgenic mouse lines that express the bicistronic reporter ubiquitously. For both lines, transgenic mice are born in Mendelian frequencies and are found to be healthy and fertile. Myr-TdTomato and H2B-GFP segregate to mutually exclusive cellular compartments in all tissues examined from a broad range of developmental stages, ranging from embryo to adult. One transgenic line shows X-linked inheritance of the transgene and mosaic expression in females but uniform expression in males, indicating

Transgenic overexpression of NanogP8 in the mouse prostate is insufficient to initiate tumorigenesis but weakly promotes tumor development in the Hi-Myc mouse model.

Science.gov (United States)

Liu, Bigang; Gong, Shuai; Li, Qiuhui; Chen, Xin; Moore, John; Suraneni, Mahipal V; Badeaux, Mark D; Jeter, Collene R; Shen, Jianjun; Mehmood, Rashid; Fan, Qingxia; Tang, Dean G

2017-08-08

This project was undertaken to address a critical cancer biology question: Is overexpression of the pluripotency molecule Nanog sufficient to initiate tumor development in a somatic tissue? Nanog1 is critical for the self-renewal and pluripotency of ES cells, and its retrotransposed homolog, NanogP8 is preferentially expressed in somatic cancer cells. Our work has shown that shRNA-mediated knockdown of NanogP8 in prostate, breast, and colon cancer cells inhibits tumor regeneration whereas inducible overexpression of NanogP8 promotes cancer stem cell phenotypes and properties. To address the key unanswered question whether tissue-specific overexpression of NanogP8 is sufficient to promote tumor development in vivo , we generated a NanogP8 transgenic mouse model, in which the ARR 2 PB promoter was used to drive NanogP8 cDNA. Surprisingly, the ARR 2 PB-NanogP8 transgenic mice were viable, developed normally, and did not form spontaneous tumors in >2 years. Also, both wild type and ARR 2 PB-NanogP8 transgenic mice responded similarly to castration and regeneration and castrated ARR 2 PB-NanogP8 transgenic mice also did not develop tumors. By crossing the ARR 2 PB-NanogP8 transgenic mice with ARR 2 PB-Myc (i.e., Hi-Myc) mice, we found that the double transgenic (i.e., ARR 2 PB-NanogP8; Hi-Myc) mice showed similar tumor incidence and histology to the Hi-Myc mice. Interestingly, however, we observed white dots in the ventral lobes of the double transgenic prostates, which were characterized as overgrown ductules/buds featured by crowded atypical Nanog-expressing luminal cells. Taken together, our present work demonstrates that transgenic overexpression of NanogP8 in the mouse prostate is insufficient to initiate tumorigenesis but weakly promotes tumor development in the Hi-Myc mouse model.

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

Directory of Open Access Journals (Sweden)

Zhang Qifa

2008-11-01

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

ANALYSIS OF IMMUNE RESPONSES ON TRANSGENIC TIGER SHRIMP (Penaeus monodon AGAINST PATHOGENIC BACTERIUM Vibrio harveyi

Directory of Open Access Journals (Sweden)

Andi Parenrengi

2014-06-01

Full Text Available Vibriosis is one of main diseases of the black tiger shrimp Penaeus monodon infected by pathogenic bioluminous bacterium Vibrio harveyi that can cause mass mortalities in shrimp culture. The bacteria can also trigger the disease white spot syndrome virus (WSSV. An effort to produce shrimp disease-resistant strains has been done through transgenesis technology with antiviral gene transfection. By this technology, it is expected an increase in the immune response of shrimp in a variety of diseasecausing pathogens. This study aimed to determine the immune responses (total haemocytes, haemocyte differentiation, and phenoloxydase activity of transgenic tiger shrimp against pathogenic bacterium V. harveyi. Research using completely randomized design, which consists of two treatments and three replications. Test animals being used were transgenic and non-transgenic shrimp with size, weight 3.93±1.25 g and a total length of 7.59±0.87 cm. Treatments being tested were the injection of bacterium V. harveyi (density of 5x106 cfu/mL of 0.1 mL/individual on transgenic (A and non-transgenic shrimp (B. Immune response parameters such as total haemocytes, haemocyte differentiation, and phenoloxydase activity were observed on day 1, 3, and 6 days after challenging. Data were analyzed using t-test by SPSS software. The results showed that the total haemocyte of transgenic shrimp was not significantly different (P>0.05 from non-transgenic shrimp, but haemocyte differentiation and phenoloxydase activity were significantly different (P<0.05 especially on sixth days after being exposed to the bioluminescent bacteria. The study results implied that transgenic shrimp has a better immune response compared than non-transgenic shrimp.

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

Directory of Open Access Journals (Sweden)

Stear Michael

2003-03-01

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

A two-fluid model for vertical flow applied to CO₂ injection wells

DEFF Research Database (Denmark)

Linga, Gaute; Lund, Halvor

2016-01-01

Flow of CO2 in wells is associated with substantial variations in thermophysical properties downhole, due to the coupled transient processes involved: complex flow patterns, density changes, phase transitions, and heat transfer to and from surroundings. Large temperature variations can lead...... the well, including tubing, packer fluid, casing, cement or drilling mud, and rock formation. This enables prediction of the temperature in the well fluid and in each layer of the well. The model is applied to sudden shut-in and blowout cases of a CO2 injection well, where we employ the highly accurate...

Cloning and expression of human deoxycytidine kinase cDNA

International Nuclear Information System (INIS)

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

1991-01-01

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

Overexpression of a tea flavanone 3-hydroxylase gene confers tolerance to salt stress and Alternaria solani in transgenic tobacco.

Science.gov (United States)

Mahajan, Monika; Yadav, Sudesh Kumar

2014-08-01

Flavan-3-ols are the major flavonoids present in tea (Camellia sinensis) leaves. These are known to have antioxidant and free radical scavenging properties in vitro. Flavanone 3-hydroxylase is considered to be an important enzyme of flavonoid pathway leading to accumulation of flavan-3-ols in tea. Expression analysis revealed the upregulation in transcript levels of C. sinensis flavanone 3-hydroxylase (CsF3H) encoding gene under salt stress. In this study, the biotechnological potential of CsF3H was evaluated by gene overexpression in tobacco (Nicotiana tabacum cv. Xanthi). Overexpression of CsF3H cDNA increased the content of flavan-3-ols in tobacco and conferred tolerance to salt stress and fungus Alternaria solani infection. Transgenic tobaccos were observed for increase in primary root length, number of lateral roots, chlorophyll content, antioxidant enzyme expression and their activities. Also, they showed lesser malondialdehyde content and electrolyte leakage compared to control tobacco plants. Further, transgenic plants produced higher degree of pectin methyl esterification via decreasing pectin methyl esterase (PME) activity in roots and leaves under unstressed and salt stressed conditions. The effect of flavan-3-ols on pectin methyl esterification under salt stressed conditions was further validated through in vitro experiments in which non-transgenic (wild) tobacco seedlings were exposed to salt stress in presence of flavan-3-ols, epicatechin and epigallocatechin. The in vitro exposed seedlings showed similar trend of increase in pectin methyl esterification through decreasing PME activity as observed in CsF3H transgenic lines. Taken together, overexpression of CsF3H provided tolerance to salt stress and fungus A. solani infection to transgenic tobacco through improved antioxidant system and enhanced pectin methyl esterification.

Hepatic oxidative stress in ovariectomized transgenic mice expressing the hepatitis C virus polyprotein is augmented through suppression of adenosine monophosphate-activated protein kinase/proliferator-activated receptor gamma co-activator 1 alpha signaling.

Science.gov (United States)

Tomiyama, Yasuyuki; Nishina, Sohji; Hara, Yuichi; Kawase, Tomoya; Hino, Keisuke

2014-10-01

Oxidative stress plays an important role in hepatocarcinogenesis of hepatitis C virus (HCV)-related chronic liver diseases. Despite the evidence of an increased proportion of females among elderly patients with HCV-related hepatocellular carcinoma (HCC), it remains unknown whether HCV augments hepatic oxidative stress in postmenopausal women. The aim of this study was to determine whether oxidative stress was augmented in ovariectomized (OVX) transgenic mice expressing the HCV polyprotein and to investigate its underlying mechanisms. OVX and sham-operated female transgenic mice expressing the HCV polyprotein and non-transgenic littermates were assessed for the production of reactive oxygen species (ROS), expression of inflammatory cytokines and antioxidant potential in the liver. Compared with OVX non-transgenic mice, OVX transgenic mice showed marked hepatic steatosis and ROS production without increased induction of inflammatory cytokines, but there was no increase in ROS-detoxifying enzymes such as superoxide dismutase 2 and glutathione peroxidase 1. In accordance with these results, OVX transgenic mice showed less activation of peroxisome proliferator-activated receptor-γ co-activator-1α (PGC-1α), which is required for the induction of ROS-detoxifying enzymes, and no activation of adenosine monophosphate-activated protein kinase-α (AMPKα), which regulates the activity of PGC-1α. Our study demonstrated that hepatic oxidative stress was augmented in OVX transgenic mice expressing the HCV polyprotein by attenuation of antioxidant potential through inhibition of AMPK/PGC-1α signaling. These results may account in part for the mechanisms by which HCV-infected women are at high risk for HCC development when some period has passed after menopause. © 2013 The Japan Society of Hepatology.

Salt tolerance and activity of antioxidative enzymes of transgenic finger millet overexpressing a vacuolar H(+)-pyrophosphatase gene (SbVPPase) from Sorghum bicolor.

Science.gov (United States)

Anjaneyulu, Ediga; Reddy, Palle Surender; Sunita, Merla Srilakshmi; Kishor, Polavarapu B Kavi; Meriga, Balaji

2014-06-15

A vacuolar proton pyrophosphatase cDNA clone was isolated from Sorghum bicolor (SbVPPase) using end-to-end gene-specific primer amplification. It showed 80-90% homology at the nucleotide and 85-95% homology at the amino acid level with other VPPases. The gene was introduced into expression vector pCAMBIA1301 under the control of the cauliflower mosaic virus 35S (CaMV35S) promoter and transformed into Agrobacterium tumifaciens strain LBA4404 to infect embryogenic calli of finger millet (Eleusine coracana). Successful transfer of SbVPPase was confirmed by a GUS histochemical assay and PCR analysis. Both, controls and transgenic plants were subjected to 100 and 200mM NaCl and certain biochemical and physiological parameters were studied. Relative water content (RWC), plant height, leaf expansion, finger length and width and grain weight were severely reduced (50-70%), and the flowering period was delayed by 20% in control plants compared to transgenic plants under salinity stress. With increasing salt stress, the proline and chlorophyll contents as well as the enzyme activities of superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), guaiacol peroxidase (GPX) and glutathione reductase (GR) increased by 25-100% in transgenics, while malondialdehyde (MDA) showed a 2-4-fold decrease. The increased activities of antioxidant enzymes and the reduction in the MDA content suggest efficient scavenging of reactive oxygen species (ROS) in transgenics and, as a consequence, probably alleviation of salt stress. Also, the leaf tissues of the transgenics accumulated 1.5-2.5-fold higher Na(+) and 0.4-0.8-fold higher K(+) levels. Together, these results clearly demonstrate that overexpression of SbVPPase in transgenic finger millet enhances the plant's performance under salt stress. Copyright © 2014 Elsevier GmbH. All rights reserved.

Expression of an osmotin-like protein from Solanum nigrum confers drought tolerance in transgenic soybean.

Science.gov (United States)

Weber, Ricardo Luís Mayer; Wiebke-Strohm, Beatriz; Bredemeier, Christian; Margis-Pinheiro, Márcia; de Brito, Giovani Greigh; Rechenmacher, Ciliana; Bertagnolli, Paulo Fernando; de Sá, Maria Eugênia Lisei; Campos, Magnólia de Araújo; de Amorim, Regina Maria Santos; Beneventi, Magda Aparecida; Margis, Rogério; Grossi-de-Sa, Maria Fátima; Bodanese-Zanettini, Maria Helena

2014-12-10

Drought is by far the most important environmental factor contributing to yield losses in crops, including soybeans [Glycine max (L.) Merr.]. To address this problem, a gene that encodes an osmotin-like protein isolated from Solanum nigrum var. americanum (SnOLP) driven by the UBQ3 promoter from Arabidopsis thaliana was transferred into the soybean genome by particle bombardment. Two independently transformed soybean lines expressing SnOLP were produced. Segregation analyses indicated single-locus insertions for both lines. qPCR analysis suggested a single insertion of SnOLP in the genomes of both transgenic lines, but one copy of the hpt gene was inserted in the first line and two in the second line. Transgenic plants exhibited no remarkable phenotypic alterations in the seven analyzed generations. When subjected to water deficit, transgenic plants performed better than the control ones. Leaf physiological measurements revealed that transgenic soybean plants maintained higher leaf water potential at predawn, higher net CO2 assimilation rate, higher stomatal conductance and higher transpiration rate than non-transgenic plants. Grain production and 100-grain weight were affected by water supply. Decrease in grain productivity and 100-grain weight were observed for both transgenic and non-transgenic plants under water deficit; however, it was more pronounced for non-transgenic plants. Moreover, transgenic lines showed significantly higher 100-grain weight than non-transgenic plants under water shortage. This is the first report showing that expression of SnOLP in transgenic soybeans improved physiological responses and yield components of plants when subjected to water deficit, highlighting the potential of this gene for biotechnological applications.

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

DEFF Research Database (Denmark)

Kristensen, Torsten; Schousboe, Inger; Boel, Espen

1991-01-01

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

Qualitative and quantitative changes in detrital reservoir rocks caused by CO2-brine-rock interactions during first injection phases (Utrillas sandstones, northern Spain)

Science.gov (United States)

Berrezueta, E.; Ordóñez-Casado, B.; Quintana, L.

