Sequencing of chemotherapy (CT) and radiotherapy (RT) following conservative surgery (CS) for patients with early-stage breast cancer: results of a randomized trial

Energy Technology Data Exchange (ETDEWEB)

Recht, A; Come, SE; Silver, B; Gelman, RS; Hayes, DF; Shulman, LN; Henderson, IC; Harris, JR

1995-07-01

Purpose: To determine the effects of different sequences of CT and RT following CS for pts with clinical Stage I or II breast carcinoma. Methods and Materials: From 6/84-12/92, 244 pts were randomized following CS to receive CT (4 cycles of CAMFP at 3-wk intervals) either before or after RT. Median pt age was 45 yrs (range, 20-68). 164 pts were pre-, 18 pts peri-, and 62 pts were postmenopausal. 209 pts were node-positive (61 had 4 or more positive nodes); after 6/88, 35 selected node-negative pts were also included. 45 Gy was prescribed to the entire breast, followed by a boost of 16 Gy; nodal RT was optional. The medians ( and ranges) of the surgery-radiotherapy interval (SRI) according to assigned arm were: RT-first, 36 days (14-234); CT-first, 126 days (98-185). Drug doses were based on ideal body weight. Doses (per square meter) and schedule were: methotrexate, 200 mg, d.1 and 15; leucovorin, 10 mg, q6H x 12, start d.2 and 16; 5-fluorouracil, 500 mg, d.1; cyclophosphamide, 500 mg, d.1; prednisone, 40 mg/day x 5d, start d.1; and doxorubicin, 45 mg, d.3. All except leucovorin and prednisone were given by IV bolus. Dose reductions were based on the granulocyte nadir from the previous cycle, with an attempt to reescalate doses in subsequent cycles. Tamoxifen 10 mg BID was prescribed to postmenopausal pts with ER+ tumors following both RT and CT. 4 pts were ineligible due to prior malignancies, 2 pts on the RT-first arm refused their assigned sequence and received CT first, 2 pts on the RT-first arm and 1 pt on the CT-first arm refused to start or complete CT, 1 pt on the CT-arm underwent elective mastectomy prior to starting RT, and 1 pt on each arm received nonprotocol CT. Possible prognostic factors including age, menopausal status, ERP status, nodal status, and lymphatic vessel invasion were distributed nearly uniformly between the 2 arms. There was an excess of pts with EIC-positive tumors on the RT-first arm (25%) compared to the CT-first arm (11%) (p-bar=0

Sequencing of chemotherapy (CT) and radiotherapy (RT) following conservative surgery (CS) for patients with early-stage breast cancer: results of a randomized trial

International Nuclear Information System (INIS)

Recht, A; Come, SE; Silver, B; Gelman, RS; Hayes, DF; Shulman, LN; Henderson, IC; Harris, JR

1995-01-01

Purpose: To determine the effects of different sequences of CT and RT following CS for pts with clinical Stage I or II breast carcinoma. Methods and Materials: From 6/84-12/92, 244 pts were randomized following CS to receive CT (4 cycles of CAMFP at 3-wk intervals) either before or after RT. Median pt age was 45 yrs (range, 20-68). 164 pts were pre-, 18 pts peri-, and 62 pts were postmenopausal. 209 pts were node-positive (61 had 4 or more positive nodes); after 6/88, 35 selected node-negative pts were also included. 45 Gy was prescribed to the entire breast, followed by a boost of 16 Gy; nodal RT was optional. The medians ( and ranges) of the surgery-radiotherapy interval (SRI) according to assigned arm were: RT-first, 36 days (14-234); CT-first, 126 days (98-185). Drug doses were based on ideal body weight. Doses (per square meter) and schedule were: methotrexate, 200 mg, d.1 and 15; leucovorin, 10 mg, q6H x 12, start d.2 and 16; 5-fluorouracil, 500 mg, d.1; cyclophosphamide, 500 mg, d.1; prednisone, 40 mg/day x 5d, start d.1; and doxorubicin, 45 mg, d.3. All except leucovorin and prednisone were given by IV bolus. Dose reductions were based on the granulocyte nadir from the previous cycle, with an attempt to reescalate doses in subsequent cycles. Tamoxifen 10 mg BID was prescribed to postmenopausal pts with ER+ tumors following both RT and CT. 4 pts were ineligible due to prior malignancies, 2 pts on the RT-first arm refused their assigned sequence and received CT first, 2 pts on the RT-first arm and 1 pt on the CT-first arm refused to start or complete CT, 1 pt on the CT-arm underwent elective mastectomy prior to starting RT, and 1 pt on each arm received nonprotocol CT. Possible prognostic factors including age, menopausal status, ERP status, nodal status, and lymphatic vessel invasion were distributed nearly uniformly between the 2 arms. There was an excess of pts with EIC-positive tumors on the RT-first arm (25%) compared to the CT-first arm (11%) (p-bar=0

BLEACHING EUCALYPTUS PULPS WITH SHORT SEQUENCES

Directory of Open Access Journals (Sweden)

Flaviana Reis Milagres

2011-03-01

Full Text Available Eucalyptus spp kraft pulp, due to its high content of hexenuronic acids, is quite easy to bleach. Therefore, investigations have been made attempting to decrease the number of stages in the bleaching process in order to minimize capital costs. This study focused on the evaluation of short ECF (Elemental Chlorine Free and TCF (Totally Chlorine Free sequences for bleaching oxygen delignified Eucalyptus spp kraft pulp to 90% ISO brightness: PMoDP (Molybdenum catalyzed acid peroxide, chlorine dioxide and hydrogen peroxide, PMoD/P (Molybdenum catalyzed acid peroxide, chlorine dioxide and hydrogen peroxide, without washing PMoD(PO (Molybdenum catalyzed acid peroxide, chlorine dioxide and pressurized peroxide, D(EPODP (chlorine dioxide, extraction oxidative with oxygen and peroxide, chlorine dioxide and hydrogen peroxide, PMoQ(PO (Molybdenum catalyzed acid peroxide, DTPA and pressurized peroxide, and XPMoQ(PO (Enzyme, molybdenum catalyzed acid peroxide, DTPA and pressurized peroxide. Uncommon pulp treatments, such as molybdenum catalyzed acid peroxide (PMo and xylanase (X bleaching stages, were used. Among the ECF alternatives, the two-stage PMoD/P sequence proved highly cost-effective without affecting pulp quality in relation to the traditional D(EPODP sequence and produced better quality effluent in relation to the reference. However, a four stage sequence, XPMoQ(PO, was required to achieve full brightness using the TCF technology. This sequence was highly cost-effective although it only produced pulp of acceptable quality.

Genome sequence of Mycobacterium yongonense RT 955-2015 isolate from a patient misdiagnosed with multi-drug resistant tuberculosis: first clinical isolate in Tanzania.

Science.gov (United States)

Mnyambwa, Nicholaus Peter; Kim, Dong-Jin; Ngadaya, Esther; Chun, Jongsik; Ha, Sung-Min; Petrucka, Pammla; Addo, Kennedy Kwasi; Kazwala, Rudovick R; Mfinanga, Sayoki G

2018-04-24

Mycobacterium yongonense is a recently described novel species belonging to Mycobacterium avium complex which is the most prevalent etiology of non-tuberculous mycobacteria associated with pulmonary infections, and posing tuberculosis diagnostic challenges in high-burden, resource-constrained settings. We used whole genome shotgun sequencing and comparative microbial genomic analyses to characterize the isolate from a patient diagnosed with multi-drug resistant tuberculosis (MDR-TB) after relapse. We present a genome sequence of the first case of M. yongonense (M. yongonense RT 955-2015) in Tanzania. Sequence analysis revealed that the RT 955-2015 strain had a high similarity to M. yongonense 05-1390(T) (98.74%) and M. chimaera DSM 44623(T) (98%). Its 16S rRNA showed similarity to M. paraintracellulare KCTC 290849(T) (100%); M. intracellulare ATCC 13950(T) (100%); M. chimaera DSM 44623(T) (99.9%); and M. yongonense 05-1390(T) (98%). The strain had a substantially different rpoB sequence from that of M. yongonense 05-1390 (95.16%) but exhibited a sequence closely related to M. chimaera DSM 44623(T) (99.86%), M. intracellulare ATCC 13950(T) (99.53%), and M. paraintracellulare KCTC 290849(T) (99.53%). In light of the OrthoANI algorithm, and phylogenetic analysis, we conclude that the isolate was M. yongonense Type II genotype, which is an indication that the patient was misdiagnosed with TB/MDR-TB and received inappropriate treatment. Copyright © 2018. Published by Elsevier Ltd.

Salt-bridging effects on short amphiphilic helical structure and introducing sequence-based short beta-turn motifs.

Science.gov (United States)

Guarracino, Danielle A; Gentile, Kayla; Grossman, Alec; Li, Evan; Refai, Nader; Mohnot, Joy; King, Daniel

2018-02-01

Determining the minimal sequence necessary to induce protein folding is beneficial in understanding the role of protein-protein interactions in biological systems, as their three-dimensional structures often dictate their activity. Proteins are generally comprised of discrete secondary structures, from α-helices to β-turns and larger β-sheets, each of which is influenced by its primary structure. Manipulating the sequence of short, moderately helical peptides can help elucidate the influences on folding. We created two new scaffolds based on a modestly helical eight-residue peptide, PT3, we previously published. Using circular dichroism (CD) spectroscopy and changing the possible salt-bridging residues to new combinations of Lys, Arg, Glu, and Asp, we found that our most helical improvements came from the Arg-Glu combination, whereas the Lys-Asp was not significantly different from the Lys-Glu of the parent scaffold, PT3. The marked 3 10 -helical contributions in PT3 were lessened in the Arg-Glu-containing peptide with the beginning of cooperative unfolding seen through a thermal denaturation. However, a unique and unexpected signature was seen for the denaturation of the Lys-Asp peptide which could help elucidate the stages of folding between the 3 10 and α-helix. In addition, we developed a short six-residue peptide with β-turn/sheet CD signature, again to help study minimal sequences needed for folding. Overall, the results indicate that improvements made to short peptide scaffolds by fine-tuning the salt-bridging residues can enhance scaffold structure. Likewise, with the results from the new, short β-turn motif, these can help impact future peptidomimetic designs in creating biologically useful, short, structured β-sheet-forming peptides.

AWIPS II Application Development, a SPoRT Perspective

Science.gov (United States)

Burks, Jason E.; Smith, Matthew; McGrath, Kevin M.

2014-01-01

The National Weather Service (NWS) is deploying its next-generation decision support system, called AWIPS II (Advanced Weather Interactive Processing System II). NASA's Short-term Prediction Research and Transition (SPoRT) Center has developed several software 'plug-ins' to extend the capabilities of AWIPS II. SPoRT aims to continue its mission of improving short-term forecasts by providing NASA and NOAA products on the decision support system used at NWS weather forecast offices (WFOs). These products are not included in the standard Satellite Broadcast Network feed provided to WFOs. SPoRT has had success in providing support to WFOs as they have transitioned to AWIPS II. Specific examples of transitioning SPoRT plug-ins to WFOs with newly deployed AWIPS II systems will be presented. Proving Ground activities (GOES-R and JPSS) will dominate SPoRT's future AWIPS II activities, including tool development as well as enhancements to existing products. In early 2012 SPoRT initiated the Experimental Product Development Team, a group of AWIPS II developers from several institutions supporting NWS forecasters with innovative products. The results of the team's spring and fall 2013 meeting will be presented. Since AWIPS II developers now include employees at WFOs, as well as many other institutions related to weather forecasting, the NWS has dealt with a multitude of software governance issues related to the difficulties of multiple remotely collaborating software developers. This presentation will provide additional examples of Research-to-Operations plugins, as well as an update on how governance issues are being handled in the AWIPS II developer community.

Unequal distribution of RT-PCR artifacts along the E1-E2 region of Hepatitis C virus.

Science.gov (United States)

Domingo-Calap, Pilar; Sentandreu, Vicente; Bracho, Maria Alma; González-Candelas, Fernando; Moya, Andrés; Sanjuán, Rafael

2009-10-01

Although viral variability studies have focused traditionally on consensus sequences, the relevance of molecular clone sequences for studying viral evolution at the intra-host level is being increasingly recognized. However, for this approach to be reliable, RT-PCR artifacts do not have to contribute excessively to the observed variability. Molecular clone sequences were obtained from an in vitro transcript to estimate the maximum error rate associated to RT-PCR for the Hepatitis C virus (HCV) E1-E2 region. On average, the frequency of RT-PCR errors was one order of magnitude lower than the level of intra-host genetic variability observed in samples from an HCV outbreak. However, RT-PCR errors were not distributed evenly along the E1-E2 region and were concentrated heavily in the hypervariable region 2 (HVR 2). Although it is concluded that RT-PCR molecular clone sequences are reliable, these results warn against extrapolation of RT-PCR error rates to different genome regions. The data suggest that the RNA sequence context or secondary structure can determine the fidelity of in vitro transcription or reverse transcription. Potentially, these factors might also modify the fidelity of the viral polymerase.

Variance to mean ratio, R(t), for poisson processes on phylogenetic trees.

Science.gov (United States)

Goldman, N

1994-09-01

The ratio of expected variance to mean, R(t), of numbers of DNA base substitutions for contemporary sequences related by a "star" phylogeny is widely seen as a measure of the adherence of the sequences' evolution to a Poisson process with a molecular clock, as predicted by the "neutral theory" of molecular evolution under certain conditions. A number of estimators of R(t) have been proposed, all predicted to have mean 1 and distributions based on the chi 2. Various genes have previously been analyzed and found to have values of R(t) far in excess of 1, calling into question important aspects of the neutral theory. In this paper, I use Monte Carlo simulation to show that the previously suggested means and distributions of estimators of R(t) are highly inaccurate. The analysis is applied to star phylogenies and to general phylogenetic trees, and well-known gene sequences are reanalyzed. For star phylogenies the results show that Kimura's estimators ("The Neutral Theory of Molecular Evolution," Cambridge Univ. Press, Cambridge, 1983) are unsatisfactory for statistical testing of R(t), but confirm the accuracy of Bulmer's correction factor (Genetics 123: 615-619, 1989). For all three nonstar phylogenies studied, attained values of all three estimators of R(t), although larger than 1, are within their true confidence limits under simple Poisson process models. This shows that lineage effects can be responsible for high estimates of R(t), restoring some limited confidence in the molecular clock and showing that the distinction between lineage and molecular clock effects is vital.(ABSTRACT TRUNCATED AT 250 WORDS)

The SPoRT-WRF: Evaluating the Impact of NASA Datasets on Convective Forecasts

Science.gov (United States)

Zavodsky, Bradley; Kozlowski, Danielle; Case, Jonathan; Molthan, Andrew

2012-01-01

Short-term Prediction Research and Transition (SPoRT) seeks to improve short-term, regional weather forecasts using unique NASA products and capabilities SPoRT has developed a unique, real-time configuration of the NASA Unified Weather Research and Forecasting (WRF)WRF (ARW) that integrates all SPoRT modeling research data: (1) 2-km SPoRT Sea Surface Temperature (SST) Composite, (2) 3-km LIS with 1-km Greenness Vegetation Fraction (GVFs) (3) 45-km AIRS retrieved profiles. Transitioned this real-time forecast to NOAA's Hazardous Weather Testbed (HWT) as deterministic model at Experimental Forecast Program (EFP). Feedback from forecasters/participants and internal evaluation of SPoRT-WRF shows a cool, dry bias that appears to suppress convection likely related to methodology for assimilation of AIRS profiles Version 2 of the SPoRT-WRF will premier at the 2012 EFP and include NASA physics, cycling data assimilation methodology, better coverage of precipitation forcing, and new GVFs

SeqEntropy: genome-wide assessment of repeats for short read sequencing.

Directory of Open Access Journals (Sweden)

Hsueh-Ting Chu

Full Text Available BACKGROUND: Recent studies on genome assembly from short-read sequencing data reported the limitation of this technology to reconstruct the entire genome even at very high depth coverage. We investigated the limitation from the perspective of information theory to evaluate the effect of repeats on short-read genome assembly using idealized (error-free reads at different lengths. METHODOLOGY/PRINCIPAL FINDINGS: We define a metric H(k to be the entropy of sequencing reads at a read length k and use the relative loss of entropy ΔH(k to measure the impact of repeats for the reconstruction of whole-genome from sequences of length k. In our experiments, we found that entropy loss correlates well with de-novo assembly coverage of a genome, and a score of ΔH(k>1% indicates a severe loss in genome reconstruction fidelity. The minimal read lengths to achieve ΔH(k<1% are different for various organisms and are independent of the genome size. For example, in order to meet the threshold of ΔH(k<1%, a read length of 60 bp is needed for the sequencing of human genome (3.2 10(9 bp and 320 bp for the sequencing of fruit fly (1.8×10(8 bp. We also calculated the ΔH(k scores for 2725 prokaryotic chromosomes and plasmids at several read lengths. Our results indicate that the levels of repeats in different genomes are diverse and the entropy of sequencing reads provides a measurement for the repeat structures. CONCLUSIONS/SIGNIFICANCE: The proposed entropy-based measurement, which can be calculated in seconds to minutes in most cases, provides a rapid quantitative evaluation on the limitation of idealized short-read genome sequencing. Moreover, the calculation can be parallelized to scale up to large euakryotic genomes. This approach may be useful to tune the sequencing parameters to achieve better genome assemblies when a closely related genome is already available.

Table 1. Primer sequences used for real-time qRT-PCR analysis of ...

Indian Academy of Sciences (India)

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TGTCCCAGTAAACCGCTC. GAATCCAGCACGATACCAGT. Figure 1. Expression analysis of candidate CsActin and CsFbox genes by qRT-PCR in response to 4°C treatment. The y-axis indicates Cq values, and error bars represent standard deviations of the mean values of four replicates. Rt, roots; St, stems; Le, leaves; Fl ...

Accurate typing of short tandem repeats from genome-wide sequencing data and its applications.

Science.gov (United States)

Fungtammasan, Arkarachai; Ananda, Guruprasad; Hile, Suzanne E; Su, Marcia Shu-Wei; Sun, Chen; Harris, Robert; Medvedev, Paul; Eckert, Kristin; Makova, Kateryna D

2015-05-01

Short tandem repeats (STRs) are implicated in dozens of human genetic diseases and contribute significantly to genome variation and instability. Yet profiling STRs from short-read sequencing data is challenging because of their high sequencing error rates. Here, we developed STR-FM, short tandem repeat profiling using flank-based mapping, a computational pipeline that can detect the full spectrum of STR alleles from short-read data, can adapt to emerging read-mapping algorithms, and can be applied to heterogeneous genetic samples (e.g., tumors, viruses, and genomes of organelles). We used STR-FM to study STR error rates and patterns in publicly available human and in-house generated ultradeep plasmid sequencing data sets. We discovered that STRs sequenced with a PCR-free protocol have up to ninefold fewer errors than those sequenced with a PCR-containing protocol. We constructed an error correction model for genotyping STRs that can distinguish heterozygous alleles containing STRs with consecutive repeat numbers. Applying our model and pipeline to Illumina sequencing data with 100-bp reads, we could confidently genotype several disease-related long trinucleotide STRs. Utilizing this pipeline, for the first time we determined the genome-wide STR germline mutation rate from a deeply sequenced human pedigree. Additionally, we built a tool that recommends minimal sequencing depth for accurate STR genotyping, depending on repeat length and sequencing read length. The required read depth increases with STR length and is lower for a PCR-free protocol. This suite of tools addresses the pressing challenges surrounding STR genotyping, and thus is of wide interest to researchers investigating disease-related STRs and STR evolution. © 2015 Fungtammasan et al.; Published by Cold Spring Harbor Laboratory Press.

DeepRT: deep learning for peptide retention time prediction in proteomics

OpenAIRE

Ma, Chunwei; Zhu, Zhiyong; Ye, Jun; Yang, Jiarui; Pei, Jianguo; Xu, Shaohang; Zhou, Ruo; Yu, Chang; Mo, Fan; Wen, Bo; Liu, Siqi

2017-01-01

Accurate predictions of peptide retention times (RT) in liquid chromatography have many applications in mass spectrometry-based proteomics. Herein, we present DeepRT, a deep learning based software for peptide retention time prediction. DeepRT automatically learns features directly from the peptide sequences using the deep convolutional Neural Network (CNN) and Recurrent Neural Network (RNN) model, which eliminates the need to use hand-crafted features or rules. After the feature learning, pr...

RT-PCR and sequence analysis of the full-length fusion protein of Canine Distemper Virus from domestic dogs.

Science.gov (United States)

Romanutti, Carina; Gallo Calderón, Marina; Keller, Leticia; Mattion, Nora; La Torre, José

2016-02-01

During 2007-2014, 84 out of 236 (35.6%) samples from domestic dogs submitted to our laboratory for diagnostic purposes were positive for Canine Distemper Virus (CDV), as analyzed by RT-PCR amplification of a fragment of the nucleoprotein gene. Fifty-nine of them (70.2%) were from dogs that had been vaccinated against CDV. The full-length gene encoding the Fusion (F) protein of fifteen isolates was sequenced and compared with that of those of other CDVs, including wild-type and vaccine strains. Phylogenetic analysis using the F gene full-length sequences grouped all the Argentinean CDV strains in the SA2 clade. Sequence identity with the Onderstepoort vaccine strain was 89.0-90.6%, and the highest divergence was found in the 135 amino acids corresponding to the F protein signal-peptide, Fsp (64.4-66.7% identity). In contrast, this region was highly conserved among the local strains (94.1-100% identity). One extra putative N-glycosylation site was identified in the F gene of CDV Argentinean strains with respect to the vaccine strain. The present report is the first to analyze full-length F protein sequences of CDV strains circulating in Argentina, and contributes to the knowledge of molecular epidemiology of CDV, which may help in understanding future disease outbreaks. Copyright © 2015 Elsevier B.V. All rights reserved.

Applications of LANCE Data at SPoRT

Science.gov (United States)

Molthan, Andrew

2014-01-01

Short term Prediction Research and Transition (SPoRT) Center: Mission: Apply NASA and NOAA measurement systems and unique Earth science research to improve the accuracy of short term weather prediction at the regional/local scale. Goals: Evaluate and assess the utility of NASA and NOAA Earth science data and products and unique research capabilities to address operational weather forecast problems; Provide an environment which enables the development and testing of new capabilities to improve short term weather forecasts on a regional scale; Help ensure successful transition of new capabilities to operational weather entities for the benefit of society

TMS Over the Cerebellum Interferes with Short-term Memory of Visual Sequences.

Science.gov (United States)

Ferrari, C; Cattaneo, Z; Oldrati, V; Casiraghi, L; Castelli, F; D'Angelo, E; Vecchi, T

2018-04-30

Growing evidence suggests that the cerebellum is not only involved in motor functions, but it significantly contributes to sensory and cognitive processing as well. In particular, it has been hypothesized that the cerebellum identifies recurrent serial events and recognizes their violations. Here we used transcranial magnetic stimulation (TMS) to shed light on the role of the cerebellum in short-term memory of visual sequences. In two experiments, we found that TMS over the right cerebellar hemisphere impaired participants' ability to recognize the correct order of appearance of geometrical stimuli varying in shape and/or size. In turn, cerebellar TMS did not affect recognition of highly familiar short sequences of letters or numbers. Overall, our data suggest that the cerebellum is involved in memorizing the order in which (concatenated) stimuli appear, this process being important for sequence learning.

Studying a free fall experiment using short sequences of images

International Nuclear Information System (INIS)

Vera, Francisco; Romanque, Cristian

2008-01-01

We discuss a new alternative for obtaining position and time coordinates from a video of a free fall experiment. In our approach, after converting the video to a short sequence of images, the images are analyzed using a web page application developed by the author. The main advantage of the setup explained in this work, is that it is simple to use, no software license fees are necessary, and can be scaled-up to be used by a big number of students in introductory physics courses. The steps involved in the full analysis of a falling object are: we grab a short digital video of the experiment and convert it to a sequence of images, then, using a web page that includes all the necessary javascript, the student can easily click on the object of interest to obtain the (x,y,t) coordinates, finally, the student analyze motion using a spreadsheet.

NASA SPoRT GOES-R Proving Ground Activities

Science.gov (United States)

Stano, Geoffrey T.; Fuell, Kevin K.; Jedloec, Gary J.

2010-01-01

The NASA Short-term Prediction Research and Transition (SPoRT) program is a partner with the GOES-R Proving Ground (PG) helping prepare forecasters understand the unique products to come from the GOES-R instrument suite. SPoRT is working collaboratively with other members of the GOES-R PG team and Algorithm Working Group (AWG) scientists to develop and disseminate a suite of proxy products that address specific forecast problems for the WFOs, Regional and National Support Centers, and other NOAA users. These products draw on SPoRT s expertise with the transition and evaluation of products into operations from the MODIS instrument and the North Alabama Lightning Mapping Array (NALMA). The MODIS instrument serves as an excellent proxy for the Advanced Baseline Imager (ABI) that will be aboard GOES-R. SPoRT has transitioned and evaluated several multi-channel MODIS products. The true and false color products are being used in natural hazard detection by several SPoRT partners to provide better observation of land features, such as fires, smoke plumes, and snow cover. Additionally, many of SPoRT s partners are coastal offices and already benefit from the MODIS sea surface temperature composite. This, along with other surface feature observations will be developed into ABI proxy products for diagnostic use in the forecast process as well as assimilation into forecast models. In addition to the MODIS instrument, the NALMA has proven very valuable to WFOs with access to these total lightning data. These data provide situational awareness and enhanced warning decision making to improve lead times for severe thunderstorm and tornado warnings. One effort by SPoRT scientists includes a lightning threat product to create short-term model forecasts of lightning activity. Additionally, SPoRT is working with the AWG to create GLM proxy data from several of the ground based total lightning networks, such as the NALMA. The evaluation will focus on the vastly improved spatial

Characterizing novel endogenous retroviruses from genetic variation inferred from short sequence reads

DEFF Research Database (Denmark)

Mourier, Tobias; Mollerup, Sarah; Vinner, Lasse

2015-01-01

From Illumina sequencing of DNA from brain and liver tissue from the lion, Panthera leo, and tumor samples from the pike-perch, Sander lucioperca, we obtained two assembled sequence contigs with similarity to known retroviruses. Phylogenetic analyses suggest that the pike-perch retrovirus belongs...... to the epsilonretroviruses, and the lion retrovirus to the gammaretroviruses. To determine if these novel retroviral sequences originate from an endogenous retrovirus or from a recently integrated exogenous retrovirus, we assessed the genetic diversity of the parental sequences from which the short Illumina reads...

SRComp: short read sequence compression using burstsort and Elias omega coding.

Directory of Open Access Journals (Sweden)

Jeremy John Selva

Full Text Available Next-generation sequencing (NGS technologies permit the rapid production of vast amounts of data at low cost. Economical data storage and transmission hence becomes an increasingly important challenge for NGS experiments. In this paper, we introduce a new non-reference based read sequence compression tool called SRComp. It works by first employing a fast string-sorting algorithm called burstsort to sort read sequences in lexicographical order and then Elias omega-based integer coding to encode the sorted read sequences. SRComp has been benchmarked on four large NGS datasets, where experimental results show that it can run 5-35 times faster than current state-of-the-art read sequence compression tools such as BEETL and SCALCE, while retaining comparable compression efficiency for large collections of short read sequences. SRComp is a read sequence compression tool that is particularly valuable in certain applications where compression time is of major concern.

SPoRT - An End-to-End R2O Activity

Science.gov (United States)

Jedlovec, Gary J.

2009-01-01

Established in 2002 to demonstrate the weather and forecasting application of real-time EOS measurements, the Short-term Prediction Research and Transition (SPoRT) program has grown to be an end-to-end research to operations activity focused on the use of advanced NASA modeling and data assimilation approaches, nowcasting techniques, and unique high-resolution multispectral observational data applications from EOS satellites to improve short-term weather forecasts on a regional and local scale. SPoRT currently partners with several universities and other government agencies for access to real-time data and products, and works collaboratively with them and operational end users at 13 WFOs to develop and test the new products and capabilities in a "test-bed" mode. The test-bed simulates key aspects of the operational environment without putting constraints on the forecaster workload. Products and capabilities which show utility in the test-bed environment are then transitioned experimentally into the operational environment for further evaluation and assessment. SPoRT focuses on a suite of data and products from MODIS, AMSR-E, and AIRS on the NASA Terra and Aqua satellites, and total lightning measurements from ground-based networks. Some of the observations are assimilated into or used with various versions of the WRF model to provide supplemental forecast guidance to operational end users. SPoRT is enhancing partnerships with NOAA / NESDIS for new product development and data access to exploit the remote sensing capabilities of instruments on the NPOESS satellites to address short term weather forecasting problems. The VIIRS and CrIS instruments on the NPP and follow-on NPOESS satellites provide similar observing capabilities to the MODIS and AIRS instruments on Terra and Aqua. SPoRT will be transitioning existing and new capabilities into the AWIIPS II environment to continue the continuity of its activities.

Optimization of short amino acid sequences classifier

Science.gov (United States)

Barcz, Aleksy; Szymański, Zbigniew

This article describes processing methods used for short amino acid sequences classification. The data processed are 9-symbols string representations of amino acid sequences, divided into 49 data sets - each one containing samples labeled as reacting or not with given enzyme. The goal of the classification is to determine for a single enzyme, whether an amino acid sequence would react with it or not. Each data set is processed separately. Feature selection is performed to reduce the number of dimensions for each data set. The method used for feature selection consists of two phases. During the first phase, significant positions are selected using Classification and Regression Trees. Afterwards, symbols appearing at the selected positions are substituted with numeric values of amino acid properties taken from the AAindex database. In the second phase the new set of features is reduced using a correlation-based ranking formula and Gram-Schmidt orthogonalization. Finally, the preprocessed data is used for training LS-SVM classifiers. SPDE, an evolutionary algorithm, is used to obtain optimal hyperparameters for the LS-SVM classifier, such as error penalty parameter C and kernel-specific hyperparameters. A simple score penalty is used to adapt the SPDE algorithm to the task of selecting classifiers with best performance measures values.

Projected Applications of a ``Climate in a Box'' Computing System at the NASA Short-term Prediction Research and Transition (SPoRT) Center

Science.gov (United States)

Jedlovec, G.; Molthan, A.; Zavodsky, B.; Case, J.; Lafontaine, F.

2010-12-01

The NASA Short-term Prediction Research and Transition (SPoRT) Center focuses on the transition of unique observations and research capabilities to the operational weather community, with a goal of improving short-term forecasts on a regional scale. Advances in research computing have lead to “Climate in a Box” systems, with hardware configurations capable of producing high resolution, near real-time weather forecasts, but with footprints, power, and cooling requirements that are comparable to desktop systems. The SPoRT Center has developed several capabilities for incorporating unique NASA research capabilities and observations with real-time weather forecasts. Planned utilization includes the development of a fully-cycled data assimilation system used to drive 36-48 hour forecasts produced by the NASA Unified version of the Weather Research and Forecasting (WRF) model (NU-WRF). The horsepower provided by the “Climate in a Box” system is expected to facilitate the assimilation of vertical profiles of temperature and moisture provided by the Atmospheric Infrared Sounder (AIRS) aboard the NASA Aqua satellite. In addition, the Moderate Resolution Imaging Spectroradiometer (MODIS) instruments aboard NASA’s Aqua and Terra satellites provide high-resolution sea surface temperatures and vegetation characteristics. The development of MODIS normalized difference vegetation index (NVDI) composites for use within the NASA Land Information System (LIS) will assist in the characterization of vegetation, and subsequently the surface albedo and processes related to soil moisture. Through application of satellite simulators, NASA satellite instruments can be used to examine forecast model errors in cloud cover and other characteristics. Through the aforementioned application of the “Climate in a Box” system and NU-WRF capabilities, an end goal is the establishment of a real-time forecast system that fully integrates modeling and analysis capabilities developed

Projected Applications of a "Climate in a Box" Computing System at the NASA Short-Term Prediction Research and Transition (SPoRT) Center

Science.gov (United States)

Jedlovec, Gary J.; Molthan, Andrew L.; Zavodsky, Bradley; Case, Jonathan L.; LaFontaine, Frank J.

2010-01-01

The NASA Short-term Prediction Research and Transition (SPoRT) Center focuses on the transition of unique observations and research capabilities to the operational weather community, with a goal of improving short-term forecasts on a regional scale. Advances in research computing have lead to "Climate in a Box" systems, with hardware configurations capable of producing high resolution, near real-time weather forecasts, but with footprints, power, and cooling requirements that are comparable to desktop systems. The SPoRT Center has developed several capabilities for incorporating unique NASA research capabilities and observations with real-time weather forecasts. Planned utilization includes the development of a fully-cycled data assimilation system used to drive 36-48 hour forecasts produced by the NASA Unified version of the Weather Research and Forecasting (WRF) model (NU-WRF). The horsepower provided by the "Climate in a Box" system is expected to facilitate the assimilation of vertical profiles of temperature and moisture provided by the Atmospheric Infrared Sounder (AIRS) aboard the NASA Aqua satellite. In addition, the Moderate Resolution Imaging Spectroradiometer (MODIS) instruments aboard NASA s Aqua and Terra satellites provide high-resolution sea surface temperatures and vegetation characteristics. The development of MODIS normalized difference vegetation index (NVDI) composites for use within the NASA Land Information System (LIS) will assist in the characterization of vegetation, and subsequently the surface albedo and processes related to soil moisture. Through application of satellite simulators, NASA satellite instruments can be used to examine forecast model errors in cloud cover and other characteristics. Through the aforementioned application of the "Climate in a Box" system and NU-WRF capabilities, an end goal is the establishment of a real-time forecast system that fully integrates modeling and analysis capabilities developed within the NASA SPoRT

Sequence quality analysis tool for HIV type 1 protease and reverse transcriptase.

Science.gov (United States)

Delong, Allison K; Wu, Mingham; Bennett, Diane; Parkin, Neil; Wu, Zhijin; Hogan, Joseph W; Kantor, Rami

2012-08-01

Access to antiretroviral therapy is increasing globally and drug resistance evolution is anticipated. Currently, protease (PR) and reverse transcriptase (RT) sequence generation is increasing, including the use of in-house sequencing assays, and quality assessment prior to sequence analysis is essential. We created a computational HIV PR/RT Sequence Quality Analysis Tool (SQUAT) that runs in the R statistical environment. Sequence quality thresholds are calculated from a large dataset (46,802 PR and 44,432 RT sequences) from the published literature ( http://hivdb.Stanford.edu ). Nucleic acid sequences are read into SQUAT, identified, aligned, and translated. Nucleic acid sequences are flagged if with >five 1-2-base insertions; >one 3-base insertion; >one deletion; >six PR or >18 RT ambiguous bases; >three consecutive PR or >four RT nucleic acid mutations; >zero stop codons; >three PR or >six RT ambiguous amino acids; >three consecutive PR or >four RT amino acid mutations; >zero unique amino acids; or 15% genetic distance from another submitted sequence. Thresholds are user modifiable. SQUAT output includes a summary report with detailed comments for troubleshooting of flagged sequences, histograms of pairwise genetic distances, neighbor joining phylogenetic trees, and aligned nucleic and amino acid sequences. SQUAT is a stand-alone, free, web-independent tool to ensure use of high-quality HIV PR/RT sequences in interpretation and reporting of drug resistance, while increasing awareness and expertise and facilitating troubleshooting of potentially problematic sequences.

Detection and Analysis of Circular RNAs by RT-PCR.

Science.gov (United States)

Panda, Amaresh C; Gorospe, Myriam

2018-03-20

Gene expression in eukaryotic cells is tightly regulated at the transcriptional and posttranscriptional levels. Posttranscriptional processes, including pre-mRNA splicing, mRNA export, mRNA turnover, and mRNA translation, are controlled by RNA-binding proteins (RBPs) and noncoding (nc)RNAs. The vast family of ncRNAs comprises diverse regulatory RNAs, such as microRNAs and long noncoding (lnc)RNAs, but also the poorly explored class of circular (circ)RNAs. Although first discovered more than three decades ago by electron microscopy, only the advent of high-throughput RNA-sequencing (RNA-seq) and the development of innovative bioinformatic pipelines have begun to allow the systematic identification of circRNAs (Szabo and Salzman, 2016; Panda et al ., 2017b; Panda et al ., 2017c). However, the validation of true circRNAs identified by RNA sequencing requires other molecular biology techniques including reverse transcription (RT) followed by conventional or quantitative (q) polymerase chain reaction (PCR), and Northern blot analysis (Jeck and Sharpless, 2014). RT-qPCR analysis of circular RNAs using divergent primers has been widely used for the detection, validation, and sometimes quantification of circRNAs (Abdelmohsen et al ., 2015 and 2017; Panda et al ., 2017b). As detailed here, divergent primers designed to span the circRNA backsplice junction sequence can specifically amplify the circRNAs and not the counterpart linear RNA. In sum, RT-PCR analysis using divergent primers allows direct detection and quantification of circRNAs.

SPoRT's Participation in the GOES-R Proving Ground Activity

Science.gov (United States)

Jedlovec, Gary; Fuell, Kevin; Smith, Matthew; Stano, Geoffrey; Molthan, Andrew

2011-01-01

The next generation geostationary satellite, GOES-R, will carry two new instruments with unique atmospheric and surface observing capabilities, the Advanced Baseline Imager (ABI) and the Geostationary Lightning Mapper (GLM), to study short-term weather processes. The ABI will bring enhanced multispectral observing capabilities with frequent refresh rates for regional and full disk coverage to geostationary orbit to address many existing and new forecast challenges. The GLM will, for the first time, provide the continuous monitoring of total lightning flashes over a hemispherical region from space. NOAA established the GOES-R Proving Ground activity several years ago to demonstrate the new capabilities of these instruments and to prepare forecasters for their day one use. Proving Ground partners work closely with algorithm developers and the end user community to develop and transition proxy data sets representing GOES-R observing capabilities. This close collaboration helps to maximize refine algorithms leading to the delivery of a product that effectively address a forecast challenge. The NASA Short-term Prediction Research and Transition (SPoRT) program has been a participant in the NOAA GOES-R Proving Ground activity by developing and disseminating selected GOES-R proxy products to collaborating WFOs and National Centers. Established in 2002 to demonstrate the weather and forecasting application of real-time EOS measurements, the SPoRT program has grown to be an end-to-end research to operations activity focused on the use of advanced NASA modeling and data assimilation approaches, nowcasting techniques, and unique high-resolution multispectral data from EOS satellites to improve short-term weather forecasts on a regional and local scale. Participation in the Proving Ground activities extends SPoRT s activities and taps its experience and expertise in diagnostic weather analysis, short-term weather forecasting, and the transition of research and experimental

Interference by clustered regularly interspaced short palindromic repeat (CRISPR) RNA is governed by a seed sequence

NARCIS (Netherlands)

Semenova, E.V.; Jore, M.M.; Westra, E.R.; Oost, van der J.; Brouns, S.J.J.

2011-01-01

Prokaryotic clustered regularly interspaced short palindromic repeat (CRISPR)/Cas (CRISPR-associated sequences) systems provide adaptive immunity against viruses when a spacer sequence of small CRISPR RNA (crRNA) matches a protospacer sequence in the viral genome. Viruses that escape CRISPR/Cas

Resonant magnetoelectric response of composite cantilevers: Theory of short vs. open circuit operation and layer sequence effects

Directory of Open Access Journals (Sweden)

Matthias C. Krantz

2015-11-01

Full Text Available The magnetoelectric effect in layered composite cantilevers consisting of strain coupled layers of magnetostrictive (MS, piezoelectric (PE, and substrate materials is investigated for magnetic field excitation at bending resonance. Analytic theories are derived for the transverse magnetoelectric (ME response in short and open circuit operation for three different layer sequences and results presented and discussed for the FeCoBSi-AlN-Si and the FeCoBSi-PZT-Si composite systems. Response optimized PE-MS layer thickness ratios are found to greatly change with operation mode shifting from near equal MS and PE layer thicknesses in the open circuit mode to near vanishing PE layer thicknesses in short circuit operation for all layer sequences. In addition the substrate layer thickness is found to differently affect the open and short circuit ME response producing shifts and reversal between ME response maxima depending on layer sequence. The observed rich ME response behavior for different layer thicknesses, sequences, operating modes, and PE materials can be explained by common neutral plane effects and different elastic compliance effects in short and open circuit operation.

Characterization of Campylobacter jejuni applying flaA short variable region sequencing, multilocus sequencing and Fourier transform infrared spectroscopy

DEFF Research Database (Denmark)

Josefsen, Mathilde Hartmann; Bonnichsen, Lise; Larsson, Jonas

flaA short variable region sequencing and phenetic Fourier transform infrared (FTIR) spectroscopy was applied on a collection of 102 Campylobacter jejuni isolated from continuous sampling of organic, free range geese and chickens. FTIR has been shown to serve as a valuable tool in typing...

Modified Polyadenylation-Based RT-qPCR Increases Selectivity of Amplification of 3′-MicroRNA Isoforms

Directory of Open Access Journals (Sweden)

Charlotte Nejad

2018-01-01

Full Text Available MicroRNA (miRNA detection by reverse transcription (RT quantitative real-time PCR (RT-qPCR is the most popular method currently used to measure miRNA expression. Although the majority of miRNA families are constituted of several 3′-end length variants (“isomiRs”, little attention has been paid to their differential detection by RT-qPCR. However, recent evidence indicates that 3′-end miRNA isoforms can exhibit 3′-length specific regulatory functions, underlining the need to develop strategies to differentiate 3′-isomiRs by RT-qPCR approaches. We demonstrate here that polyadenylation-based RT-qPCR strategies targeted to 20–21 nt isoforms amplify entire miRNA families, but that primers targeted to >22 nt isoforms were specific to >21 nt isoforms. Based on this observation, we developed a simple method to increase selectivity of polyadenylation-based RT-qPCR assays toward shorter isoforms, and demonstrate its capacity to help distinguish short RNAs from longer ones, using synthetic RNAs and biological samples with altered isomiR stoichiometry. Our approach can be adapted to many polyadenylation-based RT-qPCR technologies already exiting, providing a convenient way to distinguish long and short 3′-isomiRs.

Simple, quick and cost-efficient: A universal RT-PCR and sequencing strategy for genomic characterisation of foot-and-mouth disease viruses.