2016-01-01

The aim of this article is to describe and interpret qualitative and quantitative changes at rock matrix scale of lower-upper Cretaceous sandstones exposed to supercritical (SC) CO2 and brine. The effects of experimental injection of CO2-rich brine during the first injection phases were studied at rock matrix scale, in a potential deep sedimentary reservoir in northern Spain (Utrillas unit, at the base of the Cenozoic Duero Basin).Experimental CO2-rich brine was exposed to sandstone in a reactor chamber under realistic conditions of deep saline formations (P ≈ 7.8 MPa, T ≈ 38 °C and 24 h exposure time). After the experiment, exposed and non-exposed equivalent sample sets were compared with the aim of assessing possible changes due to the effect of the CO2-rich brine exposure. Optical microscopy (OpM) and scanning electron microscopy (SEM) aided by optical image analysis (OIA) were used to compare the rock samples and get qualitative and quantitative information about mineralogy, texture and pore network distribution. Complementary chemical analyses were performed to refine the mineralogical information and to obtain whole rock geochemical data. Brine composition was also analyzed before and after the experiment.The petrographic study of contiguous sandstone samples (more external area of sample blocks) before and after CO2-rich brine injection indicates an evolution of the pore network (porosity increase ≈ 2 %). It is probable that these measured pore changes could be due to intergranular quartz matrix detachment and partial removal from the rock sample, considering them as the early features produced by the CO2-rich brine. Nevertheless, the whole rock and brine chemical analyses after interaction with CO2-rich brine do not present important changes in the mineralogical and chemical configuration of the rock with respect to initial conditions, ruling out relevant precipitation or dissolution at these early stages to rock-block scale. These results

Intermediate Scale Laboratory Testing to Understand Mechanisms of Capillary and Dissolution Trapping during Injection and Post-Injection of CO₂ in Heterogeneous Geological Formations

Energy Technology Data Exchange (ETDEWEB)

Illangasekare, Tissa [Colorado School of Mines, Golden, CO (United States); Trevisan, Luca [Colorado School of Mines, Golden, CO (United States); Agartan, Elif [Colorado School of Mines, Golden, CO (United States); Mori, Hiroko [Colorado School of Mines, Golden, CO (United States); Vargas-Johnson, Javier [Colorado School of Mines, Golden, CO (United States); Gonzalez-Nicolas, Ana [Colorado School of Mines, Golden, CO (United States); Cihan, Abdullah [Colorado School of Mines, Golden, CO (United States); Birkholzer, Jens [Colorado School of Mines, Golden, CO (United States); Zhou, Quanlin [Colorado School of Mines, Golden, CO (United States)

2015-03-31

Carbon Capture and Storage (CCS) represents a technology aimed to reduce atmospheric loading of CO₂ from power plants and heavy industries by injecting it into deep geological formations, such as saline aquifers. A number of trapping mechanisms contribute to effective and secure storage of the injected CO₂ in supercritical fluid phase (scCO₂) in the formation over the long term. The primary trapping mechanisms are structural, residual, dissolution and mineralization. Knowledge gaps exist on how the heterogeneity of the formation manifested at all scales from the pore to the site scales affects trapping and parameterization of contributing mechanisms in models. An experimental and modeling study was conducted to fill these knowledge gaps. Experimental investigation of fundamental processes and mechanisms in field settings is not possible as it is not feasible to fully characterize the geologic heterogeneity at all relevant scales and gathering data on migration, trapping and dissolution of scCO₂. Laboratory experiments using scCO₂ under ambient conditions are also not feasible as it is technically challenging and cost prohibitive to develop large, two- or three-dimensional test systems with controlled high pressures to keep the scCO₂ as a liquid. Hence, an innovative approach that used surrogate fluids in place of scCO₂ and formation brine in multi-scale, synthetic aquifers test systems ranging in scales from centimeter to meter scale developed used. New modeling algorithms were developed to capture the processes controlled by the formation heterogeneity, and they were tested using the data from the laboratory test systems. The results and findings are expected to contribute toward better conceptual models, future improvements to DOE numerical codes, more accurate assessment of storage capacities, and optimized placement strategies. This report presents the experimental and modeling methods

Transgenic algae engineered for higher performance

Science.gov (United States)

Unkefer, Pat J; Anderson, Penelope S; Knight, Thomas J

2014-10-21

The present disclosure relates to transgenic algae having increased growth characteristics, and methods of increasing growth characteristics of algae. In particular, the disclosure relates to transgenic algae comprising a glutamine phenylpyruvate transaminase transgene and to transgenic algae comprising a glutamine phenylpyruvate transaminase transgene and a glutamine synthetase.

Numerical Simulation of Multiphase Hydromechanical Processes Induced by CO2 Injection into Deep Saline Aquifers Simulation numérique des processus hydromécaniques polyphasiques provoqués par l’injection de CO2 dans des aquifères salins profonds

Directory of Open Access Journals (Sweden)

Goerke U.-J.

2011-02-01

Full Text Available In this paper, the conceptual modeling and the numerical simulation of two-phase flow during CO2 injection into deep saline aquifers is presented. The work focuses on isothermal short-term processes in the vicinity of the injection well. Governing differential equations are based on balance laws for mass and momentum, and completed by constitutive relations for the fluid and solid phases as well as their mutual interactions. Constraint conditions for the partial saturations and the pressure fractions of CO2 and brine are defined. To characterize the stress state in the solid matrix, the effective stress principle is applied. The coupled problem is solved using the inhouse scientific code OpenGeoSys (an open source finite element code and verified with benchmarks. Cet article présente le concept de la modélisation ainsi que la simulation numérique d’écoulement biphasé lors de l’injection de CO2 dans des aquifères salins profonds. L’étude se concentre sur des processus à court terme dans la proximité de puits d’injection dans les conditions isothermes. Les équations différentielles principales sont dérivées des équations de bilan de masse et de la quantité de mouvement, et elles sont complétées par des relations constitutives pour des phases solides et fluides ainsi que leur interaction. Les conditions de contrainte sont définies pour la saturation partielle, les fractions de pression de CO2 et l’eau salée. Pour caractériser l’état de contrainte dans la matrice solide, le principe de contrainte effective est appliqué. Le problème couplé considéré est résolu en utilisant le logiciel scientifique interne OpenGeoSys (un logiciel d’éléments finis libre d’accès et vérifié avec des exemples pour les processus concernés.

Construction of a T7 Human Lung Cancer cDNA Library

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Wentao YUE

2008-10-01

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

Cloning, sequencing, and expression of cDNA for human β-glucuronidase

International Nuclear Information System (INIS)

Oshima, A.; Kyle, J.W.; Miller, R.D.

1987-01-01

The authors report here the cDNA sequence for human placental β-glucuronidase (β-D-glucuronoside glucuronosohydrolase, EC 3.2.1.31) and demonstrate expression of the human enzyme in transfected COS cells. They also sequenced a partial cDNA clone from human fibroblasts that contained a 153-base-pair deletion within the coding sequence and found a second type of cDNA clone from placenta that contained the same deletion. Nuclease S1 mapping studies demonstrated two types of mRNAs in human placenta that corresponded to the two types of cDNA clones isolated. The NH 2 -terminal amino acid sequence determined for human spleen β-glucuronidase agreed with that inferred from the DNA sequence of the two placental clones, beginning at amino acid 23, suggesting a cleaved signal sequence of 22 amino acids. When transfected into COS cells, plasmids containing either placental clone expressed an immunoprecipitable protein that contained N-linked oligosaccharides as evidenced by sensitivity to endoglycosidase F. However, only transfection with the clone containing the 153-base-pair segment led to expression of human β-glucuronidase activity. These studies provide the sequence for the full-length cDNA for human β-glucuronidase, demonstrate the existence of two populations of mRNA for β-glucuronidase in human placenta, only one of which specifies a catalytically active enzyme, and illustrate the importance of expression studies in verifying that a cDNA is functionally full-length

Comparison of the Wymark CO2 Reservoir with the Midale Beds at the Weyburn CO2 Injection Project

International Nuclear Information System (INIS)

Ryerson, F.; Johnson, J.

2010-01-01

The Devonian carbonates of the Duperow Formation on the western flank of the Williston Basin in southwest Saskatchewan contain natural accumulations of CO 2 , and may have done so for as long as 50 m.y. in the views of some investigations. These carbonate sediments are characterized by a succession of carbonate cycles capped by anhydrite-rich evaporites that are thought to act as seals to fluid migration. The Weyburn CO 2 injection site lies 400 km to the east in a series of Mississippian carbonates that were deposited in a similar depositional environment. That natural CO 2 can be stored long-term within carbonate strata has motivated the investigation of the Duperow rocks as a potential natural analogue to storage of anthropogenic CO 2 that may ultimately provide additional confidence for CO 2 sequestration in carbonate lithologies. For the Duperow strata to represent a legitimate analog for Midale injection and storage, the similarity in lithofacies, whole rock compositions, mineral compositions and porosity with the Midale Beds must be established. Previous workers have demonstrated the similarity of the lithofacies at both sites. Here we compare the whole rock compositions, mineralogy and mineral compositions. The major mineral phases at both locales are calcite, dolomite and anhydrite. In addition, accessory pyrite, fluorite and celestine are also observed. The distribution of porosity in the Midale Vuggy units is virtually identical to that of the Duperow Formation, but the Marly units of the Midale have significantly higher porosity. The Duperow Formation is topped by the Dinesmore evaporite that is particularly rich in anhydrite, and often contains authigenic K-feldspar. The chemistry of dolomite and calcite from the two localities also overlaps. Silicate minerals are in low abundance within the analyzed Duperow samples, < 3 wt% on a normative basis, with quartz the only phase identifiable in x-ray diffraction patterns. The Midale Beds contain

Leishmania mexicana Gp63 cDNA Using Gene Gun Induced Higher Immunity to L. mexicana Infection Compared to Soluble Leishmania Antigen in BALB/C

Science.gov (United States)

Rezvan, H; Rees, R; Ali, SA

2011-01-01

Background Leishmaniasis is a worldwide disease prevalent in tropical and sub tropical countries. Many attempts have been made and different strategies have been approached to develop a potent vaccine against Leishmania. DNA immunisation is a method, which is shown to be effective in Leishmania vaccination. Leishmania Soluble Antigen (SLA) has also recently been used Leishmania vaccination. Methods The immunity generated by SLA and L. mexicana gp63 cDNA was compared in groups of 6 mice, which were statistically analysed by student t- test with the P-value of 0.05. SLA was administered by two different methods; intramuscular injection and injection of dendritic cells (DCs) loaded with SLA. L. mexicana gp63 cDNA was administered by the gene gun. Results Immunisation of BALB/c mice with L. mexicana gp63 resulted in high levels of Th1-type immune response and cytotoxic T lymphocytes (CTL) activity, which were accompanied with protection induced by the immunisation against L. mexicana infection. In contrast, administration of SLA, produced a mixed Th1/Th2-type immune responses as well as a high level of CTL activity but did not protect mice from the infection. Conclusion The results indicate higher protection by DNA immunisation using L. mexicana gp63 cDNA compared to SLA, which is accompanied by a high level of Th1 immune response. However, the CTL activity does not necessarily correlate with the protection induced by the vaccine. Also, gene gun immunisation is a potential approach in Leishmania vaccination. These findings would be helpful in opening new windows in Leishmania vaccine research. PMID:22347315

An Experimental Study of Surfactant Alternating CO2 Injection for Enhanced Oil Recovery of Carbonated Reservoir

Directory of Open Access Journals (Sweden)

Asghar Gandomkar

2016-10-01

Full Text Available Core flooding experiments were conducted with the objective of evaluating near miscible surfactant alternating CO2 injection and the effect of surfactant concentrations on gas-oil and water displacements in porous media. The core samples were provided from a low permeability mixed wet oil reservoir at 156 °F and 1900 psia. In addition, very few studies of surfactant adsorption on carbonate minerals have been conducted. Hence, the surfactant adsorption on carbonate rock was determined by core flooding and crushed tests. It was found that for the crushed rock, the required equilibrium time is approximately five hours, while it is more than four days for the flow-through tests. Hysteresis effects demonstrated that the irreducible water saturations were 5 to 10% higher than the initial connate water saturation after drainage cycles during 5000 ppm surfactant solution. Furthermore, near-miscible surfactant alternating CO2 injection process led to a 4-17% increase in the recovery factor in comparison to water alternating gas process.

Sequence of a cloned cDNA encoding human ribosomal protein S11

Energy Technology Data Exchange (ETDEWEB)

Lott, J B; Mackie, G A

1988-02-11

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

Ultra-low emittance beam generation using two-color ionization injection in a CO2 laser-driven plasma accelerator

International Nuclear Information System (INIS)

Schroeder, Carl; Benedetti, Carlo; Bulanov, Stepan; Chen, Min; Esarey, Eric; Geddes, Cameron; Vay, J.; Yu, Lule; Leemans, Wim

2015-01-01

Ultra-low emittance (tens of nm) beams can be generated in a plasma accelerator using ionization injection of electrons into a wakefield. An all-optical method of beam generation uses two laser pulses of different colors. A long-wavelength drive laser pulse (with a large ponderomotive force and small peak electric field) is used to excite a large wakefield without fully ionizing a gas, and a short-wavelength injection laser pulse (with a small ponderomotive force and large peak electric field), co-propagating and delayed with respect to the pump laser, to ionize a fraction of the remaining bound electrons at a trapped wake phase, generating an electron beam that is accelerated in the wake. The trapping condition, the ionized electron distribution, and the trapped bunch dynamics are discussed. Expressions for the beam transverse emittance, parallel and orthogonal to the ionization laser polarization, are presented. An example is shown using a 10-micron CO 2 laser to drive the wake and a frequency-doubled Ti:Al 2 O 3 laser for ionization injection.

Production of germline transgenic prairie voles (Microtus ochrogaster) using lentiviral vectors.

Science.gov (United States)

Donaldson, Zoe R; Yang, Shang-Hsun; Chan, Anthony W S; Young, Larry J

2009-12-01

The study of alternative model organisms has yielded tremendous insights into the regulation of behavioral and physiological traits not displayed by more widely used animal models, such as laboratory rats and mice. In particular, comparative approaches often exploit species ideally suited for investigating specific phenomenon. For instance, comparative studies of socially monogamous prairie voles and polygamous meadow voles have been instrumental toward gaining an understanding of the genetic and neurobiological basis of social bonding. However, laboratory studies of less commonly used organisms, such as prairie voles, have been limited by a lack of genetic tools, including the ability to manipulate the genome. Here, we show that lentiviral vector-mediated transgenesis is a rapid and efficient approach for creating germline transgenics in alternative laboratory rodents. Injection of a green fluorescent protein (GFP)-expressing lentiviral vector into the perivitelline space of 23 single-cell embryos yielded three live offspring (13 %), one of which (33%) contained germline integration of a GFP transgene driven by the human ubiquitin-C promoter. In comparison, transfer of 23 uninjected embryos yielded six live offspring (26%). Green fluorescent protein is present in all tissues examined and is expressed widely in the brain. The GFP transgene is heritable and stably expressed until at least the F(2) generation. This technology has the potential to allow investigation of specific gene candidates in prairie voles and provides a general protocol to pursue germline transgenic manipulation in many different rodent species.

5'-end sequences of budding yeast full-length cDNA clones and quality scores - Budding yeast cDNA sequencing project | LSDB Archive [Life Science Database Archive metadata

Lifescience Database Archive (English)

Full Text Available east_seq_qual.zip File URL: ftp://ftp.biosciencedbc.jp/archive/yeast_cdna/LATEST/...yeast_seq_qual.zip File size: 59.9MB Simple search URL http://togodb.biosciencedbc.jp/togodb/view/budding_yeast_cdna

Hydraulic and Mechanical Effects from Gas Hydrate Conversion and Secondary Gas Hydrate Formation during Injection of CO2 into CH4-Hydrate-Bearing Sediments

Science.gov (United States)

Bigalke, N.; Deusner, C.; Kossel, E.; Schicks, J. M.; Spangenberg, E.; Priegnitz, M.; Heeschen, K. U.; Abendroth, S.; Thaler, J.; Haeckel, M.