Science.gov (United States)

Dill, V; Beer, M; Hoffmann, B

2017-08-01

Foot-and-mouth disease (FMD) is a major contributor to poverty and food insecurity in Africa and Asia, and it is one of the biggest threats to agriculture in highly developed countries. As FMD is extremely contagious, strategies for its prevention, early detection, and the immediate characterisation of outbreak strains are of great importance. The generation of whole-genome sequences enables phylogenetic characterisation, the epidemiological tracing of virus transmission pathways and is supportive in disease control strategies. This study describes the development and validation of a rapid, universal and cost-efficient RT-PCR system to generate genome sequences of FMDV, reaching from the IRES to the end of the open reading frame. The method was evaluated using twelve different virus strains covering all seven serotypes of FMDV. Additionally, samples from experimentally infected animals were tested to mimic diagnostic field samples. All primer pairs showed a robust amplification with a high sensitivity for all serotypes. In summary, the described assay is suitable for the generation of FMDV sequences from all serotypes to allow immediate phylogenetic analysis, detailed genotyping and molecular epidemiology. Copyright © 2017 Elsevier B.V. All rights reserved.

MATAM: reconstruction of phylogenetic marker genes from short sequencing reads in metagenomes.

Science.gov (United States)

Pericard, Pierre; Dufresne, Yoann; Couderc, Loïc; Blanquart, Samuel; Touzet, Hélène

2018-02-15

Advances in the sequencing of uncultured environmental samples, dubbed metagenomics, raise a growing need for accurate taxonomic assignment. Accurate identification of organisms present within a community is essential to understanding even the most elementary ecosystems. However, current high-throughput sequencing technologies generate short reads which partially cover full-length marker genes and this poses difficult bioinformatic challenges for taxonomy identification at high resolution. We designed MATAM, a software dedicated to the fast and accurate targeted assembly of short reads sequenced from a genomic marker of interest. The method implements a stepwise process based on construction and analysis of a read overlap graph. It is applied to the assembly of 16S rRNA markers and is validated on simulated, synthetic and genuine metagenomes. We show that MATAM outperforms other available methods in terms of low error rates and recovered fractions and is suitable to provide improved assemblies for precise taxonomic assignments. https://github.com/bonsai-team/matam. pierre.pericard@gmail.com or helene.touzet@univ-lille1.fr. Supplementary data are available at Bioinformatics online. © The Author (2017). Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com

Study of the fast inversion recovery pulse sequence. With reference to fast fluid attenuated inversion recovery and fast short TI inversion recovery pulse sequence

International Nuclear Information System (INIS)

Tsuchihashi, Toshio; Maki, Toshio; Suzuki, Takeshi

1997-01-01

The fast inversion recovery (fast IR) pulse sequence was evaluated. We compared the fast fluid attenuated inversion recovery (fast FLAIR) pulse sequence in which inversion time (TI) was established as equal to the water null point for the purpose of the water-suppressed T 2 -weighted image, with the fast short TI inversion recovery (fast STIR) pulse sequence in which TI was established as equal to the fat null point for purpose of fat suppression. In the fast FLAIR pulse sequence, the water null point was increased by making TR longer. In the FLAIR pulse sequence, the longitudinal magnetization contrast is determined by TI. If TI is increased, T 2 -weighted contrast improves in the same way as increasing TR for the SE pulse sequence. Therefore, images should be taken with long TR and long TI, which are longer than TR and longer than the water null point. On the other hand, the fat null point is not affected by TR in the fast STIR pulse sequence. However, effective TE was affected by variation of the null point. This increased in proportion to the increase in effective TE. Our evaluation indicated that the fast STIR pulse sequence can control the extensive signals from fat in a short time. (author)

Keeping it together: Semantic coherence stabilizes phonological sequences in short-term memory.

Science.gov (United States)

Savill, Nicola; Ellis, Rachel; Brooke, Emma; Koa, Tiffany; Ferguson, Suzie; Rojas-Rodriguez, Elena; Arnold, Dominic; Smallwood, Jonathan; Jefferies, Elizabeth

2018-04-01

Our ability to hold a sequence of speech sounds in mind, in the correct configuration, supports many aspects of communication, but the contribution of conceptual information to this basic phonological capacity remains controversial. Previous research has shown modest and inconsistent benefits of meaning on phonological stability in short-term memory, but these studies were based on sets of unrelated words. Using a novel design, we examined the immediate recall of sentence-like sequences with coherent meaning, alongside both standard word lists and mixed lists containing words and nonwords. We found, and replicated, substantial effects of coherent meaning on phoneme-level accuracy: The phonemes of both words and nonwords within conceptually coherent sequences were more likely to be produced together and in the correct order. Since nonwords do not exist as items in long-term memory, the semantic enhancement of phoneme-level recall for both item types cannot be explained by a lexically based item reconstruction process employed at the point of retrieval ("redintegration"). Instead, our data show, for naturalistic input, that when meaning emerges from the combination of words, the phonological traces that support language are reinforced by a semantic-binding process that has been largely overlooked by past short-term memory research.

Tools for analyzing genetic variants from sequencing data Case study: short tandem repeats

OpenAIRE

Gymrek, Melissa

2016-01-01

This was presented as a BitesizeBio Webinar entitled "Tools for analyzing genetic variants from sequencing data Case study: short tandem repeats"Accompanying scripts can be accessed on github:https://github.com/mgymrek/mgymrek-bitesizebio-webinar 

RNA-Seq analysis and gene discovery of Andrias davidianus using Illumina short read sequencing.

Directory of Open Access Journals (Sweden)

Fenggang Li

Full Text Available The Chinese giant salamander, Andrias davidianus, is an important species in the course of evolution; however, there is insufficient genomic data in public databases for understanding its immunologic mechanisms. High-throughput transcriptome sequencing is necessary to generate an enormous number of transcript sequences from A. davidianus for gene discovery. In this study, we generated more than 40 million reads from samples of spleen and skin tissue using the Illumina paired-end sequencing technology. De novo assembly yielded 87,297 transcripts with a mean length of 734 base pairs (bp. Based on the sequence similarities, searching with known proteins, 38,916 genes were identified. Gene enrichment analysis determined that 981 transcripts were assigned to the immune system. Tissue-specific expression analysis indicated that 443 of transcripts were specifically expressed in the spleen and skin. Among these transcripts, 147 transcripts were found to be involved in immune responses and inflammatory reactions, such as fucolectin, β-defensins and lymphotoxin beta. Eight tissue-specific genes were selected for validation using real time reverse transcription quantitative PCR (qRT-PCR. The results showed that these genes were significantly more expressed in spleen and skin than in other tissues, suggesting that these genes have vital roles in the immune response. This work provides a comprehensive genomic sequence resource for A. davidianus and lays the foundation for future research on the immunologic and disease resistance mechanisms of A. davidianus and other amphibians.

CORE-SINEs: eukaryotic short interspersed retroposing elements with common sequence motifs.

Science.gov (United States)

Gilbert, N; Labuda, D

1999-03-16

A 65-bp "core" sequence is dispersed in hundreds of thousands copies in the human genome. This sequence was found to constitute the central segment of a group of short interspersed elements (SINEs), referred to as mammalian-wide interspersed repeats, that proliferated before the radiation of placental mammals. Here, we propose that the core identifies an ancient tRNA-like SINE element, which survived in different lineages such as mammals, reptiles, birds, and fish, as well as mollusks, presumably for >550 million years. This element gave rise to a number of sequence families (CORE-SINEs), including mammalian-wide interspersed repeats, whose distinct 3' ends are shared with different families of long interspersed elements (LINEs). The evolutionary success of the generic CORE-SINE element can be related to the recruitment of the internal promoter from highly transcribed host RNA as well as to its capacity to adapt to changing retropositional opportunities by sequence exchange with actively amplifying LINEs. It reinforces the notion that the very existence of SINEs depends on the cohabitation with both LINEs and the host genome.

The analysis of novel microRNA mimic sequences in cancer cells reveals lack of specificity in stem-loop RT-qPCR-based microRNA detection.

Science.gov (United States)

Winata, Patrick; Williams, Marissa; McGowan, Eileen; Nassif, Najah; van Zandwijk, Nico; Reid, Glen

2017-11-17

MicroRNAs are frequently downregulated in cancer, and restoring expression has tumour suppressive activity in tumour cells. Our recent phase I clinical trial investigated microRNA-based therapy in patients with malignant pleural mesothelioma. Treatment with TargomiRs, microRNA mimics with novel sequence packaged in EGFR antibody-targeted bacterial minicells, revealed clear signs of clinical activity. In order to detect delivery of microRNA mimics to tumour cells in future clinical trials, we tested hydrolysis probe-based assays specific for the sequence of the novel mimics in transfected mesothelioma cell lines using RT-qPCR. The custom assays efficiently and specifically amplified the consensus mimics. However, we found that these assays gave a signal when total RNA from untransfected and control mimic-transfected cells were used as templates. Further investigation revealed that the reverse transcription step using stem-loop primers appeared to introduce substantial non-specific amplification with either total RNA or synthetic RNA templates. This suggests that reverse transcription using stem-loop primers suffers from an intrinsic lack of specificity for the detection of highly similar microRNAs in the same family, especially when analysing total RNA. These results suggest that RT-qPCR is unlikely to be an effective means to detect delivery of microRNA mimic-based drugs to tumour cells in patients.

Hybridization Capture Using Short PCR Products Enriches Small Genomes by Capturing Flanking Sequences (CapFlank)

DEFF Research Database (Denmark)

Tsangaras, Kyriakos; Wales, Nathan; Sicheritz-Pontén, Thomas

2014-01-01

, a non-negligible fraction of the resulting sequence reads are not homologous to the bait. We demonstrate that during capture, the bait-hybridized library molecules add additional flanking library sequences iteratively, such that baits limited to targeting relatively short regions (e.g. few hundred...... nucleotides) can result in enrichment across entire mitochondrial and bacterial genomes. Our findings suggest that some of the off-target sequences derived in capture experiments are non-randomly enriched, and that CapFlank will facilitate targeted enrichment of large contiguous sequences with minimal prior...

Regional Data Assimilation of AIRS Profiles and Radiances at the SPoRT Center

Science.gov (United States)

Zavodsky, Brad; Chou, Shih-hung; Jedlovec, Gary

2009-01-01

This slide presentation reviews the Short Term Prediction Research and Transition (SPoRT) Center's mission to improve short-term weather prediction at the regional and local scale. It includes information on the cold bias in Weather Research and Forcasting (WRF), troposphere recordings from the Atmospheric Infrared Sounder (AIRS), and vertical resolution of analysis grid.

Choice of reference sequence and assembler for alignment of Listeria monocytogenes short-read sequence data greatly influences rates of error in SNP analyses.

Directory of Open Access Journals (Sweden)

Arthur W Pightling

Full Text Available The wide availability of whole-genome sequencing (WGS and an abundance of open-source software have made detection of single-nucleotide polymorphisms (SNPs in bacterial genomes an increasingly accessible and effective tool for comparative analyses. Thus, ensuring that real nucleotide differences between genomes (i.e., true SNPs are detected at high rates and that the influences of errors (such as false positive SNPs, ambiguously called sites, and gaps are mitigated is of utmost importance. The choices researchers make regarding the generation and analysis of WGS data can greatly influence the accuracy of short-read sequence alignments and, therefore, the efficacy of such experiments. We studied the effects of some of these choices, including: i depth of sequencing coverage, ii choice of reference-guided short-read sequence assembler, iii choice of reference genome, and iv whether to perform read-quality filtering and trimming, on our ability to detect true SNPs and on the frequencies of errors. We performed benchmarking experiments, during which we assembled simulated and real Listeria monocytogenes strain 08-5578 short-read sequence datasets of varying quality with four commonly used assemblers (BWA, MOSAIK, Novoalign, and SMALT, using reference genomes of varying genetic distances, and with or without read pre-processing (i.e., quality filtering and trimming. We found that assemblies of at least 50-fold coverage provided the most accurate results. In addition, MOSAIK yielded the fewest errors when reads were aligned to a nearly identical reference genome, while using SMALT to align reads against a reference sequence that is ∼0.82% distant from 08-5578 at the nucleotide level resulted in the detection of the greatest numbers of true SNPs and the fewest errors. Finally, we show that whether read pre-processing improves SNP detection depends upon the choice of reference sequence and assembler. In total, this study demonstrates that researchers

Correction of echo shift in reconstruction processing for ultra-short TE pulse sequence

International Nuclear Information System (INIS)

Takizawa, Masahiro; Ootsuka, Takehiro; Abe, Takayuki; Takahashi, Tetsuhiko

2010-01-01

An ultra-short echo time (TE) pulse sequence is composed of a radial sampling that acquires echo signals radially in the K-space and a half-echo acquisition that acquires only half of the echo signal. The shift in the position of the echo signal (echo shift) caused by the timing errors in the gradient magnetic field pulses affects the image quality in the radial sampling with the half-echo acquisition. To improve image quality, we have developed a signal correction algorithm that detects and eliminates this echo shift during reconstruction by performing a pre-scan within 10 seconds. The results showed that image quality is improved under oblique and/or off-centering conditions that frequently cause image distortion due to hardware error. In conclusion, we have developed a robust ultra-short TE pulse sequence that allows wide latitude in the scan parameters, including oblique and off-centering conditions. (author)

Accurate estimation of short read mapping quality for next-generation genome sequencing

Science.gov (United States)

Ruffalo, Matthew; Koyutürk, Mehmet; Ray, Soumya; LaFramboise, Thomas

2012-01-01

Motivation: Several software tools specialize in the alignment of short next-generation sequencing reads to a reference sequence. Some of these tools report a mapping quality score for each alignment—in principle, this quality score tells researchers the likelihood that the alignment is correct. However, the reported mapping quality often correlates weakly with actual accuracy and the qualities of many mappings are underestimated, encouraging the researchers to discard correct mappings. Further, these low-quality mappings tend to correlate with variations in the genome (both single nucleotide and structural), and such mappings are important in accurately identifying genomic variants. Approach: We develop a machine learning tool, LoQuM (LOgistic regression tool for calibrating the Quality of short read mappings, to assign reliable mapping quality scores to mappings of Illumina reads returned by any alignment tool. LoQuM uses statistics on the read (base quality scores reported by the sequencer) and the alignment (number of matches, mismatches and deletions, mapping quality score returned by the alignment tool, if available, and number of mappings) as features for classification and uses simulated reads to learn a logistic regression model that relates these features to actual mapping quality. Results: We test the predictions of LoQuM on an independent dataset generated by the ART short read simulation software and observe that LoQuM can ‘resurrect’ many mappings that are assigned zero quality scores by the alignment tools and are therefore likely to be discarded by researchers. We also observe that the recalibration of mapping quality scores greatly enhances the precision of called single nucleotide polymorphisms. Availability: LoQuM is available as open source at http://compbio.case.edu/loqum/. Contact: matthew.ruffalo@case.edu. PMID:22962451

Probabilistic Motor Sequence Yields Greater Offline and Less Online Learning than Fixed Sequence.

Science.gov (United States)

Du, Yue; Prashad, Shikha; Schoenbrun, Ilana; Clark, Jane E

2016-01-01

It is well acknowledged that motor sequences can be learned quickly through online learning. Subsequently, the initial acquisition of a motor sequence is boosted or consolidated by offline learning. However, little is known whether offline learning can drive the fast learning of motor sequences (i.e., initial sequence learning in the first training session). To examine offline learning in the fast learning stage, we asked four groups of young adults to perform the serial reaction time (SRT) task with either a fixed or probabilistic sequence and with or without preliminary knowledge (PK) of the presence of a sequence. The sequence and PK were manipulated to emphasize either procedural (probabilistic sequence; no preliminary knowledge (NPK)) or declarative (fixed sequence; with PK) memory that were found to either facilitate or inhibit offline learning. In the SRT task, there were six learning blocks with a 2 min break between each consecutive block. Throughout the session, stimuli followed the same fixed or probabilistic pattern except in Block 5, in which stimuli appeared in a random order. We found that PK facilitated the learning of a fixed sequence, but not a probabilistic sequence. In addition to overall learning measured by the mean reaction time (RT), we examined the progressive changes in RT within and between blocks (i.e., online and offline learning, respectively). It was found that the two groups who performed the fixed sequence, regardless of PK, showed greater online learning than the other two groups who performed the probabilistic sequence. The groups who performed the probabilistic sequence, regardless of PK, did not display online learning, as indicated by a decline in performance within the learning blocks. However, they did demonstrate remarkably greater offline improvement in RT, which suggests that they are learning the probabilistic sequence offline. These results suggest that in the SRT task, the fast acquisition of a motor sequence is driven

The genome of flax (Linum usitatissimum) assembled de novo from short shotgun sequence reads.

Science.gov (United States)

Wang, Zhiwen; Hobson, Neil; Galindo, Leonardo; Zhu, Shilin; Shi, Daihu; McDill, Joshua; Yang, Linfeng; Hawkins, Simon; Neutelings, Godfrey; Datla, Raju; Lambert, Georgina; Galbraith, David W; Grassa, Christopher J; Geraldes, Armando; Cronk, Quentin C; Cullis, Christopher; Dash, Prasanta K; Kumar, Polumetla A; Cloutier, Sylvie; Sharpe, Andrew G; Wong, Gane K-S; Wang, Jun; Deyholos, Michael K

2012-11-01

Flax (Linum usitatissimum) is an ancient crop that is widely cultivated as a source of fiber, oil and medicinally relevant compounds. To accelerate crop improvement, we performed whole-genome shotgun sequencing of the nuclear genome of flax. Seven paired-end libraries ranging in size from 300 bp to 10 kb were sequenced using an Illumina genome analyzer. A de novo assembly, comprised exclusively of deep-coverage (approximately 94× raw, approximately 69× filtered) short-sequence reads (44-100 bp), produced a set of scaffolds with N(50) =694 kb, including contigs with N(50)=20.1 kb. The contig assembly contained 302 Mb of non-redundant sequence representing an estimated 81% genome coverage. Up to 96% of published flax ESTs aligned to the whole-genome shotgun scaffolds. However, comparisons with independently sequenced BACs and fosmids showed some mis-assembly of regions at the genome scale. A total of 43384 protein-coding genes were predicted in the whole-genome shotgun assembly, and up to 93% of published flax ESTs, and 86% of A. thaliana genes aligned to these predicted genes, indicating excellent coverage and accuracy at the gene level. Analysis of the synonymous substitution rates (K(s) ) observed within duplicate gene pairs was consistent with a recent (5-9 MYA) whole-genome duplication in flax. Within the predicted proteome, we observed enrichment of many conserved domains (Pfam-A) that may contribute to the unique properties of this crop, including agglutinin proteins. Together these results show that de novo assembly, based solely on whole-genome shotgun short-sequence reads, is an efficient means of obtaining nearly complete genome sequence information for some plant species. © 2012 The Authors. The Plant Journal © 2012 Blackwell Publishing Ltd.

Sequence composition and gene content of the short arm of rye (Secale cereale chromosome 1.

Directory of Open Access Journals (Sweden)

Silvia Fluch

Full Text Available BACKGROUND: The purpose of the study is to elucidate the sequence composition of the short arm of rye chromosome 1 (Secale cereale with special focus on its gene content, because this portion of the rye genome is an integrated part of several hundreds of bread wheat varieties worldwide. METHODOLOGY/PRINCIPAL FINDINGS: Multiple Displacement Amplification of 1RS DNA, obtained from flow sorted 1RS chromosomes, using 1RS ditelosomic wheat-rye addition line, and subsequent Roche 454FLX sequencing of this DNA yielded 195,313,589 bp sequence information. This quantity of sequence information resulted in 0.43× sequence coverage of the 1RS chromosome arm, permitting the identification of genes with estimated probability of 95%. A detailed analysis revealed that more than 5% of the 1RS sequence consisted of gene space, identifying at least 3,121 gene loci representing 1,882 different gene functions. Repetitive elements comprised about 72% of the 1RS sequence, Gypsy/Sabrina (13.3% being the most abundant. More than four thousand simple sequence repeat (SSR sites mostly located in gene related sequence reads were identified for possible marker development. The existence of chloroplast insertions in 1RS has been verified by identifying chimeric chloroplast-genomic sequence reads. Synteny analysis of 1RS to the full genomes of Oryza sativa and Brachypodium distachyon revealed that about half of the genes of 1RS correspond to the distal end of the short arm of rice chromosome 5 and the proximal region of the long arm of Brachypodium distachyon chromosome 2. Comparison of the gene content of 1RS to 1HS barley chromosome arm revealed high conservation of genes related to chromosome 5 of rice. CONCLUSIONS: The present study revealed the gene content and potential gene functions on this chromosome arm and demonstrated numerous sequence elements like SSRs and gene-related sequences, which can be utilised for future research as well as in breeding of wheat and rye.

RT-PCR for detection of all seven genotypes of Lyssavirus genus.

Science.gov (United States)

Vázquez-Morón, S; Avellón, A; Echevarría, J E

2006-08-01

The Lyssavirus genus includes seven species or genotypes named 1-7. Rabies genotypes correlate with geographical distribution and specific hosts. Co-circulation of different lyssaviruses, imported cases, and the presence of unknown viruses, such as Aravan, Khujand, Irkut and West Caucasian Bat Virus, make it necessary to use generic methods able to detect all lyssaviruses. Primer sequences were chosen from conserved regions in all genotypes in order to optimise a generic RT-PCR. Serial dilutions of 12 RNA extracts from all seven Lyssavirus genotypes were examined to compare the sensitivity of the RT-PCR standardised in this study with a published RT-PCR optimised for EBLV1 detection and capable of amplifying RNA from all seven lyssaviruses. All seven genotypes were detected by both RT-PCRs, however, the sensitivity was higher with the new version of the test. Twenty samples submitted for rabies diagnosis were tested by the new RT-PCR. Eight out of 20 samples from six dogs, one horse and one bat were found positive, in agreement with immunofluorescence results. Seven samples from terrestrial mammals were genotype 1 and one from a bat was genotype 5. In conclusion, this method can be used to complement immunofluorescence for the diagnosis of rabies, enabling the detection of unexpected lyssaviruses during rabies surveillance.

Reference gene identification for reliable normalisation of quantitative RT-PCR data in Setaria viridis.

Science.gov (United States)

Nguyen, Duc Quan; Eamens, Andrew L; Grof, Christopher P L

2018-01-01

Quantitative real-time polymerase chain reaction (RT-qPCR) is the key platform for the quantitative analysis of gene expression in a wide range of experimental systems and conditions. However, the accuracy and reproducibility of gene expression quantification via RT-qPCR is entirely dependent on the identification of reliable reference genes for data normalisation. Green foxtail ( Setaria viridis ) has recently been proposed as a potential experimental model for the study of C 4 photosynthesis and is closely related to many economically important crop species of the Panicoideae subfamily of grasses, including Zea mays (maize), Sorghum bicolor (sorghum) and Sacchurum officinarum (sugarcane). Setaria viridis (Accession 10) possesses a number of key traits as an experimental model, namely; (i) a small sized, sequenced and well annotated genome; (ii) short stature and generation time; (iii) prolific seed production, and; (iv) is amendable to Agrobacterium tumefaciens -mediated transformation. There is currently however, a lack of reference gene expression information for Setaria viridis ( S. viridis ). We therefore aimed to identify a cohort of suitable S. viridis reference genes for accurate and reliable normalisation of S. viridis RT-qPCR expression data. Eleven putative candidate reference genes were identified and examined across thirteen different S. viridis tissues. Of these, the geNorm and NormFinder analysis software identified SERINE / THERONINE - PROTEIN PHOSPHATASE 2A ( PP2A ), 5 '- ADENYLYLSULFATE REDUCTASE 6 ( ASPR6 ) and DUAL SPECIFICITY PHOSPHATASE ( DUSP ) as the most suitable combination of reference genes for the accurate and reliable normalisation of S. viridis RT-qPCR expression data. To demonstrate the suitability of the three selected reference genes, PP2A , ASPR6 and DUSP , were used to normalise the expression of CINNAMYL ALCOHOL DEHYDROGENASE ( CAD ) genes across the same tissues. This approach readily demonstrated the suitably of the three

Sequence Quality Analysis Tool for HIV Type 1 Protease and Reverse Transcriptase

OpenAIRE

DeLong, Allison K.; Wu, Mingham; Bennett, Diane; Parkin, Neil; Wu, Zhijin; Hogan, Joseph W.; Kantor, Rami

2012-01-01

Access to antiretroviral therapy is increasing globally and drug resistance evolution is anticipated. Currently, protease (PR) and reverse transcriptase (RT) sequence generation is increasing, including the use of in-house sequencing assays, and quality assessment prior to sequence analysis is essential. We created a computational HIV PR/RT Sequence Quality Analysis Tool (SQUAT) that runs in the R statistical environment. Sequence quality thresholds are calculated from a large dataset (46,802...

Study on the Image Quality Comparison between in Digital RT and Film RT

International Nuclear Information System (INIS)

Park, Sang Ki; Ahn, Yean Shik; Gil, Doo Song

2011-01-01

Conventional film radiographic test has been generally and widely used in the inspection on the weldment for quality assurance. On the other hand, since the analog RT is well known for typical time and cost consuming method with complex process of inspection, the industry has researched various ways how to improve radiographic test technology. In this study, we verified the fact that digital RT provides a lot more benefit in effectively detecting defects, ever film details, through digital processing of image enhancement, compared to film RT. As a result, we reached conclusion that digital RT is positively able to replace the film RT in industry in part or in whole

The genome of flax (Linum usitatissimum) assembled de novo from short shotgun sequence reads

DEFF Research Database (Denmark)

Wang, Zhiwen; Hobson, Neil; Galindo, Leonardo

2012-01-01

Flax (Linum usitatissimum) is an ancient crop that is widely cultivated as a source of fiber, oil and medicinally relevant compounds. To accelerate crop improvement, we performed whole-genome shotgun sequencing of the nuclear genome of flax. Seven paired-end libraries ranging in size from 300 bp...... these results show that de novo assembly, based solely on whole-genome shotgun short-sequence reads, is an efficient means of obtaining nearly complete genome sequence information for some plant species....

Conservation patterns of HIV-1 RT connection and RNase H domains: identification of new mutations in NRTI-treated patients.

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André F A Santos

Full Text Available BACKGROUND: Although extensive HIV drug resistance information is available for the first 400 amino acids of its reverse transcriptase, the impact of antiretroviral treatment in C-terminal domains of Pol (thumb, connection and RNase H is poorly understood. METHODS AND FINDINGS: We wanted to characterize conserved regions in RT C-terminal domains among HIV-1 group M subtypes and CRF. Additionally, we wished to identify NRTI-related mutations in HIV-1 RT C-terminal domains. We sequenced 118 RNase H domains from clinical viral isolates in Brazil, and analyzed 510 thumb and connection domain and 450 RNase H domain sequences collected from public HIV sequence databases, together with their treatment status and histories. Drug-naïve and NRTI-treated datasets were compared for intra- and inter-group conservation, and differences were determined using Fisher's exact tests. One third of RT C-terminal residues were found to be conserved among group M variants. Three mutations were found exclusively in NRTI-treated isolates. Nine mutations in the connection and 6 mutations in the RNase H were associated with NRTI treatment in subtype B. Some of them lay in or close to amino acid residues which contact nucleic acid or near the RNase H active site. Several of the residues pointed out herein have been recently associated to NRTI exposure or increase drug resistance to NRTI. CONCLUSIONS: This is the first comprehensive genotypic analysis of a large sequence dataset that describes NRTI-related mutations in HIV-1 RT C-terminal domains in vivo. The findings into the conservation of RT C-terminal domains may pave the way to more rational drug design initiatives targeting those regions.

Improving creativity performance by short-term meditation

Science.gov (United States)

2014-01-01

Background One form of meditation intervention, the integrative body-mind training (IBMT) has been shown to improve attention, reduce stress and change self-reports of mood. In this paper we examine whether short-term IBMT can improve performance related to creativity and determine the role that mood may play in such improvement. Methods Forty Chinese undergraduates were randomly assigned to short-term IBMT group or a relaxation training (RT) control group. Mood and creativity performance were assessed by the Positive and Negative Affect Schedule (PANAS) and Torrance Tests of Creative Thinking (TTCT) questionnaire respectively. Results As predicted, the results indicated that short-term (30 min per day for 7 days) IBMT improved creativity performance on the divergent thinking task, and yielded better emotional regulation than RT. In addition, cross-lagged analysis indicated that both positive and negative affect may influence creativity in IBMT group (not RT group). Conclusions Our results suggested that emotion-related creativity-promoting mechanism may be attributed to short-term meditation. PMID:24645871

Detection of Tomato black ring virus by real-time one-step RT-PCR.

Science.gov (United States)

Harper, Scott J; Delmiglio, Catia; Ward, Lisa I; Clover, Gerard R G

2011-01-01

A TaqMan-based real-time one-step RT-PCR assay was developed for the rapid detection of Tomato black ring virus (TBRV), a significant plant pathogen which infects a wide range of economically important crops. Primers and a probe were designed against existing genomic sequences to amplify a 72 bp fragment from RNA-2. The assay amplified all isolates of TBRV tested, but no amplification was observed from the RNA of other nepovirus species or healthy host plants. The detection limit of the assay was estimated to be around nine copies of the TBRV target region in total RNA. A comparison with conventional RT-PCR and ELISA, indicated that ELISA, the current standard test method, lacked specificity and reacted to all nepovirus species tested, while conventional RT-PCR was approximately ten-fold less sensitive than the real-time RT-PCR assay. Finally, the real-time RT-PCR assay was tested using five different RT-PCR reagent kits and was found to be robust and reliable, with no significant differences in sensitivity being found. The development of this rapid assay should aid in quarantine and post-border surveys for regulatory agencies. Copyright © 2010 Elsevier B.V. All rights reserved.

Molecular diagnosis of lyssaviruses and sequence comparison of Australian bat lyssavirus samples.

Science.gov (United States)

Foord, A J; Heine, H G; Pritchard, L I; Lunt, R A; Newberry, K M; Rootes, C L; Boyle, D B

2006-07-01

To evaluate and implement molecular diagnostic tests for the detection of lyssaviruses in Australia. A published hemi-nested reverse transcriptase polymerase chain reaction (RT-PCR) for the detection of all lyssavirus genotypes was modified to a fully nested RT-PCR format and compared with the original assay. TaqMan assays for the detection of Australian bat lyssavirus (ABLV) were compared with both the nested and hemi-nested RT-PCR assays. The sequences of RT-PCR products were determined to assess sequence variations of the target region (nucleocapsid gene) in samples of ABLV originating from different regions. The nested RT-PCR assay was highly analytically specific, and at least as analytically sensitive as the hemi-nested assay. The TaqMan assays were highly analytically specific and more analytically sensitive than either RT-PCR assay, with a detection level of approximately 10 genome equivalents per microl. Sequence of the first 544 nucleotides of the nucleocapsid protein coding sequence was obtained from all samples of ABLV received at Australian Animal Health Laboratory during the study period. The nested RT-PCR provided a means for molecular diagnosis of all tested genotypes of lyssavirus including classical rabies virus and Australian bat lyssavirus. The published TaqMan assay proved to be superior to the RT-PCR assays for the detection of ABLV in terms of analytical sensitivity. The TaqMan assay would also be faster and cross contamination is less likely. Nucleotide sequence analyses of samples of ABLV from a wide geographical range in Australia demonstrated the conserved nature of this region of the genome and therefore the suitability of this region for molecular diagnosis.

Use of short tandem repeat sequences to study Mycobacterium leprae in leprosy patients in Malawi and India.

Directory of Open Access Journals (Sweden)

Saroj K Young

2008-04-01

Full Text Available Inadequate understanding of the transmission of Mycobacterium leprae makes it difficult to predict the impact of leprosy control interventions. Genotypic tests that allow tracking of individual bacterial strains would strengthen epidemiological studies and contribute to our understanding of the disease.Genotyping assays based on variation in the copy number of short tandem repeat sequences were applied to biopsies collected in population-based epidemiological studies of leprosy in northern Malawi, and from members of multi-case households in Hyderabad, India. In the Malawi series, considerable genotypic variability was observed between patients, and also within patients, when isolates were collected at different times or from different tissues. Less within-patient variability was observed when isolates were collected from similar tissues at the same time. Less genotypic variability was noted amongst the closely related Indian patients than in the Malawi series.Lineages of M. leprae undergo changes in their pattern of short tandem repeat sequences over time. Genetic divergence is particularly likely between bacilli inhabiting different (e.g., skin and nerve tissues. Such variability makes short tandem repeat sequences unsuitable as a general tool for population-based strain typing of M. leprae, or for distinguishing relapse from reinfection. Careful use of these markers may provide insights into the development of disease within individuals and for tracking of short transmission chains.

Fine de novo sequencing of a fungal genome using only SOLiD short read data: verification on Aspergillus oryzae RIB40.

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Myco Umemura

Full Text Available The development of next-generation sequencing (NGS technologies has dramatically increased the throughput, speed, and efficiency of genome sequencing. The short read data generated from NGS platforms, such as SOLiD and Illumina, are quite useful for mapping analysis. However, the SOLiD read data with lengths of <60 bp have been considered to be too short for de novo genome sequencing. Here, to investigate whether de novo sequencing of fungal genomes is possible using only SOLiD short read sequence data, we performed de novo assembly of the Aspergillus oryzae RIB40 genome using only SOLiD read data of 50 bp generated from mate-paired libraries with 2.8- or 1.9-kb insert sizes. The assembled scaffolds showed an N50 value of 1.6 Mb, a 22-fold increase than those obtained using only SOLiD short read in other published reports. In addition, almost 99% of the reference genome was accurately aligned by the assembled scaffold fragments in long lengths. The sequences of secondary metabolite biosynthetic genes and clusters, whose products are of considerable interest in fungal studies due to their potential medicinal, agricultural, and cosmetic properties, were also highly reconstructed in the assembled scaffolds. Based on these findings, we concluded that de novo genome sequencing using only SOLiD short reads is feasible and practical for molecular biological study of fungi. We also investigated the effect of filtering low quality data, library insert size, and k-mer size on the assembly performance, and recommend for the assembly use of mild filtered read data where the N50 was not so degraded and the library has an insert size of ∼2.0 kb, and k-mer size 33.

Characterization of bovine ruminal epithelial bacterial communities using 16S rRNA sequencing, PCR-DGGE, and qRT-PCR analysis.

Science.gov (United States)

Li, Meiju; Zhou, Mi; Adamowicz, Elizabeth; Basarab, John A; Guan, Le Luo

2012-02-24

Currently, knowledge regarding the ecology and function of bacteria attached to the epithelial tissue of the rumen wall is limited. In this study, the diversity of the bacterial community attached to the rumen epithelial tissue was compared to the rumen content bacterial community using 16S rRNA gene sequencing, PCR-DGGE, and qRT-PCR analysis. Sequence analysis of 2785 randomly selected clones from six 16S rDNA (∼1.4kb) libraries showed that the community structures of three rumen content libraries clustered together and were separated from the rumen tissue libraries. The diversity index of each library revealed that ruminal content bacterial communities (4.12/4.42/4.88) were higher than ruminal tissue communities (2.90/2.73/3.23), based on 97% similarity. The phylum Firmicutes was predominant in the ruminal tissue communities, while the phylum Bacteroidetes was predominant in the ruminal content communities. The phyla Fibrobacteres, Planctomycetes, and Verrucomicrobia were only detected in the ruminal content communities. PCR-DGGE analysis of the bacterial profiles of the rumen content and ruminal epithelial tissue samples from 22 steers further confirmed that there is a distinct bacterial community that inhibits the rumen epithelium. The distinctive epimural bacterial communities suggest that Firmicutes, together with other epithelial-specific species, may have additional functions other than food digestion. Copyright © 2011 Elsevier B.V. All rights reserved.

Easy and accurate reconstruction of whole HIV genomes from short-read sequence data with shiver

Science.gov (United States)

Blanquart, François; Golubchik, Tanya; Gall, Astrid; Bakker, Margreet; Bezemer, Daniela; Croucher, Nicholas J; Hall, Matthew; Hillebregt, Mariska; Ratmann, Oliver; Albert, Jan; Bannert, Norbert; Fellay, Jacques; Fransen, Katrien; Gourlay, Annabelle; Grabowski, M Kate; Gunsenheimer-Bartmeyer, Barbara; Günthard, Huldrych F; Kivelä, Pia; Kouyos, Roger; Laeyendecker, Oliver; Liitsola, Kirsi; Meyer, Laurence; Porter, Kholoud; Ristola, Matti; van Sighem, Ard; Cornelissen, Marion; Kellam, Paul; Reiss, Peter

2018-01-01

Abstract Studying the evolution of viruses and their molecular epidemiology relies on accurate viral sequence data, so that small differences between similar viruses can be meaningfully interpreted. Despite its higher throughput and more detailed minority variant data, next-generation sequencing has yet to be widely adopted for HIV. The difficulty of accurately reconstructing the consensus sequence of a quasispecies from reads (short fragments of DNA) in the presence of large between- and within-host diversity, including frequent indels, may have presented a barrier. In particular, mapping (aligning) reads to a reference sequence leads to biased loss of information; this bias can distort epidemiological and evolutionary conclusions. De novo assembly avoids this bias by aligning the reads to themselves, producing a set of sequences called contigs. However contigs provide only a partial summary of the reads, misassembly may result in their having an incorrect structure, and no information is available at parts of the genome where contigs could not be assembled. To address these problems we developed the tool shiver to pre-process reads for quality and contamination, then map them to a reference tailored to the sample using corrected contigs supplemented with the user’s choice of existing reference sequences. Run with two commands per sample, it can easily be used for large heterogeneous data sets. We used shiver to reconstruct the consensus sequence and minority variant information from paired-end short-read whole-genome data produced with the Illumina platform, for sixty-five existing publicly available samples and fifty new samples. We show the systematic superiority of mapping to shiver’s constructed reference compared with mapping the same reads to the closest of 3,249 real references: median values of 13 bases called differently and more accurately, 0 bases called differently and less accurately, and 205 bases of missing sequence recovered. We also

Short-read reading-frame predictors are not created equal: sequence error causes loss of signal

Directory of Open Access Journals (Sweden)

Trimble William L

2012-07-01

Full Text Available Abstract Background Gene prediction algorithms (or gene callers are an essential tool for analyzing shotgun nucleic acid sequence data. Gene prediction is a ubiquitous step in sequence analysis pipelines; it reduces the volume of data by identifying the most likely reading frame for a fragment, permitting the out-of-frame translations to be ignored. In this study we evaluate five widely used ab initio gene-calling algorithms—FragGeneScan, MetaGeneAnnotator, MetaGeneMark, Orphelia, and Prodigal—for accuracy on short (75–1000 bp fragments containing sequence error from previously published artificial data and “real” metagenomic datasets. Results While gene prediction tools have similar accuracies predicting genes on error-free fragments, in the presence of sequencing errors considerable differences between tools become evident. For error-containing short reads, FragGeneScan finds more prokaryotic coding regions than does MetaGeneAnnotator, MetaGeneMark, Orphelia, or Prodigal. This improved detection of genes in error-containing fragments, however, comes at the cost of much lower (50% specificity and overprediction of genes in noncoding regions. Conclusions Ab initio gene callers offer a significant reduction in the computational burden of annotating individual nucleic acid reads and are used in many metagenomic annotation systems. For predicting reading frames on raw reads, we find the hidden Markov model approach in FragGeneScan is more sensitive than other gene prediction tools, while Prodigal, MGA, and MGM are better suited for higher-quality sequences such as assembled contigs.

Detecting Newcastle disease virus in combination of RT-PCR with red blood cell absorption

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Liu Chengqian

2011-05-01

Full Text Available Abstract Reverse transcription-polymerase chain reaction (RT-PCR has limited sensitivity when treating complicated samples, such as feces, waste-water in farms, and nucleic acids, protein rich tissue samples, all the factors may interfere with the sensitivity of PCR test or generate false results. In this study, we developed a sensitive RT-PCR, combination of red blood cell adsorption, for detecting Newcastle disease virus (NDV. One pair of primers which was highly homologous to three NDV pathotypes was designed according to the consensus nucleocapsid protein (NP gene sequence. To eliminate the interfere of microbes and toxic substances, we concentrated and purified NDV from varied samples utilizing the ability of NDV binding red blood cells (RBCs. The RT-PCR coupled with red blood cell adsorption was much more sensitive in comparison with regular RT-PCR. The approach could also be used to detect other viruses with the property of hemagglutination, such as influenza viruses.

A sensitive short read homology search tool for paired-end read sequencing data.

Science.gov (United States)

Techa-Angkoon, Prapaporn; Sun, Yanni; Lei, Jikai

2017-10-16

Homology search is still a significant step in functional analysis for genomic data. Profile Hidden Markov Model-based homology search has been widely used in protein domain analysis in many different species. In particular, with the fast accumulation of transcriptomic data of non-model species and metagenomic data, profile homology search is widely adopted in integrated pipelines for functional analysis. While the state-of-the-art tool HMMER has achieved high sensitivity and accuracy in domain annotation, the sensitivity of HMMER on short reads declines rapidly. The low sensitivity on short read homology search can lead to inaccurate domain composition and abundance computation. Our experimental results showed that half of the reads were missed by HMMER for a RNA-Seq dataset. Thus, there is a need for better methods to improve the homology search performance for short reads. We introduce a profile homology search tool named Short-Pair that is designed for short paired-end reads. By using an approximate Bayesian approach employing distribution of fragment lengths and alignment scores, Short-Pair can retrieve the missing end and determine true domains. In particular, Short-Pair increases the accuracy in aligning short reads that are part of remote homologs. We applied Short-Pair to a RNA-Seq dataset and a metagenomic dataset and quantified its sensitivity and accuracy on homology search. The experimental results show that Short-Pair can achieve better overall performance than the state-of-the-art methodology of profile homology search. Short-Pair is best used for next-generation sequencing (NGS) data that lack reference genomes. It provides a complementary paired-end read homology search tool to HMMER. The source code is freely available at https://sourceforge.net/projects/short-pair/ .

SMAP Data Assimilation at NASA SPoRT

Science.gov (United States)

Blankenship, Clay B.; Case, Jonathan L.; Zavodsky, Bradley T.

2016-01-01

The NASA Short-Term Prediction Research and Transition (SPoRT) Center maintains a near-real- time run of the Noah Land Surface Model within the Land Information System (LIS) at 3-km resolution. Soil moisture products from this model are used by several NOAA/National Weather Service Weather Forecast Offices for flood and drought situational awareness. We have implemented assimilation of soil moisture retrievals from the Soil Moisture Ocean Salinity (SMOS) and Soil Moisture Active/ Passive (SMAP) satellites, and are now evaluating the SMAP assimilation. The SMAP-enhanced LIS product is planned for public release by October 2016.

NASA SPoRT JPSS PG Activities in Alaska

Science.gov (United States)

Berndt, Emily; Molthan, Andrew; Fuell, Kevin; McGrath, Kevin; Smith, Matt; LaFontaine, Frank; Leroy, Anita; White, Kris

2018-01-01

SPoRT (NASA's Short-term Prediction Research and Transition Center) has collaboratively worked with Alaska WFOs (Weather Forecast Offices) to introduce RGB (Red/Green/Blue false color image) imagery to prepare for NOAA-20 (National Oceanic and Atmospheric Administration, JPSS (Joint Polar Satellite System) series-20 satellite) VIIRS (Visible Infrared Imaging Radiometer Suite) and improve forecasting aviation-related hazards. Last R2O/O2R (Research-to-Operations/Operations-to-Research) steps include incorporating NOAA-20 VIIRS in RGB suite and fully transitioning client-side RGB processing to GINA (Geographic Information Network of Alaska) and Alaska Region. Alaska Region WFOs have been part of the successful R2O/O2R story to assess the use of NESDIS (National Environmental Satellite, Data, and Information Service) Snowfall Rate product in operations. SPoRT introduced passive microwave rain rate and IMERG (Integrated Multi-satellitE Retrievals for GPM (Global Precipitation Measurement)) (IMERG) to Alaska WFOs for use in radar-void areas and assessing flooding potential. SPoRT has been part of the multi-organization collaborative effort to introduce Gridded NUCAPS (NOAA Unique CrIS/ATMS (Crosstrack Infrared Sounder/Advanced Technology Microwave Sounder) Processing System) to the Anchorage CWSU (Center Weather Service Unit) to assess Cold Air Aloft events, [and as part of NOAA's PG (Product Generation) effort].

GenHtr: a tool for comparative assessment of genetic heterogeneity in microbial genomes generated by massive short-read sequencing

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Yu GongXin

2010-10-01

Full Text Available Abstract Background Microevolution is the study of short-term changes of alleles within a population and their effects on the phenotype of organisms. The result of the below-species-level evolution is heterogeneity, where populations consist of subpopulations with a large number of structural variations. Heterogeneity analysis is thus essential to our understanding of how selective and neutral forces shape bacterial populations over a short period of time. The Solexa Genome Analyzer, a next-generation sequencing platform, allows millions of short sequencing reads to be obtained with great accuracy, allowing for the ability to study the dynamics of the bacterial population at the whole genome level. The tool referred to as GenHtr was developed for genome-wide heterogeneity analysis. Results For particular bacterial strains, GenHtr relies on a set of Solexa short reads on given bacteria pathogens and their isogenic reference genome to identify heterogeneity sites, the chromosomal positions with multiple variants of genes in the bacterial population, and variations that occur in large gene families. GenHtr accomplishes this by building and comparatively analyzing genome-wide heterogeneity genotypes for both the newly sequenced genomes (using massive short-read sequencing and their isogenic reference (using simulated data. As proof of the concept, this approach was applied to SRX007711, the Solexa sequencing data for a newly sequenced Staphylococcus aureus subsp. USA300 cell line, and demonstrated that it could predict such multiple variants. They include multiple variants of genes critical in pathogenesis, e.g. genes encoding a LysR family transcriptional regulator, 23 S ribosomal RNA, and DNA mismatch repair protein MutS. The heterogeneity results in non-synonymous and nonsense mutations, leading to truncated proteins for both LysR and MutS. Conclusion GenHtr was developed for genome-wide heterogeneity analysis. Although it is much more time

Comparison of ELISA and dual stage real time RT-PCR to differentiate Sabin like and non-Sabin like poliovirus isolates.

Science.gov (United States)

Kaundal, Nirmal; Sarkate, Purva; Prakash, Charu; Rishi, Narayan

2017-06-01

Environmental surveillance of polioviruses has been used as an important tool in monitoring circulation of wild polioviruses and/or Vaccine derived polioviruses in sewage samples. It is important to distinguish Sabin like isolates from non-Sabin like; ELISA & dual stage real time RT-PCR have been used for the same. Current study was carried out on sewage isolates to compare ELISA & RT-PCR with sequencing to distinguish Sabin like from non-Sabin like. Out of 468 sewage specimens, 91 (19.44%) were non-polio enteroviruses positive and 377 (80.56%) were polio positive by virus isolation method. A total of 488 polio virus isolates were detected by L20B and RD route which were further subjected to ELISA and RT-PCR. The results were compared with sequencing. On comparison, the specificity of ELISA was only 66.67% in spite of very low sensitivity (3.43%). The sensitivity of RT-PCR was 97.71% which makes it a good primary screening test for detection of non-Sabin like viruses. However, the specificity was only 33.33%. RT-PCR appears to be a sensitive tool for detecting non-Sabin like viruses however; the isolates which are non-Sabin like by RT-PCR may not necessarily be mutated viruses. ELISA cannot be used for differentiation of Sabin likes from non-Sabin likes due to low sensitivity.

Computational complexity of algorithms for sequence comparison, short-read assembly and genome alignment.

Science.gov (United States)

Baichoo, Shakuntala; Ouzounis, Christos A

A multitude of algorithms for sequence comparison, short-read assembly and whole-genome alignment have been developed in the general context of molecular biology, to support technology development for high-throughput sequencing, numerous applications in genome biology and fundamental research on comparative genomics. The computational complexity of these algorithms has been previously reported in original research papers, yet this often neglected property has not been reviewed previously in a systematic manner and for a wider audience. We provide a review of space and time complexity of key sequence analysis algorithms and highlight their properties in a comprehensive manner, in order to identify potential opportunities for further research in algorithm or data structure optimization. The complexity aspect is poised to become pivotal as we will be facing challenges related to the continuous increase of genomic data on unprecedented scales and complexity in the foreseeable future, when robust biological simulation at the cell level and above becomes a reality. Copyright © 2017 Elsevier B.V. All rights reserved.

A simplified strategy for sensitive detection of Rose rosette virus compatible with three RT-PCR chemistries.

Science.gov (United States)

Dobhal, Shefali; Olson, Jennifer D; Arif, Mohammad; Garcia Suarez, Johnny A; Ochoa-Corona, Francisco M

2016-06-01

Rose rosette disease is a disorder associated with infection by Rose rosette virus (RRV), a pathogen of roses that causes devastating effects on most garden cultivated varieties, and the wild invasive rose especially Rosa multiflora. Reliable and sensitive detection of this disease in early phases is needed to implement proper control measures. This study assesses a single primer-set based detection method for RRV and demonstrates its application in three different chemistries: Endpoint RT-PCR, TaqMan-quantitative RT-PCR (RT-qPCR) and SYBR Green RT-qPCR with High Resolution Melting analyses. A primer set (RRV2F/2R) was designed from consensus sequences of the nucleocapsid protein gene p3 located in the RNA 3 region of RRV. The specificity of primer set RRV2F/2R was validated in silico against published GenBank sequences and in-vitro against infected plant samples and an exclusivity panel of near-neighbor and other viruses that commonly infect Rosa spp. The developed assay is sensitive with a detection limit of 1fg from infected plant tissue. Thirty rose samples from 8 different states of the United States were tested using the developed methods. The developed methods are sensitive and reliable, and can be used by diagnostic laboratories for routine testing and disease management decisions. Copyright © 2016 Elsevier B.V. All rights reserved.

Development and evaluation of tailored specific real-time RT-PCR assays for detection of foot-and-mouth disease virus serotypes circulating in East Africa

DEFF Research Database (Denmark)

Bachanek-Bankowska, Katarzyna; Mero, Herieth R.; Wadsworth, Jemma

2016-01-01

Rapid, reliable and accurate diagnostic methods provide essential support to programmes that monitor and control foot-and-mouth disease (FMD). While pan-specific molecular tests for FMD virus (FMDV) detection are well established and widely used in endemic and FMD-free countries, current serotyping...... methods mainly rely either on antigen detection ELISAs or nucleotide sequencing approaches. This report describes the development of a panel of serotype-specific real-time RT-PCR assays (rRT-PCR) tailored to detect FMDV lineages currently circulating in East Africa. These assays target sequences within...... sequencing. Samples (n = 71) representing serotype A (topotype AFRICA, lineage G-I), serotype O (topotypes EA-2 and EA-4), serotype SAT 1 (topotype I (NWZ)) and serotype SAT2 (topotype IV) were correctly identified with these rRT-PCR assays. Furthermore, FMDV RNA from samples that did not contain infectious...

POLYCLONAL ANTIBODIES TO DETECT THE CP-RT PROTEIN OF Potato Mop-Top VIRUS ANTICUERPOS POLICLONALES PARA LA DETECCIÓN DE LA PROTEÍNA CP-RT DEL Potato Mop-Top VIRUS

Directory of Open Access Journals (Sweden)

Derly Andrade Molina

2012-06-01

Full Text Available Abstract. Potato mop-top virus (PMTV causes an important re-emerging disease in potato crops in Colombia due to the increased incidence of its protozoan vector Spongospora subterranea f.sp. subterranea (Sss, the causing agent of Powdery scab disease. For an accurate detection of PMTV it is recommended to combine different diagnostic tests and evaluate multiple samples per plant or tissue. In order to increase the number of available tools for detection of PMTV, antibodies targeting the RT domain of the CP-RT protein were developed in this work. Sequencing of the RT domain from the colombian strain R25 was achieved and using bioinformatic analysis, a potential antigenic region was identified. A peptide mimicking the antigenic region was inoculated in rabbits for the production of polyclonal antibodies. The antibodies were tested by ELISA using Nicotiana benthamiana bait-plants infected with Sss cystosori and potato plants collected in La Unión (Antioquia. The validity of serological tests was confirmed by RT-PCR. A complete sequence of the RT domain and 441 nt of the CP gene were obtained. Phylogenetic analysis identified strain R25 as closely associated to the PMTV lineage distributed worldwide. A total of 19.26 mg of anti-CP-RT polyclonal antibodies useful in detecting PMTV in infected plants were obtained. As CP-RT is involved in transmission of PMTV by Sss, these antibodies will be useful for supporting not only diagnostic programs but also basic and epidemiologic studies aimed at understanding interactions between PMTV and Sss.Resumen. El Potato mop-top virus (PMTV es uno de los virus re-emergentes en los cultivos de papa de Colombia, como resultado del aumento de la incidencia de su vector natural, el protozoo Spongospora subterranea f.sp. mamarinm@unal.edu.co (Sss, agente causal de la Sarna polvosa de la papa. Para la detección del virus se recomienda la realización simultánea de diferentes pruebas, así como la evaluación de m

Monitoring and improving the sensitivity of dengue nested RT-PCR used in longitudinal surveillance in Thailand.

Science.gov (United States)

Klungthong, Chonticha; Manasatienkij, Wudtichai; Phonpakobsin, Thipwipha; Chinnawirotpisan, Piyawan; Rodpradit, Prinyada; Hussem, Kittinun; Thaisomboonsuk, Butsaya; Ong-ajchaowlerd, Prapapun; Nisalak, Ananda; Kalayanarooj, Siripen; Buddhari, Darunee; Gibbons, Robert V; Jarman, Richard G; Yoon, In-Kyu; Fernandez, Stefan

2015-02-01

AFRIMS longitudinal dengue surveillance in Thailand depends on the nested RT-PCR and the dengue IgM/IgG ELISA. To examine and improve the sensitivity of the nested RT-PCR using a panel of archived samples collected during dengue surveillance. A retrospective analysis of 16,454 dengue IgM/IgG ELISA positive cases collected between 2000 and 2013 was done to investigate the sensitivity of the nested RT-PCR. From these cases, 318 acute serum specimens or extracted RNA, previously found to be negative by the nested RT-PCR, were tested using TaqMan real-time RT-PCR (TaqMan rRT-PCR). To improve the sensitivity of nested RT-PCR, we designed a new primer based on nucleotide sequences from contemporary strains found to be positive by the TaqMan rRT-PCR. Sensitivity of the new nested PCR was calculated using a panel of 87 samples collected during 2011-2013. The percentage of dengue IgM/IgG ELISA positive cases that were negative by the nested RT-PCR varied from 17% to 42% for all serotypes depending on the year. Using TaqMan rRT-PCR, dengue RNA was detected in 194 (61%) of the 318 acute sera or extracted RNA previously found to be negative by the nested RT-PCR. The newly designed DENV-1 specific primer increased the sensitivity of DENV-1 detection by the nested RT-PCR from 48% to 88%, and of all 4 serotypes from 73% to 87%. These findings demonstrate the impact of genetic diversity and signal erosion on the sensitivity of PCR-based methods. Published by Elsevier B.V.

One-step cross-genogroup multiplex RT-qPCR with an internal control system for the detection of infectious pancreatic necrosis virus (IPNV).

Science.gov (United States)

Hoferer, Marc; Braun, Anne; Skrypski, Julia; Bock, Sabine; Thalheim, Sabine; Sting, Reinhard

2017-09-01

Infectious pancreatic necrosis virus (IPNV) causes great losses in fish hatcheries world-wide. The detection of IPNV can be challenging in certain circumstances, particularly due to low viral load and the genetic variability of this RNA virus. For the first time, this project created a quantitative triplex real-time reverse transcription PCR (RT-qPCR), including an endogenous control system, for specific, sensitive and rapid detection of IPNV in routine diagnostics. Multiple sequence alignment of 46 nucleotide sequences of the segment A genome obtained from the NCBI database allowed the design of two RT-qPCR systems covering the IPNV genogroup 1 and genogroups 2-5, respectively. The completed triplex RT-qPCR including a salmonid-specific endogenous control showed high specificity and an analytical sensitivity of 20-40 oligonucleotide copies. Testing of dilution series of virus-loaded cell culture suspensions proved equality of the triplex RT-qPCR with virus detection in cell culture and a higher sensitivity than conventional RT-PCR in field samples. In comparative studies of a total of 77 field samples tested, 51 showed identical positive and 19 identical negative results in cell culture and the triplex RT-qPCR. However, seven other samples yielded positive results in the triplex RT-qPCR, but negative results in cell culture. Copyright © 2017 Elsevier B.V. All rights reserved.

Evaluation of ALK gene rearrangement in central nervous system metastases of non-small-cell lung cancer using two-step RT-PCR technique.

Science.gov (United States)

Nicoś, M; Krawczyk, P; Wojas-Krawczyk, K; Bożyk, A; Jarosz, B; Sawicki, M; Trojanowski, T; Milanowski, J

2017-12-01

RT-PCR technique has showed a promising value as pre-screening method for detection of mRNA containing abnormal ALK sequences, but its sensitivity and specificity is still discussable. Previously, we determined the incidence of ALK rearrangement in CNS metastases of NSCLC using IHC and FISH methods. We evaluated ALK gene rearrangement using two-step RT-PCR method with EML4-ALK Fusion Gene Detection Kit (Entrogen, USA). The studied group included 145 patients (45 females, 100 males) with CNS metastases of NSCLC and was heterogeneous in terms of histology and smoking status. 21% of CNS metastases of NSCLC (30/145) showed presence of mRNA containing abnormal ALK sequences. FISH and IHC tests confirmed the presence of ALK gene rearrangement and expression of ALK abnormal protein in seven patients with positive result of RT-PCR analysis (4.8% of all patients, 20% of RT-PCR positive patients). RT-PCR method compared to FISH analysis achieved 100% of sensitivity and only 82.7% of specificity. IHC method compared to FISH method indicated 100% of sensitivity and 97.8% of specificity. In comparison to IHC, RT-PCR showed identical sensitivity with high number of false positive results. Utility of RT-PCR technique in screening of ALK abnormalities and in qualification patients for molecularly targeted therapies needs further validation.

Single reaction, real time RT-PCR detection of all known avian and human metapneumoviruses.

Science.gov (United States)

Lemaitre, E; Allée, C; Vabret, A; Eterradossi, N; Brown, P A

2018-01-01

Current molecular methods for the detection of avian and human metapneumovirus (AMPV, HMPV) are specifically targeted towards each virus species or individual subgroups of these. Here a broad range SYBR Green I real time RT-PCR was developed which amplified a highly conserved fragment of sequence in the N open reading frame. This method was sufficiently efficient and specific in detecting all MPVs. Its validation according to the NF U47-600 norm for the four AMPV subgroups estimated low limits of detection between 1000 and 10copies/μL, similar with detection levels described previously for real time RT-PCRs targeting specific subgroups. RNA viruses present a challenge for the design of durable molecular diagnostic test due to the rate of change in their genome sequences which can vary substantially in different areas and over time. The fact that the regions of sequence for primer hybridization in the described method have remained sufficiently conserved since the AMPV and HMPV diverged, should give the best chance of continued detection of current subgroups and of potential unknown or future emerging MPV strains. Copyright © 2017 Elsevier B.V. All rights reserved.

A study on the establishment of practical program for the RT promotion plan

Energy Technology Data Exchange (ETDEWEB)

Kim, Seong Yen; Jin, J. H.; Beyn, M. W.; Han, H. S.; Park, C.; Lee, J. W.; Yang, M. S.; Jeong, I. H.; Nho, Y. C

2003-03-01

The present state of RT utilizations in Korea is categorized into radioisotope applications to treat incurable diseases, food irradiation, and nondestructive test, and radiation applications by radiation generators such as cyclotrons to produce short half-life radioisotopes for medical uses and electron accelerators to make better products of cables and radial tiers. In order to activate more utilizations of RT, the followings should be carried out: 1. Establishment of RT Research Institute and RT Industrial Complex - Establishment of the tentatively named [Radiation Science and Technology Research Center] and 'Cyclotron Center' in four provinces should be effectively carried out as scheduled. - Successful pursuing design and construction of the proton accelerator for research and technology development, and CNS research facility. - Radiation area workers training participating the RT projects; Participation and training of university graduates to the on-going and planned RT projects, Classes for special skills and experts to radiation could be designed at the Consociated Graduate School, being established by joining national research organizations, if necessary. - Secure minimum number of R and D man power for RT development 2. Review design and construction of RI production reactor and construct distribution and transportation of RIs - Constructions of distribution lines and a central depository site with the establishment of Cyclotron Centers and RI production reactor - Designation, as early as possible, of the organization to handle the constructions of the distribution lines and the central depository site. 3. Establishment of an organization to appeal RT industries' concerns - An organization composed of governmental officials, researchers from national research institutes and/or private research institutes, representatives from associations and RT industries should be formed to define the issues to promote radiation uses. 4. Establishment of

A study on the establishment of practical program for the RT promotion plan

International Nuclear Information System (INIS)

Kim, Seong Yen; Jin, J. H.; Beyn, M. W.; Han, H. S.; Park, C.; Lee, J. W.; Yang, M. S.; Jeong, I. H.; Nho, Y. C.

2003-03-01

The present state of RT utilizations in Korea is categorized into radioisotope applications to treat incurable diseases, food irradiation, and nondestructive test, and radiation applications by radiation generators such as cyclotrons to produce short half-life radioisotopes for medical uses and electron accelerators to make better products of cables and radial tiers. In order to activate more utilizations of RT, the followings should be carried out: 1. Establishment of RT Research Institute and RT Industrial Complex - Establishment of the tentatively named [Radiation Science and Technology Research Center] and 'Cyclotron Center' in four provinces should be effectively carried out as scheduled. - Successful pursuing design and construction of the proton accelerator for research and technology development, and CNS research facility. - Radiation area workers training participating the RT projects; Participation and training of university graduates to the on-going and planned RT projects, Classes for special skills and experts to radiation could be designed at the Consociated Graduate School, being established by joining national research organizations, if necessary. - Secure minimum number of R and D man power for RT development 2. Review design and construction of RI production reactor and construct distribution and transportation of RIs - Constructions of distribution lines and a central depository site with the establishment of Cyclotron Centers and RI production reactor - Designation, as early as possible, of the organization to handle the constructions of the distribution lines and the central depository site. 3. Establishment of an organization to appeal RT industries' concerns - An organization composed of governmental officials, researchers from national research institutes and/or private research institutes, representatives from associations and RT industries should be formed to define the issues to promote radiation uses. 4. Establishment of Pan-ministry PR

Short Interspersed Nuclear Element (SINE) Sequences in the Genome of the Human Pathogenic Fungus Aspergillus fumigatus Af293.

Science.gov (United States)

Kanhayuwa, Lakkhana; Coutts, Robert H A

2016-01-01

Novel families of short interspersed nuclear element (SINE) sequences in the human pathogenic fungus Aspergillus fumigatus, clinical isolate Af293, were identified and categorised into tRNA-related and 5S rRNA-related SINEs. Eight predicted tRNA-related SINE families originating from different tRNAs, and nominated as AfuSINE2 sequences, contained target site duplications of short direct repeat sequences (4-14 bp) flanking the elements, an extended tRNA-unrelated region and typical features of RNA polymerase III promoter sequences. The elements ranged in size from 140-493 bp and were present in low copy number in the genome and five out of eight were actively transcribed. One putative tRNAArg-derived sequence, AfuSINE2-1a possessed a unique feature of repeated trinucleotide ACT residues at its 3'-terminus. This element was similar in sequence to the I-4_AO element found in A. oryzae and an I-1_AF long nuclear interspersed element-like sequence identified in A. fumigatus Af293. Families of 5S rRNA-related SINE sequences, nominated as AfuSINE3, were also identified and their 5'-5S rRNA-related regions show 50-65% and 60-75% similarity to respectively A. fumigatus 5S rRNAs and SINE3-1_AO found in A. oryzae. A. fumigatus Af293 contains five copies of AfuSINE3 sequences ranging in size from 259-343 bp and two out of five AfuSINE3 sequences were actively transcribed. Investigations on AfuSINE distribution in the fungal genome revealed that the elements are enriched in pericentromeric and subtelomeric regions and inserted within gene-rich regions. We also demonstrated that some, but not all, AfuSINE sequences are targeted by host RNA silencing mechanisms. Finally, we demonstrated that infection of the fungus with mycoviruses had no apparent effects on SINE activity.

STAR-RT: Visual attention for real-time video game playing

OpenAIRE

Kotseruba, Iuliia; Tsotsos, John K.

2017-01-01

In this paper we present STAR-RT - the first working prototype of Selective Tuning Attention Reference (STAR) model and Cognitive Programs (CPs). The Selective Tuning (ST) model received substantial support through psychological and neurophysiological experiments. The STAR framework expands ST and applies it to practical visual tasks. In order to do so, similarly to many cognitive architectures, STAR combines the visual hierarchy (based on ST) with the executive controller, working and short-...

Rapid Multiplex Small DNA Sequencing on the MinION Nanopore Sequencing Platform

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Shan Wei

2018-05-01

Full Text Available Real-time sequencing of short DNA reads has a wide variety of clinical and research applications including screening for mutations, target sequences and aneuploidy. We recently demonstrated that MinION, a nanopore-based DNA sequencing device the size of a USB drive, could be used for short-read DNA sequencing. In this study, an ultra-rapid multiplex library preparation and sequencing method for the MinION is presented and applied to accurately test normal diploid and aneuploidy samples’ genomic DNA in under three hours, including library preparation and sequencing. This novel method shows great promise as a clinical diagnostic test for applications requiring rapid short-read DNA sequencing.

Challenges in Transitioning Research Data to Operations: The SPoRT Paradigm

Science.gov (United States)

Jedloved, Gary J.; Smith, Matt; McGrath, Kevin

2010-01-01

Established in 2002 to demonstrate the weather and forecasting application of real-time EOS measurements, the NASA Short-term Prediction Research and Transition (SPoRT) program has grown to be an end-to-end research to operations activity focused on the use of advanced NASA modeling and data assimilation approaches, nowcasting techniques, and unique high-resolution multispectral data from EOS satellites to improve short-term weather forecasts on a regional and local scale. With the ever-broadening application of real-time high resolution satellite data from current EOS and planned NPP, JPSS, and GOES-R sensors to weather forecast problems, significant challenges arise in the acquisition, delivery, and integration of the new capabilities into the decision making process of the operational weather community. For polar orbiting sensors such as MODIS, AIRS, VIIRS, and CRiS, the use of direct broadcast ground stations is key to the real-time delivery of the data and derived products in a timely fashion. With the ABI on the geostationary GOES-R satellite, the data volume will likely increase by a factor of 5- 10 from current data streams. However, the high data volume and limited bandwidth of end user facilities presents a formidable obstacle to timely access to the data. This challenge can be addressed through the use of subsetting techniques, innovative web services, and the judicious selection of data formats. Many of these approaches have been implemented by SPoRT for the delivery of real-time products to NWS forecast offices and other weather entities. Once available in decision support systems like AWIPS II, these new data and products must be integrated into existing and new displays that allow for the integration of the data with existing operational products in these systems. SPoRT is leading the way in demonstrating this enhanced capability. This paper will highlight the ways SPoRT is overcoming many of the challenges presented by the enormous data volumes of

Clinical application of RT-nested PCR integrated with RFLP in Hantavirus detection and genotyping: a prospective study in Shandong Province, PR China.

Science.gov (United States)

Liu, Yun-Xi; Zhao, Zhong-Tang; Cao, Wu-Chun; Xu, Xiao-Qun; Suo, Ji-Jiang; Xing, Yu-Bin; Jia, Ning; Du, Ming-Mei; Liu, Bo-Wei; Yao, Yuan

2013-01-01

The aim of the present study was to evaluate the clinical usefulness of applying RT-nested PCR along with RFLP as a method for diagnosis and genotypic differentiation of Hantavirus in the acute-stage sera of HFRS patients as compared to the ELISA technique. A prospective study of patients with suspected HFRS patients was carried out. Sera were collected for serological evaluation by ELISA and RT-nested PCR testing. Primers were selected from the published sequence of the S segment of HTNV strain 76-118 and SEOV strain SR-11, which made it possible to obtain an amplicon of 403 bp by RT-nested PCR. The genotypic differentiations of the RT-nested PCR amplicons were carried out by RFLP. Sequence analyses of the amplicons were used to confirm the accuracy of the results obtained by RFLP. Of the 48 acute-stage sera from suspected HFRS patients, 35 were ELISA-positive while 41 were positive by RT-nested PCR. With Hind III and Hinf I, RFLP profiles of the RT-nested PCR amplicons of the 41 positive sera exhibited two patterns. 33 had RFLP profiles similar to the reference strain R22, and thus belonged to the SEOV type. The other 8 samples which were collected during October-December had RFLP profiles similar to the reference strain 76-118, and thus belonged to the HTNV type. Sequence phylogenetic analysis of RT-nested PCR amplicons revealed sdp1, sdp2 YXL-2008, and sdp3 as close relatives of HTNV strain 76-118, while sdp22 and sdp37 as close relatives of SEOV strain Z37 and strain R22 located in two separate clusters in the phylogenetic tree. These results were identical to those acquired by RFLP. RT-nested PCR integrated with RFLP was a rapid, simple, accurate method for detecting and differentiating the genotypes of Hantavirus in the acute-stage sera of suspected HFRS patients. In Shandong province, the main genotypes of Hantavirus belonged to the SEOV types, while the HTNV types were observed during the autumn-winter season.

Analysis of gene expression in small numbers of purified hemopoietic progenitor cells by RT-PCR.

Science.gov (United States)

Ziegler, B L; Lamping, C P; Thoma, S J; Fliedner, T M

1995-05-01

Primitive hemopoietic stem cells represent the most probable targets for genetic alterations due to exposure to ionizing irradiation or chemical carcinogens. We have applied a two-step protocol for the purification of CD34+HLA-DR-/low hemopoietic progenitor cells from cord blood (CB). CD34+ cells were isolated by monoclonal antibody (mAb) against CD34 (My10) and immunomagnetic beads. Beads were cleaved off the CD34+ cells by enzymatic treatment with chymopapain. Due to chymopapain-resistance of epitopes recognized by the used mAbs purity control of CD34+ cells and separation into CD34+HLA-DR-/low and CD34+HLA-DR+ subsets could be performed by using flow cytometry. Two miniaturized procedures were applied to isolate poly(A)+ mRNA for the reverse transcription polymerase chain reaction (RT-PCR) from small numbers of CD34+HLA-DR-/low cells. In five experiments, the mean purity of immunomagnetically isolated CD34+ cells was 93.8% +/- 3.9. Flow cytometry sorting of CD34+ cells resulted in pure CD34+HLA-DR-/low populations (purity > 98.8%; range 98.8% to 99.9%; viability > 96%) with an average yield of 2600 +/- 800 cells/5 x 10(7) low density CB cells. By RT-PCR using both poly(A)+ mRNA isolation procedures, sequences corresponding to CD34 and beta 2-microglobulin were amplified from as few as 20 cells. Furthermore, a sequence-independent RT-PCR (SIP-RT-PCR) was applied to amplify the cDNA derived from five erythroblasts isolated from a burst-forming unit-erythroid (BFU-E). Upon hybridization, full-length c-fos message was detected in the SIP-RT-PCR amplified material. Our data demonstrate that gene expression can be detected at the transcriptional level in small numbers of hemopoietic progenitor cells. In addition, the SIP-RT-PCR may allow the amplification of unique mRNA species when subtractive hybridization procedures are performed. The presented data should be useful to analyze gene expression in rare subsets of radiation-exposed immature hemopoietic stem

Sequence and RT-PCR expression analysis of two peroxidases from Arabidopsis thaliana belonging to a novel evolutionary branch of plant peroxidases.

Science.gov (United States)

Kjaersgård, I V; Jespersen, H M; Rasmussen, S K; Welinder, K G

1997-03-01

cDNA clones encoding two new Arabidopsis thaliana peroxidases, ATP 1a and ATP 2a, have been identified by searching the Arabidopsis database of expressed sequence tags (dbEST). They represent a novel branch of hitherto uncharacterized plant peroxidases which is only 35% identical in amino acid sequence to the well characterized group of basic plant peroxidases represented by the horseradish (Armoracia rusticana) isoperoxidases HRP C, HRP E5 and the similar Arabidopsis isoperoxidases ATP Ca, ATP Cb, and ATP Ea. However ATP 1a is 87% identical in amino acid sequence to a peroxidase encoded by an mRNA isolated from cotton (Gossypium hirsutum). As cotton and Arabidopsis belong to rather diverse families (Malvaceae and Crucifereae, respectively), in contrast with Arabidopsis and horseradish (both Crucifereae), the high degree of sequence identity indicates that this novel type of peroxidase, albeit of unknown function, is likely to be widespread in plant species. The atp 1 and atp 2 types of cDNA sequences were the most redundant among the 28 different isoperoxidases identified among about 200 peroxidase encoding ESTs. Interestingly, 8 out of totally 38 EST sequences coding for ATP 1 showed three identical nucleotide substitutions. This variant form is designated ATP 1b. Similarly, six out of totally 16 EST sequences coding for ATP 2 showed a number of deletions and nucleotide changes. This variant form is designated ATP 2b. The selected EST clones are full-length and contain coding regions of 993 nucleotides for atp 1a, and 984 nucleotides for atp 2a. These regions show 61% DNA sequence identity. The predicted mature proteins ATP 1a, and ATP 2a are 57% identical in sequence and contain the structurally and functionally important residues, characteristic of the plant peroxidase superfamily. However, they do show two differences of importance to peroxidase catalysis: (1) the asparagine residue linked with the active site distal histidine via hydrogen bonding is absent

Identification and characterization of a novel tospovirus species using a new RT-PCR approach

NARCIS (Netherlands)

Cortez, I.; Saaijer, J.; Wonjkaew, K.S.; Pereira, A.M.; Goldbach, R.W.; Peters, D.; Kormelink, R.

2001-01-01

Summary. A novel tospovirus serologically distinct from all established tospo- virus species was found in Thailand in Physalis minima L. The S RNA of this virus was cloned by a new RT-PCR approach revealing a nucleotide sequence of 3257 nucleotides. The ambisense RNA segment encoded a nonstructural

Assessing the 5S ribosomal RNA heterogeneity in Arabidopsis thaliana using short RNA next generation sequencing data.

Science.gov (United States)

Szymanski, Maciej; Karlowski, Wojciech M

2016-01-01

In eukaryotes, ribosomal 5S rRNAs are products of multigene families organized within clusters of tandemly repeated units. Accumulation of genomic data obtained from a variety of organisms demonstrated that the potential 5S rRNA coding sequences show a large number of variants, often incompatible with folding into a correct secondary structure. Here, we present results of an analysis of a large set of short RNA sequences generated by the next generation sequencing techniques, to address the problem of heterogeneity of the 5S rRNA transcripts in Arabidopsis and identification of potentially functional rRNA-derived fragments.

Real-Time RT-PCR for the Detection of Lyssavirus Species

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A. Deubelbeiss

2014-01-01

Full Text Available The causative agents of rabies are single-stranded, negative-sense RNA viruses in the genus Lyssavirus of Rhabdoviridae, consisting of twelve classified and three as yet unclassified species including classical rabies virus (RABV. Highly neurotropic RABV causes rapidly progressive encephalomyelitis with nearly invariable fatal outcome. Rapid and reliable diagnosis of rabies is highly relevant for public and veterinary health. Due to growing variety of the genus Lyssavirus observed, the development of suitable molecular assays for diagnosis and differentiation is challenging. This work focused on the establishment of a suitable real-time RT-PCR technique for rabies diagnosis as a complement to fluorescent antibody test and rabies tissue culture infection test as gold standard for diagnosis and confirmation. The real-time RT-PCR was adapted with the goal to detect the whole spectrum of lyssavirus species, for nine of which synthesized DNA fragments were used. For the detection of species, seven probes were developed. Serial dilutions of the rabies virus strain CVS-11 showed a 100-fold higher sensitivity of real-time PCR compared to heminested RT-PCR. Using a panel of thirty-one lyssaviruses representing four species, the suitability of the protocol could be shown. Phylogenetic analysis of the sequences obtained by heminested PCR allowed correct classification of all viruses used.

Detection of avian metapneumovirus subtypes in turkeys using RT-PCR.

Science.gov (United States)

Ongor, H; Karahan, M; Kalin, R; Bulut, H; Cetinkaya, B

2010-03-20

This study investigated the prevalence of avian metapneumovirus (aMPV) and the detection of molecular subtypes of field strains of the virus using RT-PCR in clinically healthy turkeys and those showing signs of respiratory disease. In the RT-PCR examination of 624 tracheal tissue samples collected from a local turkey abattoir, 2.9 per cent (18/624) of samples tested positive. In the examination of tracheal swab samples collected from flocks with respiratory problems, 18 of 20 samples tested positive. When the results were assessed at flock level, aMPV infection was detected in only one of the 23 clinically healthy turkey flocks, whereas all four flocks with respiratory problems were infected. Molecular typing using primers specific to the attachment glycoprotein (G) gene showed that all 36 positive samples belonged to subtype B. Partial sequence analysis of DNA samples showed 95 per cent homology between the field types and the reference strain aMPV subtype B. Whereas clinically healthy turkeys had been vaccinated with a subtype A virus vaccine, the flocks with respiratory problems had been vaccinated with a subtype B virus vaccine. Despite four blind passages of RT-PCR-positive samples on Vero and chicken embryo fibroblast cells, no cytopathic effect was detected by microscopic examination.

Identification of rhabdoviral sequences in oropharyngeal swabs from German and Danish bats.

Science.gov (United States)

Fischer, Melina; Freuling, Conrad M; Müller, Thomas; Schatz, Juliane; Rasmussen, Thomas Bruun; Chriel, Mariann; Balkema-Buschmann, Anne; Beer, Martin; Hoffmann, Bernd

2014-11-25

In the frame of active lyssavirus surveillance in bats, oropharyngeal swabs from German (N = 2297) and Danish (N = 134) insectivorous bats were investigated using a newly developed generic pan-lyssavirus real-time reverse transcriptase PCR (RT-qPCR). In total, 15 RT-qPCR positive swabs were detected. Remarkably, sequencing of positive samples did not confirm the presence of bat associated lyssaviruses but revealed nine distinct novel rhabdovirus-related sequences. Several novel rhabdovirus-related sequences were detected both in German and Danish insectivorous bats. The results also prove that the novel generic pan-lyssavirus RT-qPCR offers a very broad detection range that allows the collection of further valuable data concerning the broad and complex diversity within the family Rhabdoviridae.

A novel and highly sensitive real-time nested RT-PCR assay in a single closed tube for detection of enterovirus.

Science.gov (United States)

Shen, Xin-Xin; Qiu, Fang-Zhou; Zhao, Huai-Long; Yang, Meng-Jie; Hong, Liu; Xu, Song-Tao; Zhou, Shuai-Feng; Li, Gui-Xia; Feng, Zhi-Shan; Ma, Xue-Jun

2018-03-01

The sensitivity of qRT-PCR assay is not adequate for the detection of the samples with lower viral load, particularly in the cerebrospinal fluid (CSF) of patients. Here, we present the development of a highly sensitive real-time nested RT-PCR (RTN RT-PCR) assay in a single closed tube for detection of human enterovirus (HEV). The clinical performance of both RTN RT-PCR and qRT-PCR was also tested and compared using 140 CSF and fecal specimens. The sensitivities of RTN RT-PCR assay for EV71, Coxsackievirus A (CVA)16, CVA6 and CVA10 achieved 10 -8 dilution with a corresponding Ct value of 38.20, 36.45, 36.75, and 36.45, respectively, which is equal to traditional two-step nested RT-PCR assay and approximately 2-10-fold lower than that of qRT-PCR assay. The specificity of RTN RT-PCR assay was extensively analyzed insilico and subsequently verified using the reference isolates and clinical samples. Sixteen qRT-PCR-negative samples were detected by RTN RT-PCR and a variety of enterovirus serotypes was identified by sequencing of inner PCR products. We conclude RTN RT-PCR is more sensitive than qRT-PCR for the detection of HEV in clinical samples. Copyright © 2017 Elsevier Inc. All rights reserved.

An Endogenous Murine Leukemia Viral Genome Contaminant in a Commercial RT-PCR Kit is Amplified Using Standard Primers for XMRV

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Miyazawa Takayuki

2010-12-01

Full Text Available Abstract During pilot studies to investigate the presence of viral RNA of xenotropic murine leukemia virus (MLV-related virus (XMRV infection in sera from chronic fatigue syndrome (CFS patients in Japan, a positive band was frequently detected at the expected product size in negative control samples when detecting a partial gag region of XMRV using a one-step RT-PCR kit. We suspected that the kit itself might have been contaminated with small traces of endogenous MLV genome or XMRV and attempted to evaluate the quality of the kit in two independent laboratories. We purchased four one-step RT-PCR kits from Invitrogen, TaKaRa, Promega and QIAGEN in Japan. To amplify the partial gag gene of XMRV or other MLV-related viruses, primer sets (419F and 1154R, and GAG-I-F and GAG-I-R which have been widely used in XMRV studies were employed. The nucleotide sequences of the amplicons were determined and compared with deposited sequences of a polytropic endogenous MLV (PmERV, XMRV and endogenous MLV-related viruses derived from CFS patients. We found that the enzyme mixtures of the one-step RT-PCR kit from Invitrogen were contaminated with RNA derived from PmERV. The nucleotide sequence of a partial gag region of the contaminant amplified by RT-PCR was nearly identical (99.4% identity to a PmERV on chromosome 7 and highly similar (96.9 to 97.6% to recently identified MLV-like viruses derived from CFS patients. We also determined the nucleotide sequence of a partial env region of the contaminant and found that it was almost identical (99.6% to the PmERV. In the investigation of XMRV infection in patients of CFS and prostate cancer, researchers should prudently evaluate the test kits for the presence of endogenous MLV as well as XMRV genomes prior to PCR and RT-PCR tests.

Effects of High Intensity White Noise on Short-Term Memory for Position in a List and Sequence

Science.gov (United States)

Daee, Safar; Wilding, J. M.

1977-01-01

Seven experiments are described investigating the effecy of high intensity white noise during the visual presentation of words on a number of short-term memory tasks. Examines results relative to position learning and sequence learning. (Editor/RK)

Short Interspersed Nuclear Element (SINE Sequences in the Genome of the Human Pathogenic Fungus Aspergillus fumigatus Af293.

Directory of Open Access Journals (Sweden)

Lakkhana Kanhayuwa

Full Text Available Novel families of short interspersed nuclear element (SINE sequences in the human pathogenic fungus Aspergillus fumigatus, clinical isolate Af293, were identified and categorised into tRNA-related and 5S rRNA-related SINEs. Eight predicted tRNA-related SINE families originating from different tRNAs, and nominated as AfuSINE2 sequences, contained target site duplications of short direct repeat sequences (4-14 bp flanking the elements, an extended tRNA-unrelated region and typical features of RNA polymerase III promoter sequences. The elements ranged in size from 140-493 bp and were present in low copy number in the genome and five out of eight were actively transcribed. One putative tRNAArg-derived sequence, AfuSINE2-1a possessed a unique feature of repeated trinucleotide ACT residues at its 3'-terminus. This element was similar in sequence to the I-4_AO element found in A. oryzae and an I-1_AF long nuclear interspersed element-like sequence identified in A. fumigatus Af293. Families of 5S rRNA-related SINE sequences, nominated as AfuSINE3, were also identified and their 5'-5S rRNA-related regions show 50-65% and 60-75% similarity to respectively A. fumigatus 5S rRNAs and SINE3-1_AO found in A. oryzae. A. fumigatus Af293 contains five copies of AfuSINE3 sequences ranging in size from 259-343 bp and two out of five AfuSINE3 sequences were actively transcribed. Investigations on AfuSINE distribution in the fungal genome revealed that the elements are enriched in pericentromeric and subtelomeric regions and inserted within gene-rich regions. We also demonstrated that some, but not all, AfuSINE sequences are targeted by host RNA silencing mechanisms. Finally, we demonstrated that infection of the fungus with mycoviruses had no apparent effects on SINE activity.

Isolation, sequence identification and tissue expression profile of a ...