2014-12-01

The injection of CO2 into CH4-hydrate-bearing sediments has the potential to drive natural gas production and simultaneously sequester CO2 by hydrate conversion. The process aims at maintaining the in situ hydrate saturation and structure and causing limited impact on soil hydraulic properties and geomechanical stability. However, to increase hydrate conversion yields and rates it must potentially be assisted by thermal stimulation or depressurization. Further, secondary formation of CO2-rich hydrates from pore water and injected CO2 enhances hydrate conversion and CH4 production yields [1]. Technical stimulation and secondary hydrate formation add significant complexity to the bulk conversion process resulting in spatial and temporal effects on hydraulic and geomechanical properties that cannot be predicted by current reservoir simulation codes. In a combined experimental and numerical approach, it is our objective to elucidate both hydraulic and mechanical effects of CO2 injection and CH4-CO2-hydrate conversion in CH4-hydrate bearing soils. For the experimental approach we used various high-pressure flow-through systems equipped with different online and in situ monitoring tools (e.g. Raman microscopy, MRI and ERT). One particular focus was the design of triaxial cell experimental systems, which enable us to study sample behavior even during large deformations and particle flow. We present results from various flow-through high-pressure experimental studies on different scales, which indicate that hydraulic and geomechanical properties of hydrate-bearing sediments are drastically altered during and after injection of CO2. We discuss the results in light of the competing processes of hydrate dissociation, hydrate conversion and secondary hydrate formation. Our results will also contribute to the understanding of effects of temperature and pressure changes leading to dissociation of gas hydrates in ocean and permafrost systems. [1] Deusner C, Bigalke N, Kossel E

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

International Nuclear Information System (INIS)

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

1987-01-01

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

Ultrasound Backscatter Microscopy Image-Guided Intraventricular Gene Delivery at Murine Embryonic Age 9.5 and 10.5 Produces Distinct Transgene Expression Patterns at the Adult Stage

Directory of Open Access Journals (Sweden)

Jiwon Jang

2013-11-01

Full Text Available In utero injection of a retroviral vector into the embryonic telencephalon aided by ultrasound backscatter microscopy permits introduction of a gene of interest at an early stage of development. In this study, we compared the tissue distribution of gene expression in adult mice injected with retroviral vectors at different embryonic ages in utero. Following ultrasound image-guided gene delivery (UIGD into the embryonic telencephalon, adult mice were subjected to whole-body luciferase imaging and immunohistochemical analysis at 6 weeks and 1 year postinjection. Luciferase activity was observed in a wide range of tissues in animals injected at embryonic age 9.5 (E9.5, whereas animals injected at E10.5 showed brain-localized reporter gene expression. These results suggest that mouse embryonic brain creates a closed and impermeable structure around E10. Therefore, by injecting a transgene before or after E10, transgene expression can be manipulated to be local or systemic. Our results also provide information that widens the applicability of UIGD beyond neuroscience studies.

Tamarix hispida metallothionein-like ThMT3, a reactive oxygen species scavenger, increases tolerance against Cd(2+), Zn(2+), Cu(2+), and NaCl in transgenic yeast.

Science.gov (United States)

Yang, Jingli; Wang, Yucheng; Liu, Guifeng; Yang, Chuanping; Li, Chenghao

2011-03-01

A metallothionein-like gene, ThMT3, encoding a type 3 metallothionein, was isolated from a Tamarix hispida leaf cDNA library. Expression analysis revealed that mRNA of ThMT3 was upregulated by high salinity as well as by heavy metal ions, and that ThMT3 was predominantly expressed in the leaf. Transgenic yeast (Saccharomyces cerevisiae) expressing ThMT3 showed increased tolerance to Cd(2+), Zn(2+), Cu(2+), and NaCl stress. Transgenic yeast also accumulated more Cd(2+), Zn(2+), and NaCl, but not Cu(2+). Analysis of the expression of four genes (GLR1, GTT2, GSH1, and YCF1) that aid in transporting heavy metal (Cd(2+)) from the cytoplasm to the vacuole demonstrated that none of these genes were induced under Cd(2+), Zn(2+), Cu(2+), and NaCl stress in ThMT3-transgenic yeast. H(2)O(2) levels in transgenic yeast under such stress conditions were less than half those in control yeast under the same conditions. Three antioxidant genes (SOD1, CAT1, and GPX1) were specifically expressed under Cd(2+), Zn(2+), Cu(2+), and NaCl stress in the transgenic yeast. Cd(2+), Zn(2+), and Cu(2+) increased the expression levels of SOD1, CAT1, and GPX1, respectively, whereas NaCl induced the expression of SOD1 and GPX1.

Two-frequency operation of a hybrid TEA CO2 laser and its application to two-frequency pulse injection locking

International Nuclear Information System (INIS)

Sasaki, Koichi; Ohno, Hirotaka; Fujii, Takaharu; Tsukishima, Takashige.

1990-10-01

Simultaneous two-frequency oscillation of a hybrid TEA CO 2 laser is exhibited when the cw section is operated in a 'below threshold' state. The output of the hybrid laser thus obtained is injected into a main TEA CO 2 laser to obtain a power-modulated, long-pulse output with a well suppressed gain-switched spike. (author)

Comparison of nutrition composition of transgenic maize (chitinase gene) with its non-transgenic counterpart

OpenAIRE

Ping-mei, Yan; Yu-kui, Rui; Xiao-yan, Yan; Zheng, Chai; Qing, Wang; Jian-zhong, Du; Yi, Sun

2011-01-01

In order to compare the nutrition components of transgenic maize seeds (chitinase gene), achieved by the pollen-mediated approach, with its non-transgenic counterpart, Vitamin B1, vitamin B2, fatty acids and essential amino acids of transgenic maize seeds and their counterparts were analyzed by the Chinese national standard methods or AOAC methods. The results showed that the contents of all the six kinds of fatty acids detected in transgenic maize seeds were significantly higher than those i...

Liver-specific expression of the agouti gene in transgenic mice promotes liver carcinogenesis in the absence of obesity and diabetes

Energy Technology Data Exchange (ETDEWEB)

Kuklin, Alexander [ORNL; Mynatt, Randall [ORNL; Klebig, Mitch [ORNL; Kiefer, Laura [Glaxo Wellcome, Research Triangle Park, NC; Wilkison, William O [Glaxo Wellcome, Research Triangle Park, NC; Woychik, Richard P [Jackson Laboratory, The, Bar Harbor, ME; Michaud III, Edward J [ORNL

2004-01-01

Background: The agouti protein is a paracrine factor that is normally present in the skin of many species of mammals. Agouti regulates the switch between black and yellow hair pigmentation by signalling through the melanocortin 1 receptor (Mc1r) on melanocytes. Lethal yellow (Ay) and viable yellow (Avy) are dominant regulatory mutations in the mouse agouti gene that cause the wild- ype protein to be produced at abnormally high levels throughout the body. Mice harboring these mutations exhibit a pleiotropic syndrome characterized by yellow coat color, obesity, hyperglycemia, hyperinsulinemia, and increased susceptibility to hyperplasia and carcinogenesis in numerous tissues, including the liver. The goal of this research was to determine if ectopic expression of the agouti gene in the liver alone is sufficient to recapitulate any aspect of this syndrome. For this purpose, we generated lines of transgenic mice expressing high levels of agouti in the liver under the regulatory control of the albumin promoter. Expression levels of the agouti transgene in the liver were quantified by Northern blot analysis. Functional agouti protein in the liver of transgenic mice was assayed by its ability to inhibit binding of the -melanocyte stimulating hormone ( MSH) to the Mc1r. Body weight, plasma insulin and blood glucose levels were analyzed in control and transgenic mice. Control and transgenic male mice were given a single intraperitoneal injection (10 mg/kg) of the hepatocellular carcinogen, diethylnitrosamine (DEN), at 15 days of age. Mice were euthanized at 36 or 40 weeks after DEN injection and the number of tumors per liver and total liver weights were recorded. Results: The albumin-agouti transgene was expressed at high levels in the livers of mice and produced a functional agouti protein. Albumin-agouti transgenic mice had normal body weights and normal levels of blood glucose and plasma insulin, but responded to chemical initiation of the liver with an increased number

Sequence of a cDNA encoding turtle high mobility group 1 protein.

Science.gov (United States)

Zheng, Jifang; Hu, Bi; Wu, Duansheng

2005-07-01

In order to understand sequence information about turtle HMG1 gene, a cDNA encoding HMG1 protein of the Chinese soft-shell turtle (Pelodiscus sinensis) was amplified by RT-PCR from kidney total RNA, and was cloned, sequenced and analyzed. The results revealed that the open reading frame (ORF) of turtle HMG1 cDNA is 606 bp long. The ORF codifies 202 amino acid residues, from which two DNA-binding domains and one polyacidic region are derived. The DNA-binding domains share higher amino acid identity with homologues sequences of chicken (96.5%) and mammalian (74%) than homologues sequence of rainbow trout (67%). The polyacidic region shows 84.6% amino acid homology with the equivalent region of chicken HMG1 cDNA. Turtle HMG1 protein contains 3 Cys residues located at completely conserved positions. Conservation in sequence and structure suggests that the functions of turtle HMG1 cDNA may be highly conserved during evolution. To our knowledge, this is the first report of HMG1 cDNA sequence in any reptilian.

Comparison of nutritional value of transgenic peanut expressing bar and rcg3 genes with non-transgenic counterparts

International Nuclear Information System (INIS)

Robab, U.E.; )

2014-01-01

The transgenic peanut plants expressing bar and rcg3 genes were subjected to assessment of any change in nutritional value of the crop at various locations. The protein and fat contents of transgenic lines were compared with the non-transgenic parent varieties. Protein content in the transgenic lines was higher as compared to that in non-transgenic counterparts and differences among locations for fat and protein content were significant. No differences among fatty acids were recorded for genes, events and locations. Irrespective of small differences, all the values were in range described for this crop and transgenic lines appeared to be substantially equivalent to non-transgenic parent varieties. (author)

Molecular cloning of L-methylmalonyl-CoA mutase: Gene transfer and analysis of mut cell lines

International Nuclear Information System (INIS)

Ledley, F.D.; Lumetta, M.; Nguyen, P.N.; Kolhouse, J.F.; Allen, R.H.

1988-01-01

L-Methylmalonyl-CoA mutase (MCM, EC 5.4.99.2) is a mitochondrial adenosylcobalamin-requiring enzyme that catalyzes the isomerization of L-methylmalonyl-CoA to succinyl-CoA. This enzyme is deficient in methylmalonic acidemia, an often fatal disorder of organic acid metabolism. Antibody against human placental MCM was used to screen human placenta and liver cDNA expression libraries for MCM cDNA clones. One clone expressed epitopes that could affinity-purify antibodies against MCM. A cDNA corresponding in length to the mRNA was obtained and introduced into COS cells by DNA-mediated gene transfer. Cells transformed with this clone expressed increased levels of MCM enzymatic activity. RNA blot analysis of cells genetically deficient in MCM indicates that several deficient cell lines have a specific decrease in the amount of hybridizable mRNA. These data confirm the authenticity of the MCM cDNA clone, establish the feasibility of constituting MCM activity by gene transfer for biochemical analysis and gene therapy, and provide a preliminary picture of the genotypic spectrum underlying MCM deficiency

Cloning and expression of cDNA coding for bouganin.

Science.gov (United States)

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

2002-03-01

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

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

Science.gov (United States)

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

2017-09-15

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

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

Directory of Open Access Journals (Sweden)

Ju-Yeon Yoon

2014-03-01

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

Joint inversion of time-lapse VSP data for monitoring CO2 injection at the Farnsworth EOR field in Texas

Science.gov (United States)

Zhang, M.; Gao, K.; Balch, R. S.; Huang, L.

2016-12-01

During the Development Phase (Phase III) of the U.S. Southwest Regional Partnership on Carbon Sequestration (SWP), time-lapse 3D vertical seismic profiling (VSP) data were acquired to monitor CO2 injection/migration at the Farnsworth Enhanced Oil Recovery (EOR) field, in partnership with the industrial partner Chaparral Energy. The project is to inject a million tons of carbon dioxide into the target formation, the deep oil-bearing Morrow Formation in the Farnsworth Unit EOR field. Quantitative time-lapse seismic monitoring has the potential to track CO2 movement in geologic carbon storage sites. Los Alamos National Laboratory (LANL) has recently developed new full-waveform inversion methods to jointly invert time-lapse seismic data for changes in elastic and anisotropic parameters in target monitoring regions such as a CO2 reservoir. We apply our new joint inversion methods to time-lapse VSP data acquired at the Farnsworth EOR filed, and present some preliminary results showing geophysical properties changes in the reservoir.

Variation of Se, Zn, Co, Fe and Rb distribution in rats upon sequence of injection with SeO2 and glutathione

International Nuclear Information System (INIS)

Czauderna, M.; Samochocka, K.; Kwiathkowska, J.

1984-01-01

The contents of Se, Zn, Co, Fe and Rb in several organs of Wistar rats were determined by instrumental neutron activation analysis (INAA) after injections of SeO 2 and glutathione (GSH). Se was incorporated in all the examined organs, and the efficiency of incorporation does not depend upon the sequence of injection with SeO 2 and GSH. The sequence of these injections affects the contents of the other elements in all the examined organs. (author)

Laboratory Mid-frequency (Kilohertz) Range Seismic Property Measurements and X-ray CT Imaging of Fractured Sandstone Cores During Supercritical CO2 Injection

Science.gov (United States)

Nakagawa, S.; Kneafsey, T. J.; Chang, C.; Harper, E.

2014-12-01

During geological sequestration of CO2, fractures are expected to play a critical role in controlling the migration of the injected fluid in reservoir rock. To detect the invasion of supercritical (sc-) CO2 and to determine its saturation, velocity and attenuation of seismic waves can be monitored. When both fractures and matrix porosity connected to the fractures are present, wave-induced dynamic poroelastic interactions between these two different types of rock porosity—high-permeability, high-compliance fractures and low-permeability, low-compliance matrix porosity—result in complex velocity and attenuation changes of compressional waves as scCO2 invades the rock. We conducted core-scale laboratory scCO2 injection experiments on small (diameter 1.5 inches, length 3.5-4 inches), medium-porosity/permeability (porosity 15%, matrix permeability 35 md) sandstone cores. During the injection, the compressional and shear (torsion) wave velocities and attenuations of the entire core were determined using our Split Hopkinson Resonant Bar (short-core resonant bar) technique in the frequency range of 1-2 kHz, and the distribution and saturation of the scCO2 determined via X-ray CT imaging using a medical CT scanner. A series of tests were conducted on (1) intact rock cores, (2) a core containing a mated, core-parallel fracture, (3) a core containing a sheared core-parallel fracture, and (4) a core containing a sheared, core-normal fracture. For intact cores and a core containing a mated sheared fracture, injections of scCO2 into an initially water-saturated sample resulted in large and continuous decreases in the compressional velocity as well as temporary increases in the attenuation. For a sheared core-parallel fracture, large attenuation was also observed, but almost no changes in the velocity occurred. In contrast, a sample containing a core-normal fracture exhibited complex behavior of compressional wave attenuation: the attenuation peaked as the leading edge of

Cognitive abilities of Alzheimer's disease transgenic mice are modulated by social context and circadian rhythm.

Science.gov (United States)

Kiryk, Anna; Mochol, Gabriela; Filipkowski, Robert K; Wawrzyniak, Marcin; Lioudyno, Victoria; Knapska, Ewelina; Gorkiewicz, Tomasz; Balcerzyk, Marcin; Leski, Szymon; Leuven, Fred Van; Lipp, Hans-Peter; Wojcik, Daniel K; Kaczmarek, Leszek

2011-12-01

In the present study, we used a new training paradigm in the intelliCage automatic behavioral assessment system to investigate cognitive functions of the transgenic mice harboring London mutation of the human amyloid precursor protein (APP.V717I). Three groups of animals: 5-, 12- and 18-24-month old were subjected to both Water Maze training and the IntelliCage-based appetitive conditioning. The spatial memory deficit was observed in all three groups of transgenic mice in both behavioral paradigms. However, the APP mice were capable to learn normally when co-housed with the wild-type (WT) littermates, in contrast to clearly impaired learning observed when the transgenic mice were housed alone. Furthermore, in the transgenic mice kept in the Intellicage alone, the cognitive deficit of the young animals was modulated by the circadian rhythm, namely was prominent only during the active phase of the day. The novel approach to study the transgenic mice cognitive abilities presented in this paper offers new insight into cognitive dysfunctions of the Alzheimer's disease mouse model.

Epigenetic variants of a transgenic petunia line show hypermethylation in transgene DNA: an indication for specific recognition of foreign DNA in transgenic plants.