African Journals Online (AJOL)

The complete expressed sequence tag (CDS) sequence of Banna mini-pig inbred line (BMI) ribokinase gene (RBKS) was amplified using the reverse transcription-polymerase chain reaction (RT-PCR) based on the conserved sequence information of the cattle or other mammals and known highly homologous swine ESTs.

Request for regular monitoring of the symbiotic variable RT Cru

Science.gov (United States)

Waagen, Elizabeth O.

2014-08-01

Dr. Margarita Karovska (Harvard-Smithsonian Center for Astrophysics) and colleagues have requested AAVSO observer assistance in their campaign on the symbiotic variable RT Cru (member of a new class of hard X-ray emitting symbiotic binaries). Weekly or more frequent monitoring (B, V, and visual) beginning now is requested in support of upcoming Chandra observations still to be scheduled. "We plan Chandra observations of RT Cru in the near future that will help us understand the characteristics of the accretion onto the white dwarf in this sub-class of symbiotics. This is an important step for determining the precursor conditions for formation of a fraction of asymmetric Planetary Nebulae, and the potential of symbiotic systems as progenitors of at least a fraction of Type Ia supernovae." Finder charts with sequence may be created using the AAVSO Variable Star Plotter (http://www.aavso.org/vsp). Observations should be submitted to the AAVSO International Database. See full Alert Notice for more details and observations.

JPSS Proving Ground Activities with NASA's Short-term Prediction Research and Transition (SPoRT) Center

Science.gov (United States)

Schultz, L. A.; Smith, M. R.; Fuell, K.; Stano, G. T.; LeRoy, A.; Berndt, E.

2015-12-01

Instruments aboard the Joint Polar Satellite System (JPSS) series of satellites will provide imagery and other data sets relevant to operational weather forecasts. To prepare current and future weather forecasters in application of these data sets, Proving Ground activities have been established that demonstrate future JPSS capabilities through use of similar sensors aboard NASA's Terra and Aqua satellites, and the S-NPP mission. As part of these efforts, NASA's Short-term Prediction Research and Transition (SPoRT) Center in Huntsville, Alabama partners with near real-time providers of S-NPP products (e.g., NASA, UW/CIMSS, UAF/GINA, etc.) to demonstrate future capabilities of JPSS. This includes training materials and product distribution of multi-spectral false color composites of the visible, near-infrared, and infrared bands of MODIS and VIIRS. These are designed to highlight phenomena of interest to help forecasters digest the multispectral data provided by the VIIRS sensor. In addition, forecasters have been trained on the use of the VIIRS day-night band, which provides imagery of moonlit clouds, surface, and lights emitted by human activities. Hyperspectral information from the S-NPP/CrIS instrument provides thermodynamic profiles that aid in the detection of extremely cold air aloft, helping to map specific aviation hazards at high latitudes. Hyperspectral data also support the estimation of ozone concentration, which can highlight the presence of much drier stratospheric air, and map its interaction with mid-latitude or tropical cyclones to improve predictions of their strengthening or decay. Proving Ground activities are reviewed, including training materials and methods that have been provided to forecasters, and forecaster feedback on these products that has been acquired through formal, detailed assessment of their applicability to a given forecast threat or task. Future opportunities for collaborations around the delivery of training are proposed

Node Ranking Tool - NoRT

Science.gov (United States)

2018-03-23

NoRT) that was developed as part of the Applied Network Science 6.2 base program work unit at NRL, Code 5580. We explain the theory of NoRT and how...to use it. 23-03-2018 Memorandum Report TOPSIS, Social Network, Sensor Network, Centrality, Diffusion, Disease, Virus, Expectation , Pandemic...base program work unit at NRL. We explain the theory of NoRT and how to use it. Index Terms TOPSIS, Social Network, Sensor Network, Centrality, Diffusion

Genome dynamics of short oligonucleotides: the example of bacterial DNA uptake enhancing sequences.

Directory of Open Access Journals (Sweden)

Mohammed Bakkali

Full Text Available Among the many bacteria naturally competent for transformation by DNA uptake-a phenomenon with significant clinical and financial implications- Pasteurellaceae and Neisseriaceae species preferentially take up DNA containing specific short sequences. The genomic overrepresentation of these DNA uptake enhancing sequences (DUES causes preferential uptake of conspecific DNA, but the function(s behind this overrepresentation and its evolution are still a matter for discovery. Here I analyze DUES genome dynamics and evolution and test the validity of the results to other selectively constrained oligonucleotides. I use statistical methods and computer simulations to examine DUESs accumulation in Haemophilus influenzae and Neisseria gonorrhoeae genomes. I analyze DUESs sequence and nucleotide frequencies, as well as those of all their mismatched forms, and prove the dependence of DUESs genomic overrepresentation on their preferential uptake by quantifying and correlating both characteristics. I then argue that mutation, uptake bias, and weak selection against DUESs in less constrained parts of the genome combined are sufficient enough to cause DUESs accumulation in susceptible parts of the genome with no need for other DUES function. The distribution of overrepresentation values across sequences with different mismatch loads compared to the DUES suggests a gradual yet not linear molecular drive of DNA sequences depending on their similarity to the DUES. Other genomically overrepresented sequences, both pro- and eukaryotic, show similar distribution of frequencies suggesting that the molecular drive reported above applies to other frequent oligonucleotides. Rare oligonucleotides, however, seem to be gradually drawn to genomic underrepresentation, thus, suggesting a molecular drag. To my knowledge this work provides the first clear evidence of the gradual evolution of selectively constrained oligonucleotides, including repeated, palindromic and protein

Detection and identification of dengue virus isolates from Brazil by a simplified reverse transcription - polymerase chain reaction (RT-PCR method

Directory of Open Access Journals (Sweden)

FIGUEIREDO Luiz Tadeu Moraes

1997-01-01

Full Text Available We show here a simplified RT-PCR for identification of dengue virus types 1 and 2. Five dengue virus strains, isolated from Brazilian patients, and yellow fever vaccine 17DD as a negative control, were used in this study. C6/36 cells were infected and supernatants were collected after 7 days. The RT-PCR, done in a single reaction vessel, was carried out following a 1/10 dilution of virus in distilled water or in a detergent mixture containing Nonidet P40. The 50 µl assay reaction mixture included 50 pmol of specific primers amplifying a 482 base pair sequence for dengue type 1 and 210 base pair sequence for dengue type 2. In other assays, we used dengue virus consensus primers having maximum sequence similarity to the four serotypes, amplifying a 511 base pair sequence. The reaction mixture also contained 0.1 mM of the four deoxynucleoside triphosphates, 7.5 U of reverse transcriptase, 1U of thermostable Taq DNA polymerase. The mixture was incubated for 5 minutes at 37ºC for reverse transcription followed by 30 cycles of two-step PCR amplification (92ºC for 60 seconds, 53ºC for 60 seconds with slow temperature increment. The PCR products were subjected to 1.7% agarose gel electrophoresis and visualized by UV light after staining with ethidium bromide solution. Low virus titer around 10 3, 6 TCID50/ml was detected by RT-PCR for dengue type 1. Specific DNA amplification was observed with all the Brazilian dengue strains by using dengue virus consensus primers. As compared to other RT-PCRs, this assay is less laborious, done in a shorter time, and has reduced risk of contamination

Performance of a RT-PCR Assay in Comparison to FISH and Immunohistochemistry for the Detection of ALK in Non-Small Cell Lung Cancer.

Science.gov (United States)

Hout, David R; Schweitzer, Brock L; Lawrence, Kasey; Morris, Stephan W; Tucker, Tracy; Mazzola, Rosetta; Skelton, Rachel; McMahon, Frank; Handshoe, John; Lesperance, Mary; Karsan, Aly; Saltman, David L

2017-08-01

Patients with lung cancers harboring an activating anaplastic lymphoma kinase ( ALK ) rearrangement respond favorably to ALK inhibitor therapy. Fluorescence in situ hybridization (FISH) and immunohistochemistry (IHC) are validated and widely used screening tests for ALK rearrangements but both methods have limitations. The ALK RGQ RT-PCR Kit (RT-PCR) is a single tube quantitative real-time PCR assay for high throughput and automated interpretation of ALK expression. In this study, we performed a direct comparison of formalin-fixed paraffin-embedded (FFPE) lung cancer specimens using all three ALK detection methods. The RT-PCR test (diagnostic cut-off Δ C t of ≤8) was shown to be highly sensitive (100%) when compared to FISH and IHC. Sequencing of RNA detected full-length ALK transcripts or EML4-ALK and KIF5B-ALK fusion variants in discordant cases in which ALK expression was detected by the ALK RT-PCR test but negative by FISH and IHC. The overall specificity of the RT-PCR test for the detection of ALK in cases without full-length ALK expression was 94% in comparison to FISH and sequencing. These data support the ALK RT-PCR test as a highly efficient and reliable diagnostic screening approach to identify patients with non-small cell lung cancer whose tumors are driven by oncogenic ALK.

A one-step multiplex RT-PCR assay for simultaneous detection of four viruses that infect peach.

Science.gov (United States)

Yu, Y; Zhao, Z; Jiang, D; Wu, Z; Li, S

2013-10-01

A multiplex reverse transcription polymerase chain reaction (mRT-PCR) assay was developed to enable the simultaneous detection and differentiation of four viruses that infect peach, namely Apple chlorotic leaf spot virus (ACLSV), Cherry green ring mottle virus (CGRMV), Prunus necrotic ringspot virus (PNRSV) and Apricot pseudo-chlorotic leaf spot virus (APCLSV). In this study, four pairs of primers, one specific for each virus, were designed; the corresponding PCR products were 632, 439, 346 and 282 bp in length for ACLSV, CGRMV, PNRSV and APCLSV, respectively, and the fragments could be distinguished clearly by agarose gel electrophoresis. The sensitivity and specificity of the method were tested using individual RT-PCR and enzyme-linked immunosorbent assay (ELISA), and the identity of the RT-PCR amplification products was also confirmed by DNA sequencing. The results of RT-PCR and ELISA, along with batch detection using samples collected from peach orchards, revealed that this rapid and simple technique is an effective way to identify the four viruses simultaneously. The mRT-PCR assay described in this study was developed for the simultaneous detection of four peach viruses from infected peach samples is reliable and sensitive. In contrast to conventional uniplex RT-PCR, mRT-PCR is more efficient, reducing costs, time and handling when testing large numbers of samples. This rapid and simple method is useful for large-scale surveys of viruses that infect peach. © 2013 The Society for Applied Microbiology.

Next-generation small RNA sequencing for microRNAs profiling in the honey bee Apis mellifera.

Science.gov (United States)

Chen, X; Yu, X; Cai, Y; Zheng, H; Yu, D; Liu, G; Zhou, Q; Hu, S; Hu, F

2010-12-01

MicroRNAs (miRNAs) are key regulators in various physiological and pathological processes via post-transcriptional regulation of gene expression. The honey bee (Apis mellifera) is a key model for highly social species, and its complex social behaviour can be interpreted theoretically as changes in gene regulation, in which miRNAs are thought to be involved. We used the SOLiD sequencing system to identify the repertoire of miRNAs in the honey bee by sequencing a mixed small RNA library from different developmental stages. We obtained a total of 36,796,459 raw sequences; of which 5,491,100 short sequences were fragments of mRNA and other noncoding RNAs (ncRNA), and 1,759,346 reads mapped to the known miRNAs. We predicted 267 novel honey bee miRNAs representing 380,182 short reads, including eight miRNAs of other insects in 14,107,583 genome-mapped sequences. We verified 50 of them using stem-loop reverse-transcription PCR (RT-PCR), in which 35 yielded PCR products. Cross-species analyses showed 81 novel miRNAs with homologues in other insects, suggesting that they were authentic miRNAs and have similar functions. The results of this study provide a basis for studies of the miRNA-modulating networks in development and some intriguing phenomena such as caste differentiation in A. mellifera. © 2010 The Authors. Insect Molecular Biology © 2010 The Royal Entomological Society.

Screening of SHOX gene sequence variants in Saudi Arabian children with idiopathic short stature.

Science.gov (United States)

Alharthi, Abdulla A; El-Hallous, Ehab I; Talaat, Iman M; Alghamdi, Hamed A; Almalki, Matar I; Gaber, Ahmed

2017-10-01

Short stature affects approximately 2%-3% of children, representing one of the most frequent disorders for which clinical attention is sought during childhood. Despite assumed genetic heterogeneity, mutations or deletions in the short stature homeobox-containing gene ( SHOX ) are frequently detected in subjects with short stature. Idiopathic short stature (ISS) refers to patients with short stature for various unknown reasons. The goal of this study was to screen all the exons of SHOX to identify related mutations. We screened all the exons of SHOX for mutations analysis in 105 ISS children patients (57 girls and 48 boys) living in Taif governorate, KSA using a direct DNA sequencing method. Height, arm span, and sitting height were recorded, and subischial leg length was calculated. A total of 30 of 105 ISS patients (28%) contained six polymorphic variants in exons 1, 2, 4, and 6. One mutation was found in the DNA domain binding region of exon 4. Three of these polymorphic variants were novel, while the others were reported previously. There were no significant differences in anthropometric measures in ISS patients with and without identifiable polymorphic variants in SHOX . In Saudi Arabia ISS patients, rather than SHOX , it is possible that new genes are involved in longitudinal growth. Additional molecular analysis is required to diagnose and understand the etiology of this disease.

Transitioning NPOESS Data to Weather Offices: The SPoRT Paradigm with EOS Data

Science.gov (United States)

Jedlovec, Gary

2009-01-01

Real-time satellite information provides one of many data sources used by NWS weather forecast offices (WFOs) to diagnose current weather conditions and to assist in short-term forecast preparation. While GOES satellite data provides relatively coarse spatial resolution coverage of the continental U.S. on a 10-15 minute repeat cycle, polar orbiting imagery has the potential to provide snapshots of weather conditions at high-resolution in many spectral channels. Additionally, polar orbiting sounding data can provide additional information on the thermodynamic structure of the atmosphere in data sparse regions of at asynoptic observation times. The NASA Short-term Prediction Research and Transition (SPoRT) project has demonstrated the utility of polar orbiting MODIS and AIRS data on the Terra and Aqua satellites to improve weather diagnostics and short-term forecasting on the regional and local scales. SPoRT scientists work directly forecasters at selected WFOS in the Southern Region (SR) to help them ingest these unique data streams into their AWIPS system, understand how to use the data (through on-site and distance learn techniques), and demonstrate the utility of these products to address significant forecast problems. This process also prepares forecasters for the use of similar observational capabilities from NPOESS operational sensors. NPOESS environmental data records (EDRs) from the Visible 1 Infrared Imager I Radiometer Suite (VIIRS), the Cross-track Infrared Sounder (CrlS) and Advanced Technology Microwave Sounder (ATMS) instruments and additional value-added products produced by NESDIS will be available in near real-time and made available to WFOs to extend their use of NASA EOS data into the NPOESS era. These new data streams will be integrated into the NWs's new AWIPS II decision support tools. The AWIPS I1 system to be unveiled in WFOs in 2009 will be a JAVA-based decision support system which preserves the functionality of the existing systems and

Isolation and amino acid sequence of a short-chain neurotoxin from an Australian elapid snake, Pseudechis australis.

OpenAIRE

Takasaki, C; Tamiya, N

1985-01-01

A short-chain neurotoxin Pseudechis australis a (toxin Pa a) was isolated from the venom of an Australian elapid snake Pseudechis australis (king brown snake) by sequential chromatography on CM-cellulose, Sephadex G-50 and CM-cellulose columns. Toxin Pa a has an LD50 (intravenous) value of 76 micrograms/kg body wt. in mice and consists of 62 amino acid residues. The amino acid sequence of Pa a shows considerable homology with those of short-chain neurotoxins of elapid snakes, especially of tr...

MZC Gel Inhibits SHIV-RT and HSV-2 in Macaque Vaginal Mucosa and SHIV-RT in Rectal Mucosa.

Science.gov (United States)

Calenda, Giulia; Villegas, Guillermo; Barnable, Patrick; Litterst, Claudia; Levendosky, Keith; Gettie, Agegnehu; Cooney, Michael L; Blanchard, James; Fernández-Romero, José A; Zydowsky, Thomas M; Teleshova, Natalia

2017-03-01

The Population Council's microbicide gel MZC (also known as PC-1005) containing MIV-150 and zinc acetate dihydrate (ZA) in carrageenan (CG) has shown promise as a broad-spectrum microbicide against HIV, herpes simplex virus (HSV), and human papillomavirus. Previous data show antiviral activity against these viruses in cell-based assays, prevention of vaginal and rectal simian-human immunodeficiency virus reverse transcriptase (SHIV-RT) infection, and reduction of vaginal HSV shedding in rhesus macaques and also excellent antiviral activity against HSV and human papillomavirus in murine models. Recently, we demonstrated that MZC is safe and effective against SHIV-RT in macaque vaginal explants. Here we established models of ex vivo SHIV-RT/HSV-2 coinfection of vaginal mucosa and SHIV-RT infection of rectal mucosa in macaques (challenge of rectal mucosa with HSV-2 did not result in reproducible tissue infection), evaluated antiviral activity of MZC, and compared quantitative polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay readouts for monitoring SHIV-RT infection. MZC (at nontoxic dilutions) significantly inhibited SHIV-RT in vaginal and rectal mucosas and HSV-2 in vaginal mucosa when present during viral challenge. Analysis of SHIV-RT infection and MZC activity by 1-step simian immunodeficiency virus gag quantitative RT-PCR and p27 enzyme-linked immunosorbent assay demonstrated similar virus growth dynamics and MZC activity by both methods and higher sensitivity of quantitative RT-PCR. Our data provide more evidence that MZC is a promising dual compartment multipurpose prevention technology candidate.

Evaluating the Impact of AIRS Observations on Regional Forecasts at the SPoRT Center

Science.gov (United States)

Zavodsky, Bradley

2011-01-01

NASA Short-term Prediction Research and Transition (SPoRT) Center collaborates with operational partners of different sizes and operational goals to improve forecasts using targeted projects and data sets. Modeling and DA activities focus on demonstrating utility of NASA data sets and capabilities within operational systems. SPoRT has successfully assimilated the Atmospheric Infrared Sounder (AIRS) radiance and profile data. A collaborative project is underway with the Joint Center for Satellite Data Assimilation (JCSDA) to use AIRS profiles to better understand the impact of AIRS radiances assimilated within Gridpoint Statistical Interpolation (GSI) in hopes of engaging the operational DA community in a reassessment of assimilation methodologies to more effectively assimilate hyperspectral radiances.

SPoRT Participation in the GOES-R and JPSS Proving Grounds

Science.gov (United States)

Jedlovec, Gary; Fuell, Kevin; Smith, Matthew

2013-01-01

For the last several years, the NASA Short-term Prediction Research and Transition (SPoRT) project at has been working with the various algorithm working groups and science teams to demonstrate the utility of future operational sensors for GOES-R and the suite of instruments for the JPSS observing platforms. For GOES-R, imagery and products have been developed from polar-orbiting sensors such as MODIS and geostationary observations from SEVIRI, simulated imagery, enhanced products derived from existing GOES satellites, and data from ground-based observing systems to generate pseudo or proxy products for the ABI and GLM instruments. The suite of products include GOES-POES basic and RGB hybrid imagery, total lightning flash products, quantitative precipitation estimates, and convective initiation products. SPoRT is using imagery and products from VIIRS, CrIS, ATMS, and OMPS to show the utility of data and products from their operational counterparts on JPSS. The products include VIIRS imagery in swath form, the GOES-POES hybrid, a suite of RGB products including the air mass RGB using water vapor and ozone channels from CrIS, and several DNB products. Over a dozen SPoRT collaborative WFOs and several National Centers are involved in an intensive evaluation of the operational utility of these products.

Reverse transcriptase sequences from mulberry LTR retrotransposons: characterization analysis

Directory of Open Access Journals (Sweden)

Ma Bi

2017-10-01

Full Text Available Copia and Gypsy play important roles in structural, functional and evolutionary dynamics of plant genomes. In this study, a total of 106 and 101, Copia and Gypsy reverse transcriptase (rt were amplified respectively in the Morus notabilis genome using degenerate primers. All sequences exhibited high levels of heterogeneity, were rich in AT and possessed higher sequence divergence of Copia rt in comparison to Gypsy rt. Two reasons are likely to account for this phenomenon: a these elements often experience deletions or fragmentation by illegitimate or unequal homologous recombination in the transposition process; b strong purifying selective pressure drives the evolution of these elements through “selective silencing” with random mutation and eventual deletion from the host genome. Interestingly, mulberry rt clustered with other rt from distantly related taxa according to the phylogenetic analysis. This phenomenon did not result from horizontal transposable element transfer. Results obtained from fluorescence in situ hybridization revealed that most of the hybridization signals were preferentially concentrated in pericentromeric and distal regions of chromosomes, and these elements may play important roles in the regions in which they are found. Results of this study support the continued pursuit of further functional studies of Copia and Gypsy in the mulberry genome.

SPoRT: Transitioning NASA and NOAA Experimental Data to the Operational Weather Community

Science.gov (United States)

Jedlovec, Gary J.

2013-01-01

Established in 2002 to demonstrate the weather and forecasting application of real-time EOS measurements, the NASA Short-term Prediction Research and Transition (SPoRT) program has grown to be an end-to-end research to operations activity focused on the use of advanced NASA modeling and data assimilation approaches, nowcasting techniques, and unique high-resolution multispectral data from EOS satellites to improve short-term weather forecasts on a regional and local scale. With the ever-broadening application of real-time high resolution satellite data from current EOS, Suomi NPP, and planned JPSS and GOES-R sensors to weather forecast problems, significant challenges arise in the acquisition, delivery, and integration of the new capabilities into the decision making process of the operational weather community. For polar orbiting sensors such as MODIS, AIRS, VIIRS, and CRiS, the use of direct broadcast ground stations is key to the real-time delivery of the data and derived products in a timely fashion. With the ABI on the geostationary GOES-R satellite, the data volumes will likely increase by a factor of 5-10 from current data streams. However, the high data volume and limited bandwidth of end user facilities presents a formidable obstacle to timely access to the data. This challenge can be addressed through the use of subsetting techniques, innovative web services, and the judicious selection of data formats. Many of these approaches have been implemented by SPoRT for the delivery of real-time products to NWS forecast offices and other weather entities. Once available in decision support systems like AWIPS II, these new data and products must be integrated into existing and new displays that allow for the integration of the data with existing operational products in these systems. SPoRT is leading the way in demonstrating this enhanced capability. This paper will highlight the ways SPoRT is overcoming many of the challenges presented by the enormous data

A simple RT-PCR-based strategy for screening connexin identity

Directory of Open Access Journals (Sweden)

M. Urban

1999-08-01

Full Text Available Vertebrate gap junctions are aggregates of transmembrane channels which are composed of connexin (Cx proteins encoded by at least fourteen distinct genes in mammals. Since the same Cx type can be expressed in different tissues and more than one Cx type can be expressed by the same cell, the thorough identification of which connexin is in which cell type and how connexin expression changes after experimental manipulation has become quite laborious. Here we describe an efficient, rapid and simple method by which connexin type(s can be identified in mammalian tissue and cultured cells using endonuclease cleavage of RT-PCR products generated from "multi primers" (sense primer, degenerate oligonucleotide corresponding to a region of the first extracellular domain; antisense primer, degenerate oligonucleotide complementary to the second extracellular domain that amplify the cytoplasmic loop regions of all known connexins except Cx36. In addition, we provide sequence information on RT-PCR primers used in our laboratory to screen individual connexins and predictions of extension of the "multi primer" method to several human connexins.

Conventional and real time RT-PCR assays for the detection and differentiation of variant rabbit hemorrhagic disease virus (RHDVb) and its recombinants.

Science.gov (United States)

Dalton, K P; Arnal, J L; Benito, A A; Chacón, G; Martín Alonso, J M; Parra, F

2018-01-01

Since its emergence, variant RHDV (RHDVb/RHDV2) has spread throughout the Iberian Peninsula aided by the apparent lack of cross protection provided by classic (genogroup 1; G1) strain derived vaccines. In addition to RHDVb, full-length genome sequencing of RHDV strains has recently revealed the circulation of recombinant viruses on the Iberian Peninsula. These recombinant viruses contain the RHDVb structural protein encoding sequences and the non-structural coding regions of either pathogenic RHDV-G1 strains or non-pathogenic (np) rabbit caliciviruses. The aim of the work was twofold: firstly to validate a diagnostic real time RT-PCR developed in 2012 for the detection of RHDVb strains and secondly, to design a conventional RT-PCR for the differentiation of RHDVb strains from RHDVb recombinants by subsequent sequencing of the amplicon. Copyright © 2017 Elsevier B.V. All rights reserved.

Recent Upgrades to NASA SPoRT Initialization Datasets for the Environmental Modeling System

Science.gov (United States)

Case, Jonathan L.; Lafontaine, Frank J.; Molthan, Andrew L.; Zavodsky, Bradley T.; Rozumalski, Robert A.

2012-01-01

The NASA Short-term Prediction Research and Transition (SPoRT) Center has developed several products for its NOAA/National Weather Service (NWS) partners that can initialize specific fields for local model runs within the NOAA/NWS Science and Training Resource Center Environmental Modeling System (EMS). The suite of SPoRT products for use in the EMS consists of a Sea Surface Temperature (SST) composite that includes a Lake Surface Temperature (LST) analysis over the Great Lakes, a Great Lakes sea-ice extent within the SST composite, a real-time Green Vegetation Fraction (GVF) composite, and NASA Land Information System (LIS) gridded output. This paper and companion poster describe each dataset and provide recent upgrades made to the SST, Great Lakes LST, GVF composites, and the real-time LIS runs.

Long Terminal Repeat Retrotransposon Content in Eight Diploid Sunflower Species Inferred from Next-Generation Sequence Data

Science.gov (United States)

Tetreault, Hannah M.; Ungerer, Mark C.

2016-01-01

The most abundant transposable elements (TEs) in plant genomes are Class I long terminal repeat (LTR) retrotransposons represented by superfamilies gypsy and copia. Amplification of these superfamilies directly impacts genome structure and contributes to differential patterns of genome size evolution among plant lineages. Utilizing short-read Illumina data and sequence information from a panel of Helianthus annuus (sunflower) full-length gypsy and copia elements, we explore the contribution of these sequences to genome size variation among eight diploid Helianthus species and an outgroup taxon, Phoebanthus tenuifolius. We also explore transcriptional dynamics of these elements in both leaf and bud tissue via RT-PCR. We demonstrate that most LTR retrotransposon sublineages (i.e., families) display patterns of similar genomic abundance across species. A small number of LTR retrotransposon sublineages exhibit lineage-specific amplification, particularly in the genomes of species with larger estimated nuclear DNA content. RT-PCR assays reveal that some LTR retrotransposon sublineages are transcriptionally active across all species and tissue types, whereas others display species-specific and tissue-specific expression. The species with the largest estimated genome size, H. agrestis, has experienced amplification of LTR retrotransposon sublineages, some of which have proliferated independently in other lineages in the Helianthus phylogeny. PMID:27233667

Two RNAs or DNAs May Artificially Fuse Together at a Short Homologous Sequence (SHS) during Reverse Transcription or Polymerase Chain Reactions, and Thus Reporting an SHS-Containing Chimeric RNA Requires Extra Caution

Science.gov (United States)

Xie, Bingkun; Yang, Wei; Ouyang, Yongchang; Chen, Lichan; Jiang, Hesheng; Liao, Yuying; Liao, D. Joshua

2016-01-01

Tens of thousands of chimeric RNAs have been reported. Most of them contain a short homologous sequence (SHS) at the joining site of the two partner genes but are not associated with a fusion gene. We hypothesize that many of these chimeras may be technical artifacts derived from SHS-caused mis-priming in reverse transcription (RT) or polymerase chain reactions (PCR). We cloned six chimeric complementary DNAs (cDNAs) formed by human mitochondrial (mt) 16S rRNA sequences at an SHS, which were similar to several expression sequence tags (ESTs).These chimeras, which could not be detected with cDNA protection assay, were likely formed because some regions of the 16S rRNA are reversely complementary to another region to form an SHS, which allows the downstream sequence to loop back and anneal at the SHS to prime the synthesis of its complementary strand, yielding a palindromic sequence that can form a hairpin-like structure.We identified a 16S rRNA that ended at the 4th nucleotide(nt) of the mt-tRNA-leu was dominant and thus should be the wild type. We also cloned a mouse Bcl2-Nek9 chimeric cDNA that contained a 5-nt unmatchable sequence between the two partners, contained two copies of the reverse primer in the same direction but did not contain the forward primer, making it unclear how this Bcl2-Nek9 was formed and amplified. Moreover, a cDNA was amplified because one primer has 4 nts matched to the template, suggesting that there may be many more artificial cDNAs than we have realized, because the nuclear and mt genomes have many more 4-nt than 5-nt or longer homologues. Altogether, the chimeric cDNAs we cloned are good examples suggesting that many cDNAs may be artifacts due to SHS-caused mis-priming and thus greater caution should be taken when new sequence is obtained from a technique involving DNA polymerization. PMID:27148738

Detection of Coconut cadang-cadang viroid (CCCVd) in oil palm by reverse transcription loop-mediated isothermal amplification (RT-LAMP).

Science.gov (United States)

Thanarajoo, Sathis Sri; Kong, Lih Ling; Kadir, Jugah; Lau, Wei Hongi; Vadamalai, Ganesan

2014-06-01

A reverse transcription loop-mediated isothermal amplification (RT-LAMP) detected Coconut cadang-cadang viroid (CCCVd) within 60 min at 60 °C in total nucleic acid extracted from oil palm leaves infected with CCCVd. Positive reactions showed colour change from orange to green in the reaction mix after the addition of fluorescent reagent, and a laddering pattern band on 2% agarose gel electrophoresis. Conventional RT-PCR with LAMP primers produced amplicons with a sequence identical to the 297-nt CCCVd oil palm variant with the primers being specific for CCCVd and not for other viroids such as PSTVd and CEVd. RT-LAMP was found to be rapid and specific for detecting oil palm CCCVd. Copyright © 2014 Elsevier B.V. All rights reserved.

NASA SPoRT Modeling and Data Assimilation Research and Transition Activities Using WRF, LIS and GSI

Science.gov (United States)

Case, Jonathan L.; Blankenship, Clay B.; Zavodsky, Bradley T.; Srikishen, Jayanthi; Berndt, Emily B.

2014-01-01

weather research and forecasting ===== The NASA Short-term Prediction Research and Transition (SPoRT) program has numerous modeling and data assimilation (DA) activities in which the WRF model is a key component. SPoRT generates realtime, research satellite products from the MODIS and VIIRS instruments, making the data available to NOAA/NWS partners running the WRF/EMS, including: (1) 2-km northwestern-hemispheric SST composite, (2) daily, MODIS green vegetation fraction (GVF) over CONUS, and (3) NASA Land Information System (LIS) runs of the Noah LSM over the southeastern CONUS. Each of these datasets have been utilized by specific SPoRT partners in local EMS model runs, with select offices evaluating the impacts using a set of automated scripts developed by SPoRT that manage data acquisition and run the NCAR Model Evaluation Tools verification package. SPoRT is engaged in DA research with the Gridpoint Statistical Interpolation (GSI) and Ensemble Kalman Filter in LIS for soil moisture DA. Ongoing DA projects using GSI include comparing the impacts of assimilating Atmospheric Infrared Sounder (AIRS) radiances versus retrieved profiles, and an analysis of extra-tropical cyclones with intense non-convective winds. As part of its Early Adopter activities for the NASA Soil Moisture Active Passive (SMAP) mission, SPoRT is conducting bias correction and soil moisture DA within LIS to improve simulations using the NASA Unified-WRF (NU-WRF) for both the European Space Agency's Soil Moisture Ocean Salinity and upcoming SMAP mission data. SPoRT has also incorporated real-time global GVF data into LIS and WRF from the VIIRS product being developed by NOAA/NESDIS. This poster will highlight the research and transition activities SPoRT conducts using WRF, NU-WRF, EMS, LIS, and GSI.

Gold nanoparticle-based RT-PCR and real-time quantitative RT-PCR assays for detection of Japanese encephalitis virus

International Nuclear Information System (INIS)

Huang, S-H; Tsai, M-H; Lin, C-W; Yang, T-C; Chuang, P-H; Tsai, I-S; Lu, H-C; Wan Lei; Lin, Y-J; Lai, C-H

2008-01-01

Virus isolation and antibody detection are routinely used for diagnosis of Japanese encephalitis virus (JEV) infection, but the low level of transient viremia in some JE patients makes JEV isolation from clinical and surveillance samples very difficult. We describe the use of gold nanoparticle-based RT-PCR and real-time quantitative RT-PCR assays for detection of JEV from its RNA genome. We tested the effect of gold nanoparticles on four different PCR systems, including conventional PCR, reverse-transcription PCR (RT-PCR), and SYBR green real-time PCR and RT-PCR assays for diagnosis in the acute phase of JEV infection. Gold nanoparticles increased the amplification yield of the PCR product and shortened the PCR time compared to the conventional reaction. In addition, nanogold-based real-time RT-PCR showed a linear relationship between Ct and template amount using ten-fold dilutions of JEV. The nanogold-based RT-PCR and real-time quantitative RT-PCR assays were able to detect low levels (1-10 000 copies) of the JEV RNA genomes extracted from culture medium or whole blood, providing early diagnostic tools for the detection of low-level viremia in the acute-phase infection. The assays described here were simple, sensitive, and rapid approaches for detection and quantitation of JEV in tissue cultured samples as well as clinical samples

Selection of reference genes for quantitative real time RT-PCR during dimorphism in the zygomycete Mucor circinelloides.

Science.gov (United States)

Valle-Maldonado, Marco I; Jácome-Galarza, Irvin E; Gutiérrez-Corona, Félix; Ramírez-Díaz, Martha I; Campos-García, Jesús; Meza-Carmen, Víctor

2015-03-01

Mucor circinelloides is a dimorphic fungal model for studying several biological processes including cell differentiation (yeast-mold transitions) as well as biodiesel and carotene production. The recent release of the first draft sequence of the M. circinelloides genome, combined with the availability of analytical methods to determine patterns of gene expression, such as quantitative Reverse transcription-Polymerase chain reaction (qRT-PCR), and the development of molecular genetic tools for the manipulation of the fungus, may help identify M. circinelloides gene products and analyze their relevance in different biological processes. However, no information is available on M. circinelloides genes of stable expression that could serve as internal references in qRT-PCR analyses. One approach to solve this problem consists in the use of housekeeping genes as internal references. However, validation of the usability of these reference genes is a fundamental step prior to initiating qRT-PCR assays. This work evaluates expression of several constitutive genes by qRT-PCR throughout the morphological differentiation stages of M. circinelloides; our results indicate that tfc-1 and ef-1 are the most stable genes for qRT-PCR assays during differentiation studies and they are proposed as reference genes to carry out gene expression studies in this fungus.

Global Dynamics and International Cooperation Needs of RT Development and Utilization for the Establishment of the Northeast Asia RT Hub in Korea

Energy Technology Data Exchange (ETDEWEB)

Kim, Y. K.; Oh, G. B.; Yun, S. W.; Yang, M. H.; Ryu, J. S.; Choi, S. J.; Choi, S. G

2005-11-15

In an effort to achieve the objectives, the following scopes were categorized for in-depth study. First, analysis framework was developed for RT capacity development and international technology cooperation strategy. RT survey checklist and international technology cooperation was analyzed with interviewing and reports of domestic participants recently. Second, RT strategic environment was analyze for East-Asia hub competition/cooperation and developing nations using analysis framework. Korean RT was analyzed using SWOT analysis for establishment of RT hub in Korea. Third, East-Asian nations were classified analyzed by RT categories in standpoint of our country. Technology status and future cooperation plan were discussed about RT application for bio-medicine. Products/technology seminar related to an export was hold about support plan of admission/sales for functional food HemoHIM. This study can be utilized in the establishment of RT hub and development strategy. And it can be also utilized in promotion devising of domestic RT and planning setup for obtaing the international competitive power.

Global Dynamics and International Cooperation Needs of RT Development and Utilization for the Establishment of the Northeast Asia RT Hub in Korea

International Nuclear Information System (INIS)

Kim, Y. K.; Oh, G. B.; Yun, S. W.; Yang, M. H.; Ryu, J. S.; Choi, S. J.; Choi, S. G.

2005-11-01

In an effort to achieve the objectives, the following scopes were categorized for in-depth study. First, analysis framework was developed for RT capacity development and international technology cooperation strategy. RT survey checklist and international technology cooperation was analyzed with interviewing and reports of domestic participants recently. Second, RT strategic environment was analyze for East-Asia hub competition/cooperation and developing nations using analysis framework. Korean RT was analyzed using SWOT analysis for establishment of RT hub in Korea. Third, East-Asian nations were classified analyzed by RT categories in standpoint of our country. Technology status and future cooperation plan were discussed about RT application for bio-medicine. Products/technology seminar related to an export was hold about support plan of admission/sales for functional food HemoHIM. This study can be utilized in the establishment of RT hub and development strategy. And it can be also utilized in promotion devising of domestic RT and planning setup for obtaing the international competitive power

Expansion of the Real-Time SPoRT-Land Information System for NOAA/National Weather Service Situational Awareness and Local Modeling Applications

Science.gov (United States)

Case, Jonathan L; White, Kristopher D.

2014-01-01

The NASA Short-term Prediction Research and Transition (SPoRT) Center in Huntsville, AL is running a real-time configuration of the Noah land surface model (LSM) within the NASA Land Information System (LIS) framework (hereafter referred to as the "SPoRT-LIS"). Output from the real-time SPoRT-LIS is used for (1) initializing land surface variables for local modeling applications, and (2) displaying in decision support systems for situational awareness and drought monitoring at select NOAA/National Weather Service (NWS) partner offices. The experimental CONUS run incorporates hourly quantitative precipitation estimation (QPE) from the National Severe Storms Laboratory Multi- Radar Multi-Sensor (MRMS) which will be transitioned into operations at the National Centers for Environmental Prediction (NCEP) in Fall 2014.This paper describes the current and experimental SPoRT-LIS configurations, and documents some of the limitations still remaining through the advent of MRMS precipitation analyses in the SPoRT-LIS land surface model (LSM) simulations.

Use of alternative alkali chlorides in RT and PCR of polynucleotides containing G quadruplex structures.

Science.gov (United States)

Ramos-Alemán, Fabiola; González-Jasso, Eva; Pless, Reynaldo C

2018-02-15

Several alkali chlorides were compared for their use in reverse transcription (RT) and PCR of different types of nucleic acid templates. On a test region of biological DNA incapable of forming G quadruplex (G4) structures, Taq DNA polymerase showed similar PCR performance with 50 mM KCl, CsCl, LiCl, and NaCl. In contrast, on a synthetic model polydeoxyribonucleotide prone to G4 formation, good PCR amplification was obtained with 50 mM CsCl, but little or none with LiCl or KCl. Similarly, in RT of a G4-prone model polyribonucleotide, MMLV reverse transcriptase produced a good yield with 50 mM CsCl, mediocre yields with LiCl or without added alkali chloride, and a poor yield with 50 mM KCl. The full RT-PCR assay starting from the G4-prone polyribonucleotide, showed good results with CsCl in both stages, poor results with LiCl, and no product formation with KCl. The model polynucleotides showed fast G quadruplex formation under PCR or RT conditions with 50 mM KCl, but not with CsCl or LiCl. The results argue for the use of CsCl instead of KCl for RT and PCR of G4-prone sequences. No advantage was observed when using the 7-deaza type nucleotide analog c 7 dGTP in PCR amplification of the G4-prone polydeoxyribonucleotide. Copyright © 2017 Elsevier Inc. All rights reserved.

Molecular analysis of dolphin morbillivirus: A new sensitive detection method based on nested RT-PCR.

Science.gov (United States)

Centelleghe, Cinzia; Beffagna, Giorgia; Zanetti, Rossella; Zappulli, Valentina; Di Guardo, Giovanni; Mazzariol, Sandro

2016-09-01

Cetacean Morbillivirus (CeMV) has been identified as the most pathogenic virus for cetaceans. Over the past three decades, this RNA virus has caused several outbreaks of lethal disease in odontocetes and mysticetes worldwide. Isolation and identification of CeMV RNA is very challenging in whales because of the poor preservation status frequently shown by tissues from stranded animals. Nested reverse transcription polymerase chain reaction (nested RT-PCR) is used instead of conventional RT-PCR when it is necessary to increase the sensitivity and the specificity of the reaction. This study describes a new nested RT-PCR technique useful to amplify small amounts of the cDNA copy of Cetacean morbillivirus (CeMV) when it is present in scant quantity in whales' biological specimens. This technique was used to analyze different tissues (lung, brain, spleen and other lymphoid tissues) from one under human care seal and seven cetaceans stranded along the Italian coastline between October 2011 and September 2015. A well-characterized, 200 base pair (bp) fragment of the dolphin Morbillivirus (DMV) haemagglutinin (H) gene, obtained by nested RT-PCR, was sequenced and used to confirm DMV positivity in all the eight marine mammals under study. In conclusion, this nested RT-PCR protocol can represent a sensitive detection method to identify CeMV-positive, poorly preserved tissue samples. Furthermore, this is also a rather inexpensive molecular technique, relatively easy to apply. Copyright © 2016 Elsevier B.V. All rights reserved.

De novo assembly of a 40 Mb eukaryotic genome from short sequence reads: Sordaria macrospora, a model organism for fungal morphogenesis.

Science.gov (United States)

Nowrousian, Minou; Stajich, Jason E; Chu, Meiling; Engh, Ines; Espagne, Eric; Halliday, Karen; Kamerewerd, Jens; Kempken, Frank; Knab, Birgit; Kuo, Hsiao-Che; Osiewacz, Heinz D; Pöggeler, Stefanie; Read, Nick D; Seiler, Stephan; Smith, Kristina M; Zickler, Denise; Kück, Ulrich; Freitag, Michael

2010-04-08

Filamentous fungi are of great importance in ecology, agriculture, medicine, and biotechnology. Thus, it is not surprising that genomes for more than 100 filamentous fungi have been sequenced, most of them by Sanger sequencing. While next-generation sequencing techniques have revolutionized genome resequencing, e.g. for strain comparisons, genetic mapping, or transcriptome and ChIP analyses, de novo assembly of eukaryotic genomes still presents significant hurdles, because of their large size and stretches of repetitive sequences. Filamentous fungi contain few repetitive regions in their 30-90 Mb genomes and thus are suitable candidates to test de novo genome assembly from short sequence reads. Here, we present a high-quality draft sequence of the Sordaria macrospora genome that was obtained by a combination of Illumina/Solexa and Roche/454 sequencing. Paired-end Solexa sequencing of genomic DNA to 85-fold coverage and an additional 10-fold coverage by single-end 454 sequencing resulted in approximately 4 Gb of DNA sequence. Reads were assembled to a 40 Mb draft version (N50 of 117 kb) with the Velvet assembler. Comparative analysis with Neurospora genomes increased the N50 to 498 kb. The S. macrospora genome contains even fewer repeat regions than its closest sequenced relative, Neurospora crassa. Comparison with genomes of other fungi showed that S. macrospora, a model organism for morphogenesis and meiosis, harbors duplications of several genes involved in self/nonself-recognition. Furthermore, S. macrospora contains more polyketide biosynthesis genes than N. crassa. Phylogenetic analyses suggest that some of these genes may have been acquired by horizontal gene transfer from a distantly related ascomycete group. Our study shows that, for typical filamentous fungi, de novo assembly of genomes from short sequence reads alone is feasible, that a mixture of Solexa and 454 sequencing substantially improves the assembly, and that the resulting data can be used for

De novo assembly of a 40 Mb eukaryotic genome from short sequence reads: Sordaria macrospora, a model organism for fungal morphogenesis.

Directory of Open Access Journals (Sweden)

Minou Nowrousian

2010-04-01

Full Text Available Filamentous fungi are of great importance in ecology, agriculture, medicine, and biotechnology. Thus, it is not surprising that genomes for more than 100 filamentous fungi have been sequenced, most of them by Sanger sequencing. While next-generation sequencing techniques have revolutionized genome resequencing, e.g. for strain comparisons, genetic mapping, or transcriptome and ChIP analyses, de novo assembly of eukaryotic genomes still presents significant hurdles, because of their large size and stretches of repetitive sequences. Filamentous fungi contain few repetitive regions in their 30-90 Mb genomes and thus are suitable candidates to test de novo genome assembly from short sequence reads. Here, we present a high-quality draft sequence of the Sordaria macrospora genome that was obtained by a combination of Illumina/Solexa and Roche/454 sequencing. Paired-end Solexa sequencing of genomic DNA to 85-fold coverage and an additional 10-fold coverage by single-end 454 sequencing resulted in approximately 4 Gb of DNA sequence. Reads were assembled to a 40 Mb draft version (N50 of 117 kb with the Velvet assembler. Comparative analysis with Neurospora genomes increased the N50 to 498 kb. The S. macrospora genome contains even fewer repeat regions than its closest sequenced relative, Neurospora crassa. Comparison with genomes of other fungi showed that S. macrospora, a model organism for morphogenesis and meiosis, harbors duplications of several genes involved in self/nonself-recognition. Furthermore, S. macrospora contains more polyketide biosynthesis genes than N. crassa. Phylogenetic analyses suggest that some of these genes may have been acquired by horizontal gene transfer from a distantly related ascomycete group. Our study shows that, for typical filamentous fungi, de novo assembly of genomes from short sequence reads alone is feasible, that a mixture of Solexa and 454 sequencing substantially improves the assembly, and that the resulting data

Highly Specific Detection of Five Exotic Quarantine Plant Viruses using RT-PCR

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Hoseong Choi

2013-03-01

Full Text Available To detect five plant viruses (Beet black scorch virus, Beet necrotic yellow vein virus, Eggplant mottled dwarf virus, Pelargonium zonate spot virus, and Rice yellow mottle virus for quarantine purposes, we designed 15 RT-PCR primer sets. Primer design was based on the nucleotide sequence of the coat protein gene, which is highly conserved within species. All but one primer set successfully amplified the targets, and gradient PCRs indicated that the optimal temperature for the 14 useful primer sets was 51.9°C. Some primer sets worked well regardless of annealing temperature while others required a very specific annealing temperature. A primer specificity test using plant total RNAs and cDNAs of other plant virus-infected samples demonstrated that the designed primer sets were highly specific and generated reproducible results. The newly developed RT-PCR primer sets would be useful for quarantine inspections aimed at preventing the entry of exotic plant viruses into Korea.

sRNAnalyzer-a flexible and customizable small RNA sequencing data analysis pipeline.

Science.gov (United States)

Wu, Xiaogang; Kim, Taek-Kyun; Baxter, David; Scherler, Kelsey; Gordon, Aaron; Fong, Olivia; Etheridge, Alton; Galas, David J; Wang, Kai

2017-12-01

Although many tools have been developed to analyze small RNA sequencing (sRNA-Seq) data, it remains challenging to accurately analyze the small RNA population, mainly due to multiple sequence ID assignment caused by short read length. Additional issues in small RNA analysis include low consistency of microRNA (miRNA) measurement results across different platforms, miRNA mapping associated with miRNA sequence variation (isomiR) and RNA editing, and the origin of those unmapped reads after screening against all endogenous reference sequence databases. To address these issues, we built a comprehensive and customizable sRNA-Seq data analysis pipeline-sRNAnalyzer, which enables: (i) comprehensive miRNA profiling strategies to better handle isomiRs and summarization based on each nucleotide position to detect potential SNPs in miRNAs, (ii) different sequence mapping result assignment approaches to simulate results from microarray/qRT-PCR platforms and a local probabilistic model to assign mapping results to the most-likely IDs, (iii) comprehensive ribosomal RNA filtering for accurate mapping of exogenous RNAs and summarization based on taxonomy annotation. We evaluated our pipeline on both artificial samples (including synthetic miRNA and Escherichia coli cultures) and biological samples (human tissue and plasma). sRNAnalyzer is implemented in Perl and available at: http://srnanalyzer.systemsbiology.net/. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

sRNAnalyzer—a flexible and customizable small RNA sequencing data analysis pipeline

Science.gov (United States)

Kim, Taek-Kyun; Baxter, David; Scherler, Kelsey; Gordon, Aaron; Fong, Olivia; Etheridge, Alton; Galas, David J.

2017-01-01

Abstract Although many tools have been developed to analyze small RNA sequencing (sRNA-Seq) data, it remains challenging to accurately analyze the small RNA population, mainly due to multiple sequence ID assignment caused by short read length. Additional issues in small RNA analysis include low consistency of microRNA (miRNA) measurement results across different platforms, miRNA mapping associated with miRNA sequence variation (isomiR) and RNA editing, and the origin of those unmapped reads after screening against all endogenous reference sequence databases. To address these issues, we built a comprehensive and customizable sRNA-Seq data analysis pipeline—sRNAnalyzer, which enables: (i) comprehensive miRNA profiling strategies to better handle isomiRs and summarization based on each nucleotide position to detect potential SNPs in miRNAs, (ii) different sequence mapping result assignment approaches to simulate results from microarray/qRT-PCR platforms and a local probabilistic model to assign mapping results to the most-likely IDs, (iii) comprehensive ribosomal RNA filtering for accurate mapping of exogenous RNAs and summarization based on taxonomy annotation. We evaluated our pipeline on both artificial samples (including synthetic miRNA and Escherichia coli cultures) and biological samples (human tissue and plasma). sRNAnalyzer is implemented in Perl and available at: http://srnanalyzer.systemsbiology.net/. PMID:29069500

Archaeogenetics of Late Iron Age Çemialo Sırtı, Batman: Investigating maternal genetic continuity in north Mesopotamia since the Neolithic.

Science.gov (United States)

Yaka, Reyhan; Birand, Ayşegül; Yılmaz, Yasemin; Caner, Ceren; Açan, Sinan Can; Gündüzalp, Sidar; Parvizi, Poorya; Erim Özdoğan, Aslı; Togan, İnci; Somel, Mehmet

2018-05-01

North Mesopotamia has witnessed dramatic social change during the Holocene, but the impact of these events on its demographic history is poorly understood. Here, we study this question by analysing genetic data from the recently excavated Late Iron Age settlement of Çemialo Sırtı in Batman, southeast Turkey. Archaeological and radiocarbon evidence indicate that the site was inhabited during the second and first millennia BCE. Çemialo Sırtı reveals nomadic items of the Early Iron Age, as well as items associated with the Late Achaemenid and subsequent Hellenistic Periods. We compare Çemialo Sırtı mitochondrial DNA profiles with earlier and later populations from west Eurasia to describe genetic continuity patterns in the region. A total of 16 Çemialo Sırtı individuals' remains were studied. PCR and Sanger sequencing were used to obtain mitochondrial DNA HVRI-HVRII sequences. We studied haplotype diversity and pairwise genetic distances using F ST , comparing the Çemialo Sırtı population with ancient and modern-day populations from west Eurasia. Coalescent simulations were carried out to test continuity for specific population comparisons. Mitochondrial DNA (mtDNA) haplotypes from 12 Çemialo Sırtı individuals reveal high haplotype diversity in this population, conspicuously higher than early Holocene west Eurasian populations, which supports the notion of increasing population admixture in west Eurasia through the Holocene. In its mtDNA composition, Çemialo Sırtı shows highest affinity to Neolithic north Syria and Neolithic Anatolia among ancient populations studied, and to modern-day southwest Asian populations. Based on population genetic simulations we cannot reject continuity between Neolithic and Iron Age, or between Iron Age and present-day populations of the region. Despite the region's complex sociopolitical history and indication for increased genetic diversity over time, we find no evidence for sharp shifts in north Mesopotamian

The SPoRT concept of bracing for idiopathic scoliosis.

Science.gov (United States)

Zaina, Fabio; Fusco, Claudia; Atanasio, Salvatore; Negrini, Stefano

2011-01-01

The SPoRT (acronym: Symmetrical, Patient-oriented, Rigid, Three-dimensional, active) concept of bracing is a new way to build braces based on our 20 years of experience and the biomechanical principles of scoliosis correction, inclusive of the Sibilla and Sforzesco braces. The concept always requires a custom brace, which is made according to the patient's individual requirements. New technologies such as CAD-CAM can be applied, and often for better results, without the customary use of prebuilt forms whose measurements are stored in databases. Once the initial draft brace is completed, a final test must be made on the patient to modify and adapt it, depending on his or her real interaction between the body and the brace. The results that are today available on the SPoRT concept relate to the Sforzesco brace and are necessarily short-term, because the first treated patients are now reaching the fourth-year follow-up examination and haven't yet completed their treatments. On the basis of the initial evaluations, we can state that the Sforzesco brace is more effective than the Lyon brace after 6 months of treatment and that the Sforzesco brace is equally effective as the Risser Plast brace.

Deep sequencing discovery of novel and conserved microRNAs in trifoliate orange (Citrus trifoliata

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Yu Huaping

2010-07-01

Full Text Available Abstract Background MicroRNAs (miRNAs play a critical role in post-transcriptional gene regulation and have been shown to control many genes involved in various biological and metabolic processes. There have been extensive studies to discover miRNAs and analyze their functions in model plant species, such as Arabidopsis and rice. Deep sequencing technologies have facilitated identification of species-specific or lowly expressed as well as conserved or highly expressed miRNAs in plants. Results In this research, we used Solexa sequencing to discover new microRNAs in trifoliate orange (Citrus trifoliata which is an important rootstock of citrus. A total of 13,106,753 reads representing 4,876,395 distinct sequences were obtained from a short RNA library generated from small RNA extracted from C. trifoliata flower and fruit tissues. Based on sequence similarity and hairpin structure prediction, we found that 156,639 reads representing 63 sequences from 42 highly conserved miRNA families, have perfect matches to known miRNAs. We also identified 10 novel miRNA candidates whose precursors were all potentially generated from citrus ESTs. In addition, five miRNA* sequences were also sequenced. These sequences had not been earlier described in other plant species and accumulation of the 10 novel miRNAs were confirmed by qRT-PCR analysis. Potential target genes were predicted for most conserved and novel miRNAs. Moreover, four target genes including one encoding IRX12 copper ion binding/oxidoreductase and three genes encoding NB-LRR disease resistance protein have been experimentally verified by detection of the miRNA-mediated mRNA cleavage in C. trifoliata. Conclusion Deep sequencing of short RNAs from C. trifoliata flowers and fruits identified 10 new potential miRNAs and 42 highly conserved miRNA families, indicating that specific miRNAs exist in C. trifoliata. These results show that regulatory miRNAs exist in agronomically important trifoliate orange

Combination of RT-PCR and proteomics for the identification of Crimean-Congo hemorrhagic fever virus in ticks

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Isabel G. Fernández de Mera

2017-07-01

Full Text Available Crimean-Congo hemorrhagic fever (CCHF is an emerging tick-borne zoonotic disease caused by the CCHF virus (CCHFV. In this study, an experimental approach combining RT-PCR and proteomics was used for the identification and characterization of CCHFV in 106 ticks from 7 species that were collected from small ruminants in Greece. The methodological approach included an initial screening for CCHFV by RT-PCR followed by proteomics analysis of positive and control negative tick samples. This novel approach allowed the identification of CCHFV-positive ticks and provided additional information to corroborate the RT-PCR findings using a different approach. Two ticks, Dermacentor marginatus and Haemaphysalis parva collected from a goat and a sheep, respectively were positive for CCHFV. The sequences for CCHFV RNA segments S and L were characterized by RT-PCR and proteomics analysis of tick samples, respectively. These results showed the possibility of combining analyses at the RNA and protein levels using RT-PCR and proteomics for the characterization of CCHFV in ticks. The results supported that the CCHFV identified in ticks are genetic variants of the AP92 strain. Although the AP92-like strains probably do not represent a high risk of CCHF to the population, the circulation of genetically diverse CCHFV strains could potentially result in the appearance of novel viral genotypes with increased pathogenicity and fitness.

Enhancing the sensitivity of Dengue virus serotype detection by RT-PCR among infected children in India.

Science.gov (United States)

Ahamed, Syed Fazil; Vivek, Rosario; Kotabagi, Shalini; Nayak, Kaustuv; Chandele, Anmol; Kaja, Murali-Krishna; Shet, Anita

2017-06-01

Dengue surveillance relies on reverse transcription-polymerase chain reaction (RT-PCR), for confirmation of dengue virus (DENV) serotypes. We compared efficacies of published and modified primer sets targeting envelope (Env) and capsid-premembrane (C-prM) genes for detection of circulating DENV serotypes in southern India. Acute samples from children with clinically-diagnosed dengue were used for RT-PCR testing. All samples were also subjected to dengue serology (NS1 antigen and anti-dengue-IgM/IgG rapid immunochromatographic assay). Nested RT-PCR was performed on viral RNA using three methods targeting 654bp C-prM, 511bp C-prM and 641bp Env regions, respectively. RT-PCR-positive samples were validated by population sequencing. Among 171 children with suspected dengue, 121 were dengue serology-positive and 50 were dengue serology-negative. Among 121 serology-positives, RT-PCR detected 91 (75.2%) by CprM654, 72 (59.5%) by CprM511, and 74 (61.1%) by Env641. Among 50 serology-negatives, 10 (20.0%) were detected by CprM654, 12 (24.0%) by CprM511, and 11 (22.0%) by Env641. Overall detection rate using three methods sequentially was 82.6% (100/121) among serology-positive and 40.0% (20/50) among serology-negative samples; 6.6% (8/120) had co-infection with multiple DENV serotypes. We conclude that detection of acute dengue was enhanced by a modified RT-PCR method targeting the 654bp C-prM region, and further improved by using all three methods sequentially. Copyright © 2017 Elsevier B.V. All rights reserved.

Construction of an adult barnacle (Balanus amphitrite cDNA library and selection of reference genes for quantitative RT-PCR studies

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Burgess J Grant

2009-06-01

Full Text Available Abstract Background Balanus amphitrite is a barnacle commonly used in biofouling research. Although many aspects of its biology have been elucidated, the lack of genetic information is impeding a molecular understanding of its life cycle. As part of a wider multidisciplinary approach to reveal the biogenic cues influencing barnacle settlement and metamorphosis, we have sequenced and annotated the first cDNA library for B. amphitrite. We also present a systematic validation of potential reference genes for normalization of quantitative real-time PCR (qRT-PCR data obtained from different developmental stages of this animal. Results We generated a cDNA library containing expressed sequence tags (ESTs from adult B. amphitrite. A total of 609 unique sequences (comprising 79 assembled clusters and 530 singlets were derived from 905 reliable unidirectionally sequenced ESTs. Bioinformatics tools such as BLAST, HMMer and InterPro were employed to allow functional annotation of the ESTs. Based on these analyses, we selected 11 genes to study their ability to normalize qRT-PCR data. Total RNA extracted from 7 developmental stages was reverse transcribed and the expression stability of the selected genes was compared using geNorm, BestKeeper and NormFinder. These software programs produced highly comparable results, with the most stable gene being mt-cyb, while tuba, tubb and cp1 were clearly unsuitable for data normalization. Conclusion The collection of B. amphitrite ESTs and their annotation has been made publically available representing an important resource for both basic and applied research on this species. We developed a qRT-PCR assay to determine the most reliable reference genes. Transcripts encoding cytochrome b and NADH dehydrogenase subunit 1 were expressed most stably, although other genes also performed well and could prove useful to normalize gene expression studies.

Genome-wide analysis of replication timing by next-generation sequencing with E/L Repli-seq.

Science.gov (United States)

Marchal, Claire; Sasaki, Takayo; Vera, Daniel; Wilson, Korey; Sima, Jiao; Rivera-Mulia, Juan Carlos; Trevilla-García, Claudia; Nogues, Coralin; Nafie, Ebtesam; Gilbert, David M

2018-05-01

This protocol is an extension to: Nat. Protoc. 6, 870-895 (2014); doi:10.1038/nprot.2011.328; published online 02 June 2011Cycling cells duplicate their DNA content during S phase, following a defined program called replication timing (RT). Early- and late-replicating regions differ in terms of mutation rates, transcriptional activity, chromatin marks and subnuclear position. Moreover, RT is regulated during development and is altered in diseases. Here, we describe E/L Repli-seq, an extension of our Repli-chip protocol. E/L Repli-seq is a rapid, robust and relatively inexpensive protocol for analyzing RT by next-generation sequencing (NGS), allowing genome-wide assessment of how cellular processes are linked to RT. Briefly, cells are pulse-labeled with BrdU, and early and late S-phase fractions are sorted by flow cytometry. Labeled nascent DNA is immunoprecipitated from both fractions and sequenced. Data processing leads to a single bedGraph file containing the ratio of nascent DNA from early versus late S-phase fractions. The results are comparable to those of Repli-chip, with the additional benefits of genome-wide sequence information and an increased dynamic range. We also provide computational pipelines for downstream analyses, for parsing phased genomes using single-nucleotide polymorphisms (SNPs) to analyze RT allelic asynchrony, and for direct comparison to Repli-chip data. This protocol can be performed in up to 3 d before sequencing, and requires basic cellular and molecular biology skills, as well as a basic understanding of Unix and R.

Hyperthermic Fibrinolysis with rt-PA: In Vitro Results

International Nuclear Information System (INIS)

Schwarzenberg, Helmut; Mueller-Huelsbeck, Stefan; Brossman, Joachim; Christian Glueer, Claus; Bruhn, Hans Dieter; Heller, Martin

1998-01-01

Purpose: To investigate the influence of hyperthermia up to 45 deg. C on fibrinolysis with recombinant tissue-type plasminogen activator (rt-PA). Methods: Standardized fibrin clots were incubated in a water bath for 5 hr with either rt-PA (test group) or 0.9% sodium chloride (control group) and blood plasma at temperatures of 30-45 deg. C. Concentrations of D-dimer and time to complete clot lysis were measured.Results: The activity of fibrinolysis with rt-PA rose with increasing temperature: time to lysis approximately halved from 30 deg. C to 40 deg. C and the concentration of D-dimer tripled. In the control group clot size did not change.Conclusions: Activity of rt-PA-induced fibrinolysis rises distinctly with higher temperatures. Since even healthy subjects show a physiologic decline in body temperature in the extremities, in patients with occlusive arterial disease decreased activity of fibrinolysis with rt-PA can be expected. Controlled hyperthermia may improve fibrinolysis with rt-PA and should be investigated in vivo

GOES-R Proving Ground Activities at the NASA Short-Term Prediction Research and Transition (SPoRT) Center

Science.gov (United States)

Molthan, Andrew

2011-01-01

SPoRT is actively involved in GOES-R Proving Ground activities in a number of ways: (1) Applying the paradigm of product development, user training, and interaction to foster interaction with end users at NOAA forecast offices national centers. (2) Providing unique capabilities in collaboration with other GOES-R Proving Ground partners (a) Hybrid GOES-MODIS imagery (b) Pseudo-GLM via regional lightning mapping arrays (c) Developing new RGB imagery from EUMETSAT guidelines

Draft Genome Sequence of Lactobacillus delbrueckii Strain #22 Isolated from a Patient with Short Bowel Syndrome and Previous d-Lactic Acidosis and Encephalopathy.

Science.gov (United States)

Domann, Eugen; Fischer, Florence; Glowatzki, Fabian; Fritzenwanker, Moritz; Hain, Torsten; Zechel-Gran, Silke; Giffhorn-Katz, Susanne; Neubauer, Bernd A

2016-07-28

d-Lactic acidosis with associated encephalopathy caused by overgrowth of intestinal lactic acid bacteria is a rarely diagnosed neurological complication of patients with short bowel syndrome. Here, we report the draft genome sequence of Lactobacillus delbrueckii strain #22 isolated from a patient with short bowel syndrome and previous d-lactic acidosis/encephalopathy. Copyright © 2016 Domann et al.

Sequence analysis and over-expression of ribosomal protein S28 ...

African Journals Online (AJOL)

RPS28 is a component of the 40S small ribosomal subunit encoded by RPS28 gene, which is specific to eukaryotes. The cDNA and the genomic sequence of RPS28 were cloned successfully from the Giant Panda using RT-PCR technology and Touchdown-PCR, respectively. Both sequences were analyzed preliminarily ...

Simultaneous Detection of Mixed Infection of Onion yellow dwarf virus and an Allexivirus in RT-PCR for Ensuring Virus Free Onion Bulbs.

Science.gov (United States)

Kumar, Sandeep; Baranwal, V K; Joshi, Subodh; Arya, Meenakshi; Majumder, S

2010-06-01

Reduced seed production in onion is associated with Onion yellow dwarf virus (OYDV), a filamentous Potyvirus. Onion is also infected with other filamentous virus particles suspected to be Allexivirus. RT-PCR was used to detect mixed infection of both the viruses in leaves and bulbs. A duplex RT-PCR was developed, which simultaneously detected the presence of these two viruses in winter (Rabi) onion bulb. In summer (Kharif) onion bulbs only Allexivirus was detected. The absence of OYDV in summer crop is discussed. The sequencing of RT-PCR amplified products confirmed the identity of OYDV and Allexivirus, the latter showing closer identity to Garlic virus C (GVC)/Garlic mite-borne mosaic virus. This makes the first detection of an Allexivirus in onion crop in India. The duplex RT-PCR to detect these viruses (OYDV and Allexivirus) would be an improvement for indexing of viruses in onion bulbs for seed production.

Musicians' and nonmusicians' short-term memory for verbal and musical sequences: comparing phonological similarity and pitch proximity.

Science.gov (United States)

Williamson, Victoria J; Baddeley, Alan D; Hitch, Graham J

2010-03-01

Language-music comparative studies have highlighted the potential for shared resources or neural overlap in auditory short-term memory. However, there is a lack of behavioral methodologies for comparing verbal and musical serial recall. We developed a visual grid response that allowed both musicians and nonmusicians to perform serial recall of letter and tone sequences. The new method was used to compare the phonological similarity effect with the impact of an operationalized musical equivalent-pitch proximity. Over the course of three experiments, we found that short-term memory for tones had several similarities to verbal memory, including limited capacity and a significant effect of pitch proximity in nonmusicians. Despite being vulnerable to phonological similarity when recalling letters, however, musicians showed no effect of pitch proximity, a result that we suggest might reflect strategy differences. Overall, the findings support a limited degree of correspondence in the way that verbal and musical sounds are processed in auditory short-term memory.

Simple, specific molecular typing of dengue virus isolates using one-step RT-PCR and restriction fragment length polymorphism.

Science.gov (United States)

Ortiz, Alma; Capitan, Zeuz; Mendoza, Yaxelis; Cisneros, Julio; Moreno, Brechla; Zaldivar, Yamitzel; Garcia, Mariana; Smith, Rebecca E; Motta, Jorge; Pascale, Juan Miguel

2012-10-01

A one-step RT-PCR and one-enzyme RFLP was used to detect and distinguish among flaviviruses, including the four serotypes of dengue and the St. Louis Encephalitis, West Nile and Yellow Fever viruses in cultured virus samples or acute-phase human serum. Using a previously described RT-PCR, but novel RFLP procedure, results are obtained in 24 h with basic PCR and electrophoresis equipment. There is 95% agreement between RT-PCR/RFLP results and those achieved by indirect immunofluorescence assays, and 100% agreement between RT-PCR/RFLP results and gene sequencing. This method is more rapid than tests of cytopathic effect based on virus isolation in tissue culture, and simpler than real-time PCR. It does not require specialized equipment, radioisotopes or computer analysis and is a method that can be applied widely in the developing world. It allows for prompt determination of whether a flavivirus is the cause of illness in a febrile patient, rapid identification of dengue serotypes in circulation, and improved patient management in cases where prior dengue exposure make dengue hemorrhagic fever or dengue shock syndrome a risk. Copyright © 2012 Elsevier B.V. All rights reserved.

A community resource for high-throughput quantitative RT-PCR analysis of transcription factor gene expression in Medicago truncatula

Directory of Open Access Journals (Sweden)

Redman Julia C

2008-07-01

Full Text Available Abstract Background Medicago truncatula is a model legume species that is currently the focus of an international genome sequencing effort. Although several different oligonucleotide and cDNA arrays have been produced for genome-wide transcript analysis of this species, intrinsic limitations in the sensitivity of hybridization-based technologies mean that transcripts of genes expressed at low-levels cannot be measured accurately with these tools. Amongst such genes are many encoding transcription factors (TFs, which are arguably the most important class of regulatory proteins. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR is the most sensitive method currently available for transcript quantification, and one that can be scaled up to analyze transcripts of thousands of genes in parallel. Thus, qRT-PCR is an ideal method to tackle the problem of TF transcript quantification in Medicago and other plants. Results We established a bioinformatics pipeline to identify putative TF genes in Medicago truncatula and to design gene-specific oligonucleotide primers for qRT-PCR analysis of TF transcripts. We validated the efficacy and gene-specificity of over 1000 TF primer pairs and utilized these to identify sets of organ-enhanced TF genes that may play important roles in organ development or differentiation in this species. This community resource will be developed further as more genome sequence becomes available, with the ultimate goal of producing validated, gene-specific primers for all Medicago TF genes. Conclusion High-throughput qRT-PCR using a 384-well plate format enables rapid, flexible, and sensitive quantification of all predicted Medicago transcription factor mRNAs. This resource has been utilized recently by several groups in Europe, Australia, and the USA, and we expect that it will become the 'gold-standard' for TF transcript profiling in Medicago truncatula.

The development and application of the two real-time RT-PCR assays to detect the pathogen of HFMD.

Directory of Open Access Journals (Sweden)

Aili Cui

Full Text Available Large-scale Hand, Foot, and Mouth Disease (HFMD outbreaks have frequently occurred in China since 2008, affecting more than one million children and causing several hundred children deaths every year. The pathogens of HFMD are mainly human enteroviruses (HEVs. Among them, human enterovirus 71 (HEV71 and coxsackievirus A16 (CVA16 are the most common pathogens of HFMD. However, other HEVs could also cause HFMD. To rapidly detect HEV71 and CVA16, and ensure detection of all HEVs causing HFMD, two real-time hybridization probe-based RT-PCR assays were developed in this study. One is a multiplex real-time RT-PCR assay, which was developed to detect and differentiate HEV71 specifically from CVA16 directly from clinical specimens within 1-2 h, and the other is a broad-spectrum real-time RT-PCR assay, which targeted almost all HEVs. The experiments confirmed that the two assays have high sensitivity and specificity, and the sensitivity was up to 0.1 TCID50/ml for detection of HEVs, HEV71, and CVA16, respectively. A total of 213 clinical specimens were simultaneously detected by three kinds of assays, including the two real-time RT-PCR assays, direct conventional RT-PCR assay, and virus isolation assay on human rhabdomyosarcoma cells (RD cells. The total positive rate of both HEV71 and CVA16 was 69.48% with real-time RT-PCR assay, 47.42% with RT-PCR assay, and 34.58% with virus isolation assay. One HFMD clinical specimen was positive for HEV, but negative for HEV71 or CVA16, which was identified as Echovirus 11 (Echo11 by virus isolation, RT-PCR, and sequencing for the VP1 gene. The two real-time RT-PCR assays had been applied in 31 provincial HFMD labs to detect the pathogens of HFMD, which has contributed to the rapid identification of the pathogens in the early stages of HFMD outbreaks, and helped to clarify the etiologic agents of HFMD in China.

Analytical and clinical performance of the CDC real time RT-PCR assay for detection and typing of dengue virus.

Science.gov (United States)

Santiago, Gilberto A; Vergne, Edgardo; Quiles, Yashira; Cosme, Joan; Vazquez, Jesus; Medina, Juan F; Medina, Freddy; Colón, Candimar; Margolis, Harold; Muñoz-Jordán, Jorge L

2013-01-01

Dengue is an acute illness caused by the positive-strand RNA dengue virus (DENV). There are four genetically distinct DENVs (DENV-1-4) that cause disease in tropical and subtropical countries. Most patients are viremic when they present with symptoms; therefore, RT-PCR has been increasingly used in dengue diagnosis. The CDC DENV-1-4 RT-PCR Assay has been developed as an in-vitro diagnostic platform and was recently approved by the US Food and Drug Administration (FDA) for detection of dengue in patients with signs or symptoms of mild or severe dengue. The primers and probes of this test have been designed to detect currently circulating strains of DENV-1-4 from around the world at comparable sensitivity. In a retrospective study with 102 dengue cases confirmed by IgM anti-DENV seroconversion in the convalescent sample, the RT-PCR Assay detected DENV RNA in 98.04% of the paired acute samples. Using sequencing as a positive indicator, the RT-PCR Assay had a 97.92% positive agreement in 86 suspected dengue patients with a single acute serum sample. After extensive validations, the RT-PCR Assay performance was highly reproducible when evaluated across three independent testing sites, did not produce false positive results for etiologic agents of other febrile illnesses, and was not affected by pathological levels of potentially interfering biomolecules. These results indicate that the CDC DENV-1-4 RT-PCR Assay provides a reliable diagnostic platform capable for confirming dengue in suspected cases.

Detection, quantitation and identification of enteroviruses from surface waters and sponge tissue from the Florida Keys using real-time RT-PCR

Science.gov (United States)

Donaldson, K.A.; Griffin, Dale W.; Paul, J.H.

2002-01-01

A method was developed for the quantitative detection of pathogenic human enteroviruses from surface waters in the Florida Keys using Taqman (R) one-step Reverse transcription (RT)-PCR with the Model 7700 ABI Prism (R) Sequence Detection System. Viruses were directly extracted from unconcentrated grab samples of seawater, from seawater concentrated by vortex flow filtration using a 100kD filter and from sponge tissue. Total RNA was extracted from the samples, purified and concentrated using spin-column chromatography. A 192-196 base pair portion of the 5??? untranscribed region was amplified from these extracts. Enterovirus concentrations were estimated using real-time RT-PCR technology. Nine of 15 sample sites or 60% were positive for the presence of pathogenic human enteroviruses. Considering only near-shore sites, 69% were positive with viral concentrations ranging from 9.3viruses/ml to 83viruses/g of sponge tissue (uncorrected for extraction efficiency). Certain amplicons were selected for cloning and sequencing for identification. Three strains of waterborne enteroviruses were identified as Coxsackievirus A9, Coxsackievirus A16, and Poliovirus Sabin type 1. Time and cost efficiency of this one-step real-time RT-PCR methodology makes this an ideal technique to detect, quantitate and identify pathogenic enteroviruses in recreational waters. Copyright ?? 2002 Elsevier Science Ltd.

A Special Pre-Service-Inspection Using Radiographic Testing(RT) for Brazing Fitting used in Aircraft Hydraulic System

International Nuclear Information System (INIS)

Kim, Gyu Ho

2010-01-01

Brazing fitting which is one of the aircraft hydraulic power system components is widely used for saving weight and achieving higher reliability. Any inherent defects or damage of fitting can cause system failure and/or physical damage of human body due to highly pressurized fluid. Radiographic testing(RT) technique and additional micro-structure investigation on cut-away surfaces have been accomplished to find out some defect-like-inhomogeneity in the fittings. The radiography results showed that some defect-like-inhomogeneity existed inside body. Additional micro-structure investigation on cut-away surface reveals that the inhomogeneity is due to internal voids. In this study, it can be is said that RT technique can be a useful tool for field acceptance test of hydraulic brazing fitting in short time

An RT distribution analysis of relatedness proportion effects in lexical decision and semantic categorization reveals different mechanisms.

Science.gov (United States)

de Wit, Bianca; Kinoshita, Sachiko

2015-01-01

The magnitude of the semantic priming effect is known to increase as the proportion of related prime-target pairs in an experiment increases. This relatedness proportion (RP) effect was studied in a lexical decision task at a short prime-target stimulus onset asynchrony (240 ms), which is widely assumed to preclude strategic prospective usage of the prime. The analysis of the reaction time (RT) distribution suggested that the observed RP effect reflected a modulation of a retrospective semantic matching process. The pattern of the RP effect on the RT distribution found here is contrasted to that reported in De Wit and Kinoshita's (2014) semantic categorization study, and it is concluded that the RP effect is driven by different underlying mechanisms in lexical decision and semantic categorization.

[Possibilities in the differential diagnosis of brain neoplasms using the long and short time sequences of proton magnetic resonance spectroscopy

NARCIS (Netherlands)

Gajewicz, W.; Goraj, B.M.

2004-01-01

Currently to perform proton magnetic resonance spectroscopy (1H MRS) with single voxel spectroscopy (SVS) technique long and/or short echo time sequences are used in order to provide complementary information. PURPOSE: The aim of the study was to compare the usefulness of STEAM (time echo, TE, 20

Short-course palliative radiotherapy for uterine cervical cancer

Energy Technology Data Exchange (ETDEWEB)

Kim, Dong Hyun; Lee, Ju Hye; Ki, Yong Kan; Kim, Won Taek; Park, Dahl; Kim, Dong Won [Dept. of Radiation Oncology, Biomedical Research Institute, Pusan National University Hospital, Busan (Korea, Republic of); Nam, Ji Ho; Jeon, Sang Ho [Dept. of Radiation Oncology, Pusan National University Yangsan Hospital, Yangsan (Korea, Republic of)

2013-12-15

The purpose of this retrospective study was to evaluate the efficacy and feasibility of short-course hypofractionated radiotherapy (RT) for the palliation of uterine cervical cancer. Seventeen patients with cancer of the uterine cervix, who underwent palliative hypofractionated 3-dimensional conformal radiotherapy between January 2002 and June 2012, were retrospectively analyzed. RT was delivered to symptomatic lesions (both the primary mass and/or metastatic regional lymph nodes). The total dose was 20 to 25 Gy (median, 25 Gy) in 5 Gy daily fractions. The median follow-up duration was 12.2 months (range, 4 to 24 months). The median survival time was 7.8 months (range, 4 to 24 months). Vaginal bleeding was the most common presenting symptom followed by pelvic pain (9 patients). The overall response rates were 93.8% and 66.7% for vaginal bleeding control and pelvic pain, respectively. Nine patients did not have any acute side effects and 7 patients showed minor gastrointestinal toxicity. Only 1 patient had grade 3 diarrhea 1 week after completion of treatment, which was successfully treated conservatively. Late complications occurred in 4 patients; however, none of these were of grade 3 or higher severity. Short-course hypofractionated RT was effective and well tolerated as palliative treatment for uterine cervical cancer.

Short-course palliative radiotherapy for uterine cervical cancer

International Nuclear Information System (INIS)

Kim, Dong Hyun; Lee, Ju Hye; Ki, Yong Kan; Kim, Won Taek; Park, Dahl; Kim, Dong Won; Nam, Ji Ho; Jeon, Sang Ho

2013-01-01

The purpose of this retrospective study was to evaluate the efficacy and feasibility of short-course hypofractionated radiotherapy (RT) for the palliation of uterine cervical cancer. Seventeen patients with cancer of the uterine cervix, who underwent palliative hypofractionated 3-dimensional conformal radiotherapy between January 2002 and June 2012, were retrospectively analyzed. RT was delivered to symptomatic lesions (both the primary mass and/or metastatic regional lymph nodes). The total dose was 20 to 25 Gy (median, 25 Gy) in 5 Gy daily fractions. The median follow-up duration was 12.2 months (range, 4 to 24 months). The median survival time was 7.8 months (range, 4 to 24 months). Vaginal bleeding was the most common presenting symptom followed by pelvic pain (9 patients). The overall response rates were 93.8% and 66.7% for vaginal bleeding control and pelvic pain, respectively. Nine patients did not have any acute side effects and 7 patients showed minor gastrointestinal toxicity. Only 1 patient had grade 3 diarrhea 1 week after completion of treatment, which was successfully treated conservatively. Late complications occurred in 4 patients; however, none of these were of grade 3 or higher severity. Short-course hypofractionated RT was effective and well tolerated as palliative treatment for uterine cervical cancer.

Arvo Pärt: Kui kannatab üks inimene, siis kannatab kogu maailm! / Arvo Pärt, Nora Pärt ; intervjueerinud Yana Toom, Maarja-Liis Arujärv, Aleksandr Zukerman ; kommenteerinud Edgar Savisaar

Index Scriptorium Estoniae

Pärt, Arvo, 1935-

2010-01-01

Arvo Pärt ja tema abikaasa Nora Pärt andsid Türgis enne Pärdi uudisteose "Aadama itk" esiettekannet intervjuu, kus rääkisid nii nende raskustest Läände elama asumisel kui uudisteose sügavamast mõttest

CoverageAnalyzer (CAn: A Tool for Inspection of Modification Signatures in RNA Sequencing Profiles

Directory of Open Access Journals (Sweden)

Ralf Hauenschild

2016-11-01

Full Text Available Combination of reverse transcription (RT and deep sequencing has emerged as a powerful instrument for the detection of RNA modifications, a field that has seen a recent surge in activity because of its importance in gene regulation. Recent studies yielded high-resolution RT signatures of modified ribonucleotides relying on both sequence-dependent mismatch patterns and reverse transcription arrests. Common alignment viewers lack specialized functionality, such as filtering, tailored visualization, image export and differential analysis. Consequently, the community will profit from a platform seamlessly connecting detailed visual inspection of RT signatures and automated screening for modification candidates. CoverageAnalyzer (CAn was developed in response to the demand for a powerful inspection tool. It is freely available for all three main operating systems. With SAM file format as standard input, CAn is an intuitive and user-friendly tool that is generally applicable to the large community of biomedical users, starting from simple visualization of RNA sequencing (RNA-Seq data, up to sophisticated modification analysis with significance-based modification candidate calling.

Standard (60 Gy) or Short-Course (40 Gy) Irradiation Plus Concomitant and Adjuvant Temozolomide for Elderly Patients With Glioblastoma: A Propensity-Matched Analysis

Energy Technology Data Exchange (ETDEWEB)

Minniti, Giuseppe, E-mail: gminniti@ospedalesantandrea.it [Radiation Oncology Unit, Sant' Andrea Hospital, University Sapienza, Rome (Italy); Scientific Institute for Research, Hospitalization and Health Care (IRCCS) Neuromed, Pozzilli (Italy); Scaringi, Claudia [Radiation Oncology Unit, Sant' Andrea Hospital, University Sapienza, Rome (Italy); Lanzetta, Gaetano [Scientific Institute for Research, Hospitalization and Health Care (IRCCS) Neuromed, Pozzilli (Italy); Terrenato, Irene [Biostatistic Unit, Regina Elena National Cancer Institute, Rome (Italy); Esposito, Vincenzo; Arcella, Antonella [Scientific Institute for Research, Hospitalization and Health Care (IRCCS) Neuromed, Pozzilli (Italy); Pace, Andrea [Neurology Unit, Regina Elena National Cancer Institute, Rome (Italy); Giangaspero, Felice [Scientific Institute for Research, Hospitalization and Health Care (IRCCS) Neuromed, Pozzilli (Italy); Bozzao, Alessandro [Neuroradiology Unit, Sant' Andrea Hospital, University Sapienza, Rome (Italy); Enrici, Riccardo Maurizi [Radiation Oncology Unit, Sant' Andrea Hospital, University Sapienza, Rome (Italy)

2015-01-01

Purpose: To evaluate 2 specific radiation schedules, each combined with temozolomide (TMZ), assessing their efficacy and safety in patients aged ≥65 years with newly diagnosed glioblastoma (GBM). Methods and Materials: Patients aged ≥65 years with Karnofsky performance status (KPS) ≥60 who received either standard (60 Gy) or short-course (40 Gy) radiation therapy (RT) with concomitant and adjuvant TMZ between June 2004 and October 2013 were retrospectively analyzed. A propensity score analysis was executed for a balanced comparison of treatment outcomes. Results: A total of 127 patients received standard RT-TMZ, whereas 116 patients underwent short-course RT-TMZ. Median overall survival and progression-free survival times were similar: 12 months and 5.6 months for the standard RT-TMZ group and 12.5 months and 6.7 months for the short-course RT-TMZ group, respectively. Radiation schedule was associated with similar survival outcomes in either unadjusted or adjusted analysis. O{sup 6}-methylguanine-DNA methyltransferase promoter methylation was the most favorable prognostic factor (P=.0001). Standard RT-TMZ therapy was associated with a significant rise in grade 2 and 3 neurologic toxicity (P=.01), lowering of KPS scores during the study (P=.01), and higher posttreatment dosing of corticosteroid (P=.02). Conclusions: In older adults with GBM, survival outcomes of standard and short-course RT-TMZ were similar. An abbreviated course of RT plus TMZ may represent a reasonable therapeutic approach for these patients, without loss of survival benefit and acceptable toxicity.