Science.gov (United States)

Meyer, P; Heidmann, I

1994-05-25

We analysed de novo DNA methylation occurring in plants obtained from the transgenic petunia line R101-17. This line contains one copy of the maize A1 gene that leads to the production of brick-red pelargonidin pigment in the flowers. Due to its integration into an unmethylated genomic region the A1 transgene is hypomethylated and transcriptionally active. Several epigenetic variants of line 17 were selected that exhibit characteristic and somatically stable pigmentation patterns, displaying fully coloured, marbled or colourless flowers. Analysis of the DNA methylation patterns revealed that the decrease in pigmentation among the epigenetic variants was correlated with an increase in methylation, specifically of the transgene DNA. No change in methylation of the hypomethylated integration region could be detected. A similar increase in methylation, specifically in the transgene region, was also observed among progeny of R101-17del, a deletion derivative of R101-17 that no longer produces pelargonidin pigments due to a deletion in the A1 coding region. Again de novo methylation is specifically directed to the transgene, while the hypomethylated character of neighbouring regions is not affected. Possible mechanisms for transgene-specific methylation and its consequences for long-term use of transgenic material are discussed.

GH/IGF-I Transgene Expression on Muscle Homeostasis

Science.gov (United States)

Schwartz, Robert J.

1999-01-01

We propose to test the hypothesis that the growth hormone/ insulin like growth factor-I axis through autocrine/paracrine mechanisms may provide long term muscle homeostasis under conditions of prolonged weightlessness. As a key alternative to hormone replacement therapy, ectopic production of hGH, growth hormone releasing hormone (GHRH), and IGF-I will be studied for its potential on muscle mass impact in transgenic mice under simulated microgravity. Expression of either hGH or IGF-I would provide a chronic source of a growth-promoting protein whose biosynthesis or secretion is shut down in space. Muscle expression of the IGF-I transgene has demonstrated about a 20% increase in hind limb muscle mass over control nontransgenic litter mates. These recent experiments, also establish the utility of hind-limb suspension in mice as a workable model to study atrophy in weight bearing muscles. Thus, transgenic mice will be used in hind-limb suspension models to determine the role of GH/IGF-I on maintenance of muscle mass and whether concentric exercises might act in synergy with hormone treatment. As a means to engineer and ensure long-term protein production that would be workable in humans, gene therapy technology will be used by to monitor muscle mass preservation during hind-limb suspension, after direct intramuscular injection of a genetically engineered muscle-specific vector expressing GHRH. Effects of this gene-based therapy will be assessed in both fast twitch (medial gastrocnemius) and slow twitch muscle (soleus). End-points include muscle size, ultrastructure, fiber type, and contractile function, in normal animals, hind limb suspension, and reambutation.

DESIGN AND IMPLEMENTATION OF A CO2 FLOOD UTILIZING ADVANCED RESERVOIR CHARACTERIZATION AND HORIZONTAL INJECTION WELLS IN A SHALLOW SHELF CARBONATE APPROACHING WATERFLOOD DEPLETION

Energy Technology Data Exchange (ETDEWEB)

K.J. Harpole; Ed G. Durrett; Susan Snow; J.S. Bles; Carlon Robertson; C.D. Caldwell; D.J. Harms; R.L. King; B.A. Baldwin; D. Wegener; M. Navarrette

2002-09-01

The purpose of this project was to economically design an optimum carbon dioxide (CO{sub 2}) flood for a mature waterflood nearing its economic abandonment. The original project utilized advanced reservoir characterization and CO{sub 2} horizontal injection wells as the primary methods to redevelop the South Cowden Unit (SCU). The development plans; project implementation and reservoir management techniques were to be transferred to the public domain to assist in preventing premature abandonment of similar fields. The Unit was a mature waterflood with water cut exceeding 95%. Oil must be mobilized through the use of a miscible or near-miscible fluid to recover significant additional reserves. Also, because the unit was relatively small, it did not have the benefit of economies of scale inherent in normal larger scale projects. Thus, new and innovative methods were required to reduce investment and operating costs. Two primary methods used to accomplish improved economics were use of reservoir characterization to restrict the flood to the higher quality rock in the unit and use of horizontal injection wells to cut investment and operating costs. The project consisted of two budget phases. Budget Phase I started in June 1994 and ended late June 1996. In this phase Reservoir Analysis, Characterization Tasks and Advanced Technology Definition Tasks were completed. Completion enabled the project to be designed, evaluated, and an Authority for Expenditure (AFE) for project implementation submitted to working interest owners for approval. Budget Phase II consisted of the implementation and execution of the project in the field. Phase II was completed in July 2001. Performance monitoring, during Phase II, by mid 1998 identified the majority of producing wells which under performed their anticipated withdrawal rates. Newly drilled and re-activated wells had lower offtake rates than originally forecasted. As a result of poor offtake, higher reservoir pressure was a concern

Co3O4 protective coatings prepared by Pulsed Injection Metal Organic Chemical Vapour Deposition

DEFF Research Database (Denmark)

Burriel, M.; Garcia, G.; Santiso, J.

2005-01-01

of deposition temperature. Pure Co3O4 spinel structure was found for deposition temperatures ranging from 360 to 540 degreesC. The optimum experimental parameters to prepare dense layers with a high growth rate were determined and used to prepare corrosion protective coatings for Fe-22Cr metallic interconnects......Cobalt oxide films were grown by Pulsed Injection Metal Organic Chemical Vapour Deposition (PI-MOCVD) using Co(acac)(3) (acac=acetylacetonate) precursor dissolved in toluene. The structure, morphology and growth rate of the layers deposited on silicon substrates were studied as a function......, to be used in Intermediate Temperature Solid Oxide Fuel Cells. (C) 2004 Elsevier B.V. All rights reserved....

Combining M-FISH and Quantum Dot technology for fast chromosomal assignment of transgenic insertions

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Yusuf Mohammed

2011-12-01

Full Text Available Abstract Background Physical mapping of transgenic insertions by Fluorescence in situ Hybridization (FISH is a reliable and cost-effective technique. Chromosomal assignment is commonly achieved either by concurrent G-banding or by a multi-color FISH approach consisting of iteratively co-hybridizing the transgenic sequence of interest with one or more chromosome-specific probes at a time, until the location of the transgenic insertion is identified. Results Here we report a technical development for fast chromosomal assignment of transgenic insertions at the single cell level in mouse and rat models. This comprises a simplified 'single denaturation mixed hybridization' procedure that combines multi-color karyotyping by Multiplex FISH (M-FISH, for simultaneous and unambiguous identification of all chromosomes at once, and the use of a Quantum Dot (QD conjugate for the transgene detection. Conclusions Although the exploitation of the unique optical properties of QD nanocrystals, such as photo-stability and brightness, to improve FISH performance generally has been previously investigated, to our knowledge this is the first report of a purpose-designed molecular cytogenetic protocol in which the combined use of QDs and standard organic fluorophores is specifically tailored to assist gene transfer technology.

Final Scientific/Technical Report for project “Geomechanical Monitoring for CO₂ Hub Storage: Production and Injection at Kevin Dome

Energy Technology Data Exchange (ETDEWEB)

Daley, Thomas M. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Vasco, Don [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Ajo-Franklin, Jonathan [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States); Dobeck, Laura [Montana State Univ., Bozeman, MT (United States); Spangler, Lee [Montana State Univ., Bozeman, MT (United States); Leonti, Michelle [Montana State Univ., Bozeman, MT (United States)

2017-06-27

After learning that the TDS value in the target injection formation at the Kevin Dome site is too low to qualify for an EPA Class VI CO2 injection permit, the BSCSP project was re-scoped such that injection of CO2 is no longer planned. With no injection planned, the Geomechanics project was closed. In this final report, we describe the objective and approach of the project as proposed, and the limited results obtained before stopping work. The objective of the proposed research was the development & validation of an integrated monitoring approach for quantifying the interactions between large-scale geological carbon storage (GCS) and subsurface geomechanical state, particularly perturbations relevant to reservoir integrity such as fault reactivation and induced fracturing. In the short period of work before knowing the fate of the Kevin Dome project, we (1) researched designs for both the proposed InSAR corner reflectors as well as the near-surface 3C seismic stations; (2) developed preliminary elastic geomechanical models; (3) developed a second generation deformation prediction for the BSCSP Kevin Dome injection site; and (4) completed a preliminary map of InSAR monuments and shallow MEQ wells in the vicinity of the BSCSP injection pad.

Impact of hydrodynamic injection and phiC31 integrase on tumor latency in a mouse model of MYC-induced hepatocellular carcinoma.

Directory of Open Access Journals (Sweden)

Lauren E Woodard

2010-06-01

Full Text Available Hydrodynamic injection is an effective method for DNA delivery in mouse liver and is being translated to larger animals for possible clinical use. Similarly, phiC31 integrase has proven effective in mediating long-term gene therapy in mice when delivered by hydrodynamic injection and is being considered for clinical gene therapy applications. However, chromosomal aberrations have been associated with phiC31 integrase expression in tissue culture, leading to questions about safety.To study whether hydrodynamic delivery alone, or in conjunction with delivery of phiC31 integrase for long-term transgene expression, could facilitate tumor formation, we used a transgenic mouse model in which sustained induction of the human C-MYC oncogene in the liver was followed by hydrodynamic injection. Without injection, mice had a median tumor latency of 154 days. With hydrodynamic injection of saline alone, the median tumor latency was significantly reduced, to 105 days. The median tumor latency was similar, 106 days, when a luciferase donor plasmid and backbone plasmid without integrase were administered. In contrast, when active or inactive phiC31 integrase and donor plasmid were supplied to the mouse liver, the median tumor latency was 153 days, similar to mice receiving no injection.Our data suggest that phiC31 integrase does not facilitate tumor formation in this C-MYC transgenic mouse model. However, in groups lacking phiC31 integrase, hydrodynamic injection appeared to contribute to C-MYC-induced hepatocellular carcinoma in adult mice. Although it remains to be seen to what extent these findings may be extrapolated to catheter-mediated hydrodynamic delivery in larger species, they suggest that caution should be used during translation of hydrodynamic injection to clinical applications.

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

Science.gov (United States)

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

2008-09-01

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

Development of transgenic cotton lines expressing Allium sativum agglutinin (ASAL) for enhanced resistance against major sap-sucking pests.

Science.gov (United States)

Vajhala, Chakravarthy S K; Sadumpati, Vijaya Kumar; Nunna, Hariprasad Rao; Puligundla, Sateesh Kumar; Vudem, Dashavantha Reddy; Khareedu, Venkateswara Rao

2013-01-01

Mannose-specific Allium sativum leaf agglutinin encoding gene (ASAL) and herbicide tolerance gene (BAR) were introduced into an elite cotton inbred line (NC-601) employing Agrobacterium-mediated genetic transformation. Cotton transformants were produced from the phosphinothricin (PPT)-resistant shoots obtained after co-cultivation of mature embryos with the Agrobacterium strain EHA105 harbouring recombinant binary vector pCAMBIA3300-ASAL-BAR. PCR and Southern blot analysis confirmed the presence and stable integration of ASAL and BAR genes in various transformants of cotton. Basta leaf-dip assay, northern blot, western blot and ELISA analyses disclosed variable expression of BAR and ASAL transgenes in different transformants. Transgenes, ASAL and BAR, were stably inherited and showed co-segregation in T1 generation in a Mendelian fashion for both PPT tolerance and insect resistance. In planta insect bioassays on T2 and T3 homozygous ASAL-transgenic lines revealed potent entomotoxic effects of ASAL on jassid and whitefly insects, as evidenced by significant decreases in the survival, development and fecundity of the insects when compared to the untransformed controls. Furthermore, the transgenic cotton lines conferred higher levels of resistance (1-2 score) with minimal plant damage against these major sucking pests when bioassays were carried out employing standard screening techniques. The developed transgenics could serve as a potential genetic resource in recombination breeding aimed at improving the pest resistance of cotton. This study represents the first report of its kind dealing with the development of transgenic cotton resistant to two major sap-sucking insects.

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

Science.gov (United States)

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

2016-08-02

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

Transgenic Expression of the Anti-parasitic Factor TEP1 in the Malaria Mosquito Anopheles gambiae.

Directory of Open Access Journals (Sweden)

Gloria Volohonsky

2017-01-01

Full Text Available Mosquitoes genetically engineered to be resistant to Plasmodium parasites represent a promising novel approach in the fight against malaria. The insect immune system itself is a source of anti-parasitic genes potentially exploitable for transgenic designs. The Anopheles gambiae thioester containing protein 1 (TEP1 is a potent anti-parasitic protein. TEP1 is secreted and circulates in the mosquito hemolymph, where its activated cleaved form binds and eliminates malaria parasites. Here we investigated whether TEP1 can be used to create malaria resistant mosquitoes. Using a GFP reporter transgene, we determined that the fat body is the main site of TEP1 expression. We generated transgenic mosquitoes that express TEP1r, a potent refractory allele of TEP1, in the fat body and examined the activity of the transgenic protein in wild-type or TEP1 mutant genetic backgrounds. Transgenic TEP1r rescued loss-of-function mutations, but did not increase parasite resistance in the presence of a wild-type susceptible allele. Consistent with previous reports, TEP1 protein expressed from the transgene in the fat body was taken up by hemocytes upon a challenge with injected bacteria. Furthermore, although maturation of transgenic TEP1 into the cleaved form was impaired in one of the TEP1 mutant lines, it was still sufficient to reduce parasite numbers and induce parasite melanization. We also report here the first use of Transcription Activator Like Effectors (TALEs in Anopheles gambiae to stimulate expression of endogenous TEP1. We found that artificial elevation of TEP1 expression remains moderate in vivo and that enhancement of endogenous TEP1 expression did not result in increased resistance to Plasmodium. Taken together, our results reveal the difficulty of artificially influencing TEP1-mediated Plasmodium resistance, and contribute to further our understanding of the molecular mechanisms underlying mosquito resistance to Plasmodium parasites.

CDNA encoding a polypeptide including a hevein sequence

Science.gov (United States)

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

1995-03-21

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

INVESTIGATION OF EFFICIENCY IMPROVEMENTS DURING CO2 INJECTION IN HYDRAULICALLY AND NATURALLY FRACTURED RESERVOIRS

Energy Technology Data Exchange (ETDEWEB)

David S. Schechter

2005-04-27

This report describes the work performed during the fourth year of the project, ''Investigating of Efficiency Improvements during CO{sub 2} Injection in Hydraulically and Naturally Fractured Reservoirs.'' The objective of this project is to perform unique laboratory experiments with artificially fractured cores (AFCs) and X-ray CT scanner to examine the physical mechanisms of bypassing in hydraulically fractured reservoirs (HFR) and naturally fractured reservoirs (NFR) that eventually result in more efficient CO{sub 2} flooding in heterogeneous or fracture-dominated reservoirs. In Chapter 1, we worked with DOE-RMOTC to investigate fracture properties in the Tensleep Formation at Teapot Dome Naval Reserve as part of their CO{sub 2} sequestration project. In Chapter 2, we continue our investigation to determine the primary oil recovery mechanism in a short vertically fractured core. Finally in Chapter 3, we report our numerical modeling efforts to develop compositional simulator with irregular grid blocks.

Small RNAs were involved in homozygous state-associated silencing of a marker gene (Neomycin phosphotransferase II: nptII) in transgenic tomato plants.