Identification and differentiation of the twenty six bluetongue virus serotypes by RT-PCR amplification of the serotype-specific genome segment 2.

Directory of Open Access Journals (Sweden)

Narender S Maan

Full Text Available Bluetongue (BT is an arthropod-borne viral disease, which primarily affects ruminants in tropical and temperate regions of the world. Twenty six bluetongue virus (BTV serotypes have been recognised worldwide, including nine from Europe and fifteen in the United States. Identification of BTV serotype is important for vaccination programmes and for BTV epidemiology studies. Traditional typing methods (virus isolation and serum or virus neutralisation tests (SNT or VNT are slow (taking weeks, depend on availability of reference virus-strains or antisera and can be inconclusive. Nucleotide sequence analyses and phylogenetic comparisons of genome segment 2 (Seg-2 encoding BTV outer-capsid protein VP2 (the primary determinant of virus serotype were completed for reference strains of BTV-1 to 26, as well as multiple additional isolates from different geographic and temporal origins. The resulting Seg-2 database has been used to develop rapid (within 24 h and reliable RT-PCR-based typing assays for each BTV type. Multiple primer-pairs (at least three designed for each serotype were widely tested, providing an initial identification of serotype by amplification of a cDNA product of the expected size. Serotype was confirmed by sequencing of the cDNA amplicons and phylogenetic comparisons to previously characterised reference strains. The results from RT-PCR and sequencing were in perfect agreement with VNT for reference strains of all 26 BTV serotypes, as well as the field isolates tested. The serotype-specific primers showed no cross-amplification with reference strains of the remaining 25 serotypes, or multiple other isolates of the more closely related heterologous BTV types. The primers and RT-PCR assays developed in this study provide a rapid, sensitive and reliable method for the identification and differentiation of the twenty-six BTV serotypes, and will be updated periodically to maintain their relevance to current BTV distribution and

Molecular Cloning And Sequencing Of Disintegrin Like Domain ...

African Journals Online (AJOL)

Disintegrin-like domain was cloned and sequenced from Cerastes cerastes venom gland tissue. Nested RT-PCR was performed using initial primers designed based on the homology of disintegrins from Trimeresurus flavoviridis, Glodius halys , Agkistrodon halys and Trimeresurus macrosquamatus. The homology was ...

Sequence recombination and conservation of Varroa destructor virus-1 and deformed wing virus in field collected honey bees (Apis mellifera.

Directory of Open Access Journals (Sweden)

Hui Wang

Full Text Available We sequenced small (s RNAs from field collected honeybees (Apis mellifera and bumblebees (Bombuspascuorum using the Illumina technology. The sRNA reads were assembled and resulting contigs were used to search for virus homologues in GenBank. Matches with Varroadestructor virus-1 (VDV1 and Deformed wing virus (DWV genomic sequences were obtained for A. mellifera but not B. pascuorum. Further analyses suggested that the prevalent virus population was composed of VDV-1 and a chimera of 5'-DWV-VDV1-DWV-3'. The recombination junctions in the chimera genomes were confirmed by using RT-PCR, cDNA cloning and Sanger sequencing. We then focused on conserved short fragments (CSF, size > 25 nt in the virus genomes by using GenBank sequences and the deep sequencing data obtained in this study. The majority of CSF sites confirmed conservation at both between-species (GenBank sequences and within-population (dataset of this study levels. However, conserved nucleotide positions in the GenBank sequences might be variable at the within-population level. High mutation rates (Pi>10% were observed at a number of sites using the deep sequencing data, suggesting that sequence conservation might not always be maintained at the population level. Virus-host interactions and strategies for developing RNAi treatments against VDV1/DWV infections are discussed.

Outcome on EPIZONE Extension on VER/ VNN: Diagnostics, proficiency test and qRT-PCR validation

DEFF Research Database (Denmark)

Bigarré, Laurent; Panzarin, Valentina; Baud, Marine

2012-01-01

1 is to bring valuable genetic information on the second genome component of a given isolate. ANSES has developed new DNA probes targeting RNA1 with the goal of detecting all genotypes of nodaviruses. The aim of the project is thus: - To organize, conduct and report an inter-laboratory proficiency...... test for detection of aquatic nodaviruses by real time RT-PCR targeting RNA1 and RNA2, respectively. - To test a newly developed real-time RT-PCR targeting RNA1: sensitivity, specificity, range of detection and genetic information provided by sequencing the PCR product. Materials & methods Primers...... to be extracted by each partner. In the meantime, ANSES produced and distributed RNA extracted from healthy or infected fish (2 genogroups), or cell culture; one sample from cell culture had to be serially diluted to test the sensitivity of each method in partners’ hands. Samples were tested in duplicates...

Comparative evaluation of conventional RT-PCR and real-time RT-PCR (RRT-PCR) for detection of avian metapneumovirus subtype A

OpenAIRE

Ferreira, HL; Spilki, FR; dos Santos, MMAB; de Almeida, RS; Arns, CW

2009-01-01

Avian metapneumovirus (AMPV) belongs to Metapneumovirus genus of Paramyxoviridae family. Virus isolation, serology, and detection of genomic RNA are used as diagnostic methods for AMPV. The aim of the present study was to compare the detection of six subgroup A AMPV isolates (AMPV/A) viral RNA by using different conventional and real time RT-PCR methods. Two new RT-PCR tests and two real time RT-PCR tests, both detecting fusion (F) gene and nucleocapsid (N) gene were compared with an establis...

Yersinia enterocolitica-Specific Infection by Bacteriophages TG1 and ϕR1-RT Is Dependent on Temperature-Regulated Expression of the Phage Host Receptor OmpF.

Science.gov (United States)

Leon-Velarde, Carlos G; Happonen, Lotta; Pajunen, Maria; Leskinen, Katarzyna; Kropinski, Andrew M; Mattinen, Laura; Rajtor, Monika; Zur, Joanna; Smith, Darren; Chen, Shu; Nawaz, Ayesha; Johnson, Roger P; Odumeru, Joseph A; Griffiths, Mansel W; Skurnik, Mikael

2016-09-01

Bacteriophages present huge potential both as a resource for developing novel tools for bacterial diagnostics and for use in phage therapy. This potential is also valid for bacteriophages specific for Yersinia enterocolitica To increase our knowledge of Y. enterocolitica-specific phages, we characterized two novel yersiniophages. The genomes of the bacteriophages vB_YenM_TG1 (TG1) and vB_YenM_ϕR1-RT (ϕR1-RT), isolated from pig manure in Canada and from sewage in Finland, consist of linear double-stranded DNA of 162,101 and 168,809 bp, respectively. Their genomes comprise 262 putative coding sequences and 4 tRNA genes and share 91% overall nucleotide identity. Based on phylogenetic analyses of their whole-genome sequences and large terminase subunit protein sequences, a genus named Tg1virus within the family Myoviridae is proposed, with TG1 and ϕR1-RT (R1RT in the ICTV database) as member species. These bacteriophages exhibit a host range restricted to Y. enterocolitica and display lytic activity against the epidemiologically significant serotypes O:3, O:5,27, and O:9 at and below 25°C. Adsorption analyses of lipopolysaccharide (LPS) and OmpF mutants demonstrate that these phages use both the LPS inner core heptosyl residues and the outer membrane protein OmpF as phage receptors. Based on RNA sequencing and quantitative proteomics, we also demonstrate that temperature-dependent infection is due to strong repression of OmpF at 37°C. In addition, ϕR1-RT was shown to be able to enter into a pseudolysogenic state. Together, this work provides further insight into phage-host cell interactions by highlighting the importance of understanding underlying factors which may affect the abundance of phage host receptors on the cell surface. Only a small number of bacteriophages infecting Y. enterocolitica, the predominant causative agent of yersiniosis, have been previously described. Here, two newly isolated Y. enterocolitica phages were studied in detail, with the aim of

The genomic sequence of cowpea aphid-borne mosaic virus and its similarities with other potyviruses

NARCIS (Netherlands)

Mlotshwa, S.; Verver, J.; Sithole-Niang, I.; Kampen, van T.; Kammen, van A.; Wellink, J.

2002-01-01

The genomic sequence of a Zimbabwe isolate of Cowpea aphid-borne mosaic virus (CABMV-Z) was determined by sequencing overlapping viral cDNA clones generated by RT-PCR using degenerate and/or specific primers. The sequence is 9465 nucleotides in length excluding the 3' terminal poly (A) tail and

Efficient and controllable thermal ablation induced by short-pulsed HIFU sequence assisted with perfluorohexane nanodroplets.

Science.gov (United States)

Chang, Nan; Lu, Shukuan; Qin, Dui; Xu, Tianqi; Han, Meng; Wang, Supin; Wan, Mingxi

2018-07-01

A HIFU sequence with extremely short pulse duration and high pulse repetition frequency can achieve thermal ablation at a low acoustic power using inertial cavitation. Because of its cavitation-dependent property, the therapeutic outcome is unreliable when the treatment zone lacks cavitation nuclei. To overcome this intrinsic limitation, we introduced perfluorocarbon nanodroplets as extra cavitation nuclei into short-pulsed HIFU-mediated thermal ablation. Two types of nanodroplets were used with perfluorohexane (PFH) as the core material coated with bovine serum albumin (BSA) or an anionic fluorosurfactant (FS) to demonstrate the feasibility of this study. The thermal ablation process was recorded by high-speed photography. The inertial cavitation activity during the ablation was revealed by sonoluminescence (SL). The high-speed photography results show that the thermal ablation volume increased by ∼643% and 596% with BSA-PFH and FS-PFH, respectively, than the short-pulsed HIFU alone at an acoustic power of 19.5 W. Using nanodroplets, much larger ablation volumes were created even at a much lower acoustic power. Meanwhile, the treatment time for ablating a desired volume significantly reduced in the presence of nanodroplets. Moreover, by adjusting the treatment time, lesion migration towards the HIFU transducer could also be avoided. The SL results show that the thermal lesion shape was significantly dependent on the inertial cavitation in this short-pulsed HIFU-mediated thermal ablation. The inertial cavitation activity became more predictable by using nanodroplets. Therefore, the introduction of PFH nanodroplets as extra cavitation nuclei made the short-pulsed HIFU thermal ablation more efficient by increasing the ablation volume and speed, and more controllable by reducing the acoustic power and preventing lesion migration. Copyright © 2018. Published by Elsevier B.V.

Engineering of a DNA Polymerase for Direct m6 A Sequencing.

Science.gov (United States)

Aschenbrenner, Joos; Werner, Stephan; Marchand, Virginie; Adam, Martina; Motorin, Yuri; Helm, Mark; Marx, Andreas

2018-01-08

Methods for the detection of RNA modifications are of fundamental importance for advancing epitranscriptomics. N 6 -methyladenosine (m 6 A) is the most abundant RNA modification in mammalian mRNA and is involved in the regulation of gene expression. Current detection techniques are laborious and rely on antibody-based enrichment of m 6 A-containing RNA prior to sequencing, since m 6 A modifications are generally "erased" during reverse transcription (RT). To overcome the drawbacks associated with indirect detection, we aimed to generate novel DNA polymerase variants for direct m 6 A sequencing. Therefore, we developed a screen to evolve an RT-active KlenTaq DNA polymerase variant that sets a mark for N 6 -methylation. We identified a mutant that exhibits increased misincorporation opposite m 6 A compared to unmodified A. Application of the generated DNA polymerase in next-generation sequencing allowed the identification of m 6 A sites directly from the sequencing data of untreated RNA samples. © 2017 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA.

Tantsuime: märjaks nutetud Märt Agu saab 8000 uut sõpra / Märt Agu ; intervjueerinud Rein Sikk

Index Scriptorium Estoniae

Agu, Märt, 1980-

2011-01-01

11. Noorte tantsupeo "Maa ja ilm" üldjuht ja Tallinna Tantsuakadeemia kunstiline juht Märt Agu Tallinna lauluväljakul toimuvast tantsupeost, ülevaatustest, eesti meeste rahvatantsust, oma isast Mait Agust, tema tantsust "Põhjamaa", tantsuõpetaja elukutsest, eesti tantsust, Tallinna Tantsuakadeemiast. Andmeid Märt Agu loomingu kohta

Analytical and clinical performance of the CDC real time RT-PCR assay for detection and typing of dengue virus.

Directory of Open Access Journals (Sweden)

Gilberto A Santiago

Full Text Available Dengue is an acute illness caused by the positive-strand RNA dengue virus (DENV. There are four genetically distinct DENVs (DENV-1-4 that cause disease in tropical and subtropical countries. Most patients are viremic when they present with symptoms; therefore, RT-PCR has been increasingly used in dengue diagnosis. The CDC DENV-1-4 RT-PCR Assay has been developed as an in-vitro diagnostic platform and was recently approved by the US Food and Drug Administration (FDA for detection of dengue in patients with signs or symptoms of mild or severe dengue. The primers and probes of this test have been designed to detect currently circulating strains of DENV-1-4 from around the world at comparable sensitivity. In a retrospective study with 102 dengue cases confirmed by IgM anti-DENV seroconversion in the convalescent sample, the RT-PCR Assay detected DENV RNA in 98.04% of the paired acute samples. Using sequencing as a positive indicator, the RT-PCR Assay had a 97.92% positive agreement in 86 suspected dengue patients with a single acute serum sample. After extensive validations, the RT-PCR Assay performance was highly reproducible when evaluated across three independent testing sites, did not produce false positive results for etiologic agents of other febrile illnesses, and was not affected by pathological levels of potentially interfering biomolecules. These results indicate that the CDC DENV-1-4 RT-PCR Assay provides a reliable diagnostic platform capable for confirming dengue in suspected cases.

Development and evaluation of tailored specific real-time RT-PCR assays for detection of foot-and-mouth disease virus serotypes circulating in East Africa.

Science.gov (United States)

Bachanek-Bankowska, Katarzyna; Mero, Herieth R; Wadsworth, Jemma; Mioulet, Valerie; Sallu, Raphael; Belsham, Graham J; Kasanga, Christopher J; Knowles, Nick J; King, Donald P

2016-11-01

Rapid, reliable and accurate diagnostic methods provide essential support to programmes that monitor and control foot-and-mouth disease (FMD). While pan-specific molecular tests for FMD virus (FMDV) detection are well established and widely used in endemic and FMD-free countries, current serotyping methods mainly rely either on antigen detection ELISAs or nucleotide sequencing approaches. This report describes the development of a panel of serotype-specific real-time RT-PCR assays (rRT-PCR) tailored to detect FMDV lineages currently circulating in East Africa. These assays target sequences within the VP1-coding region that share high intra-lineage identity, but do not cross-react with FMD viruses from other serotypes that circulate in the region. These serotype-specific assays operate with the same thermal profile as the pan-diagnostic tests making it possible to run them in parallel to produce C T values comparable to the pan-diagnostic test detecting the 3D-coding region. These assays were evaluated alongside the established pan-specific molecular test using field samples and virus isolates collected from Tanzania, Kenya and Ethiopia that had been previously characterised by nucleotide sequencing. Samples (n=71) representing serotype A (topotype AFRICA, lineage G-I), serotype O (topotypes EA-2 and EA-4), serotype SAT 1 (topotype I (NWZ)) and serotype SAT2 (topotype IV) were correctly identified with these rRT-PCR assays. Furthermore, FMDV RNA from samples that did not contain infectious virus could still be serotyped using these assays. These serotype-specific real-time RT-PCR assays can detect and characterise FMDVs currently circulating in East Africa and hence improve disease control in this region. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

The use of short and long PCR products for improved detection of prunus necrotic ringspot virus in woody plants.

Science.gov (United States)

Rosner, A; Maslenin, L; Spiegel, S

1997-09-01

The reverse transcriptase-polymerase chain reaction (RT-PCR) was used for detection of prunus necrotic ringspot virus (PNRSV) in dormant peach and almond trees by the application of two different pairs of primers yielding a short and a long product, respectively. The relative amount of the short (200 base pair, bp) product was higher than the longer (785 bp) product. PNRSV was detected better in plant tissues with a low virus concentration (e.g. dormant trees) by amplification of the short PCR product, whereas the long product was product was produced at higher virus titers. Simultaneous amplification of both short and long products was demonstrated using a three-primer mixture in a single reaction tube. In this assay, amplification of either PCR product indicated the presence of PNRSV-specific sequences in the plant tissue examined, thus covering a wide range of virus concentrations in a single test. Dilution of the RNA extracted from infected plant material resulted in a steep decline in the amplification of both short and long PCR products. In contrast, serial dilutions of the intermediate cDNA template differentially affected the amplification patterns: the relative amount of the short product increased whereas that of the long product decreased. These results may explain the preferential amplification of the short PCR product observed in samples containing low virus concentrations.

Thrombolysis with Intravenous Tissue Plasminogen Activator (rt-PA) Predicts Favorable Discharge Disposition in Patients with Acute Ischemic Stroke

Science.gov (United States)

Ifejika-Jones, Nneka L.; Harun, Nusrat; Mohammed-Rajput, Nareesa A.; Noser, Elizabeth A.; Grotta, James C.

2011-01-01

Background and Purpose Acute ischemic stroke patients receiving IV tissue plasminogen activator (rt-PA) within 3 hours of symptom onset are 30% more likely to have minimal disability at three months. During hospitalization, short-term disability is subjectively measured by discharge disposition, whether to home, Inpatient Rehabilitation (IR), Skilled Nursing Facility (SNF) or Subacute Care (Sub). There are no studies assessing the role of rt-PA use as a predictor of post-stroke disposition. Methods Retrospective analysis of all ischemic stroke patients admitted to the University of Texas Houston Medical School (UTHMS) Stroke Service between Jan 2004 and Oct 2009. Baseline demographics and National Institute of Health Stroke Scale (NIHSS) score were collected. Cerebrovascular disease risk factors were used for risk stratification. Results Home vs. IR, SNF, Sub Of 2225 acute ischemic stroke patients, 1019 were discharged home, 1206 to another level of care. Patients who received rt-PA therapy were 1.9 times more likely to be discharged home (P = stroke patients, 719 patients were discharged to acute IR, 371 were discharged to SNF, 116 to Sub. There were no differences in disposition between patients who received rt-PA therapy. Conclusions Stroke patients who receive IV rt-PA for acute ischemic stroke are more 1.9 times more likely to be discharged directly home after hospitalization. This study is limited by its retrospective nature and the undetermined role of psychosocial factors related to discharge. PMID:21293014

A duplex real-time RT-PCR assay for detecting H5N1 avian influenza virus and pandemic H1N1 influenza virus

Directory of Open Access Journals (Sweden)

Qin E-de

2010-06-01

Full Text Available Abstract A duplex real-time reverse transcriptase polymerase chain reaction (RT-PCR assay was improved for simultaneous detection of highly pathogenic H5N1 avian influenza virus and pandemic H1N1 (2009 influenza virus, which is suitable for early diagnosis of influenza-like patients and for epidemiological surveillance. The sensitivity of this duplex real-time RT-PCR assay was 0.02 TCID50 (50% tissue culture infective dose for H5N1 and 0.2 TCID50 for the pandemic H1N1, which was the same as that of each single-target RT-PCR for pandemic H1N1 and even more sensitive for H5N1 with the same primers and probes. No cross reactivity of detecting other subtype influenza viruses or respiratory tract viruses was observed. Two hundred and thirty-six clinical specimens were tested by comparing with single real-time RT-PCR and result from the duplex assay was 100% consistent with the results of single real-time RT-PCR and sequence analysis.

NASA SPoRT Initialization Datasets for Local Model Runs in the Environmental Modeling System

Science.gov (United States)

Case, Jonathan L.; LaFontaine, Frank J.; Molthan, Andrew L.; Carcione, Brian; Wood, Lance; Maloney, Joseph; Estupinan, Jeral; Medlin, Jeffrey M.; Blottman, Peter; Rozumalski, Robert A.

2011-01-01

The NASA Short-term Prediction Research and Transition (SPoRT) Center has developed several products for its National Weather Service (NWS) partners that can be used to initialize local model runs within the Weather Research and Forecasting (WRF) Environmental Modeling System (EMS). These real-time datasets consist of surface-based information updated at least once per day, and produced in a composite or gridded product that is easily incorporated into the WRF EMS. The primary goal for making these NASA datasets available to the WRF EMS community is to provide timely and high-quality information at a spatial resolution comparable to that used in the local model configurations (i.e., convection-allowing scales). The current suite of SPoRT products supported in the WRF EMS include a Sea Surface Temperature (SST) composite, a Great Lakes sea-ice extent, a Greenness Vegetation Fraction (GVF) composite, and Land Information System (LIS) gridded output. The SPoRT SST composite is a blend of primarily the Moderate Resolution Imaging Spectroradiometer (MODIS) infrared and Advanced Microwave Scanning Radiometer for Earth Observing System data for non-precipitation coverage over the oceans at 2-km resolution. The composite includes a special lake surface temperature analysis over the Great Lakes using contributions from the Remote Sensing Systems temperature data. The Great Lakes Environmental Research Laboratory Ice Percentage product is used to create a sea-ice mask in the SPoRT SST composite. The sea-ice mask is produced daily (in-season) at 1.8-km resolution and identifies ice percentage from 0 100% in 10% increments, with values above 90% flagged as ice.

Development of a one-step RT-PCR assay for detection of pancoronaviruses (α-, β-, γ-, and δ-coronaviruses) using newly designed degenerate primers for porcine and avian `fecal samples.

Science.gov (United States)

Hu, Hui; Jung, Kwonil; Wang, Qiuhong; Saif, Linda J; Vlasova, Anastasia N

2018-06-01

Coronaviruses (CoVs) are critical human and animal pathogens because of their potential to cause severe epidemics of respiratory or enteric diseases. In pigs, the newly emerged porcine deltacoronavirus (PDCoV) and re-emerged porcine epidemic diarrhea virus (PEDV) reported in the US and Asia, as well as the discovery of novel CoVs in wild bats or birds, has necessitated development of improved detection and control measures for these CoVs. Because the previous pancoronavirus (panCoV) RT-PCR established in our laboratory in 2007-2011 did not detect deltacoronaviruses (δ-CoVs) in swine fecal and serum samples, our goal was to develop a new panCoV RT-PCR assay to detect known human and animal CoVs, including δ-CoVs. In this study, we designed a new primer set to amplify a 668 bp-region within the RNA-dependent RNA polymerase (RdRP) gene that encodes the most conserved protein domain of α-, β-, γ-, and δ-CoVs. We established a one-step panCoV RT-PCR assay and standardized the assay conditions. The newly established panCoV RT-PCR assay was demonstrated to have a high sensitivity and specificity. Using a panel of 60 swine biological samples (feces, intestinal contents, and sera) characterized by PEDV, PDCoV and transmissible gastroenteritis virus-specific RT-PCR assays, we demonstrated that sensitivity and specificity of the newly established panCoV RT-PCR assay were 100%. 400 avian fecal (RNA) samples were further tested simultaneously for CoV by the new panCoV RT-PCR and a one-step RT-PCR assay with the δ-CoV nucleocapsid-specific universal primers. Four of 400 avian samples were positive for CoV, three of which were positive for δ-CoV by the conventional RT-PCR. PanCoV RT-PCR fragments for 3 of the 4 CoVs were sequenced. Phylogenetic analysis revealed the presence of one γ-CoV and two δ-CoV in the sequenced samples. The newly designed panCoV RT-PCR assay should be useful for the detection of currently known CoVs in animal biological samples. Copyright © 2018

Reverse transcriptase-quantitative polymerase chain reaction (RT ...

African Journals Online (AJOL)

The reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) is a highly specific polymerase chain reaction (PCR) method that allows one to detect very low transcription levels of functional gene(s) in soil. RT-qPCR helps us to know the active members of the microbial community, and their activities can be ...

Development and evaluation of novel one-step TaqMan realtime RT-PCR assays for the detection and direct genotyping of genogroup I and II noroviruses

DEFF Research Database (Denmark)

Schultz, Anna Charlotte; Vega, Everado; Dalsgaard, Anders

2011-01-01

BackgroundCurrent detection and genotyping methods of genogroup (G) I and II noroviruses (NoVs) consist of a 2-step approach including detection of viral RNA by TaqMan realtime RT-PCR (RT-qPCR) followed by conventional RT-PCR and sequencing of partial regions of ORF1 or ORF2. ObjectiveTo develop ......Man RT-qPCR assays for the sensitive detection and direct genotyping of GI and GII NoVs from clinical and environmental matrices...... novel long-template one-step TaqMan assays (L-RT-qPCR) for the rapid detection and direct genotyping of GI and GII NoVs and to evaluate the sensitivity and specificity of the assays. Study designGI and GII-specific broadly reactive L-RT-qPCR assays were developed by combining existing NoV primers...... and probes targeting the open reading frame (ORF)1–ORF2 junction as well as region C at the 5′–ORF2. The assays were validated using GI and GII RNA transcripts and a coded panel of 75 stool samples containing NoV strains representing 9 GI genotypes and 12 GII genotypes, as well as sapoviruses, astroviruses...

A safe an easy method for building consensus HIV sequences from 454 massively parallel sequencing data.

Science.gov (United States)

Fernández-Caballero Rico, Jose Ángel; Chueca Porcuna, Natalia; Álvarez Estévez, Marta; Mosquera Gutiérrez, María Del Mar; Marcos Maeso, María Ángeles; García, Federico

2018-02-01

To show how to generate a consensus sequence from the information of massive parallel sequences data obtained from routine HIV anti-retroviral resistance studies, and that may be suitable for molecular epidemiology studies. Paired Sanger (Trugene-Siemens) and next-generation sequencing (NGS) (454 GSJunior-Roche) HIV RT and protease sequences from 62 patients were studied. NGS consensus sequences were generated using Mesquite, using 10%, 15%, and 20% thresholds. Molecular evolutionary genetics analysis (MEGA) was used for phylogenetic studies. At a 10% threshold, NGS-Sanger sequences from 17/62 patients were phylogenetically related, with a median bootstrap-value of 88% (IQR83.5-95.5). Association increased to 36/62 sequences, median bootstrap 94% (IQR85.5-98)], using a 15% threshold. Maximum association was at the 20% threshold, with 61/62 sequences associated, and a median bootstrap value of 99% (IQR98-100). A safe method is presented to generate consensus sequences from HIV-NGS data at 20% threshold, which will prove useful for molecular epidemiological studies. Copyright © 2016 Elsevier España, S.L.U. and Sociedad Española de Enfermedades Infecciosas y Microbiología Clínica. All rights reserved.

Experiment vs simulation RT WFNDEC 2014 benchmark: CIVA results

International Nuclear Information System (INIS)

Tisseur, D.; Costin, M.; Rattoni, B.; Vienne, C.; Vabre, A.; Cattiaux, G.; Sollier, T.

2015-01-01

The French Atomic Energy Commission and Alternative Energies (CEA) has developed for years the CIVA software dedicated to simulation of NDE techniques such as Radiographic Testing (RT). RT modelling is achieved in CIVA using combination of a determinist approach based on ray tracing for transmission beam simulation and a Monte Carlo model for the scattered beam computation. Furthermore, CIVA includes various detectors models, in particular common x-ray films and a photostimulable phosphor plates. This communication presents the results obtained with the configurations proposed in the World Federation of NDEC 2014 RT modelling benchmark with the RT models implemented in the CIVA software

Experiment vs simulation RT WFNDEC 2014 benchmark: CIVA results

Energy Technology Data Exchange (ETDEWEB)

Tisseur, D., E-mail: david.tisseur@cea.fr; Costin, M., E-mail: david.tisseur@cea.fr; Rattoni, B., E-mail: david.tisseur@cea.fr; Vienne, C., E-mail: david.tisseur@cea.fr; Vabre, A., E-mail: david.tisseur@cea.fr; Cattiaux, G., E-mail: david.tisseur@cea.fr [CEA LIST, CEA Saclay 91191 Gif sur Yvette Cedex (France); Sollier, T. [Institut de Radioprotection et de Sûreté Nucléaire, B.P.17 92262 Fontenay-Aux-Roses (France)

2015-03-31

The French Atomic Energy Commission and Alternative Energies (CEA) has developed for years the CIVA software dedicated to simulation of NDE techniques such as Radiographic Testing (RT). RT modelling is achieved in CIVA using combination of a determinist approach based on ray tracing for transmission beam simulation and a Monte Carlo model for the scattered beam computation. Furthermore, CIVA includes various detectors models, in particular common x-ray films and a photostimulable phosphor plates. This communication presents the results obtained with the configurations proposed in the World Federation of NDEC 2014 RT modelling benchmark with the RT models implemented in the CIVA software.

A putative peroxidase cDNA from turnip and analysis of the encoded protein sequence.

Science.gov (United States)

Romero-Gómez, S; Duarte-Vázquez, M A; García-Almendárez, B E; Mayorga-Martínez, L; Cervantes-Avilés, O; Regalado, C

2008-12-01

A putative peroxidase cDNA was isolated from turnip roots (Brassica napus L. var. purple top white globe) by reverse transcriptase-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE). Total RNA extracted from mature turnip roots was used as a template for RT-PCR, using a degenerated primer designed to amplify the highly conserved distal motif of plant peroxidases. The resulting partial sequence was used to design the rest of the specific primers for 5' and 3' RACE. Two cDNA fragments were purified, sequenced, and aligned with the partial sequence from RT-PCR, and a complete overlapping sequence was obtained and labeled as BbPA (Genbank Accession No. AY423440, named as podC). The full length cDNA is 1167bp long and contains a 1077bp open reading frame (ORF) encoding a 358 deduced amino acid peroxidase polypeptide. The putative peroxidase (BnPA) showed a calculated Mr of 34kDa, and isoelectric point (pI) of 4.5, with no significant identity with other reported turnip peroxidases. Sequence alignment showed that only three peroxidases have a significant identity with BnPA namely AtP29a (84%), and AtPA2 (81%) from Arabidopsis thaliana, and HRPA2 (82%) from horseradish (Armoracia rusticana). Work is in progress to clone this gene into an adequate host to study the specific role and possible biotechnological applications of this alternative peroxidase source.

Power lies in interdisciplinary cooperation / Kärt Summatavet ; interv. Kärt Blumberg

Index Scriptorium Estoniae

Summatavet, Kärt, 1963-

2008-01-01

Ehtekunstnik Kärt Summatavet oma uudsest meetodist ehete valmistamisel - paberil oleva kujutise ülekandmisest arvuti abil metallile, mille ta töötas väljas oma doktoritöö raames Soomes Helsingi Kunsti- ja Disainiülikoolis ning mis pälvis Soulis toimunud ülemaailmsel naisleiutajate konkursil hõbemedali

Using the SPoRT POES/GOES Hybrid Product in OCONUS Forecasting

Science.gov (United States)

Smith, Matt; Fuell, Kevin; Nelson, Jim

2014-01-01

The SPoRT (Short-term Prediction and Research Transition) Program at the NASA/Marshall Space Flight Center has been providing unique NASA and NOAA data and techniques to partner Weather Forecast Offices (WFOs) for ten years. Data are provided in the Decision Support System used by WFO forecasters: AWIPS. For the last couple of years, SPoRT has been producing the POES/GOES Hybrid. This suite of products combines the strength ofl5- minute animations of GOES imagery - providing temporal continuity, with the higher resolution, relatively random availability, of polar orbiting (POES) imagery data. The product was first introduced with only MODIS data from NASA's Terra and Aqua satellites, but recently the VIIRS instrument onboard the Suomi-NPP satellite was added, providing better high-resolution coverage. These products represent SPoRT's efforts to prepare for higher resolution, higher frequency GOES-R imagery - as well as helping to move VIIRS (JPSS) data into the mainstream of weather forecasting. SPoRT generates 5 products for this dataset: Visible, Longwave Infrared (11 micrometers), Shortwave IR (3.7 micrometers), Water Vapor (6.7 micrometers), and Fog (Difference of 11 micrometer and 3.7 micrometer channels). The Water Vapor hybrid product has a Red-Blue-Green image from MODIS inlaid, since it provides even more qualitative information than water vapor alone. Animated examples of the products will be shown in this presentation. While the resolution at nadir of GOES imagery is nominally Han (4km for IR channels), the inlaid polar orbiter imagery has a resolution of 250m (lkm for IR channels). This has tremendous application in the continental US. However, in high latitudes, since the usefulness of GOES degrades poleward rapidly, the contrast of GOES and POES data is stark. The consistent temporal nature of GOES, even though at a reduced resolution at high latitudes, provides basic situational awareness, but the introduction of polar data is very helpful in seeing

Breath-hold technique in conventional APPA or intensity-modulated radiotherapy for Hodgkin's lymphoma. Comparison of ILROG IS-RT and the GHSG IF-RT

Energy Technology Data Exchange (ETDEWEB)

Kriz, Jan; Spickermann, Max; Lehrich, Philipp; Reinartz, Gabriele; Eich, Hans; Haverkamp, Uwe [University of Muenster, Department of Radiation Oncology, Muenster (Germany); Schmidberger, Heinz [University Mainz, Department of Radiation Oncology, Mainz (Germany)

2015-09-15

The present study addresses the role of intensity-modulated radiotherapy (IMRT) in contrast to standard RT (APPA) for patients with Hodgkin's lymphoma (HL) with a focus on deep inspiration breath-hold (DIBH) technique and a comparison between the International Lymphoma Radiation Oncology Group (ILROG) Involved Site Radiotherapy (IS-RT) versus the German Hodgkin Study Group (GHSG) Involved Field Radiotherapy (IF-RT). APPA treatment and 2 IMRT plans were compared for 11 patients with HL. Furthermore, treatment with DIBH versus free breathing (FB) and two different treatment volumes, i.e. IF-RT versus IS-RT, were compared. IMRT was planned as a sliding-window technique with 5 and 7 beam angles. For each patient 12 different treatment plans were calculated (132 plans). Following organs at risk (OAR) were analysed: lung, heart, spinal cord, oesophagus, female breast and skin. Comparisons of the different values with regard to dose-volume histograms (DVH), conformity and homogeneity indices were made. IS-RT reduces treatment volumes. With respect to the planning target volume (PTV), IMRT achieves better conformity but the same homogeneity. Regarding the D{sub mean} for the lung, IMRT shows increased doses, while RT in DIBH reduces doses. The IMRT shows improved values for D{sub max} concerning the spinal cord, whereas the APPA shows an improved D{sub mean} of the lung and the female breast. IS-RT reduces treatment volumes. Intensity-modulated radiotherapy shows advantages in the conformity. Treatment in DIBH also reduces the dose applied to the lungs and the heart. (orig.) [German] Ziel dieser Auswertung ist es, die konventionelle APPA-Feldanordnung mit der Intensitaetsmodulierten Radiotherapie (IMRT) bei Patienten mit Hodgkin-Lymphom (HL) zu vergleichen. Ein besonderer Fokus liegt hierbei auf der Bestrahlung in tiefer Inspiration und Atemanhaltetechnik (DIBH). Des Weiteren wurde die ''Involved-site''-Radiotherapie (IS-RT) der International

RT-PCR Protocols - Methods in Molecular Biology

Directory of Open Access Journals (Sweden)

Manuela Monti

2011-03-01

Full Text Available “The first record I have of it, is when I made a computer file which I usually did whenever I had an idea, that would have been on the Monday when I got back, and I called it Chain Reaction.POL, meaning polymerase. That was the identifier for it and later I called the thing the Polymerase Chain Reaction, which a lot of people thought was a dumb name for it, but it stuck, and it became PCR”. With these words the Nobel prize winner, Kary Mullis, explains how he named the PCR: one of the most important techniques ever invented and currently used in molecular biology. This book “RT-PCR Protocols” covers a wide range of aspects important for the setting of a PCR experiment for both beginners and advanced users. In my opinion the book is very well structured in three different sections. The first one describes the different technologies now available, like competitive RT-PCR, nested RT-PCR or RT-PCR for cloning. An important part regards the usage of PCR in single cell mouse embryos, stressing how important...........

Detection of enteroviruses and hepatitis a virus in water by consensus primer multiplex RT-PCR

Science.gov (United States)

Li, Jun-Wen; Wang, Xin-Wei; Yuan, Chang-Qing; Zheng, Jin-Lai; Jin, Min; Song, Nong; Shi, Xiu-Quan; Chao, Fu-Huan

2002-01-01

AIM: To develop a rapid detection method of enteroviruses and Hepatitis A virus (HAV). METHODS: A one-step, single-tube consensus primers multiplex RT-PCR was developed to simultaneously detect Poliovirus, Coxsackie virus, Echovirus and HAV. A general upstream primer and a HAV primer and four different sets of primers (5 primers) specific for Poliovirus, Coxsacki evirus, Echovirus and HAV cDNA were mixed in the PCR mixture to reverse transcript and amplify the target DNA. Four distinct amplified DNA segments representing Poliovirus, Coxsackie virus, Echovirus and HAV were identified by gel electrophoresis as 589-, 671-, 1084-, and 1128 bp sequences, respectively. Semi-nested PCR was used to confirm the amplified products for each enterovirus and HAV. RESULTS: All four kinds of viral genome RNA were detected, and producing four bands which could be differentiated by the band size on the gel. To confirm the specificity of the multiplex PCR products, semi-nested PCR was performed. For all the four strains tested gave positive results. The detection sensitivity of multiplex PCR was similar to that of monoplex RT-PCR which was 24 PFU for Poliovrus, 21 PFU for Coxsackie virus, 60 PFU for Echovirus and 105 TCID50 for HAV. The minimum amount of enteric viral RNA detected by semi-nested PCR was equivalent to 2.4 PFU for Poliovrus, 2.1 PFU for Coxsackie virus, 6.0 PFU for Echovirus and 10.5 TCID50 for HAV. CONCLUSION: The consensus primers multiplex RT-PCR has more advantages over monoplex RT-PCR for enteric viruses detection, namely, the rapid turnaround time and cost effectiveness. PMID:12174381

MINIX4RT: Real-Time Semaphores

OpenAIRE

Pessolani, Pablo Andrés

2007-01-01

MINIX4RT es una extensión del conocido Sistema Operativo MINIX que incorpora servicios de Tiempo Real Estricto en un nuevo microkernel pero manteniendo compatibilidad con las versiones anteriores del MINIX estándar. Los semáforos son el mecanismo primitivo para la sincronización y exclusion mutua en varios sistemas operativos, pero MINIX no brinda esa facilidad. Se adicionaron semáforos a MINIX4RT y, como éste es un Sistema Operativo de Tiempo Real, deben reunir ciertos requisitos de procesam...

Evaluating the Impacts of NASA/SPoRT Daily Greenness Vegetation Fraction on Land Surface Model and Numerical Weather Forecasts

Science.gov (United States)

Bell, Jordan R.; Case, Jonathan L.; Molthan, Andrew L.

2011-01-01

The NASA Short-term Prediction Research and Transition (SPoRT) Center develops new products and techniques that can be used in operational meteorology. The majority of these products are derived from NASA polar-orbiting satellite imagery from the Earth Observing System (EOS) platforms. One such product is a Greenness Vegetation Fraction (GVF) dataset, which is produced from Moderate Resolution Imaging Spectroradiometer (MODIS) data aboard the NASA EOS Aqua and Terra satellites. NASA SPoRT began generating daily real-time GVF composites at 1-km resolution over the Continental United States (CONUS) on 1 June 2010. The purpose of this study is to compare the National Centers for Environmental Prediction (NCEP) climatology GVF product (currently used in operational weather models) to the SPoRT-MODIS GVF during June to October 2010. The NASA Land Information System (LIS) was employed to study the impacts of the new SPoRT-MODIS GVF dataset on land surface models apart from a full numerical weather prediction (NWP) model. For the 2010 warm season, the SPoRT GVF in the western portion of the CONUS was generally higher than the NCEP climatology. The eastern CONUS GVF had variations both above and below the climatology during the period of study. These variations in GVF led to direct impacts on the rates of heating and evaporation from the land surface. The second phase of the project is to examine the impacts of the SPoRT GVF dataset on NWP using the Weather Research and Forecasting (WRF) model. Two separate WRF model simulations were made for individual severe weather case days using the NCEP GVF (control) and SPoRT GVF (experimental), with all other model parameters remaining the same. Based on the sensitivity results in these case studies, regions with higher GVF in the SPoRT model runs had higher evapotranspiration and lower direct surface heating, which typically resulted in lower (higher) predicted 2-m temperatures (2-m dewpoint temperatures). The opposite was true

A novel RT-PCR for the detection of Helicobacter pylori and identification of clarithromycin resistance mediated by mutations in the 23S rRNA gene.

Science.gov (United States)

Redondo, Javier Jareño; Keller, Peter M; Zbinden, Reinhard; Wagner, Karoline

2018-01-01

In this study we evaluated the commercially available LightMix® RT-PCR assay for Helicobacter pylori detection and identification of clarithromycin (CLR) resistance in culture and clinical specimens (gastric biopsies and stool). The H. pylori LightMix® RT-PCR detects a 97bp long fragment of the 23S rRNA gene and allows the identification of 3 distinct point mutations conferring CLR resistance via melting curve analysis. The performance of the H. pylori LightMix® RT-PCR was evaluated using a set of 60 H. pylori strains showing phenotypical CLR susceptibility or CLR resistance (Minimum inhibitory concentrations from 0.016 to 256mg/L). We found high concordance (95%) between phenotypical CLR resistance screening by E-Test® and the Lightmix® RT-PCR. Discrepant results were verified by sequencing of the 23S rRNA gene that always confirmed the results obtained by Lightmix® RT-PCR. Furthermore, H. pylori was detected in clinical biopsy and stool specimens by Lightmix® RT-PCR that identified the correct H. pylori genotype. The LightMix® RT-PCR is an accurate, sensitive and easy to use test for H. pylori and CLR resistance detection and can therefore be readily implemented in any diagnostic laboratory. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Method and apparatus for biological sequence comparison

Science.gov (United States)

Marr, T.G.; Chang, W.I.

1997-12-23

A method and apparatus are disclosed for comparing biological sequences from a known source of sequences, with a subject (query) sequence. The apparatus takes as input a set of target similarity levels (such as evolutionary distances in units of PAM), and finds all fragments of known sequences that are similar to the subject sequence at each target similarity level, and are long enough to be statistically significant. The invention device filters out fragments from the known sequences that are too short, or have a lower average similarity to the subject sequence than is required by each target similarity level. The subject sequence is then compared only to the remaining known sequences to find the best matches. The filtering member divides the subject sequence into overlapping blocks, each block being sufficiently large to contain a minimum-length alignment from a known sequence. For each block, the filter member compares the block with every possible short fragment in the known sequences and determines a best match for each comparison. The determined set of short fragment best matches for the block provide an upper threshold on alignment values. Regions of a certain length from the known sequences that have a mean alignment value upper threshold greater than a target unit score are concatenated to form a union. The current block is compared to the union and provides an indication of best local alignment with the subject sequence. 5 figs.