Science.gov (United States)

Deng, Lei; Pan, Yu; Chen, Xuqing; Chen, Guoping; Hu, Zongli

2013-07-01

Homozygous state-associated co-suppression is not a very common phenomenon. In our experiments, two transgenic plants 3A29 and 1195A were constructed by being transformed with the constructs pBIN-353A and pBIN119A containing nptII gene as a marker respectively. The homozygous progeny from these two independent transgenic lines 3A29 and 1195A, displayed kanamycin-sensitivity and produced a short main root without any lateral roots as untransformed control (wild-type) seedlings when germinated on kanamycin media. For the seedlings derived from putative hemizygous plants, the percentage of the seedlings showing normal growth on kanamycin media was about 50% and lower than the expected percentage (75%). Southern analysis of the genomic DNA confirmed that the homozygous and hemizygous plants derived from the same lines contained the same multiple nptII transgenes, which were located on the same site of chromosome. Northern analysis suggested that the marker nptII gene was expressed in the primary and the hemizygous transformants, but it was silenced in the homozygous transgenic plants. Further Northern analysis indicated that antisense and sense small nptII-derived RNAs were present in the transgenic plants and the blotting signal of nptII-derived small RNA was much higher in the homozygous transgenic plants than that of hemizygous transgenic plants. Additionally, read-through transcripts from the TRAMP gene to the nptII gene were detected. These results suggest that the read-through transcripts may be involved in homozygous state-associated silencing of the nptII transgene in transgenic tomato plants and a certain threshold level of the nptII-derived small RNAs is required for the homozygous state-associated co-suppression of the nptII transgene. Copyright © 2013 Elsevier Masson SAS. All rights reserved.

Glyphostate-drift but not herbivory alters the rate of transgene flow from single and stacked trait transgenic canola (Brassica napus L.) to non-transgenic B. napus and B. rapa

Science.gov (United States)

While transgenic plants can offer agricultural benefits, the escape of transgenes out of crop fields is a major environmental concern. Escape of transgenic herbicide resistance has occurred between transgenic Brassica napus (canola) and weedy species in numerous locations. In t...

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

African Journals Online (AJOL)

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

Measuring oxygen yields of a thermal conversion/elemental analyzer-isotope ratio mass spectrometer for organic and inorganic materials through injection of CO.

Science.gov (United States)

Yin, Xijie; Chen, Zhigang

2014-12-01

The thermal conversion/elemental analyzer-isotope ratio mass spectrometer (TC/EA-IRMS) is widely used to measure the δ(18) O value of various substances. A premise for accurate δ(18) O measurement is that the oxygen in the sample can be converted into carbon monoxide (CO) quantitatively or at least proportionally. Therefore, a precise method to determine the oxygen yield of TC/EA-IRMS measurements is needed. Most studies have used the CO peak area obtained from a known amount of a solid reference material (for example, benzoic acid) to calibrate the oxygen yield of the sample. Although it was assumed that the oxygen yield of the solid reference material is 100%, no direct evidence has been provided. As CO is the analyte gas for δ(18) O measurement by IRMS, in this study, we use a six-port valve to inject CO gas into the TC/EA. The CO is carried to the IRMS by the He carrier gas and the CO peak area is measured by the IRMS. The CO peak area thus obtained from a known amount of the injected CO is used to calibrate the oxygen yield of the sample. The oxygen yields of commonly used organic and inorganic reference materials such as benzoic acid (C6 H5 COOH), silver phosphate (Ag3 PO4 ), calcium carbonate (CaCO3 ) and silicon dioxide (SiO2 ) are investigated at different reactor temperatures and sample sizes. We obtained excellent linear correlation between the peak area for the injected CO and its oxygen atom amount. C6 H5 COOH has the highest oxygen yield, followed by Ag3 PO4 , CaCO3 and SiO2 . The oxygen yields of TC/EA-IRMS are less than 100% for both organic and inorganic substances, but the yields are relatively stable at the specified reactor temperature and for a given quantity of sample. Copyright © 2014 John Wiley & Sons, Ltd.

Deep microbial life in the Altmark natural gas reservoir: baseline characterization prior CO2 injection

Science.gov (United States)

Morozova, Daria; Shaheed, Mina; Vieth, Andrea; Krüger, Martin; Kock, Dagmar; Würdemann, Hilke

2010-05-01

Within the framework of the CLEAN project (CO2 Largescale Enhanced gas recovery in the Altmark Natural gas field) technical basics with special emphasis on process monitoring are explored by injecting CO2 into a gas reservoir. Our study focuses on the investigation of the in-situ microbial community of the Rotliegend natural gas reservoir in the Altmark, located south of the city Salzwedel, Germany. In order to characterize the microbial life in the extreme habitat we aim to localize and identify microbes including their metabolism influencing the creation and dissolution of minerals. The ability of microorganisms to speed up dissolution and formation of minerals might result in changes of the local permeability and the long-term safety of CO2 storage. However, geology, structure and chemistry of the reservoir rock and the cap rock as well as interaction with saline formation water and natural gases and the injected CO2 affect the microbial community composition and activity. The reservoir located at the depth of about 3500m, is characterised by high salinity fluid and temperatures up to 127° C. It represents an extreme environment for microbial life and therefore the main focus is on hyperthermophilic, halophilic anaerobic microorganisms. In consequence of the injection of large amounts of CO2 in the course of a commercial EGR (Enhanced Gas Recovery) the environmental conditions (e.g. pH, temperature, pressure and solubility of minerals) for the autochthonous microorganisms will change. Genetic profiling of amplified 16S rRNA genes are applied for detecting structural changes in the community by using PCR- SSCP (PCR-Single-Strand-Conformation Polymorphism) and DGGE (Denaturing Gradient Gel Electrophoresis). First results of the baseline survey indicate the presence of microorganisms similar to representatives from other saline, hot, anoxic, deep environments. However, due to the hypersaline and hyperthermophilic reservoir conditions, cell numbers are low, so that

Method of controlling injection of oxygen into hydrogen-rich fuel cell feed stream

Science.gov (United States)

Meltser, Mark Alexander; Gutowski, Stanley; Weisbrod, Kirk

2001-01-01

A method of operating a H.sub.2 --O.sub.2 fuel cell fueled by hydrogen-rich fuel stream containing CO. The CO content is reduced to acceptable levels by injecting oxygen into the fuel gas stream. The amount of oxygen injected is controlled in relation to the CO content of the fuel gas, by a control strategy that involves (a) determining the CO content of the fuel stream at a first injection rate, (b) increasing the O.sub.2 injection rate, (c) determining the CO content of the stream at the higher injection rate, (d) further increasing the O.sub.2 injection rate if the second measured CO content is lower than the first measured CO content or reducing the O.sub.2 injection rate if the second measured CO content is greater than the first measured CO content, and (e) repeating steps a-d as needed to optimize CO consumption and minimize H.sub.2 consumption.

Groundwater Chemistry Changes as a Result of CO2 Injection at the ZERT Field Site in Bozeman, Montana

Energy Technology Data Exchange (ETDEWEB)

Apps, J.A.; Birkholzer, J.T.; Spycher, N.; Zheng, L.; Herkelrath, W.N.; Kharaka, Y.K.; Thordsen, J.J.; Kakouros, E.; Beers, S; Gullickson, K.S.; Spangler, L.H.; Ambats, G.

2009-11-01

Combustion of fossil fuels produces CO{sub 2}, a common greenhouse gas linked to global climate change. Separation of CO{sub 2}from emissions produced by large industrial point sources like power plants, cement kilns and refineries, and injection deep nderground into geologic formations is one method of preventing CO{sub 2} releases into the atmosphere. This process is referred to as Carbon Capture and Storage (CCS). CCS is one of several solutions being considered to mitigate global climate change. Other solutions nclude increased energy efficiency, renewables, nuclear power, advanced coal, and plug-in hybrid electric vehicles.

A Semi-Analytical Method for Rapid Estimation of Near-Well Saturation, Temperature, Pressure and Stress in Non-Isothermal CO2 Injection

Science.gov (United States)

LaForce, T.; Ennis-King, J.; Paterson, L.

2015-12-01

Reservoir cooling near the wellbore is expected when fluids are injected into a reservoir or aquifer in CO2 storage, enhanced oil or gas recovery, enhanced geothermal systems, and water injection for disposal. Ignoring thermal effects near the well can lead to under-prediction of changes in reservoir pressure and stress due to competition between increased pressure and contraction of the rock in the cooled near-well region. In this work a previously developed semi-analytical model for immiscible, nonisothermal fluid injection is generalised to include partitioning of components between two phases. Advection-dominated radial flow is assumed so that the coupled two-phase flow and thermal conservation laws can be solved analytically. The temperature and saturation profiles are used to find the increase in reservoir pressure, tangential, and radial stress near the wellbore in a semi-analytical, forward-coupled model. Saturation, temperature, pressure, and stress profiles are found for parameters representative of several CO2 storage demonstration projects around the world. General results on maximum injection rates vs depth for common reservoir parameters are also presented. Prior to drilling an injection well there is often little information about the properties that will determine the injection rate that can be achieved without exceeding fracture pressure, yet injection rate and pressure are key parameters in well design and placement decisions. Analytical solutions to simplified models such as these can quickly provide order of magnitude estimates for flow and stress near the well based on a range of likely parameters.

cDNA encoding a polypeptide including a hevein sequence

Energy Technology Data Exchange (ETDEWEB)

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

1993-02-16

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

Modeling of time-lapse multi-scale seismic monitoring of CO2 injected into a fault zone to enhance the characterization of permeability in enhanced geothermal systems

Science.gov (United States)

Zhang, R.; Borgia, A.; Daley, T. M.; Oldenburg, C. M.; Jung, Y.; Lee, K. J.; Doughty, C.; Altundas, B.; Chugunov, N.; Ramakrishnan, T. S.

2017-12-01

Subsurface permeable faults and fracture networks play a critical role for enhanced geothermal systems (EGS) by providing conduits for fluid flow. Characterization of the permeable flow paths before and after stimulation is necessary to evaluate and optimize energy extraction. To provide insight into the feasibility of using CO2 as a contrast agent to enhance fault characterization by seismic methods, we model seismic monitoring of supercritical CO2 (scCO2) injected into a fault. During the CO2 injection, the original brine is replaced by scCO2, which leads to variations in geophysical properties of the formation. To explore the technical feasibility of the approach, we present modeling results for different time-lapse seismic methods including surface seismic, vertical seismic profiling (VSP), and a cross-well survey. We simulate the injection and production of CO2 into a normal fault in a system based on the Brady's geothermal field and model pressure and saturation variations in the fault zone using TOUGH2-ECO2N. The simulation results provide changing fluid properties during the injection, such as saturation and salinity changes, which allow us to estimate corresponding changes in seismic properties of the fault and the formation. We model the response of the system to active seismic monitoring in time-lapse mode using an anisotropic finite difference method with modifications for fracture compliance. Results to date show that even narrow fault and fracture zones filled with CO2 can be better detected using the VSP and cross-well survey geometry, while it would be difficult to image the CO2 plume by using surface seismic methods.

[New advances in animal transgenic technology].

Science.gov (United States)

Sun, Zhen-Hong; Miao, Xiang-Yang; Zhu, Rui-Liang

2010-06-01

Animal transgenic technology is one of the fastest growing biotechnology in the 21st century. It is used to integrate foreign genes into the animal genome by genetic engineering technology so that foreign genes can be expressed and inherited to the offspring. The transgenic efficiency and precise control of gene expression are the key limiting factors on preparation of transgenic animals. A variety of transgenic techniques are available, each of which has its own advantages and disadvantages and still needs further study because of unresolved technical and safety issues. With the in-depth research, the transgenic technology will have broad application prospects in the fields of exploration of gene function, animal genetic improvement, bioreactor, animal disease models, organ transplantation and so on. This article reviews the recently developed animal gene transfer techniques, including germline stem cell mediated method to improve the efficiency, gene targeting to improve the accuracy, RNA interference (RNAi)-mediated gene silencing technology, and the induced pluripotent stem cells (iPS) transgenic technology. The new transgenic techniques can provide a better platform for the study of trans-genic animals and promote the development of medical sciences, livestock production, and other fields.

Petrophysical and rock-mechanics effects of CO2 injection for enhanced oil recovery

DEFF Research Database (Denmark)

Alam, Mohammad Monzurul; Hjuler, Morten Leth; Christensen, Helle Foged

2014-01-01

this issue we studied two types of chalk from South Arne field, North Sea: (1) Ekofisk Formation having >12% non-carbonate and (2) Tor Formation, which has less than 5% non-carbonate. We performed a series of laboratory experiments to reveal the changes in petrophysical and rock-mechanics properties due...... reservoirs. North Sea chalk is characterized by high porosity but also high specific surface causing low permeability. A high porosity provides room for CO2 storage, while a high specific surface causes a high risk for chemical reaction and consequently for mechanical weakening. In order to address...... to the injection of CO2 at supercritical state. We analyzed these changes with respect to the differences in porosity, specific surface, pore stiffness, wettability, mineralogy and mechanical strength. We observed a 2–3% increase in porosity, a minor decrease of specific surface and consequently a small increase...

D22S15 - a fetal brain cDNA with BanII and SacI RFLP

Energy Technology Data Exchange (ETDEWEB)

Rouleau, G A; Kurnit, D M; Neve, R L; Bazanowsky, A; Patterson, D; Gusella, J F

1988-02-25

A .58 kb single copy EcoRI fragment was isolated from a human fetal brain cDNA library and cloned into pBR322. This fragment recognizes a two allele polymorphism when used to probe human genomic DNA digested with SacI. There are no constant bands. Additional polymorphisms recognized by BanII and Bsp1286 are in disequilibrium with the BanII polymorphism. It has been localized to chromosome 22 by somatic cell hybrid analysis and linkage analysis. Co-dominant segregation has been observed in 15 informative families.

CO2 injection into fractured peridotites: a reactive percolation experiment

Science.gov (United States)

Escario, S.; Godard, M.; Gouze, P.; Leprovost, R.; Luquot, L.; Garcia-Rios, M.

2017-12-01

Mantle peridotites have the potential to trap CO2 as carbonates. This process observed in ophiolites and in oceanic environments provides a long term and safe storage for CO2. It occurs as a part of a complex suite of fluid-rock reactions involving silicate dissolution and precipitation of hydrous phases, carbonates and minor phases that may in turn modify the hydrodynamic properties and the reactivity of the reacted rocks. The efficiency and lastingness of the process require the renewal of fluids at the mineral-fluid interface. Fractures are dominant flow paths in exhumed mantle sections. This study aims at better understanding the effect of CO2-enriched saline fluids on hydrodynamic and chemical processes through fractured peridotites. Experiments were performed using the reactive percolation bench ICARE Lab 3 - Géosciences Montpellier. It allows monitoring the permeability changes during experiments. Effluents are recurrently sampled for analysing cation concentration, pH and alkalinity. Reacted rock samples were characterized by high resolution X-ray microtomography (ESRF ID19, Grenoble, France) and SEM. Experiments consisted in injecting CO2-enriched brines (NaCl 0.5 M) at a rate of 6 mL.h-1 into artificially fractured cores (9 mm diameter × 20 mm length) of Oman harzburgites at T=170°C and Ptotal = 25 MPa for up to 2 weeks. Fractures are of few µm apertures with rough walls. Three sets of experiments were performed at increasing value of [CO2] (0, 0.1 and 1 mol/kg). All experiments showed a decrease in permeability followed by steady state regime that can be caused by a decrease in the roughness of fracture walls (dissolution dominated process), thus favouring fracture closing, or by the precipitation of secondary phases. Maximum enrichments in Mg, Fe and Ca of the effluent fluids occur during the first 2 hours of the experiments whereas Si displays a maximum enrichment at t = 20 h, suggesting extensive dissolution. Maximum enrichments are observed with

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

DEFF Research Database (Denmark)

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

1991-01-01

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

The SbASR-1 gene cloned from an extreme halophyte Salicornia brachiata enhances salt tolerance in transgenic tobacco.

Science.gov (United States)

Jha, Bhavanath; Lal, Sanjay; Tiwari, Vivekanand; Yadav, Sweta Kumari; Agarwal, Pradeep K

2012-12-01

Salinity severely affects plant growth and development. Plants evolved various mechanisms to cope up stress both at molecular and cellular levels. Halophytes have developed better mechanism to alleviate the salt stress than glycophytes, and therefore, it is advantageous to study the role of different genes from halophytes. Salicornia brachiata is an extreme halophyte, which grows luxuriantly in the salty marshes in the coastal areas. Earlier, we have isolated SbASR-1 (abscisic acid stress ripening-1) gene from S. brachiata using cDNA subtractive hybridisation library. ASR-1 genes are abscisic acid (ABA) responsive, whose expression level increases under abiotic stresses, injury, during fruit ripening and in pollen grains. The SbASR-1 transcript showed up-regulation under salt stress conditions. The SbASR-1 protein contains 202 amino acids of 21.01-kDa molecular mass and has 79 amino acid long signatures of ABA/WDS gene family. It has a maximum identity (73 %) with Solanum chilense ASR-1 protein. The SbASR-1 has a large number of disorder-promoting amino acids, which make it an intrinsically disordered protein. The SbASR-1 gene was over-expressed under CaMV 35S promoter in tobacco plant to study its physiological functions under salt stress. T(0) transgenic tobacco seeds showed better germination and seedling growth as compared to wild type (Wt) in a salt stress condition. In the leaf tissues of transgenic lines, Na(+) and proline contents were significantly lower, as compared to Wt plant, under salt treatment, suggesting that transgenic plants are better adapted to salt stress.

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

Science.gov (United States)

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

2017-04-12

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

Heterogeneity of rat tropoelastin mRNA revealed by cDNA cloning

International Nuclear Information System (INIS)

Pierce, R.A.; Deak, S.B.; Stolle, C.A.; Boyd, C.D.

1990-01-01

A λgt11 library constructed from poly(A+) RNA isolated from aortic tissue of neonatal rats was screened for rat tropoelastin cDNAs. The first, screen, utilizing a human tropoelastin cDNA clone, provided rat tropoelastin cDNAs spanning 2.3 kb of carboxy-terminal coding sequence and extended into the 3'-untranslated region. A subsequent screen using a 5' rat tropoelastin cDNA clone yielded clones extending into the amino-terminal signal sequence coding region. Sequence analysis of these clones has provided the complete derived amino acid sequence of rat tropoelastin and allowed alignment and comparison with published bovine cDNA sequence. While the overall structure of rat tropoelastin is similar to bovine sequence, numerous substitutions, deletions, and insertions demonstrated considerable heterogeneity between species. In particular, the pentapeptide repeat VPGVG, characteristic of all tropoelastins analyzed to date, is replaced in rat tropoelastin by a repeating pentapeptide, IPGVG. The hexapeptide repeat VGVAPG, the bovine elastin receptor binding peptide, is not encoded by rat tropoelastin cDNAs. Variations in coding sequence between rat tropoelastin CDNA clones were also found which may represent mRNA heterogeneity produced by alternative splicing of the rat tropoelastin pre-mRNA

Neuroanatomy and transgenic technologies

Science.gov (United States)

This is a short review that introduces recent advances of neuroanatomy and transgenic technologies. The anatomical complexity of the nervous system remains a subject of tremendous fascination among neuroscientists. In order to tackle this extraordinary complexity, powerful transgenic technologies a...

Peptidomics combined with cDNA library unravel the diversity of centipede venom

DEFF Research Database (Denmark)

Rong, Mingqiang; Yang, Shilong; Wen, Bo

2015-01-01

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

Biotechnology network promotes knowledge of transgenics

International Nuclear Information System (INIS)

Blanco Picado, Patricia; Valdez Melara, Marta

2015-01-01

Red de Ingenieria Genetica Aplicada al Mejoramiento de Cultivos Tropicales (Rigatrop) integrated by a group of scientists from the Universidad de Costa Rica (UCR), Universidad Nacional (UNA) and of the Instituto Tecnologico de Costa Rica (TEC) have organized two forums on the topic of transgenics. The first forum has shown successful experiences of development of transgenic crops in Latin America, as for example: the transgenic bean, project realized in Brazil and transgenic eggplant in Bangladesh. The second forum has been about transgenics and environment effected at the UCR, on the occasion of World Environment Day. Rigatrop members are working currently in two projects applying biotechnological tools to coffee [es

Molecular cloning and nucleotide sequence of cDNA for human liver arginase

International Nuclear Information System (INIS)

Haraguchi, Y.; Takiguchi, M.; Amaya, Y.; Kawamoto, S.; Matsuda, I.; Mori, M.

1987-01-01

Arginase (EC3.5.3.1) catalyzes the last step of the urea cycle in the liver of ureotelic animals. Inherited deficiency of the enzyme results in argininemia, an autosomal recessive disorder characterized by hyperammonemia. To facilitate investigation of the enzyme and gene structures and to elucidate the nature of the mutation in argininemia, the authors isolated cDNA clones for human liver arginase. Oligo(dT)-primed and random primer human liver cDNA libraries in λ gt11 were screened using isolated rat arginase cDNA as a probe. Two of the positive clones, designated λ hARG6 and λ hARG109, contained an overlapping cDNA sequence with an open reading frame encoding a polypeptide of 322 amino acid residues (predicted M/sub r/, 34,732), a 5'-untranslated sequence of 56 base pairs, a 3'-untranslated sequence of 423 base pairs, and a poly(A) segment. Arginase activity was detected in Escherichia coli cells transformed with the plasmid carrying λ hARG6 cDNA insert. RNA gel blot analysis of human liver RNA showed a single mRNA of 1.6 kilobases. The predicted amino acid sequence of human liver arginase is 87% and 41% identical with those of the rat liver and yeast enzymes, respectively. There are several highly conserved segments among the human, rat, and yeast enzymes

CoJet technology for oxygen injection injection in EAFS - industrial gases supply and customers productivity improvements; Tecnologia CoJet para injecao de oxigenio em FEAS - fornecimento de gases e aumento da produtividade dos clientes

Energy Technology Data Exchange (ETDEWEB)

Marques, Izaias Lucas; Zagury, Pedro Athias [White Martins Gases Industriais Ltda., Rio de Janeiro, RJ (Brazil)

2010-07-01

PRAXAIR is one of the largest industrial gases producers of the World, being present in more than 40 countries and strongly connected to the steel industry. PRAXAIR has experience to supply products and services to all kinds of steel mills. Several technologies developed by the Company, such as CoJet (a system to coherent oxygen jet injection in EAF and BOF furnaces), Slag Splashing (splashing slag to cover BOF furnaces walls increasing refractory wear campaign life), AOD (a process for specialty steel production) and others, are currently in commercial operation in different parts of the world. In Brazil, Praxair is the owner of White Martins, the largest industrial gases company in South America, present in nine countries of the continent. Company's portfolio includes atmospheric gases, carbonic gas production, acetylene, hydrogen, specialty and health and care gases, mixtures for welding, gas seamless steel cylinders, applications equipment, and storage and gases transportation. The Company also operates a natural gas liquefaction unit. In addition to technologies and process, Praxair / White Martins can also supply solutions for measurement, control and gases injection, according to strict safety and quality standards. The present work has the objective to show the benefits of the CoJet EAF System, oxygen injectors positioned in the furnace walls. In 2009, two systems have been installed in South American mills and the results achieved will be following discussed. (author)

Electron injection mechanisms of green organic light-emitting devices fabricated utilizing a double electron injection layer consisting of cesium carbonate and fullerene

International Nuclear Information System (INIS)

Yang, J.S.; Choo, D.C.; Kim, T.W.; Jin, Y.Y.; Seo, J.H.; Kim, Y.K.

2010-01-01

Electron injection mechanisms of the luminance efficiency of green organic light-emitting devices (OLEDs) fabricated utilizing a cesium carbonate (Cs 2 CO 3 )/fullerene (C 60 ) heterostructure acting as an electron injection layer (EIL) were investigated. Current density-voltage and luminance-voltage measurements showed that the current densities and the luminances of the OLEDs with a Cs 2 CO 3 or Cs 2 CO 3 /C 60 EIL were higher than that of the OLEDs with a Liq EIL. The luminance efficiency of the OLEDs with a Cs 2 CO 3 EIL was almost three times higher than that of the OLEDs with a Liq EIL. Because the electron injection efficiency of the Cs 2 CO 3 layer in OLEDs was different from that of the C 60 layer, the luminance efficiency of the OLEDs with a double EIL consisting of a Cs 2 CO 3 layer and a C 60 layer was smaller than that of the OLEDs with a Cs 2 CO 3 EIL. The electron injection mechanisms of OLEDs with a Cs 2 CO 3 and C 60 double EIL are described on the basis of the experimental results.

Modified composite microspheres of hydroxyapatite and poly(lactide-co-glycolide) as an injectable scaffold

Energy Technology Data Exchange (ETDEWEB)

Hu, Xixue [BNLMS, State Key Laboratory of Polymer Physics and Chemistry, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 (China); CAS Key Laboratory for Biomedical Effects of Nanomaterials and Nanosafety, National Center for Nanoscience and Technology, Beijing 100190 (China); Shen, Hong, E-mail: shenhong516@iccas.ac.cn [BNLMS, State Key Laboratory of Polymer Physics and Chemistry, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 (China); Yang, Fei [BNLMS, State Key Laboratory of Polymer Physics and Chemistry, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 (China); Liang, Xinjie [CAS Key Laboratory for Biomedical Effects of Nanomaterials and Nanosafety, National Center for Nanoscience and Technology, Beijing 100190 (China); Wang, Shenguo, E-mail: wangsg@iccas.ac.cn [BNLMS, State Key Laboratory of Polymer Physics and Chemistry, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 (China); Wu, Decheng, E-mail: dcwu@iccas.ac.cn [BNLMS, State Key Laboratory of Polymer Physics and Chemistry, Institute of Chemistry, Chinese Academy of Sciences, Beijing 100190 (China)

2014-02-15

The compound of hydroxyapatite-poly(lactide-co-glycolide) (HA-PLGA) was prepared by ionic bond between HA and PLGA. HA-PLGA was more stable than the simple physical blend of hydroxyapatite and poly(lactide-co-glycolide) (HA/PLGA). The surface of HA-PLGA microsphere fabricated by an emulsion–solvent evaporation method was rougher than that of HA/PLGA microspheres. Moreover, surface HA content of HA-PLGA microspheres was more than that of HA/PLGA microspheres. In vitro mouse OCT-1 osteoblast-like cell culture results showed that the HA-PLGA microspheres clearly promoted osteoblast attachment, proliferation and alkaline phosphatase activity. It was considered that surface rich HA component and rough surface of HA-PLGA microsphere enhanced cell growth and differentiation. The good cell affinity of the HA-PLGA microspheres indicated that they could be used as an injectable scaffold for bone tissue engineering.

Modified composite microspheres of hydroxyapatite and poly(lactide-co-glycolide) as an injectable scaffold

International Nuclear Information System (INIS)

Hu, Xixue; Shen, Hong; Yang, Fei; Liang, Xinjie; Wang, Shenguo; Wu, Decheng

2014-01-01

The compound of hydroxyapatite-poly(lactide-co-glycolide) (HA-PLGA) was prepared by ionic bond between HA and PLGA. HA-PLGA was more stable than the simple physical blend of hydroxyapatite and poly(lactide-co-glycolide) (HA/PLGA). The surface of HA-PLGA microsphere fabricated by an emulsion–solvent evaporation method was rougher than that of HA/PLGA microspheres. Moreover, surface HA content of HA-PLGA microspheres was more than that of HA/PLGA microspheres. In vitro mouse OCT-1 osteoblast-like cell culture results showed that the HA-PLGA microspheres clearly promoted osteoblast attachment, proliferation and alkaline phosphatase activity. It was considered that surface rich HA component and rough surface of HA-PLGA microsphere enhanced cell growth and differentiation. The good cell affinity of the HA-PLGA microspheres indicated that they could be used as an injectable scaffold for bone tissue engineering.

Interfacial, electrical, and spin-injection properties of epitaxial Co2MnGa grown on GaAs(100)

DEFF Research Database (Denmark)

Damsgaard, Christian Danvad; Hickey, M. C.; Holmes, S. N.

2009-01-01

The interfacial, electrical, and magnetic properties of the Heusler alloy Co2MnGa grown epitaxially on GaAs(100) are presented with an emphasis on the use of this metal-semiconductor combination for a device that operates on the principles of spin-injection between the two materials. Through...... was monitored in situ by reflection high energy electron diffraction and the bulk composition was measured ex situ with inductively coupled plasma optical emission spectroscopy. The Co2MnGa L21 cubic structure is strained below a thickness of 20 nm on GaAs(100) but relaxed in films thicker than 20 nm...

Display of a Maize cDNA library on baculovirus infected insect cells

Directory of Open Access Journals (Sweden)

Jones Ian M

2008-08-01

Full Text Available Abstract Background Maize is a good model system for cereal crop genetics and development because of its rich genetic heritage and well-characterized morphology. The sequencing of its genome is well advanced, and new technologies for efficient proteomic analysis are needed. Baculovirus expression systems have been used for the last twenty years to express in insect cells a wide variety of eukaryotic proteins that require complex folding or extensive posttranslational modification. More recently, baculovirus display technologies based on the expression of foreign sequences on the surface of Autographa californica (AcMNPV have been developed. We investigated the potential of a display methodology for a cDNA library of maize young seedlings. Results We constructed a full-length cDNA library of young maize etiolated seedlings in the transfer vector pAcTMVSVG. The library contained a total of 2.5 × 105 independent clones. Expression of two known maize proteins, calreticulin and auxin binding protein (ABP1, was shown by western blot analysis of protein extracts from insect cells infected with the cDNA library. Display of the two proteins in infected insect cells was shown by selective biopanning using magnetic cell sorting and demonstrated proof of concept that the baculovirus maize cDNA display library could be used to identify and isolate proteins. Conclusion The maize cDNA library constructed in this study relies on the novel technology of baculovirus display and is unique in currently published cDNA libraries. Produced to demonstrate proof of principle, it opens the way for the development of a eukaryotic in vivo display tool which would be ideally suited for rapid screening of the maize proteome for binding partners, such as proteins involved in hormone regulation or defence.

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

DEFF Research Database (Denmark)

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

2006-01-01

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

Numerical simulation methods applied to injection and storage of CO{sub 2} in saline aquifers; Metodos de simulacion numerica aplicados a la inyeccion y almacenamiento de CO{sub 2} en formaciones salinas

Energy Technology Data Exchange (ETDEWEB)

Arjona Garcia-Borreguero, J.; Rodriguez Pons-Esparver, R.; Iglesias Lopez, A.

2015-07-01

One of the Climate Change mitigation proposals suggested by the IPCC (Intergovernmental Panel on Climate Change) in its Synthesis Report 2007 involves the launch of applications for capturing and storing carbon dioxide, existing three different geological structures suitable for gas storage: oil and gas depleted reservoirs, useless coal layers and deep saline structures. In case of deep saline structures, the main problem to prepare a study of CO{sub 2} storage is the difficulty of obtaining geological data for some selected structure with characteristics that could be suitable for injection and gas storage. According to this situation, the solution to analyze the feasibility of a storage project in a geological structure will need numerical simulation from a 3D terrain model. Numerical methods allow the simulation of the carbon dioxide filling in saline structures from a well, used to inject gas with a particular flow. This paper presents a methodology to address the modeling and simulation process of CO{sub 2} injection into deep saline aquifers. (Author)

Effects of CO2 injection and Kerogen Maturation on Low-Field Nuclear Magnetic Resonance Response

Science.gov (United States)

Prasad, M.; Livo, K.