Detection of short repeated genomic sequences on metaphase chromosomes using padlock probes and target primed rolling circle DNA synthesis

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Stougaard Magnus

2007-11-01

Full Text Available Abstract Background In situ detection of short sequence elements in genomic DNA requires short probes with high molecular resolution and powerful specific signal amplification. Padlock probes can differentiate single base variations. Ligated padlock probes can be amplified in situ by rolling circle DNA synthesis and detected by fluorescence microscopy, thus enhancing PRINS type reactions, where localized DNA synthesis reports on the position of hybridization targets, to potentially reveal the binding of single oligonucleotide-size probe molecules. Such a system has been presented for the detection of mitochondrial DNA in fixed cells, whereas attempts to apply rolling circle detection to metaphase chromosomes have previously failed, according to the literature. Methods Synchronized cultured cells were fixed with methanol/acetic acid to prepare chromosome spreads in teflon-coated diagnostic well-slides. Apart from the slide format and the chromosome spreading everything was done essentially according to standard protocols. Hybridization targets were detected in situ with padlock probes, which were ligated and amplified using target primed rolling circle DNA synthesis, and detected by fluorescence labeling. Results An optimized protocol for the spreading of condensed metaphase chromosomes in teflon-coated diagnostic well-slides was developed. Applying this protocol we generated specimens for target primed rolling circle DNA synthesis of padlock probes recognizing a 40 nucleotide sequence in the male specific repetitive satellite I sequence (DYZ1 on the Y-chromosome and a 32 nucleotide sequence in the repetitive kringle IV domain in the apolipoprotein(a gene positioned on the long arm of chromosome 6. These targets were detected with good efficiency, but the efficiency on other target sites was unsatisfactory. Conclusion Our aim was to test the applicability of the method used on mitochondrial DNA to the analysis of nuclear genomes, in particular as

Identification of microRNA-Like RNAs in the filamentous fungus Trichoderma reesei by solexa sequencing.

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Kang Kang

Full Text Available microRNAs (miRNAs are non-coding small RNAs (sRNAs capable of negatively regulating gene expression. Recently, microRNA-like small RNAs (milRNAs were discovered in several filamentous fungi but not yet in Trichoderma reesei, an industrial filamentous fungus that can secrete abundant hydrolases. To explore the presence of milRNA in T. reesei and evaluate their expression under induction of cellulose, two T. reesei sRNA libraries of cellulose induction (IN and non-induction (CON were generated and sequenced using Solexa sequencing technology. A total of 726 and 631 sRNAs were obtained from the IN and CON samples, respectively. Global expression analysis showed an extensively differential expression of sRNAs in T. reesei under the two conditions. Thirteen predicted milRNAs were identified in T. reesei based on the short hairpin structure analysis. The milRNA profiles obtained in deep sequencing were further validated by RT-qPCR assay. Computational analysis predicted a number of potential targets relating to many processes including regulation of enzyme expression. The presence and differential expression of T. reesei milRNAs imply that milRNA might play a role in T. reesei growth and cellulase induction. This work lays foundation for further functional study of fungal milRNAs and their industrial application.

Evaluation of RIDA®GENE norovirus GI/GII real time RT-PCR using stool specimens collected from children and adults with acute gastroenteritis.

Science.gov (United States)

Kanwar, N; Hassan, F; Barclay, L; Langley, C; Vinjé, J; Bryant, P W; George, K St; Mosher, L; Matthews-Greer, J M; Rocha, M A; Beenhouwer, D O; Harrison, C J; Moffatt, M; Shastri, N; Selvarangan, R

2018-04-10

Norovirus is the leading cause of epidemic and sporadic acute gastroenteritis (AGE) in the United States. Widespread prevalence necessitates implementation of accurate norovirus detection assays in clinical diagnostic laboratories. To evaluate RIDA ® GENE norovirus GI/GII real-time RT-PCR assay (RGN RT-PCR) using stool samples from patients with sporadic AGE. Patients between 14 days to 101 years of age with symptoms of AGE were enrolled prospectively at four sites across the United States during 2014-2015. Stool specimens were screened for the presence of norovirus RNA by the RGN RT-PCR assay. Results were compared with a reference method that included conventional RT-PCR and sequencing of a partial region of the 5'end of the norovirus ORF2 gene. A total of 259 (36.0%) of 719 specimens tested positive for norovirus by the reference method. The RGN RT-PCR assay detected norovirus in 244 (94%) of these 259 norovirus positive specimens. The sensitivity and specificity (95% confidence interval) of the RGN RT-PCR assay for detecting norovirus genogroup (G) I was 82.8% (63.5-93.5) and 99.1% (98.0-99.6) and for GII was 94.8% (90.8-97.2) and 98.6% (96.9-99.4), respectively. Seven specimens tested positive by the RGN-RT PCR that were negative by the reference method. The fifteen false negative samples were typed as GII.4 Sydney, GII.13, GI.3, GI.5, GI.2, GII.1, and GII.3 in the reference method. The RGN RT-PCR assay had a high sensitivity and specificity for the detection of norovirus in stool specimens from patients with sporadic AGE. Copyright © 2018. Published by Elsevier B.V.

[Topographic mapping of retinal function with a scanning laser ophthalmoscope and multifocal electroretinography using short M-sequences].

Science.gov (United States)

Rudolph, G; Bechmann, M; Berninger, T; Kutschbach, E; Held, U; Tornow, R P; Kalpadakis, P; Zol'nikova, I V; Shamshinova, A M

2001-01-01

A new method of multifocal electroretinography making use of scanning laser ophthalmoscope with a wavelength of 630 nm (SLO-m-ERG), evoking short spatial visual stimuli on the retina, is proposed. Algorithm of presenting the visual stimuli and analysis of distribution of local electroretinograms on the surface of the retina is based on short m-sequences. Mathematical cross correlation analysis shows a three-dimensional distribution of bioelectrical activity of the retina in the central visual field. In normal subjects the cone bioelectrical activity is the maximum in the macular area (corresponding to the density of cone distribution) and absent in the blind spot. The method detects the slightest pathological changes in the retina under control of the site of stimulation and ophthalmoscopic picture of the fundus oculi. The site of the pathological process correlates with the topography of changes in bioelectrical activity of the examined retinal area in diseases of the macular area and pigmented retinitis detectable by ophthalmoscopy.

The incidence and functional consequences of RT-associated cardiac perfusion defects

International Nuclear Information System (INIS)

Marks, Lawrence B.; Yu Xiaoli; Prosnitz, Robert G.; Zhou Sumin; Hardenbergh, Patricia H.; Blazing, Michael; Hollis, Donna; Lind, Pehr; Tisch, Andrea; Wong, Terence Z.; Borges-Neto, Salvador

2005-01-01

Purpose: Radiation therapy (RT) for left-sided breast cancer has been associated with cardiac dysfunction. We herein assess the temporal nature and volume dependence of RT-induced left ventricular perfusion defects and whether these perfusion defects are related to changes in cardiac wall motion or alterations in ejection fraction. Methods: From 1998 to 2001, 114 patients were enrolled onto an IRB-approved prospective clinical study to assess changes in regional and global cardiac function after RT for left-sided breast cancer. Patients were imaged 30 to 60 minutes after injection of technetium 99m sestamibi or tetrofosmin. Post-RT perfusion scans were compared with the pre-RT studies to assess for RT-induced perfusion defects as well as functional changes in wall motion and ejection fraction. Two-tailed Fisher's exact test and the Cochran-Armitage test for linear trends were used for statistical analysis. Results: The incidence of new perfusion defects 6, 12, 18, and 24 months after RT was 27%, 29%, 38%, and 42%, respectively. New defects occurred in approximately 10% to 20% and 50% to 60% of patients with less than 5%, and greater than 5%, of their left ventricle included within the RT fields, respectively (p = 0.33 to 0.00008). The rates of wall motion abnormalities in patients with and without perfusion defects were 12% to 40% versus 0% to 9%, respectively; p values were 0.007 to 0.16, depending on the post-RT interval. Conclusions: Radiation therapy causes volume-dependent perfusion defects in approximately 40% of patients within 2 years of RT. These perfusion defects are associated with corresponding wall-motion abnormalities. Additional study is necessary to better define the long-term functional consequences of RT-induced perfusion defects

Pärt: Fratres / Robert Cowan

Index Scriptorium Estoniae

Cowan, Robert

1995-01-01

Uuest heliplaadist "Pärt: Fratres (seven versions). Festina Lente. Cantus in Memory of Benjamin Britten. Summa. Peter Manning (vn.), France Springuel (vc.), Mireille Gleizes (pf.), I Fiamminghi. Telare CD CD80387 (79 minutes)

The novel as short story

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Kirk Schlueter

2013-06-01

Full Text Available In recent history, the novel has been thought of and defined primarily as a long prose narrative. However, this has not been the case historically, as the original meaning of "novel" was for "a piece of news" or "a short story or novella." Returning to this original definition, I propose a new way of viewing the work known contemporarily as the novel as a collection, or sequence, of united short stories rather than a single indivisible work, with the component short stories or novellas comprising the sequence renamed as "novels." A brief examination of several classic works traditionally considered novels serves to illustrate how this change in definition will affect reading.

MRI in multiple sclerosis of the spinal cord: evaluation of fast short-tan inversion-recovery and spin-echo sequences

International Nuclear Information System (INIS)

Dietemann, J.L.; Thibaut-Menard, A.; Neugroschl, C.; Gillis, C.; Abu Eid, M.; Bogorin, A.; Warter, J.M.; Tranchant, C.

2000-01-01

We compared the sensitivity of T2-weighted spin-echo (FSE) and fast short-tau inversion-recovery (fSTIR) sequences in detection of multiple sclerosis of the spinal cord in 100 consecutive patients with clinically confirmed multiple sclerosis (MS); 86 patients underwent also brain MRI. In all, 310 focal lesions were detected on fSTIR and 212 on T2-weighted FSE, spinal cord lesions were seen better on fSTIR images, with a higher contrast between the lesion and the normal spinal cord. In 24 patients in whom cord plaques were shown with both sequences, the cranial study was normal or inconclusive. Assessment of spinal plaques can be particularly important when MRI of the brain is inconclusive, and in there situations fSTIR can be helpful. (orig.)

RT-qPCR normalization genes in the red alga Chondrus crispus.

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Nathalie Kowalczyk

Full Text Available Chondrus crispus is a common red macroalga living on the rocky shores of the North Atlantic Ocean. It has a long research history, being a major source of carrageenan, a thickener widely used in the food industry, but also for physiological and ecological studies. To establish it as a model for red algae, its genome has been sequenced, allowing the development of molecular tools such as quantification of gene expression, including RNAseq and RT-qPCR. To determine appropriate genes for RT-qPCR normalization, the expression of 14 genes was monitored in 18 conditions using two sets of algal samples: samples from the sequenced strain, cultured and stressed in laboratory conditions and C. crispus collected on the shore and stressed in situ. The expression stability of the genes between the samples was evaluated by comparing the Ct range and using the programs geNorm and NormFinder. The candidate genes encoded translation related proteins (initiation factors IF4A-1 and IF4A-2, elongation factor EF1α and eRF3, an eukaryotic polypeptide chain release factor, cytoskeleton proteins (two β-tubulins, α-tubulin and actin, enzymes involved in the pentose phosphate pathway (glucose 6-phosphate deshydrogenase, protein recycling process (ubiquitin and ubiquitin-conjugating enzyme and glycolysis (isocitrate dehydrogenase. The two sets of samples showed different expression patterns. Most of the genes were stable in the algae cultivated in the laboratory, whereas environmental samples showed a more important variation in gene expression. When analyzing the two sets separately, the ranking of the most stables genes were different from one method to another. When considering all samples, the two statistical methods were concordant, revealing translation initiation factor 4A-2 and eukaryotic polypeptide chain release factor 3 as pertinent normalization genes. This study highlights thus the importance of testing reference genes according to the experiments as well

Whole Genome Sequencing Identifies a Missense Mutation in HES7 Associated with Short Tails in Asian Domestic Cats.

Science.gov (United States)

Xu, Xiao; Sun, Xin; Hu, Xue-Song; Zhuang, Yan; Liu, Yue-Chen; Meng, Hao; Miao, Lin; Yu, He; Luo, Shu-Jin

2016-08-25

Domestic cats exhibit abundant variations in tail morphology and serve as an excellent model to study the development and evolution of vertebrate tails. Cats with shortened and kinked tails were first recorded in the Malayan archipelago by Charles Darwin in 1868 and remain quite common today in Southeast and East Asia. To elucidate the genetic basis of short tails in Asian cats, we built a pedigree of 13 cats segregating at the trait with a founder from southern China and performed linkage mapping based on whole genome sequencing data from the pedigree. The short-tailed trait was mapped to a 5.6 Mb region of Chr E1, within which the substitution c. 5T > C in the somite segmentation-related gene HES7 was identified as the causal mutation resulting in a missense change (p.V2A). Validation in 245 unrelated cats confirmed the correlation between HES7-c. 5T > C and Chinese short-tailed feral cats as well as the Japanese Bobtail breed, indicating a common genetic basis of the two. In addition, some of our sampled kinked-tailed cats could not be explained by either HES7 or the Manx-related T-box, suggesting at least three independent events in the evolution of domestic cats giving rise to short-tailed traits.

Characterization of Sri Lanka rabies virus isolates using nucleotide sequence analysis of nucleoprotein gene.

Science.gov (United States)

Arai, Y T; Takahashi, H; Kameoka, Y; Shiino, T; Wimalaratne, O; Lodmell, D L

2001-01-01

Thirty-four suspected rabid brain samples from 2 humans, 24 dogs, 4 cats, 2 mongooses, I jackal and I water buffalo were collected in 1995-1996 in Sri Lanka. Total RNA was extracted directly from brain suspensions and examined using a one-step reverse transcription-polymerase chain reaction (RT-PCR) for the rabies virus nucleoprotein (N) gene. Twenty-eight samples were found positive for the virus N gene by RT-PCR and also for the virus antigens by fluorescent antibody (FA) test. Rabies virus isolates obtained from different animal species in different regions of Sri Lanka were genetically homogenous. Sequences of 203 nucleotides (nt)-long RT-PCR products obtained from 16 of 27 samples were found identical. Sequences of 1350 nt of N genes of 14 RT-PCR products were determined. The Sri Lanka isolates under study formed a specific cluster that included also an earlier isolate from India but did not include the known isolates from China, Thailand, Malaysia, Israel, Iran, Oman, Saudi Arabia, Russia, Nepal, Philippines, Japan and from several other countries. These results suggest that one type of rabies virus is circulating among human, dog, cat, mongoose, jackal and water buffalo living near Colombo City and in other five remote regions in Sri Lanka.

Analysis of sequences from field samples reveals the presence of the recently described pepper vein yellows virus (genus Polerovirus) in six additional countries.

Science.gov (United States)

Knierim, Dennis; Tsai, Wen-Shi; Kenyon, Lawrence

2013-06-01

Polerovirus infection was detected by reverse transcription polymerase chain reaction (RT-PCR) in 29 pepper plants (Capsicum spp.) and one black nightshade plant (Solanum nigrum) sample collected from fields in India, Indonesia, Mali, Philippines, Thailand and Taiwan. At least two representative samples for each country were selected to generate a general polerovirus RT-PCR product of 1.4 kb length for sequencing. Sequence analysis of the partial genome sequences revealed the presence of pepper vein yellows virus (PeVYV) in all 13 samples. A 1990 Australian herbarium sample of pepper described by serological means as infected with capsicum yellows virus (CYV) was identified by sequence analysis of a partial CP sequence as probably infected with a potato leaf roll virus (PLRV) isolate.

Design of Long Period Pseudo-Random Sequences from the Addition of -Sequences over

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Ren Jian

2004-01-01

Full Text Available Pseudo-random sequence with good correlation property and large linear span is widely used in code division multiple access (CDMA communication systems and cryptology for reliable and secure information transmission. In this paper, sequences with long period, large complexity, balance statistics, and low cross-correlation property are constructed from the addition of -sequences with pairwise-prime linear spans (AMPLS. Using -sequences as building blocks, the proposed method proved to be an efficient and flexible approach to construct long period pseudo-random sequences with desirable properties from short period sequences. Applying the proposed method to , a signal set is constructed.

Artist - analytical RT inspection simulation tool

International Nuclear Information System (INIS)

Bellon, C.; Jaenisch, G.R.

2007-01-01

The computer simulation of radiography is applicable for different purposes in NDT such as for the qualification of NDT systems, the prediction of its reliability, the optimization of system parameters, feasibility analysis, model-based data interpretation, education and training of NDT/NDE personnel, and others. Within the framework of the integrated project FilmFree the radiographic testing (RT) simulation software developed by BAM is being further developed to meet practical requirements for inspection planning in digital industrial radiology. It combines analytical modelling of the RT inspection process with the CAD-orientated object description applicable to various industrial sectors such as power generation, railways and others. (authors)

Arvo Pärt jagab preemia koolidele / Rebekka Lotman, Eva Kübar ; kommenteerinud Arvo Pärt

Index Scriptorium Estoniae

Lotman, Rebekka, 1978-

2009-01-01

Kultuuriministeerium tegi ettepaneku anda Arvo Pärdile elutööpreemia, millega kaasneva rahasumma lubas Pärt kinkida Rakvere ja Paide muusikakoolile. Elutööpreemiad saavad veel Ellen Niit ja Aarne Üksküla

A real time Taqman RT-PCR for the detection of rabbit hemorrhagic disease virus 2 (RHDV2).

Science.gov (United States)

Duarte, Margarida Dias; Carvalho, Carina L; Barros, Silvia C; Henriques, Ana M; Ramos, Fernanda; Fagulha, Teresa; Luís, Tiago; Duarte, Elsa L; Fevereiro, Miguel

2015-07-01

A specific real time RT-PCR for the detection of RHDV2 was developed and validated using RHDV and RHDV2 RNA preparations from positive field samples. The system was designed to amplify a 127 nucleotide-long RNA region located within the vp60 gene, based on the alignment of six sequences originated in Portugal, obtained in our laboratory, and 11 sequences from France and Italy. The primers and probe target sequences are highly conserved in the vast majority of the RHDV2 sequences presently known. In the sequences showing variability, only one mismatch is found per strain, usually outlying the 3' end of the primer or probe hybridization sequences. The specificity of the method was demonstrated in vitro with a panel of common rabbit pathogens. Standardization was performed with RNA transcripts obtained from a recombinant plasmid harboring the target sequence. The method was able to detected nine RNA molecules with an efficiency of 99.4% and a R(2) value of 1. Repeatability and reproducibility of the method were very high, with coefficients of variation lower than 2.40%. The assay was proven a valuable tool to diagnose most of RDVH2 circulating strains, and may be also useful to monitor viral loads, and consequently, disease progression and vaccination efficacy. Copyright © 2015 Elsevier B.V. All rights reserved.

Atucha II NPP full scope simulator modelling with the thermal hydraulic code TRAC{sub R}T

Energy Technology Data Exchange (ETDEWEB)

Alonso, Pablo Rey; Ruiz, Jose Antonio; Rivero, Norberto, E-mail: prey@tecnatom.e, E-mail: jaruiz@tecnatom.e, E-mail: nrivero@tecnatom.e [Tecnatom S.A., Madrid (Spain)

2011-07-01

In February 2010 NA-SA (Nucleoelectrica Argentina S.A.) awarded Tecnatom the Atucha II full scope simulator project. NA-SA is a public company owner of the Argentinean nuclear power plants. Atucha II is due to enter in operation shortly. Atucha II NPP is a PHWR type plant cooled by the water of the Parana River and has the same design as the Atucha I unit, doubling its power capacity. Atucha II will produce 745 MWe utilizing heavy water as coolant and moderator, and natural uranium as fuel. A plant singular feature is the permanent core refueling. TRAC{sub R}T is the first real time thermal hydraulic six-equations code used in the training simulation industry for NSSS modeling. It is the result from adapting to real time the best estimate code TRACG. TRAC{sub R}T is based on first principle conservation equations for mass, energy and momentum for liquid and steam phases, with two phase flows under non homogeneous and non equilibrium conditions. At present, it has been successfully implemented in twelve full scope replica simulators in different training centers throughout the world. To ease the modeling task, TRAC{sub R}T includes a graphical pre-processing tool designed to optimize this process and alleviate the burden of entering alpha numerical data in an input file. (author)

Evaluating the Impacts of NASA/SPoRT Daily Greenness Vegetation Fraction on Land Surface Model and Numerical Weather Forecasts

Science.gov (United States)

Bell, Jordan R.; Case, Jonathan L.; LaFontaine, Frank J.; Kumar, Sujay V.

2012-01-01

The NASA Short-term Prediction Research and Transition (SPoRT) Center has developed a Greenness Vegetation Fraction (GVF) dataset, which is updated daily using swaths of Normalized Difference Vegetation Index data from the Moderate Resolution Imaging Spectroradiometer (MODIS) data aboard the NASA EOS Aqua and Terra satellites. NASA SPoRT began generating daily real-time GVF composites at 1-km resolution over the Continental United States (CONUS) on 1 June 2010. The purpose of this study is to compare the National Centers for Environmental Prediction (NCEP) climatology GVF product (currently used in operational weather models) to the SPoRT-MODIS GVF during June to October 2010. The NASA Land Information System (LIS) was employed to study the impacts of the SPoRT-MODIS GVF dataset on a land surface model (LSM) apart from a full numerical weather prediction (NWP) model. For the 2010 warm season, the SPoRT GVF in the western portion of the CONUS was generally higher than the NCEP climatology. The eastern CONUS GVF had variations both above and below the climatology during the period of study. These variations in GVF led to direct impacts on the rates of heating and evaporation from the land surface. In the West, higher latent heat fluxes prevailed, which enhanced the rates of evapotranspiration and soil moisture depletion in the LSM. By late Summer and Autumn, both the average sensible and latent heat fluxes increased in the West as a result of the more rapid soil drying and higher coverage of GVF. The impacts of the SPoRT GVF dataset on NWP was also examined for a single severe weather case study using the Weather Research and Forecasting (WRF) model. Two separate coupled LIS/WRF model simulations were made for the 17 July 2010 severe weather event in the Upper Midwest using the NCEP and SPoRT GVFs, with all other model parameters remaining the same. Based on the sensitivity results, regions with higher GVF in the SPoRT model runs had higher evapotranspiration and

Simultaneous detection of three lily viruses using Triplex IC-RT-PCR.

Science.gov (United States)

Zhang, Yubao; Wang, Yajun; Xie, Zhongkui; Yang, Guo; Guo, Zhihong; Wang, Le

2017-11-01

Viruses commonly infecting lily (Lilium spp.) include: Lily symptomless virus (LSV), Cucumber mosaic virus (CMV) and Lily mottle virus (LMoV). These viruses usually co-infect lilies causing severe economic losses in terms of quantity and quality of flower and bulb production around the world. Reliable and precise detection systems need to be developed for virus identification. We describe the development of a triplex immunocapture (IC) reverse transcription (RT) polymerase chain reaction (PCR) assay for the simultaneous detection of LSV, CMV and LMoV. The triplex IC-RT-PCR was compared with a quadruplex RT-PCR assay. Relative to the quadruplex RT-PCR, the specificity of the triplex IC-RT-PCR system for LSV, CMV and LMoV was 100% for field samples. The sensitivity of the triplex IC-RT-PCR system was 99.4%, 81.4% and 98.7% for LSV, CMV and LMoV, respectively. Agreement (κ) between the results obtained from the two tests was 0.968, 0.844 and 0.984 for LSV, CMV and LMoV, respectively. This is the first report of the simultaneous detection of LSV, CMV and LMoV in a triplex IC-RT-PCR assay. In particular we believe this convenient and reliable triplex IC-RT-PCR method could be used routinely for large-scale field surveys or crop health monitoring of lily. Copyright © 2017. Published by Elsevier B.V.

Comparative Evaluation Of Conventional Rt-pcr And Real-time Rt-pcr (rrt-pcr) For Detection Of Avian Metapneumovirus Subtype A [comparação Entre As Técnicas De Rt-pcr Convencional E Rt-pcr Em Tempo Real Para A Detecção Do Metapneumovírus Aviários Subtipo A

OpenAIRE

Ferreira H.L.; Spilki F.R.; dos Santos M.M.A.B.; de Almeida R.S.; Arns C.W.

2009-01-01

Avian metapneumovirus (AMPV) belongs to Metapneumovirus genus of Paramyxoviridae family. Virus isolation, serology, and detection of genomic RNA are used as diagnostic methods for AMPV. The aim of the present study was to compare the detection of six subgroup A AMPV isolates (AMPV/A) viral RNA by using different conventional and real time RT-PCR methods. Two new RT-PCR tests and two real time RT-PCR tests, both detecting fusion (F) gene and nucleocapsid (N) gene were compared with an establis...

Comparative evaluation of conventional RT-PCR and real-time RT-PCR (RRT-PCR for detection of avian metapneumovirus subtype A Comparação entre as técnicas de RT-PCR convencional e RT-PCR em tempo real para a detecção do metapneumovírus aviários subtipo A

Directory of Open Access Journals (Sweden)

Helena Lage Ferreira

2009-08-01

Full Text Available Avian metapneumovirus (AMPV belongs to Metapneumovirus genus of Paramyxoviridae family. Virus isolation, serology, and detection of genomic RNA are used as diagnostic methods for AMPV. The aim of the present study was to compare the detection of six subgroup A AMPV isolates (AMPV/A viral RNA by using different conventional and real time RT-PCR methods. Two new RT-PCR tests and two real time RT-PCR tests, both detecting fusion (F gene and nucleocapsid (N gene were compared with an established test for the attachment (G gene. All the RT-PCR tested assays were able to detect the AMPV/A. The lower detection limits were observed using the N-, F- based RRT-PCR and F-based conventional RT-PCR (10(0.3 to 10¹ TCID50 mL-1. The present study suggests that the conventional F-based RT-PCR presented similar detection limit when compared to N- and F-based RRT-PCR and they can be successfully used for AMPV/A detection.O metapneumovírus aviário (AMPV pertence ao gênero Metapneumovirus, família Paramyxoviridae. Isolamento viral, sorologia e detecção do RNA genômico são atualmente as técnicas utilizadas para o diagnóstico desse agente. O objetivo do presente estudo foi comparar a detecção de RNA viral de seis isolados de AMPV, subtipo A (AMPV/A, utilizando diferentes métodos de RT-PCR convencional e real time RT-PCR (RRT-PCR. Duas novas técnicas de RT-PCR convencional e duas técnicas de RRT-PCR, ambas para a detecção dos genes da nucleoproteína (N e da proteína de fusão (F, foram comparadas com um RT-PCR previamente estabelecido para a detecção do AMPV (gene da glicoproteína -G. Todos esses métodos foram capazes de detectar os isolados AMPV/A. As técnicas RRT-PCR (genes F e N mostraram os menores limites de detecção (10(0.3 to 10¹ TCID50 mL-1. Os resultados sugerem que as técnicas RT-PCR convencional (gene F e as técnicas de RRT-PCR (gene F e N desenvolvidas no presente estudo podem ser utilizadas com sucesso para a detecção do

Märt Haamer: Tsahknal pole Overalliga mingisugust seost / Märt Haamer ; intervjueerinud Jan Jõgis-Laats

Index Scriptorium Estoniae

Haamer, Märt, 1973-

2009-01-01

Overall Eesti juhi Märt Haameri väitel pole Anders Tsahkna ühtegi riigihanget Overalli ettevõtetele suunanud. Overalli omandusse kuuluv firma Gennet Lab on olnud alltöövõtja paaris E-tervise Sihtasutuse korraldatud hankes

Identification and chromosomal distribution of copia-like retrotransposon sequences in the coffee (Coffea L. genome

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Juan-Carlos Herrera

2013-12-01

Full Text Available The presence of copia-like transposable elements in seven coffee (Coffea sp. species, including the cultivated Coffea arabica, was investigated. The highly conserved domains of the reverse transcriptase (RT present in the copia retrotransposons were amplified by PCR using degenerated primers. Fragments of roughly 300 bp were obtained and the nucleotide sequence was determined for 36 clones, 19 of which showed good quality. The deduced amino acid sequences were compared by multiple alignment analysis. The data suggested two distinct coffee RT groups, designated as CRTG1 and CRTG2. The sequence identities among the groups ranged from 52 to 60% for CRTG1 and 74 to 85% for CRTG2. The multiple alignment analysis revealed that some of the clones in CRTG1 were closely related to the representative elements present in other plant species such as Brassica napus, Populus ciliata and Picea abis. Furthermore, the chromosomal localization of the RT domains in C. arabica and their putative ancestors was investigated by fluorescence in situ hybridization (FISH analysis. FISH signals were observed throughout the chromosomes following a similar dispersed pattern with some localized regions exhibiting higher concentrations of those elements, providing new evidence of their relative conservation and stability in the coffee genome

WE-G-17A-03: MRIgRT: Quantification of Organ Motion

International Nuclear Information System (INIS)

Stanescu, T; Tadic, T; Jaffray, D

2014-01-01

Purpose: To develop an MRI-based methodology and tools required for the quantification of organ motion on a dedicated MRI-guided radiotherapy system. A three-room facility, consisting of a TrueBeam 6X linac vault, a 1.5T MR suite and a brachytherapy interventional room, is currently under commissioning at our institution. The MR scanner can move and image in either room for diagnostic and treatment guidance purposes. Methods: A multi-imaging modality (MR, kV) phantom, featuring programmable 3D simple and complex motion trajectories, was used for the validation of several image sorting algorithms. The testing was performed on MRI (e.g. TrueFISP, TurboFLASH), 4D CT and 4D CBCT. The image sorting techniques were based on a) direct image pixel manipulation into columns or rows, b) single and aggregated pixel data tracking and c) using computer vision techniques for global pixel analysis. Subsequently, the motion phantom and sorting algorithms were utilized for commissioning of MR fast imaging techniques for 2D-cine and 4D data rendering. MR imaging protocols were optimized (e.g. readout gradient strength vs. SNR) to minimize the presence of susceptibility-induced distortions, which were reported through phantom experiments and numerical simulations. The system-related distortions were also quantified (dedicated field phantom) and treated as systematic shifts where relevant. Results: Image sorting algorithms were validated for specific MR-based applications such as quantification of organ motion, local data sampling, and 4D MRI for pre-RT delivery with accuracy better than the raw image pixel size (e.g. 1 mm). MR fast imaging sequences were commissioning and imaging strategies were developed to mitigate spatial artifacts with minimal penalty on the image spatial and temporal sampling. Workflows (e.g. liver) were optimized to include the new motion quantification tools for RT planning and daily patient setup verification. Conclusion: Comprehensive methods were developed

Preparation of rT3 in human serum ria kit

International Nuclear Information System (INIS)

Zhang Peixuan; Han Shiquan; Zhang Liling

1991-02-01

A method for the measurement of rT 3 in serum by radioimmunoassay is presented. The double antibody plus polyethylene glycol was used as separating reagent. The method is simple, precise and sensitive. Serum rT 3 values of normal (n = 61) were 47.7 ± 11.34 x 10 -8 g/L (X-bar ± SD). Patients with hyperthyroidism (n = 8) serum rT 3 values were 272.2 ± 91.1 x 10 -8 g/L (X-bar ± SD) and patients with hypothyroidism (n = 2) serum rT 3 values were 11.4 ± 8.3 x 10 -8 g/L (X-bar ± SD). It appears to be a useful diagnostic procedure for thyroid diseases

Evaluation of gamma-sterilization (60Co) by RT-PCR by DHFR expression detection

International Nuclear Information System (INIS)

Converso, Ana Paula G.; Andrade Junior, Heitor F. de; Vieira, Daniel P.

2007-01-01

The improvement of techniques to detect pathogen agents in blood had reduced significantly the contamination mechanisms by hemocomponents in blood transfusion procedures. Ionizing radiation is a method that has presented several applications on medicine and in currently days has been showing special attention on blood banks which has been applied to avoid TA-GVHD development. DHFR is an enzyme constitutive in Plasmodium protozoa and has an important role in folate metabolism on these parasites. Detecting the expression of RNAm coder for this enzyme is possible to evaluate the viability of this parasite in blood samples. Plasmodium chabaudi AJ is a parasite that induces lethal malaria in rodents similar to human malaria In this work, the objective was to detect the presence of plasmodium protozoa in irradiated blood samples, infected experimentally, through the application of a RT-PCR using primers for the coder sequence of DHFR's mRNA. We studied doses of ionizing radiation between 0 and 75 Gy. The irradiation procedures were accomplished in Center of Radiation Technology of IPEN-CNEN in a 60 Co panoramic source. Our results had demonstrated that RT-PCR is a sensible method to evaluate the viability of plasmodium in blood samples because the technique could detect low parasite burden in all tested samples. (author)

Soft-Rt: software for IMRT simulations based on MCNPX

International Nuclear Information System (INIS)

Ferreira F, T. C.; Campos, T.

2015-10-01

Intensity Modulated Radiation Therapy (IMRT) is an advanced treatment technique, widely used in external radiotherapy. This paper presents the Soft-Rt which allows the simulation of an entire IMRT treatment protocol. The Soft-Rt performs a full three-dimensional rendering of a set of patient images, including the definitions of region of interest with organs in risk, and the target tumor volume and margins (PTV). Thus, a more accurate analysis and planning can be performed, taking into account the features and orientation of the radiation beams. The exposed tissues as well as the amount of absorbed dose is depicted in healthy and/or cancerous tissues. As conclusion, Soft-Rt can predict dose on the PTV accurately, preserving the surrounding healthy tissues. Soft-Rt is coupled with SISCODES code. The SISCODES code is firstly applied to segment the set of CT or MRI patient images in distinct tissues pointing out its respective density and chemical compositions. Later, the voxel model is export to the Soft-Rt IMRT planning module in which a full treatment planning is created. All geometrical parameters are sent to the general purpose Monte Carlo transport code - MCNP - to simulate the interaction of each incident beam towards to the PTV avoiding organs in risk. The normalized dose results are exported to the Soft-Rt out-module, in which the three-dimensional model visualization is shown in a transparent glass procedure adopting gray scale for the dependence on the mass density of the correlated tissue; while, a color scale to depict dose values in a superimpose protocol. (Author)

Soft-Rt: software for IMRT simulations based on MCNPX

Energy Technology Data Exchange (ETDEWEB)

Ferreira F, T. C. [Centro de Desenvolvimento da Tecnologia Nuclear / CNEN, Av. Pte. Antonio Carlos 6627, 31270-901 Belo Horizonte, Minas Gerais (Brazil); Campos, T., E-mail: tcff01@gmail.com [Universidade Federal de Minas Gerais, Departamento de Engenharia Nuclear, Programa de Pos Graduacao em Ciencias e Tecnicas Nucleares, Av. Pte. Antonio Carlos 6627, 31270-901 Belo Horizonte, Minas Gerais (Brazil)

2015-10-15

Intensity Modulated Radiation Therapy (IMRT) is an advanced treatment technique, widely used in external radiotherapy. This paper presents the Soft-Rt which allows the simulation of an entire IMRT treatment protocol. The Soft-Rt performs a full three-dimensional rendering of a set of patient images, including the definitions of region of interest with organs in risk, and the target tumor volume and margins (PTV). Thus, a more accurate analysis and planning can be performed, taking into account the features and orientation of the radiation beams. The exposed tissues as well as the amount of absorbed dose is depicted in healthy and/or cancerous tissues. As conclusion, Soft-Rt can predict dose on the PTV accurately, preserving the surrounding healthy tissues. Soft-Rt is coupled with SISCODES code. The SISCODES code is firstly applied to segment the set of CT or MRI patient images in distinct tissues pointing out its respective density and chemical compositions. Later, the voxel model is export to the Soft-Rt IMRT planning module in which a full treatment planning is created. All geometrical parameters are sent to the general purpose Monte Carlo transport code - MCNP - to simulate the interaction of each incident beam towards to the PTV avoiding organs in risk. The normalized dose results are exported to the Soft-Rt out-module, in which the three-dimensional model visualization is shown in a transparent glass procedure adopting gray scale for the dependence on the mass density of the correlated tissue; while, a color scale to depict dose values in a superimpose protocol. (Author)

Alignment of Short Reads: A Crucial Step for Application of Next-Generation Sequencing Data in Precision Medicine

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Hao Ye

2015-11-01

Full Text Available Precision medicine or personalized medicine has been proposed as a modernized and promising medical strategy. Genetic variants of patients are the key information for implementation of precision medicine. Next-generation sequencing (NGS is an emerging technology for deciphering genetic variants. Alignment of raw reads to a reference genome is one of the key steps in NGS data analysis. Many algorithms have been developed for alignment of short read sequences since 2008. Users have to make a decision on which alignment algorithm to use in their studies. Selection of the right alignment algorithm determines not only the alignment algorithm but also the set of suitable parameters to be used by the algorithm. Understanding these algorithms helps in selecting the appropriate alignment algorithm for different applications in precision medicine. Here, we review current available algorithms and their major strategies such as seed-and-extend and q-gram filter. We also discuss the challenges in current alignment algorithms, including alignment in multiple repeated regions, long reads alignment and alignment facilitated with known genetic variants.

Identification of protoplast-isolation responsive microRNAs in Citrus reticulata Blanco by high-throughput sequencing.

Science.gov (United States)

Xu, Xiaoyong; Xu, Xiaoling; Zhou, Yipeng; Zeng, Shaohua; Kong, Weiwen

2017-01-01

Protoplast isolation is a stress-inducing process, during which a variety of physiological and molecular alterations take place. Such stress response affects the expression of totipotency of cultured protoplasts. MicroRNAs (miRNAs) play important roles in plant growth, development and stress responses. However, the underlying mechanism of miRNAs involved in the protoplast totipotency remains unclear. In this study, high-throughput sequencing technology was used to sequence two populations of small RNA from calli and callus-derived protoplasts in Citrus reticulata Blanco. A total of 67 known miRNAs from 35 families and 277 novel miRNAs were identified. Among these miRNAs, 18 known miRNAs and 64 novel miRNAs were identified by differentially expressed miRNAs (DEMs) analysis. The expression patterns of the eight DEMs were verified by qRT-PCR. Target prediction showed most targets of the miRNAs were transcription factors. The expression levels of half targets showed a negative correlation to those of the miRNAs. Furthermore, the physiological analysis showed high levels of antioxidant activities in isolated protoplasts. In short, our results indicated that miRNAs may play important roles in protoplast-isolation response.

Data Analysis of Sequences and qPCR for Microbial Communities during Algal Blooms

Science.gov (United States)

A training opportunity is open to a highly microbial-research-motivated student to conduct sequence analysis, explore novel genes and metabolic pathways, validate resultant findings using qPCR/RT-qPCR and summarize the findings

Kuidas sünnib pealkiri? / Pärt Lias

Index Scriptorium Estoniae

Lias, Pärt

2003-01-01

Eesti kirjanikud vastavad Pärt Liase küsimusele oma teoste pealkirjade saamisloo kohta: Doris Kareva, Ilona Laaman, Eha Lättemäe, Priidu Beier, Oskar Kruus, Jaan Kruusvall, Paul-Eerik Rummo ja Andres Vanapa (1); Triin Soomets, Priit Aimla, Aarne Puu, Einar Sander, Jüri Tuulik ja Ülo Tuulik (2); Nikolai Baturin, Leo Metsar, Toomas Vint, Leonid Stolovitš ja Pärt Lias (3)

Real time RT-PCR assay for detection of different serotypes of FMDV in Egypt

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Laila El-Shehawy

these localities as 19 out of 30 OP samples were positive by rRT-PCR while in contrast there were only 16 out of 30 samples positive by CFT. Conclusion: In conclusion, this study demonstrates that real-time RT-PCR currently used at the WRL for FMD provides an extremely sensitive and rapid additional procedure for improved laboratory diagnosis of FMD especially in-contact and carrier cases. The rRT-PCR generated results in less than one day from test commencement, in contrast to up to four days to define some positive and all negative samples by combined use of CFT and virus isolation. This is an important feature when definitive diagnostic results are required in a short timescale during emergencies. Also this study demonstrates the current situation of FMDV circulating in EL-Fayoum governorate and the introduction of new SAT2 serotype beside type A and O. [Vet World 2012; 5(12.000: 732-737

Avaliação das técnicas de RT-PCR e heminested RT-PCR em cérebros de cães com sinais neurológicos compatíveis com cinomose

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Adriana Cortez

2015-12-01

Full Text Available The diagnostic value of RT-PCR and hemi-nested RT-PCR (hnRT-PCR was compared in brain samples of dogs presenting neurological signs compatible with canine distemper. Samples of central nervous system (CNS were collected from 68 dogs and tested by direct immunofluorescence test (RFID and, independent of the results, they were stored at -20°C for at least three years. They were submitted to the RT-PCR and hnRT-PCR techniques aiming to determine the gene responsible for the viral nucleoprotein decoding. Fifty-nine samples were positive for RIFD, 40 for RT-PCR (Kappa = 0.358 and 54 for hnRT-PCR (Kappa = 0.740. All nine RIFD negative samples were also negative for RT-PCR and hnRT-PCR. In spite of the storage duration and proper sample conditions, the estimated accordance between hnRT-PCR and RIFD demonstrated that hnRT-PCR technique can be applied in retrospective studies.

Post-RT CT results as a predictive model for the necessity of planned post-RT neck dissection in patients with cervical metastatic disease from squamous cell carcinoma

International Nuclear Information System (INIS)

Ojiri, Hiroya; Mendenhall, William M.; Stringer, Scott P.; Johnson, Patches L.; Mancuso, Anthony A.