2017-12-01

Low-field Nuclear Magnetic Resonance (NMR) is commonly used in petrophysical analysis of petroleum reservoir rocks. NMR experiments record the relaxation and polarization of in-situ hydrogen protons present in gaseous phases such as free-gas intervals and solution gas fluids, bulk fluid phases such as oil and aquifer intervals, and immovable fractions of kerogen and bitumen. Analysis of NMR relaxation spectra is performed to record how fluid composition, maturity, and viscosity change NMR experimental results. We present T1-T2 maps as thermal maturity of a water-saturated, sub-mature Woodford shale is increased at temperatures from 125 to 400 degrees Celsius. Experiments with applied fluid pressure in paraffinic mineral oil and DI water with varying fluid pH have been performed to mimic reservoir conditions in analysis of the relaxation of bulk fluid phases. We have recorded NMR spectra, T1-T2 maps, and fluid diffusion coefficients using a low-field (2 MHz) MagritekTM NMR. CO2 was injected at a pressure of 900 psi in an in house developed NMR pressure vessel made of torlon plastic. Observable 2D NMR shifts in immature kerogen formations as thermal maturity is increased show generation of lighter oils with increased maturity. CO2 injection leads to a decrease in bulk fluid relaxation time that is attributed to viscosity modification with gas presence. pH variation with increased CO2 presence were shown to not effect NMR spectra. From this, fluid properties have been shown to greatly affect NMR readings and must be taken into account for more accurate NMR reservoir characterization.

Ubiquitous overexpression of a transgene encoding the extracellular portion of the Drosophila roughest-irregular chiasm C protein induces early embryonic lethality.

Science.gov (United States)

Moda, L; Machado, R C; Ramos, R G

2000-09-01

The cell adhesion molecule Rst-irreC is a transmembrane glycoprotein of the immunoglobulin superfamily involved in several important developmental processes in Drosophila, including axonal pathfinding in the optic lobe and programmed cell death and pigment cell differentiation in the pupal retina. As an initial step towards the "in vivo" functional analysis of this protein we have generated transgenic fly stocks carrying a truncated cDNA construct encoding only the extracellular domain of Rst-IrreC under the transcriptional control of the heat shock inducible promoter hsp70. We show that heat-shocking embryos bearing the transgene during the first 8hs of development lead to a 3-4 fold reduction in their viability compared to wild type controls. The embryonic lethality can already be produced by applying the heat pulse in the first 3hs of embryonic development, does not seem to be suppressed in the absence of wildtype product and is progressively reduced as the heat treatment is applied later in embryogenesis. These results are compatible with the hypothesis of the lethal phenotype being primarily due to heterophilic interactions between Rst-IrreC extracellular domain and an yet unknown ligand.

Correlated Inflammatory Responses and Neurodegeneration in Peptide-Injected Animal Models of Alzheimer’s Disease

Directory of Open Access Journals (Sweden)

James G. McLarnon

2014-01-01

Full Text Available Animal models of Alzheimer’s disease (AD which emphasize activation of microglia may have particular utility in correlating proinflammatory activity with neurodegeneration. This paper reviews injection of amyloid-β (Aβ into rat brain as an alternative AD animal model to the use of transgenic animals. In particular, intrahippocampal injection of Aβ1-42 peptide demonstrates prominent microglial mobilization and activation accompanied by a significant loss of granule cell neurons. Furthermore, pharmacological inhibition of inflammatory reactivity is demonstrated by a broad spectrum of drugs with a common endpoint in conferring neuroprotection in peptide-injected animals. Peptide-injection models provide a focus on glial cell responses to direct peptide injection in rat brain and offer advantages in the study of the mechanisms underlying neuroinflammation in AD brain.

cDNA encoding a polypeptide including a hevein sequence

Energy Technology Data Exchange (ETDEWEB)

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

2000-07-04

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

cDNA encoding a polypeptide including a hevein sequence

Energy Technology Data Exchange (ETDEWEB)

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

1999-05-04

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

cDNA encoding a polypeptide including a hevein sequence

Energy Technology Data Exchange (ETDEWEB)

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

1999-05-04

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

cDNA encoding a polypeptide including a hevein sequence

Energy Technology Data Exchange (ETDEWEB)

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

1995-03-21

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

Multi-Organ Damage in Human Dipeptidyl Peptidase 4 Transgenic Mice Infected with Middle East Respiratory Syndrome-Coronavirus.

Directory of Open Access Journals (Sweden)

Guangyu Zhao

Full Text Available The Middle East Respiratory Syndrome Coronavirus (MERS-CoV causes severe acute respiratory failure and considerable extrapumonary organ dysfuction with substantial high mortality. For the limited number of autopsy reports, small animal models are urgently needed to study the mechanisms of MERS-CoV infection and pathogenesis of the disease and to evaluate the efficacy of therapeutics against MERS-CoV infection. In this study, we developed a transgenic mouse model globally expressing codon-optimized human dipeptidyl peptidase 4 (hDPP4, the receptor for MERS-CoV. After intranasal inoculation with MERS-CoV, the mice rapidly developed severe pneumonia and multi-organ damage, with viral replication being detected in the lungs on day 5 and in the lungs, kidneys and brains on day 9 post-infection. In addition, the mice exhibited systemic inflammation with mild to severe pneumonia accompanied by the injury of liver, kidney and spleen with neutrophil and macrophage infiltration. Importantly, the mice exhibited symptoms of paralysis with high viral burden and viral positive neurons on day 9. Taken together, this study characterizes the tropism of MERS-CoV upon infection. Importantly, this hDPP4-expressing transgenic mouse model will be applicable for studying the pathogenesis of MERS-CoV infection and investigating the efficacy of vaccines and antiviral agents designed to combat MERS-CoV infection.

Transgene mobilization and regulatory uncertainty for non-GE fruit products of transgenic rootstocks.

Science.gov (United States)

Haroldsen, Victor M; Chi-Ham, Cecilia L; Bennett, Alan B

2012-10-31

Genetically engineered (GE) rootstocks may offer some advantages for biotechnology applications especially in woody perennial crops such as grape or walnut. Transgrafting combines horticultural grafting practices with modern GE methods for crop improvement. Here, a non-GE conventional scion (upper stem portion) is grafted onto a transgenic GE rootstock. Thus, the scion does not contain the genetic modification present in the rootstock genome. We examined transgene presence in walnut and tomato GE rootstocks and non-GE fruit-bearing scions. Mobilization of transgene DNA, protein, and mRNA across the graft was not detected. Though transgenic siRNA mobilization was not observed in grafted tomatoes or walnut scions, transgenic siRNA signal was detected in walnut kernels. Prospective benefits from transgrafted plants include minimized risk of GE pollen flow (Lev-Yadun and Sederoff, 2001), possible use of more than one scion per approved GE rootstock which could help curb the estimated US$136 million (CropLife International, 2011) cost to bring a GE crop to international markets, as well as potential for improved consumer and market acceptance since the consumable product is not itself GE. Thus, transgrafting provides an alternative option for agricultural industries wishing to expand their biotechnology portfolio. Copyright © 2012 Elsevier B.V. All rights reserved.

Studies on chemically crosslinkable carboxy terminated-poly(propylene fumarate-co-ethylene glycol)-acrylamide hydrogel as an injectable biomaterial

International Nuclear Information System (INIS)

Kallukalam, B C; Jayabalan, M; Sankar, V

2009-01-01

Carboxy terminated-poly(propylene fumarate)-co-ethylene glycol) (CT-PPF-co-PEG) was prepared and set into crosslinked hydrogel material with acrylamide. The setting studies reveal that this copolymer system can be used as an injectable material. The hydrogel material exhibits a higher degree of swelling, good mechanical strength and flexibility. The hydrogel favours adhesion of L929 fibroblast cells without proliferation on the surface. However, cardiac fibroblast cells (isolated from new born rat (Wistar) hearts) adhere and proliferate on the hydrogel due to the formation of synergistic hydrophilic-hydrophobic surface-by-surface reorganization.

Studies on chemically crosslinkable carboxy terminated-poly(propylene fumarate-co-ethylene glycol)-acrylamide hydrogel as an injectable biomaterial

Energy Technology Data Exchange (ETDEWEB)

Kallukalam, B C; Jayabalan, M [Polymer Division, Biomedical Technology Wing, Sree Chitra Tirunal Institute for Medical Sciences and Technology, Thiruvananthapuram 695 012 (India); Sankar, V, E-mail: muthujayabalan@rediffmail.co [Division of Cellular and Molecular Cardiology, Sree Chitra Tirunal Institute for Medical Sciences and Technology, Thiruvananthapuram 695 012 (India)

2009-02-15

Carboxy terminated-poly(propylene fumarate)-co-ethylene glycol) (CT-PPF-co-PEG) was prepared and set into crosslinked hydrogel material with acrylamide. The setting studies reveal that this copolymer system can be used as an injectable material. The hydrogel material exhibits a higher degree of swelling, good mechanical strength and flexibility. The hydrogel favours adhesion of L929 fibroblast cells without proliferation on the surface. However, cardiac fibroblast cells (isolated from new born rat (Wistar) hearts) adhere and proliferate on the hydrogel due to the formation of synergistic hydrophilic-hydrophobic surface-by-surface reorganization.

Construction and Cloning of Reporter-Tagged Replicon cDNA for an In Vitro Replication Study of Murine Norovirus-1 (MNV-1).

Science.gov (United States)

Ahmad, Muhammad Khairi; Tabana, Yasser M; Ahmed, Mowaffaq Adam; Sandai, Doblin Anak; Mohamed, Rafeezul; Ismail, Ida Shazrina; Zulkiflie, Nurulisa; Yunus, Muhammad Amir

2017-12-01

A norovirus maintains its viability, infectivity and virulence by its ability to replicate. However, the biological mechanisms of the process remain to be explored. In this work, the NanoLuc™ Luciferase gene was used to develop a reporter-tagged replicon system to study norovirus replication. The NanoLuc™ Luciferase reporter protein was engineered to be expressed as a fusion protein for MNV-1 minor capsid protein, VP2. The foot-and-mouth disease virus 2A (FMDV2A) sequence was inserted between the 3'end of the reporter gene and the VP2 start sequence to allow co-translational 'cleavage' of fusion proteins during intracellular transcript expression. Amplification of the fusion gene was performed using a series of standard and overlapping polymerase chain reactions. The resulting amplicon was then cloned into three readily available backbones of MNV-1 cDNA clones. Restriction enzyme analysis indicated that the NanoLucTM Luciferase gene was successfully inserted into the parental MNV-1 cDNA clone. The insertion was further confirmed by using DNA sequencing. NanoLuc™ Luciferase-tagged MNV-1 cDNA clones were successfully engineered. Such clones can be exploited to develop robust experimental assays for in vitro assessments of viral RNA replication.

The nucleotide sequence of human transition protein 1 cDNA

Energy Technology Data Exchange (ETDEWEB)

Luerssen, H; Hoyer-Fender, S; Engel, W [Universitaet Goettingen (West Germany)

1988-08-11

The authors have screened a human testis cDNA library with an oligonucleotide of 81 mer prepared according to a part of the published nucleotide sequence of the rat transition protein TP 1. They have isolated a cDNA clone with the length of 441 bp containing the coding region of 162 bp for human transition protein 1. There is about 84% homology in the coding region of the sequence compared to rat. The human cDNA-clone encodes a polypeptide of 54 amino acids of which 7 are different to that of rat.

The temporal expression pattern of alpha-synuclein modulates olfactory neurogenesis in transgenic mice.

Directory of Open Access Journals (Sweden)

Sebastian R Schreglmann

Full Text Available Adult neurogenesis mirrors the brain´s endogenous capacity to generate new neurons throughout life. In the subventricular zone/ olfactory bulb system adult neurogenesis is linked to physiological olfactory function and has been shown to be impaired in murine models of neuronal alpha-Synuclein overexpression. We analyzed the degree and temporo-spatial dynamics of adult olfactory bulb neurogenesis in transgenic mice expressing human wild-type alpha-Synuclein (WTS under the murine Thy1 (mThy1 promoter, a model known to have a particularly high tg expression associated with impaired olfaction.Survival of newly generated neurons (NeuN-positive in the olfactory bulb was unchanged in mThy1 transgenic animals. Due to decreased dopaminergic differentiation a reduction in new dopaminergic neurons within the olfactory bulb glomerular layer was present. This is in contrast to our previously published data on transgenic animals that express WTS under the control of the human platelet-derived growth factor β (PDGF promoter, that display a widespread decrease in survival of newly generated neurons in regions of adult neurogenesis, resulting in a much more pronounced neurogenesis deficit. Temporal and quantitative expression analysis using immunofluorescence co-localization analysis and Western blots revealed that in comparison to PDGF transgenic animals, in mThy1 transgenic animals WTS is expressed from later stages of neuronal maturation only but at significantly higher levels both in the olfactory bulb and cortex.The dissociation between higher absolute expression levels of alpha-Synuclein but less severe impact on adult olfactory neurogenesis in mThy1 transgenic mice highlights the importance of temporal expression characteristics of alpha-Synuclein on the maturation of newborn neurons.

Secondary and tertiary gas injection in fractured carbonate rock: Experimental study

Energy Technology Data Exchange (ETDEWEB)

Karimaie, H.; Torsaeter, O. [SPE, NTNU (Norway); Darvish, G.R. [SPE, STATOIL (Norway); Lindeberg, E. [SPE, SINTEF (Norway)

2008-09-15

The use of CO{sub 2} has received considerable interest as a method of EOR but a major drawback is its availability and increasing cost. Therefore, as the number of CO{sub 2} injection projects increase, an alternative must be considered to meet the economic considerations. For this reason attention has been directed to nitrogen injection which may be a good substitute for CO{sub 2}. The purpose of the experiments described in this paper was to investigate the efficiency of oil recovery by CO{sub 2} and N{sub 2} in fractured carbonate rock. The combined effects of gravity drainage and component exchange between gas in fracture and oil in matrix on oil recovery in fractured reservoirs subjected to CO{sub 2} or nitrogen gas injection are experimentally studied. Laboratory experiments have been carried out on a low permeable outcrop chalk, as an analogue to a North Sea reservoir rock. This was surrounded by a fracture, established with a novel experimental set-up. The experiments aimed to investigate the potential of oil recovery by secondary and tertiary CO{sub 2} and nitrogen gas injection at high pressure high temperature condition. The matrix block was saturated using recombined binary mixture live oil (C{sub 1}-C{sub 7}), while the fracture was filled with a sealing material to obtain a homogeneous saturation. The sealing material was then removed by increasing the temperature which in turn creates the fracture surrounding the core. Gas was injected into the fracture at pressures above the bubble point of the oil. Oil recovery as a function of time was monitored during the experiments. Results from secondary gas injection experiments indicate that CO{sub 2} injection at elevated pressure and temperature is more efficient than N{sub 2} injection. Results from tertiary gas injection experiments also show that injection of CO{sub 2} could significantly recover the oil, even after waterflooding, compared to N{sub 2} injection. (author)

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

CSIR Research Space (South Africa)

Van den Berg, N

2004-11-01

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

Construction and identification of subtracted cDNA library in bone marrow cells of radon-exposed mice

International Nuclear Information System (INIS)

Li Jianxiang; Nie Jihua; Tong Jian; Fu Chunling; Zhou Jianwei

2008-01-01

Objective: To construct and identify subtracted cDNA library in bone marrow cells of mice exposed to radon inhalation. Methods: Adult male BALB/c mice, weighing 18-22 g, were placed in a multi- functional radon chamber. One group of mice was exposed to radon up to the accumulative dose of 105 work level month (WLM). The control group of mice was housed in a room with an accumulative dose of 1 WLM. To construct a subtracted cDNA library enriched with differentially expressed genes, the SMART technique and the suppression subtractive hybridization were performed. The obtained forward and reverse cDNA fragments were directly inserted into pMD18-T vector and transformed into E. coli JM109. The inserting cDNA fragments were screened by the blue-and-white blot screening and nested PCR of bacterium liquid. Results: The 244 of 285 white bacteria clones obtained randomly were positive clones contained 100-1100 bp inserted cDNA fragments. Conclusions: The forward and reverse subtracted cDNA library in bone marrow cells of mice exposed to radon inhalation is successfully constructed. (authors)

Enhanced motivation to alcohol in transgenic mice expressing human α-synuclein.

Science.gov (United States)

Rotermund, Carola; Reolon, Gustavo K; Leixner, Sarah; Boden, Cindy; Bilbao, Ainhoa; Kahle, Philipp J

2017-11-01

α-Synuclein (αSYN) is the neuropathological hallmark protein of Parkinson's disease (PD) and related neurodegenerative disorders. Moreover, the gene encoding αSYN (SNCA) is a major genetic contributor to PD. Interestingly, independent genome-wide association studies also identified SNCA as the most important candidate gene for alcoholism. Furthermore, single-nucleotide-polymorphisms have been associated with alcohol-craving behavior and alcohol-craving patients showed augmented αSYN expression in blood. To investigate the effect of αSYN on the addictive properties of chronic alcohol use, we examined consumption, motivation, and seeking responses induced by environmental stimuli and relapse behavior in transgenic mice expressing the human mutant [A30P]αSYN throughout the brain. The primary reinforcing effects of alcohol under operant self-administration conditions were increased, while consumption and the alcohol deprivation effect were not altered in the transgenic mice. The same mice were subjected to immunohistochemical measurements of immediate-early gene inductions in brain regions involved in addiction-related behaviors. Acute ethanol injection enhanced immunostaining for the phosphorylated form of cAMP response element binding protein in both amygdala and nucleus accumbens of αSYN transgenic mice, while in wild-type mice no effect was visible. However, at the same time, levels of cFos remain unchanged in both genotypes. These results provide experimental confirmation of SNCA as a candidate gene for alcoholism in addition to its known link to PD. © 2017 International Society for Neurochemistry.

Molecular characterization of medium-chain acyl-CoA dehydrogenase (MCAD) deficiency

DEFF Research Database (Denmark)

Gregersen, N; Andresen, B S; Bross, P

1991-01-01

. All clones sequenced from the patient exhibited a single base substitution from adenine (A) to guanine (G) at position 985 in the MCAD cDNA as the only consistent base-variation compared with control cDNA. In contrast, the parents contained cDNA with the normal and the mutated sequence, revealing......A series of experiments has established the molecular defect in the medium-chain acyl-coenzyme A (CoA) dehydrogenase (MCAD) gene in a family with MCAD deficiency. Demonstration of intra-mitochondrial mature MCAD indistinguishable in size (42.5-kDa) from control MCAD, and of mRNA with the correct...... size of 2.4 kb, indicated a point-mutation in the coding region of the MCAD gene to be disease-causing. Consequently, cloning and DNA sequencing of polymerase chain reaction (PCR) amplified complementary DNA (cDNA) from messenger RNA of fibroblasts from the patient and family members were performed...

The cDNA sequence of a neutral horseradish peroxidase.

Science.gov (United States)

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

1991-02-16

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

[Construction of fetal mesenchymal stem cell cDNA subtractive library].

Science.gov (United States)

Yang, Li; Wang, Dong-Mei; Li, Liang; Bai, Ci-Xian; Cao, Hua; Li, Ting-Yu; Pei, Xue-Tao

2002-04-01

To identify differentially expressed genes between fetal mesenchymal stem cell (MSC) and adult MSC, especially specified genes expressed in fetal MSC, a cDNA subtractive library of fetal MSC was constructed using suppression subtractive hybridization (SSH) technique. At first, total RNA was isolated from fetal and adult MSC. Using SMART PCR synthesis method, single-strand and double-strand cDNAs were synthesized. After Rsa I digestion, fetal MSC cDNAs were divided into two groups and ligated to adaptor 1 and adaptor 2 respectively. Results showed that the amplified library contains 890 clones. Analysis of 890 clones with PCR demonstrated that 768 clones were positive. The positive rate is 86.3%. The size of inserted fragments in these positive clones was between 0.2 - 1 kb, with an average of 400 - 600 bp. SSH is a convenient and effective method for screening differentially expressed genes. The constructed cDNA subtractive library of fetal MSC cDNA lays solid foundation for screening and cloning new and specific function related genes of fetal MSC.

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

International Nuclear Information System (INIS)

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

2009-01-01

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

Styl RFLP recognized by a human IRBP cDNA localized to chromosome 10

Energy Technology Data Exchange (ETDEWEB)

Chin, K S; Mathew, C G.P.; Fong, S L; Bridges, C D; Ponder, B A.J.

1988-02-25

A 2184 bp cDNA (H.4 IRBP) encoding human interstitial retinol-biding protein isolated from a human retina cDNA library in lambdagt10 by screening with a bovine IRBP cDNA probe. Styl identifies a 2-allele polymorphism with bands at 2.3 kb (Cl) and 1.95 kb (C2) and invariant bands at 1.1, 1.0 and 0.8kb. Codominant segregation was observed in two informative families. The RFLP was mapped to chromosome 10 using somatic cell hybrids. In situ hybridization suggests regional assignments near p11.2 -q11.2 with a secondary site of hybridization at q24-25.

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

Science.gov (United States)

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

2017-01-01

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

Agrobacterium rhizogenes rolB gene affects photosynthesis and chlorophyll content in transgenic tomato (Solanum lycopersicum L.) plants.

Science.gov (United States)

Bettini, Priscilla P; Marvasi, Massimiliano; Fani, Fabiola; Lazzara, Luigi; Cosi, Elena; Melani, Lorenzo; Mauro, Maria Luisa

2016-10-01

Insertion of Agrobacterium rhizogenes rolB gene into plant genome affects plant development, hormone balance and defence. However, beside the current research, the overall transcriptional response and gene expression of rolB as a modulator in plant is unknown. Transformed rolB tomato plant (Solanum lycopersicum L.) cultivar Tondino has been used to investigate the differential expression profile. Tomato is a well-known model organism both at the genetic and molecular level, and one of the most important commercial food crops in the world. Through the construction and characterization of a cDNA subtracted library, we have investigated the differential gene expression between transgenic clones of rolB and control tomato and have evaluated genes specifically transcribed in transgenic rolB plants. Among the selected genes, five genes encoding for chlorophyll a/b binding protein, carbonic anhydrase, cytochrome b 6 /f complex Fe-S subunit, potassium efflux antiporter 3, and chloroplast small heat-shock protein, all involved in chloroplast function, were identified. Measurement of photosynthesis efficiency by the level of three different photosynthetic parameters (F v /F m , rETR, NPQ) showed rolB significant increase in non-photochemical quenching and a, b chlorophyll content. Our results point to highlight the role of rolB on plant fitness by improving photosynthesis. Copyright © 2016 Elsevier GmbH. All rights reserved.

Functional imaging of interleukin 1 beta expression in inflammatory process using bioluminescence imaging in transgenic mice

Directory of Open Access Journals (Sweden)

Liu Zhihui

2008-08-01

Full Text Available Abstract Background Interleukin 1 beta (IL-1β plays an important role in a number of chronic and acute inflammatory diseases. To understand the role of IL-1β in disease processes and develop an in vivo screening system for anti-inflammatory drugs, a transgenic mouse line was generated which incorporated the transgene firefly luciferase gene driven by a 4.5-kb fragment of the human IL-1β gene promoter. Luciferase gene expression was monitored in live mice under anesthesia using bioluminescence imaging in a number of inflammatory disease models. Results In a LPS-induced sepsis model, dramatic increase in luciferase activity was observed in the mice. This transgene induction was time dependent and correlated with an increase of endogenous IL-1β mRNA and pro-IL-1β protein levels in the mice. In a zymosan-induced arthritis model and an oxazolone-induced skin hypersensitivity reaction model, luciferase expression was locally induced in the zymosan injected knee joint and in the ear with oxazolone application, respectively. Dexamethasone suppressed the expression of luciferase gene both in the acute sepsis model and in the acute arthritis model. Conclusion Our data suggest that the transgenic mice model could be used to study transcriptional regulation of the IL-1β gene expression in the inflammatory process and evaluation the effect of anti-inflammatory drug in vivo.

Enhancement of cell-based therapeutic angiogenesis using a novel type of injectable scaffolds of hydroxyapatite-polymer nanocomposite microspheres.

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Yohei Mima

Full Text Available BACKGROUND: Clinical trials demonstrate the effectiveness of cell-based therapeutic angiogenesis in patients with severe ischemic diseases; however, their success remains limited. Maintaining transplanted cells in place are expected to augment the cell-based therapeutic angiogenesis. We have reported that nano-hydroxyapatite (HAp coating on medical devices shows marked cell adhesiveness. Using this nanotechnology, HAp-coated poly(l-lactic acid (PLLA microspheres, named nano-scaffold (NS, were generated as a non-biological, biodegradable and injectable cell scaffold. We investigate the effectiveness of NS on cell-based therapeutic angiogenesis. METHODS AND RESULTS: Bone marrow mononuclear cells (BMNC and NS or control PLLA microspheres (LA were intramuscularly co-implanted into mice ischemic hindlimbs. When BMNC derived from enhanced green fluorescent protein (EGFP-transgenic mice were injected into ischemic muscle, the muscle GFP level in NS+BMNC group was approximate fivefold higher than that in BMNC or LA+BMNC groups seven days after operation. Kaplan-Meier analysis demonstrated that NS+BMNC markedly prevented hindlimb necrosis (P<0.05 vs. BMNC or LA+BMNC. NS+BMNC revealed much higher induction of angiogenesis in ischemic tissues and collateral blood flow confirmed by three-dimensional computed tomography angiography than those of BMNC or LA+BMNC groups. NS-enhanced therapeutic angiogenesis and arteriogenesis showed good correlations with increased intramuscular levels of vascular endothelial growth factor and fibroblast growth factor-2. NS co-implantation also prevented apoptotic cell death of transplanted cells, resulting in prolonged cell retention. CONCLUSION: A novel and feasible injectable cell scaffold potentiates cell-based therapeutic angiogenesis, which could be extremely useful for the treatment of severe ischemic disorders.

Radioactive cDNA microarray in neurospsychiatry

International Nuclear Information System (INIS)

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

2003-01-01

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

Radioactive cDNA microarray in neurospsychiatry

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

2003-02-01

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

Cattle mammary bioreactor generated by a novel procedure of transgenic cloning for large-scale production of functional human lactoferrin.

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Penghua Yang

Full Text Available Large-scale production of biopharmaceuticals by current bioreactor techniques is limited by low transgenic efficiency and low expression of foreign proteins. In general, a bacterial artificial chromosome (BAC harboring most regulatory elements is capable of overcoming the limitations, but transferring BAC into donor cells is difficult. We describe here the use of cattle mammary bioreactor to produce functional recombinant human lactoferrin (rhLF by a novel procedure of transgenic cloning, which employs microinjection to generate transgenic somatic cells as donor cells. Bovine fibroblast cells were co-microinjected for the first time with a 150-kb BAC carrying the human lactoferrin gene and a marker gene. The resulting transfection efficiency of up to 15.79 x 10(-2 percent was notably higher than that of electroporation and lipofection. Following somatic cell nuclear transfer, we obtained two transgenic cows that secreted rhLF at high levels, 2.5 g/l and 3.4 g/l, respectively. The rhLF had a similar pattern of glycosylation and proteolytic susceptibility as the natural human counterpart. Biochemical analysis revealed that the iron-binding and releasing properties of rhLF were identical to that of native hLF. Importantly, an antibacterial experiment further demonstrated that rhLF was functional. Our results indicate that co-microinjection with a BAC and a marker gene into donor cells for somatic cell cloning indeed improves transgenic efficiency. Moreover, the cattle mammary bioreactors generated with this novel procedure produce functional rhLF on an industrial scale.

Human tissue factor: cDNA sequence and chromosome localization of the gene

International Nuclear Information System (INIS)

Scarpati, E.M.; Wen, D.; Broze, G.J. Jr.; Miletich, J.P.; Flandermeyer, R.R.; Siegel, N.R.; Sadler, J.E.

1987-01-01

A human placenta cDNA library in λgt11 was screened for the expression of tissue factor antigens with rabbit polyclonal anti-human tissue factor immunoglobulin G. Among 4 million recombinant clones screened, one positive, λHTF8, expressed a protein that shared epitopes with authentic human brain tissue factor. The 1.1-kilobase cDNA insert of λHTF8 encoded a peptide that contained the amino-terminal protein sequence of human brain tissue factor. Northern blotting identified a major mRNA species of 2.2 kilobases and a minor species of ∼ 3.2 kilobases in poly(A) + RNA of placenta. Only 2.2-kilobase mRNA was detected in human brain and in the human monocytic U937 cell line. In U937 cells, the quantity of tissue factor mRNA was increased several fold by exposure of the cells to phorbol 12-myristate 13-acetate. Additional cDNA clones were selected by hybridization with the cDNA insert of λHTF8. These overlapping isolates span 2177 base pairs of the tissue factor cDNA sequence that includes a 5'-noncoding region of 75 base pairs, an open reading frame of 885 base pairs, a stop codon, a 3'-noncoding region of 1141 base pairs, and a poly(a) tail. The open reading frame encodes a 33-kilodalton protein of 295 amino acids. The predicted sequence includes a signal peptide of 32 or 34 amino acids, a probable extracellular factor VII binding domain of 217 or 219 amino acids, a transmembrane segment of 23 acids, and a cytoplasmic tail of 21 amino acids. There are three potential glycosylation sites with the sequence Asn-X-Thr/Ser. The 3'-noncoding region contains an inverted Alu family repetitive sequence. The tissue factor gene was localized to chromosome 1 by hybridization of the cDNA insert of λHTF8 to flow-sorted human chromosomes

Verification of zinc injection applicability to Japanese BWRs

International Nuclear Information System (INIS)

Hosokawa, H.; Uetake, N.; Ishida, K.; Nakamura, M.; Mochizuki, K.; Nagata, T.; Ogawa, N.; Baba, T.; Ono, S.; Ishigure, K.

2002-01-01

The verification test program on zinc injection applicability to Japanese BWRs was started in 1997. Laboratory tests using high temperature water loops under BWR reactor water conditions are in progress. This paper is an interim report on results obtained so far. Co-58 and Zn-65 were simultaneously used in the Co radioactivity buildup test to evaluate zinc injection suppression effects towards cobalt deposition on pre-oxidized stainless steel. The following results were obtained. The Co deposition was suppressed effectively by Zn injection, even when there was a pre-oxide film. For the test piping that had the pre-oxide film formed under the NWC (normal water chemistry) condition, when soaked under the HWC (hydrogen water chemistry) condition a large amount of Co-58 was taken into a small part of the inner layer. The distribution ratio of Co-58 in the inner layer and outer layer of the oxide film was almost the same for both the pre-oxidized test piping and the non pre-oxidized test piping under the HWC condition. Zn injection decreased the Co concentration in the inner layer of the oxide film under all conditions. The effect of gamma ray irradiation on the incorporation of Co into the oxide film is small. Gamma ray irradiation influenced the Co incorporation behavior complexly. The change of the Co amount incorporated into the non-pre-filmed specimen were small relatively, but the incorporation into the pre-filmed specimen promotes distinctly. Zn addition effectively suppresses the Co incorporation even gamma ray irradiation. (authors)