2002-01-01

Purpose: To establish whether the extent of neck disease on postradiation therapy (RT) computed tomography (CT) can predict the likelihood of positive neck nodes and, thereby, the necessity of planned post-RT neck dissection. Methods and Materials: Ninety-five patients who underwent post-RT neck dissection within 2 months for squamous cell carcinoma of the head and neck were eligible. Of the 95 patients, 37 (32.7%) of 113 hemineck specimens were pathologically positive. On post-RT CT imaging studies, the number and size of lymph nodes >1 cm were recorded. Internal focal defects and the likelihood of extracapsular spread were graded. Results: If lymph nodes on post-RT CT were ≤15 mm, free of significant internal focal low-attenuation or calcification, and without imaging evidence of extracapsular spread, the surgical hemineck specimen was positive in 1 (3.4%) of the 29 hemineck specimens. A focal low-attenuation defect (p=0.0078) and evidence of extracapsular spread (p=0.0721) seen in the residual nodal mass on CT were independent predictors of a positive surgical specimen by multivariate analysis. Conclusion: CT findings on post-RT neck studies can help predict the likelihood of residual disease and, thereby, the necessity of planned post-RT neck dissection

Pärt: Berliner Messe / Rob Cowan

Index Scriptorium Estoniae

Cowan, Rob

1998-01-01

Uuest heliplaadist "Pärt: Berliner Messe. The Beatitudes. Annum per Annum. Magnificat. Seven Magnificat Antiphons. De profundis. Polyphony / Steven Layton with Andrew Lucas" Hyperion CDA66960 (74 minutes:DDD)

Arvo Pärt. Litany / Peter Korfmacher

Index Scriptorium Estoniae

Korfmacher, Peter

1996-01-01

Uuest heliplaadist "Pärt, Arvo. Litany; Psalom; Trisagion. Hilliard Ensemble, Estonian Philharmonic Chamber Choir, Tallinn Chamber Orchestra, Tõnu Kaljuste"; Lithuanian Chamber Orchestra, Saulius Sondeckis. ECM 1592/449 810-2

Profiling structured product labeling with NDF-RT and RxNorm

Science.gov (United States)

2012-01-01

Background Structured Product Labeling (SPL) is a document markup standard approved by Health Level Seven (HL7) and adopted by United States Food and Drug Administration (FDA) as a mechanism for exchanging drug product information. The SPL drug labels contain rich information about FDA approved clinical drugs. However, the lack of linkage to standard drug ontologies hinders their meaningful use. NDF-RT (National Drug File Reference Terminology) and NLM RxNorm as standard drug ontology were used to standardize and profile the product labels. Methods In this paper, we present a framework that intends to map SPL drug labels with existing drug ontologies: NDF-RT and RxNorm. We also applied existing categorical annotations from the drug ontologies to classify SPL drug labels into corresponding classes. We established the classification and relevant linkage for SPL drug labels using the following three approaches. First, we retrieved NDF-RT categorical information from the External Pharmacologic Class (EPC) indexing SPLs. Second, we used the RxNorm and NDF-RT mappings to classify and link SPLs with NDF-RT categories. Third, we profiled SPLs using RxNorm term type information. In the implementation process, we employed a Semantic Web technology framework, in which we stored the data sets from NDF-RT and SPLs into a RDF triple store, and executed SPARQL queries to retrieve data from customized SPARQL endpoints. Meanwhile, we imported RxNorm data into MySQL relational database. Results In total, 96.0% SPL drug labels were mapped with NDF-RT categories whereas 97.0% SPL drug labels are linked to RxNorm codes. We found that the majority of SPL drug labels are mapped to chemical ingredient concepts in both drug ontologies whereas a relatively small portion of SPL drug labels are mapped to clinical drug concepts. Conclusions The profiling outcomes produced by this study would provide useful insights on meaningful use of FDA SPL drug labels in clinical applications through

Profiling structured product labeling with NDF-RT and RxNorm

Directory of Open Access Journals (Sweden)

Zhu Qian

2012-12-01

Full Text Available Abstract Background Structured Product Labeling (SPL is a document markup standard approved by Health Level Seven (HL7 and adopted by United States Food and Drug Administration (FDA as a mechanism for exchanging drug product information. The SPL drug labels contain rich information about FDA approved clinical drugs. However, the lack of linkage to standard drug ontologies hinders their meaningful use. NDF-RT (National Drug File Reference Terminology and NLM RxNorm as standard drug ontology were used to standardize and profile the product labels. Methods In this paper, we present a framework that intends to map SPL drug labels with existing drug ontologies: NDF-RT and RxNorm. We also applied existing categorical annotations from the drug ontologies to classify SPL drug labels into corresponding classes. We established the classification and relevant linkage for SPL drug labels using the following three approaches. First, we retrieved NDF-RT categorical information from the External Pharmacologic Class (EPC indexing SPLs. Second, we used the RxNorm and NDF-RT mappings to classify and link SPLs with NDF-RT categories. Third, we profiled SPLs using RxNorm term type information. In the implementation process, we employed a Semantic Web technology framework, in which we stored the data sets from NDF-RT and SPLs into a RDF triple store, and executed SPARQL queries to retrieve data from customized SPARQL endpoints. Meanwhile, we imported RxNorm data into MySQL relational database. Results In total, 96.0% SPL drug labels were mapped with NDF-RT categories whereas 97.0% SPL drug labels are linked to RxNorm codes. We found that the majority of SPL drug labels are mapped to chemical ingredient concepts in both drug ontologies whereas a relatively small portion of SPL drug labels are mapped to clinical drug concepts. Conclusions The profiling outcomes produced by this study would provide useful insights on meaningful use of FDA SPL drug labels in clinical

Profiling structured product labeling with NDF-RT and RxNorm.

Science.gov (United States)

Zhu, Qian; Jiang, Guoqian; Chute, Christopher G

2012-12-20

Structured Product Labeling (SPL) is a document markup standard approved by Health Level Seven (HL7) and adopted by United States Food and Drug Administration (FDA) as a mechanism for exchanging drug product information. The SPL drug labels contain rich information about FDA approved clinical drugs. However, the lack of linkage to standard drug ontologies hinders their meaningful use. NDF-RT (National Drug File Reference Terminology) and NLM RxNorm as standard drug ontology were used to standardize and profile the product labels. In this paper, we present a framework that intends to map SPL drug labels with existing drug ontologies: NDF-RT and RxNorm. We also applied existing categorical annotations from the drug ontologies to classify SPL drug labels into corresponding classes. We established the classification and relevant linkage for SPL drug labels using the following three approaches. First, we retrieved NDF-RT categorical information from the External Pharmacologic Class (EPC) indexing SPLs. Second, we used the RxNorm and NDF-RT mappings to classify and link SPLs with NDF-RT categories. Third, we profiled SPLs using RxNorm term type information. In the implementation process, we employed a Semantic Web technology framework, in which we stored the data sets from NDF-RT and SPLs into a RDF triple store, and executed SPARQL queries to retrieve data from customized SPARQL endpoints. Meanwhile, we imported RxNorm data into MySQL relational database. In total, 96.0% SPL drug labels were mapped with NDF-RT categories whereas 97.0% SPL drug labels are linked to RxNorm codes. We found that the majority of SPL drug labels are mapped to chemical ingredient concepts in both drug ontologies whereas a relatively small portion of SPL drug labels are mapped to clinical drug concepts. The profiling outcomes produced by this study would provide useful insights on meaningful use of FDA SPL drug labels in clinical applications through standard drug ontologies such as NDF-RT and

Evaluation of NASA SPoRT's Pseudo-Geostationary Lightning Mapper Products in the 2011 Spring Program

Science.gov (United States)

Stano, Geoffrey T.; Carcione, Brian; Siewert, Christopher; Kuhlman, Kristin M.

2012-01-01

NASA's Short-term Prediction Research and Transition (SPoRT) program is a contributing partner with the GOES-R Proving Ground (PG) preparing forecasters to understand and utilize the unique products that will be available in the GOES-R era. This presentation emphasizes SPoRT s actions to prepare the end user community for the Geostationary Lightning Mapper (GLM). This preparation is a collaborative effort with SPoRT's National Weather Service partners, the National Severe Storms Laboratory (NSSL), and the Hazardous Weather Testbed s Spring Program. SPoRT continues to use its effective paradigm of matching capabilities to forecast problems through collaborations with our end users and working with the developers at NSSL to create effective evaluations and visualizations. Furthermore, SPoRT continues to develop software plug-ins so that these products will be available to forecasters in their own decision support system, AWIPS and eventually AWIPS II. In 2009, the SPoRT program developed the original pseudo geostationary lightning mapper (PGLM) flash extent product to demonstrate what forecasters may see with GLM. The PGLM replaced the previous GLM product and serves as a stepping-stone until the AWG s official GLM proxy is ready. The PGLM algorithm is simple and can be applied to any ground-based total lightning network. For 2011, the PGLM used observations from four ground-based networks (North Alabama, Kennedy Space Center, Oklahoma, and Washington D.C.). While the PGLM is not a true proxy product, it is intended as a tool to train forecasters about total lightning as well as foster discussions on product visualizations and incorporating GLM-resolution data into forecast operations. The PGLM has been used in 2010 and 2011 and is likely to remain the primary lightning training tool for the GOES-R program for the near future. This presentation will emphasize the feedback received during the 2011 Spring Program. This will discuss several topics. Based on feedback

Selection of reference genes for quantitative RT-PCR studies in striped dolphin (Stenella coeruleoalba skin biopsies

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Casini Silvia

2006-09-01

Full Text Available Abstract Background Odontocete cetaceans occupy the top position of the marine food-web and are particularly sensitive to the bioaccumulation of lipophilic contaminants. The effects of environmental pollution on these species are highly debated and various ecotoxicological studies have addressed the impact of xenobiotic compounds on marine mammals, raising conservational concerns. Despite its sensitivity, quantitative real-time PCR (qRT-PCR has never been used to quantify gene induction caused by exposure of cetaceans to contaminants. A limitation for the application of qRT-PCR is the need for appropriate reference genes which allow the correct quantification of gene expression. A systematic evaluation of potential reference genes in cetacean skin biopsies is presented, in order to validate future qRT-PCR studies aiming at using the expression of selected genes as non-lethal biomarkers. Results Ten commonly used housekeeping genes (HKGs were partially sequenced in the striped dolphin (Stenella coeruleoalba and, for each gene, PCR primer pairs were specifically designed and tested in qRT-PCR assays. The expression of these potential control genes was examined in 30 striped dolphin skin biopsy samples, obtained from specimens sampled in the north-western Mediterranean Sea. The stability of selected control genes was determined using three different specific VBA applets (geNorm, NormFinder and BestKeeper which produce highly comparable results. Glyceraldehyde-3P-dehydrogenase (GAPDH and tyrosine 3-monooxygenase (YWHAZ always rank as the two most stably expressed HKGs according to the analysis with geNorm and Normfinder, and are defined as optimal control genes by BestKepeer. Ribosomal protein L4 (RPL4 and S18 (RPS18 also exhibit a remarkable stability of their expression levels. On the other hand, transferrin receptor (TFRC, phosphoglycerate kinase 1 (PGK1, hypoxanthine ribosyltransferase (HPRT1 and β-2-microglobin (B2M show variable expression

Molecular Properties of Poliovirus Isolates: Nucleotide Sequence Analysis, Typing by PCR and Real-Time RT-PCR.

Science.gov (United States)

Burns, Cara C; Kilpatrick, David R; Iber, Jane C; Chen, Qi; Kew, Olen M

2016-01-01

Virologic surveillance is essential to the success of the World Health Organization initiative to eradicate poliomyelitis. Molecular methods have been used to detect polioviruses in tissue culture isolates derived from stool samples obtained through surveillance for acute flaccid paralysis. This chapter describes the use of realtime PCR assays to identify and serotype polioviruses. In particular, a degenerate, inosine-containing, panpoliovirus (panPV) PCR primer set is used to distinguish polioviruses from NPEVs. The high degree of nucleotide sequence diversity among polioviruses presents a challenge to the systematic design of nucleic acid-based reagents. To accommodate the wide variability and rapid evolution of poliovirus genomes, degenerate codon positions on the template were matched to mixed-base or deoxyinosine residues on both the primers and the TaqMan™ probes. Additional assays distinguish between Sabin vaccine strains and non-Sabin strains. This chapter also describes the use of generic poliovirus specific primers, along with degenerate and inosine-containing primers, for routine VP1 sequencing of poliovirus isolates. These primers, along with nondegenerate serotype-specific Sabin primers, can also be used to sequence individual polioviruses in mixtures.

Rapid detection and subtyping of European swine influenza viruses in porcine clinical samples by haemagglutinin- and neuraminidase-specific tetra- and triplex real-time RT-PCRs.

Science.gov (United States)

Henritzi, Dinah; Zhao, Na; Starick, Elke; Simon, Gaelle; Krog, Jesper S; Larsen, Lars Erik; Reid, Scott M; Brown, Ian H; Chiapponi, Chiara; Foni, Emanuela; Wacheck, Silke; Schmid, Peter; Beer, Martin; Hoffmann, Bernd; Harder, Timm C

2016-11-01

A diversifying pool of mammalian-adapted influenza A viruses (IAV) with largely unknown zoonotic potential is maintained in domestic swine populations worldwide. The most recent human influenza pandemic in 2009 was caused by a virus with genes originating from IAV isolated from swine. Swine influenza viruses (SIV) are widespread in European domestic pig populations and evolve dynamically. Knowledge regarding occurrence, spread and evolution of potentially zoonotic SIV in Europe is poorly understood. Efficient SIV surveillance programmes depend on sensitive and specific diagnostic methods which allow for cost-effective large-scale analysis. New SIV haemagglutinin (HA) and neuraminidase (NA) subtype- and lineage-specific multiplex real-time RT-PCRs (RT-qPCR) have been developed and validated with reference virus isolates and clinical samples. A diagnostic algorithm is proposed for the combined detection in clinical samples and subtyping of SIV strains currently circulating in Europe that is based on a generic, M-gene-specific influenza A virus RT-qPCR. In a second step, positive samples are examined by tetraplex HA- and triplex NA-specific RT-qPCRs to differentiate the porcine subtypes H1, H3, N1 and N2. Within the HA subtype H1, lineages "av" (European avian-derived), "hu" (European human-derived) and "pdm" (human pandemic A/H1N1, 2009) are distinguished by RT-qPCRs, and within the NA subtype N1, lineage "pdm" is differentiated. An RT-PCR amplicon Sanger sequencing method of small fragments of the HA and NA genes is also proposed to safeguard against failure of multiplex RT-qPCR subtyping. These new multiplex RT-qPCR assays provide adequate tools for sustained SIV monitoring programmes in Europe. © 2016 The Authors. Influenza and Other Respiratory Viruses Published by John Wiley & Sons Ltd.

Optimisation of the RT-PCR detection of immunomagnetically enriched carcinoma cells

International Nuclear Information System (INIS)

Raynor, Michael; Stephenson, Sally-Anne; Walsh, David CA; Pittman, Kenneth B; Dobrovic, Alexander

2002-01-01

Immunomagnetic enrichment followed by RT-PCR (immunobead RT-PCR) is an efficient methodology to identify disseminated carcinoma cells in the blood and bone marrow. The RT-PCR assays must be both specific for the tumor cells and sufficiently sensitive to enable detection of single tumor cells. We have developed a method to test RT-PCR assays for any cancer. This has been investigated using a panel of RT-PCR markers suitable for the detection of breast cancer cells. In the assay, a single cell line-derived tumor cell is added to 100 peripheral blood mononuclear cells (PBMNCs) after which mRNA is isolated and reverse transcribed for RT-PCR analysis. PBMNCs without added tumor cells are used as specificity controls. The previously studied markers epidermal growth factor receptor (EGFR), mammaglobin 1 (MGB1), epithelial cell adhesion molecule (EpCAM/TACSTD1), mucin 1 (MUC1), carcinoembryonic antigen (CEA) were tested. Two new epithelial-specific markers ELF3 and EphB4 were also tested. MUC1 was unsuitable as strong amplification was detected in 100 cell PBMNC controls. Expression of ELF3, EphB4, EpCAM, EGFR, CEA and MGB1 was found to be both specific for the tumor cell, as demonstrated by the absence of a signal in most 100 cell PBMNC controls, and sensitive enough to detect a single tumor cell in 100 PBMNCs using a single round of RT-PCR. ELF3, EphB4, EpCAM, EGFR, CEA and MGB1 are appropriate RT-PCR markers for use in a marker panel to detect disseminated breast cancer cells after immunomagnetic enrichment

MetaVelvet: An Extension of Velvet Assembler to de novo Metagenome Assembly from Short Sequence Reads (Metagenomics Informatics Challenges Workshop: 10K Genomes at a Time)

Energy Technology Data Exchange (ETDEWEB)

Sakakibara, Yasumbumi

2011-10-13

Keio University's Yasumbumi Sakakibara on "MetaVelvet: An Extension of Velvet Assembler to de novo Metagenome Assembly from Short Sequence Reads" at the Metagenomics Informatics Challenges Workshop held at the DOE JGI on October 12-13, 2011.

Modelling Dynamic Topologies via Extensions of VDM-RT

DEFF Research Database (Denmark)

Nielsen, Claus Ballegård

Only a few formal methods include descriptions of the network topology that the modelled system is deployed onto. In VDM Real-Time (VDM-RT) this has been enabled for distributed systems that have a static structure. However, when modelling dynamic systems this fixed topology becomes an issue....... Systems with highly distributed and alternating relationships cannot be expressed correctly in a static model. This document describes how VDM-RT can be extended with new language constructs to enable the description of dynamic reconfiguration of the network topology during the runtime execution...... of a model. The extension is developed on the basis of a case study involving a dynamic system that has a constant changing system topology. With a basis in the case study a model is developed that uses the static version of VDM-RT in order to reveal the limitations of the language. The case study...

Evaluation of gamma-sterilization ({sup 60}Co) by RT-PCR by DHFR expression detection

Energy Technology Data Exchange (ETDEWEB)

Converso, Ana Paula G.; Andrade Junior, Heitor F. de [Instituto de Medicina Tropical de Sao Paulo, Sao Paulo, SP (Brazil)]. E-mail: anapaulagconverso@gmail.com; hfandrad@usp.br; Vieira, Daniel P. [Instituto de Pesquisas Energeticas e Nucleares (IPEN/CNEN-SP), Sao Paulo, SP (Brazil); Instituto de Medicina Tropical de Sao Paulo, Sao Paulo, SP (Brazil); E-mail: dperezv@usp.br

2007-07-01

The improvement of techniques to detect pathogen agents in blood had reduced significantly the contamination mechanisms by hemocomponents in blood transfusion procedures. Ionizing radiation is a method that has presented several applications on medicine and in currently days has been showing special attention on blood banks which has been applied to avoid TA-GVHD development. DHFR is an enzyme constitutive in Plasmodium protozoa and has an important role in folate metabolism on these parasites. Detecting the expression of RNAm coder for this enzyme is possible to evaluate the viability of this parasite in blood samples. Plasmodium chabaudi AJ is a parasite that induces lethal malaria in rodents similar to human malaria In this work, the objective was to detect the presence of plasmodium protozoa in irradiated blood samples, infected experimentally, through the application of a RT-PCR using primers for the coder sequence of DHFR's mRNA. We studied doses of ionizing radiation between 0 and 75 Gy. The irradiation procedures were accomplished in Center of Radiation Technology of IPEN-CNEN in a {sup 60}Co panoramic source. Our results had demonstrated that RT-PCR is a sensible method to evaluate the viability of plasmodium in blood samples because the technique could detect low parasite burden in all tested samples. (author)

Identification of microRNAs and their targets in Finger millet by high throughput sequencing.

Science.gov (United States)

Usha, S; Jyothi, M N; Sharadamma, N; Dixit, Rekha; Devaraj, V R; Nagesh Babu, R

2015-12-15

MicroRNAs are short non-coding RNAs which play an important role in regulating gene expression by mRNA cleavage or by translational repression. The majority of identified miRNAs were evolutionarily conserved; however, others expressed in a species-specific manner. Finger millet is an important cereal crop; nonetheless, no practical information is available on microRNAs to date. In this study, we have identified 95 conserved microRNAs belonging to 39 families and 3 novel microRNAs by high throughput sequencing. For the identified conserved and novel miRNAs a total of 507 targets were predicted. 11 miRNAs were validated and tissue specificity was determined by stem loop RT-qPCR, Northern blot. GO analyses revealed targets of miRNA were involved in wide range of regulatory functions. This study implies large number of known and novel miRNAs found in Finger millet which may play important role in growth and development. Copyright © 2015 Elsevier B.V. All rights reserved.

Short- and long-term evolutionary dynamics of bacterial insertion sequences: insights from Wolbachia endosymbionts.

Science.gov (United States)

Cerveau, Nicolas; Leclercq, Sébastien; Leroy, Elodie; Bouchon, Didier; Cordaux, Richard

2011-01-01

Transposable elements (TE) are one of the major driving forces of genome evolution, raising the question of the long-term dynamics underlying their evolutionary success. Long-term TE evolution can readily be reconstructed in eukaryotes, thanks to many degraded copies constituting genomic fossil records of past TE proliferations. By contrast, bacterial genomes usually experience high sequence turnover and short TE retention times, thereby obscuring ancient TE evolutionary patterns. We found that Wolbachia bacterial genomes contain 52-171 insertion sequence (IS) TEs. IS account for 11% of Wolbachia wRi, which is one of the highest IS genomic coverage reported in prokaryotes to date. We show that many IS groups are currently expanding in various Wolbachia genomes and that IS horizontal transfers are frequent among strains, which can explain the apparent synchronicity of these IS proliferations. Remarkably, >70% of Wolbachia IS are nonfunctional. They constitute an unusual bacterial IS genomic fossil record providing direct empirical evidence for a long-term IS evolutionary dynamics following successive periods of intense transpositional activity. Our results show that comprehensive IS annotations have the potential to provide new insights into prokaryote TE evolution and, more generally, prokaryote genome evolution. Indeed, the identification of an important IS genomic fossil record in Wolbachia demonstrates that IS elements are not always of recent origin, contrary to the conventional view of TE evolution in prokaryote genomes. Our results also raise the question whether the abundance of IS fossils is specific to Wolbachia or it may be a general, albeit overlooked, feature of prokaryote genomes.

RT-PCR Detection of HIV in Republic of Macedonia

Directory of Open Access Journals (Sweden)

Golubinka Bosevska

2008-11-01

Full Text Available The aim of the study was to detect HIV RNA in seropositive patients using RT-PCR method and thus, to establish PCR methodology in the routine laboratory works.The total of 33 examined persons were divided in two groups: 1 13 persons seropositive for HIV; and 2 20 healthy persons - randomly selected blood donors that made the case control group. The subjects age was between 25 and 52 years (average 38,5.ELFA test for combined detection of HIV p24 antigen and anti HIV-1 + 2 IgG and ELISA test for detection of antibodies against HIV-1 and HIV-2, were performed for each examined person. RNA from the whole blood was extracted using a commercial kit based on salt precipitation. Detection of HIV RNA was performed using RT-PCR kit. Following nested PCR, the product was separated by electrophoresis in 1,5 % agarose gel. The result was scored positive if the band of 210bp was visible regardless of intensity Measures of precaution were taken during all the steps of the work and HIV infected materials were disposed of accordingly.In the group of blood donors ELFA, ELISA and RT-PCR were negative. Assuming that prevalence of HIV infection is zero, the clinical specificity of RT-PCR is 100 %. The analytical specificity of RT-PCR method was tested against Hepatitis C and B, Human Papiloma Virus, Cytomegalovirus, Herpes Simplex Virus, Rubella Virus, Mycobacterium tuberculosis, Chlamydia trachomatis. None of these templates yielded amplicon. In the group of 13 seropositive persons, 33 samples were analyzed. HIV RNA was detected in 15 samples. ELISA and ELFA test were positive in all samples. Different aliquots of the samples were tested independently and showed the same results. After different periods of storing the RNA samples at -70°C, RT-PCR reaction was identical to the one performed initially. The obtained amplicons were maintained frozen at -20°C for a week and the subsequently performed electrophoresis was identical to the previous one. The reaction is

Integrated analysis of 454 and Illumina transcriptomic sequencing characterizes carbon flux and energy source for fatty acid synthesis in developing Lindera glauca fruits for woody biodiesel.

Science.gov (United States)

Lin, Zixin; An, Jiyong; Wang, Jia; Niu, Jun; Ma, Chao; Wang, Libing; Yuan, Guanshen; Shi, Lingling; Liu, Lili; Zhang, Jinsong; Zhang, Zhixiang; Qi, Ji; Lin, Shanzhi

2017-01-01

Lindera glauca fruit with high quality and quantity of oil has emerged as a novel potential source of biodiesel in China, but the molecular regulatory mechanism of carbon flux and energy source for oil biosynthesis in developing fruits is still unknown. To better develop fruit oils of L. glauca as woody biodiesel, a combination of two different sequencing platforms (454 and Illumina) and qRT-PCR analysis was used to define a minimal reference transcriptome of developing L. glauca fruits, and to construct carbon and energy metabolic model for regulation of carbon partitioning and energy supply for FA biosynthesis and oil accumulation. We first analyzed the dynamic patterns of growth tendency, oil content, FA compositions, biodiesel properties, and the contents of ATP and pyridine nucleotide of L. glauca fruits from seven different developing stages. Comprehensive characterization of transcriptome of the developing L. glauca fruit was performed using a combination of two different next-generation sequencing platforms, of which three representative fruit samples (50, 125, and 150 DAF) and one mixed sample from seven developing stages were selected for Illumina and 454 sequencing, respectively. The unigenes separately obtained from long and short reads (201, and 259, respectively, in total) were reconciled using TGICL software, resulting in a total of 60,031 unigenes (mean length = 1061.95 bp) to describe a transcriptome for developing L. glauca fruits. Notably, 198 genes were annotated for photosynthesis, sucrose cleavage, carbon allocation, metabolite transport, acetyl-CoA formation, oil synthesis, and energy metabolism, among which some specific transporters, transcription factors, and enzymes were identified to be implicated in carbon partitioning and energy source for oil synthesis by an integrated analysis of transcriptomic sequencing and qRT-PCR. Importantly, the carbon and energy metabolic model was well established for oil biosynthesis of developing L

Forført af hjernen?

DEFF Research Database (Denmark)

Lieberoth, Andreas

2013-01-01

Blandingen af offentlig interesse, gode mediehistorier og smarte sælgere har ført til sejlivede neuromyter, der ikke bare distraherer almindelige mennesker, men også fører til udbredte misforståelser blandt professionelle som lærere og plejepersonale. I bedste fald leder neuromyter til spild af t...

Novel expressed sequences identified in a model of androgen independent prostate cancer

Directory of Open Access Journals (Sweden)

Jones Steven JM

2007-01-01

Full Text Available Abstract Background Prostate cancer is the most frequently diagnosed cancer in American men, and few effective treatment options are available to patients who develop hormone-refractory prostate cancer. The molecular changes that occur to allow prostate cells to proliferate in the absence of androgens are not fully understood. Results Subtractive hybridization experiments performed with samples from an in vivo model of hormonal progression identified 25 expressed sequences representing novel human transcripts. Intriguingly, these 25 sequences have small open-reading frames and are not highly conserved through evolution, suggesting many of these novel expressed sequences may be derived from untranslated regions of novel transcripts or from non-coding transcripts. Examination of a large metalibrary of human Serial Analysis of Gene Expression (SAGE tags demonstrated that only three of these novel sequences had been previously detected. RT-PCR experiments confirmed that the 6 sequences tested were expressed in specific human tissues, as well as in clinical samples of prostate cancer. Further RT-PCR experiments for five of these fragments indicated they originated from large untranslated regions of unannotated transcripts. Conclusion This study underlines the value of using complementary techniques in the annotation of the human genome. The tissue-specific expression of 4 of the 6 clones tested indicates the expression of these novel transcripts is tightly regulated, and future work will determine the possible role(s these novel transcripts may play in the progression of prostate cancer.

A prospective randomized study of postoperative adjuvant chemo-radiotherapy (CT+RT) vs. radiotherapy(RT) alone in resected stage II and IIIA non-small cell lung cancer (NSCLC)

International Nuclear Information System (INIS)

Chang, Geol Lee; Joo, Hang Kim; Kyung, Young Chung; Doo, Yun Lee; Kil, Dong Kim; Won, Young Lee; Sung, Kyu Kim; Sei, Kyu Kim; Gwi, Eon Kim

1995-01-01

Objective: A prospective randomized study has been conducted to compare the results of treatment between CT+RT and RT alone as an adjuvant setting in completely resected stage II and IIIA NSCLC. Materials and Methods: Patients who had completely resected stage II and IIIA NSCLC were randomized into a CT+RT arm(arm A) and a RT alone arm(arm B) as an adjuvant setting after stratification according to cell type(squamous vs. non-squamous) and stage(II vs. IIIA). CT(Etoposide 100mg/m2 I.V. infusion d1-3, Cisplatin 20mg/m2 I.V. infusion d1-5, total 6cycles) was started in postop. 3 weeks with a 4 weeks interval. RT(5040cGy/5-6wks, 180cGy/fr) was started in postoperative 5 weeks after the first cycle of CT for group A and in postoperative 4 weeks for group B. A total of 69 patients were registered from Sep. 1990 to Jun. 1993. Sixty five of these patients were evaluable because 4 patients were ineligible due to distant metastasis before adjuvant treatment. Two patients who refused adjuvant treatment were included in this study to avoid selection bias. Results: Sixteen patients (48%) have received CT of more than 3 cycles and 51 patients(78%) have received RT of more than 50Gy. Four patients died due to treatment-related complications [broncho-pleural fistula 3(arm A:B=2:1), pneumonia 1(arm A)]. Survival and the patterns of failure are as follows: Conclusion: There is no statistical significance in either the overall survival or the patterns of failure between the CT+RT arm and RT alone arm as an adjuvant setting in resected stage II and IIIA NSCLC

Does tonality boost short-term memory in congenital amusia?

Science.gov (United States)

Albouy, Philippe; Schulze, Katrin; Caclin, Anne; Tillmann, Barbara

2013-11-06

Congenital amusia is a neuro-developmental disorder of music perception and production. Recent findings have demonstrated that this deficit is linked to an impaired short-term memory for tone sequences. As it has been shown before that non-musicians' implicit knowledge of musical regularities can improve short-term memory for tone information, the present study investigated if this type of implicit knowledge could also influence amusics' short-term memory performance. Congenital amusics and their matched controls, who were non-musicians, had to indicate whether sequences of five tones, presented in pairs, were the same or different; half of the pairs respected musical regularities (tonal sequences) and the other half did not (atonal sequences). As previously reported for non-musician participants, the control participants showed better performance (as measured with d') for tonal sequences than for atonal ones. While this improvement was not observed in amusics, both control and amusic participants showed faster response times for tonal sequences than for atonal sequences. These findings suggest that some implicit processing of tonal structures is potentially preserved in congenital amusia. This observation is encouraging as it strengthens the perspective to exploit implicit knowledge to help reducing pitch perception and memory deficits in amusia. © 2013 Elsevier B.V. All rights reserved.

De novo sequencing, assembly and characterization of antennal transcriptome of Anomala corpulenta Motschulsky (Coleoptera: Rutelidae.

Directory of Open Access Journals (Sweden)

Haoliang Chen

Full Text Available Anomala corpulenta is an important insect pest and can cause enormous economic losses in agriculture, horticulture and forestry. It is widely distributed in China, and both larvae and adults can cause serious damage. It is difficult to control this pest because the larvae live underground. Any new control strategy should exploit alternatives to heavily and frequently used chemical insecticides. However, little genetic research has been carried out on A. corpulenta due to the lack of genomic resources. Genomic resources could be produced by next generation sequencing technologies with low cost and in a short time. In this study, we performed de novo sequencing, assembly and characterization of the antennal transcriptome of A. corpulenta.Illumina sequencing technology was used to sequence the antennal transcriptome of A. corpulenta. Approximately 76.7 million total raw reads and about 68.9 million total clean reads were obtained, and then 35,656 unigenes were assembled. Of these unigenes, 21,463 of them could be annotated in the NCBI nr database, and, among the annotated unigenes, 11,154 and 6,625 unigenes could be assigned to GO and COG, respectively. Additionally, 16,350 unigenes could be annotated in the Swiss-Prot database, and 14,499 unigenes could map onto 258 pathways in the KEGG Pathway database. We also found 24 unigenes related to OBPs, 6 to CSPs, and in total 167 unigenes related to chemodetection. We analyzed 4 OBPs and 3CSPs sequences and their RT-qPCR results agreed well with their FPKM values.We produced the first large-scale antennal transcriptome of A. corpulenta, which is a species that has little genomic information in public databases. The identified chemodetection unigenes can promote the molecular mechanistic study of behavior in A. corpulenta. These findings provide a general sequence resource for molecular genetics research on A. corpulenta.

Pärt, Arvo. Fratres / Rainer Wagner

Index Scriptorium Estoniae

Wagner, Rainer

1995-01-01

Uuest heliplaadist "Pärt, Arvo. Fratres, Cantus (in Memory of Benjamin Britten), summa, Festina Lente; Peter Manning, France Springuel, Mireille Gleizes, Huub Righarts, I Fiamminghi, Rudolf Werthen" (AD: 1994) Telarc/in-akustik CD 80 387 (WD: 79'00")

Car sequencing is NP-hard: a short proof

OpenAIRE

B Estellon; F Gardi

2013-01-01

In this note, a new proof is given that the car sequencing (CS) problem is NP-hard. Established from the Hamiltonian Path problem, the reduction is direct while closing some gaps remaining in the previous NP-hardness results. Since CS is studied in many operational research courses, this result and its proof are particularly interesting for teaching purposes.

Whole Genome Sequencing of Enterovirus species C Isolates by High-throughput Sequencing: Development of Generic Primers

Directory of Open Access Journals (Sweden)

Maël Bessaud

2016-08-01

Full Text Available Enteroviruses are among the most common viruses infecting humans and can cause diverse clinical syndromes ranging from minor febrile illness to severe and potentially fatal diseases. Enterovirus species C (EV-C consists of more than 20 types, among which the 3 serotypes of polioviruses, the etiological agents of poliomyelitis, are included. Biodiversity and evolution of EV-C genomes are shaped by frequent recombination events. Therefore, identification and characterization of circulating EV-C strains require the sequencing of different genomic regions.A simple method was developed to sequence quickly the entire genome of EV-C isolates. Four overlapping fragments were produced separately by RT-PCR performed with generic primers. The four amplicons were then pooled and purified prior to be sequenced by high-throughput technique.The method was assessed on a panel of EV-Cs belonging to a wide-range of types. It can be used to determine full-length genome sequences through de novo assembly of thousands of reads. It was also able to discriminate reads from closely related viruses in mixtures.By decreasing the workload compared to classical Sanger-based techniques, this method will serve as a precious tool for sequencing large panels of EV-Cs isolated in cell cultures during environmental surveillance or from patients, including vaccine-derived polioviruses.

Multiple regulatory mechanisms of hepatocyte growth factor expression in malignant cells with a short poly(dA) sequence in the HGF gene promoter.

Science.gov (United States)

Sakai, Kazuko; Takeda, Masayuki; Okamoto, Isamu; Nakagawa, Kazuhiko; Nishio, Kazuto

2015-01-01

Hepatocyte growth factor (HGF) expression is a poor prognostic factor in various types of cancer. Expression levels of HGF have been reported to be regulated by shorter poly(dA) sequences in the promoter region. In the present study, the poly(dA) mononucleotide tract in various types of human cancer cell lines was examined and compared with the HGF expression levels in those cells. Short deoxyadenosine repeat sequences were detected in five of the 55 cell lines used in the present study. The H69, IM95, CCK-81, Sui73 and H28 cells exhibited a truncated poly(dA) sequence in which the number of poly(dA) repeats was reduced by ≥5 bp. Two of the cell lines exhibited high HGF expression, determined by reverse transcription quantitative polymerase chain reaction and enzyme-linked immunosorbent assay. The CCK-81, Sui73 and H28 cells with shorter poly(dA) sequences exhibited low HGF expression. The cause of the suppression of HGF expression in the CCK-81, Sui73 and H28 cells was clarified by two approaches, suppression by methylation and single nucleotide polymorphisms in the HGF gene. Exposure to 5-Aza-dC, an inhibitor of DNA methyltransferase 1, induced an increased expression of HGF in the CCK-81 cells, but not in the other cells. Single-nucleotide polymorphism (SNP) rs72525097 in intron 1 was detected in the Sui73 and H28 cells. Taken together, it was found that the defect of poly(dA) in the HGF promoter was present in various types of cancer, including lung, stomach, colorectal, pancreas and mesothelioma. The present study proposes the negative regulation mechanisms by methylation and SNP in intron 1 of HGF for HGF expression in cancer cells with short poly(dA).

Short term reproducibility of a high contrast 3-D isotropic optic nerve imaging sequence in healthy controls

Science.gov (United States)

Harrigan, Robert L.; Smith, Alex K.; Mawn, Louise A.; Smith, Seth A.; Landman, Bennett A.

2016-03-01

The optic nerve (ON) plays a crucial role in human vision transporting all visual information from the retina to the brain for higher order processing. There are many diseases that affect the ON structure such as optic neuritis, anterior ischemic optic neuropathy and multiple sclerosis. Because the ON is the sole pathway for visual information from the retina to areas of higher level processing, measures of ON damage have been shown to correlate well with visual deficits. Increased intracranial pressure has been shown to correlate with the size of the cerebrospinal fluid (CSF) surrounding the ON. These measures are generally taken at an arbitrary point along the nerve and do not account for changes along the length of the ON. We propose a high contrast and high-resolution 3-D acquired isotropic imaging sequence optimized for ON imaging. We have acquired scan-rescan data using the optimized sequence and a current standard of care protocol for 10 subjects. We show that this sequence has superior contrast-to-noise ratio to the current standard of care while achieving a factor of 11 higher resolution. We apply a previously published automatic pipeline to segment the ON and CSF sheath and measure the size of each individually. We show that these measures of ON size have lower short- term reproducibility than the population variance and the variability along the length of the nerve. We find that the proposed imaging protocol is (1) useful in detecting population differences and local changes and (2) a promising tool for investigating biomarkers related to structural changes of the ON.

Comparison of RT-PCR-Dot blot hybridization based on radioisotope 32P with conventional RT-PCR and commercial ELISA Assays for blood screening of HIV-1

International Nuclear Information System (INIS)

Maria Lina R; Andi Yasmon

2011-01-01

There are many commercial ELISA and rapid test kits that have been used for blood screening; however, the kits can give false positive and negative results. Therefore, RT-PCR (Reverse Transcription Polymerase Chain Reaction) - Dot Blot Hybridization based on radioisotope 32 P (RDBR) method was developed in this research, to compare the method with the conventional RT-PCR and commercial ELISA Enzyme-Linked lmmunosorbent Assay) kit. This method is efficient for screening of large blood specimens and surveillance study. Eighty seven samples were used and serum of the samples were tested by ELISA to detect HIV-1. The HIV-l RNA genome was extracted from plasma samples and tested using the RT-PCR and RDBR methods. Of 87 samples that were tested, the rates of positive testing of the RT-PCR, the RDBR, and the ELISA were 71.26%, 74.71%, and 80.46%, respectively. The RDBR (a combination of RTPCR and dot blot hybridization) was more sensitive than conventional RT-PCR by showing 3.45% in increase number of positive specimens. The results showed that of 9 samples (10.34%) were negative RDBR and positive ELISA, while 4 samples (4.60%) were negative ELISA and positive RDBR. The two methods showed slightly difference in the results but further validation is still needed. However, RDBR has high potential as an alternative method for screening of blood in large quantities when compared to method of conventional RT-PCR and ELISA. (author)

NASA/SPoRT's GOES-R Activities in Support of Product Development, Management, and Training

Science.gov (United States)

Fuell, K. K.; Jedlovec, G.; Molthan, A.; Stano, G. T.

2012-12-01

The NASA Short-term Prediction Research and Transition (SPoRT) Center supports many activities within the GOES-R Proving Grounds (PG). These include the development of imagery from existing instrumentation as a proxy to future Advanced Baseline Imager (ABI) capabilities on GOES-R. The Moderate Resolution Imaging Spectroradiometer (MODIS) and the Visible/Infrared Imager/Radiometer Suite (VIIRS) instruments are used to provide a glimpse of the multi-spectral capabilities that will become the norm as the number of channels and data rate dramatically increase with GOES-R. The NOAA/NWS has plans to provide operational users with all ABI channels at the highest resolution. Data fusion of individual channels into composite red, green, and blue imagery products will assist the end user with this future wave of information. While increasing the efficiency in the operational use of ABI channels, these composites provide only qualitative information. Within the GOES-R PG, SPoRT and other partners are exploring ways to include quantitative information as part of the composite imagery. However, limitations in local hardware processing and/or data bandwidth for users of the GOES-R data stream are challenges to overcome. This presentation will discuss the creation of these composite images as well as possible solutions to address these processing challenges. In a similar manner the Geostationary Lightning Mapper (GLM) to be launched on GOES-R presents several data management challenges. The GLM is a pioneering instrument to quantify total lightning from a geostationary platform. The expected data frequency from the GLM is to be at a sub-minute interval. Users of such a data set may have little experience in handling such a rapid update of information. To assist users, SPoRT is working with the NWS to develop tools within the user's decision support system to allow tracking and analysis of total lightning from a storm-based perspective. This presentation will discuss the

NASA/SPoRT's GOES-R Activities in Support of Product Development, Management, and Training

Science.gov (United States)

Fuell, Kevin K.; Jedlovec, Gary; Molthan, Andrew L.; Stano, Geoffrey T.

2012-01-01

The NASA Short-term Prediction Research and Transition (SPoRT) Center supports many activities within the GOES-R Proving Grounds (PG). These include the development of imagery from existing instrumentation as a proxy to future Advanced Baseline Imager (ABI) capabilities on GOES-R. The Moderate Resolution Imaging Spectroradiometer (MODIS) and the Visible/Infrared Imager/Radiometer Suite (VIIRS) instruments are used to provide a glimpse of the multi-spectral capabilities that will become the norm as the number of channels and data rate dramatically increase with GOES-R. The NOAA/NWS has plans to provide operational users with all ABI channels at the highest resolution. Data fusion of individual channels into composite red, green, and blue imagery products will assist the end user with this future wave of information. While increasing the efficiency in the operational use of ABI channels, these composites provide only qualitative information. Within the GOES-R PG, SPoRT and other partners are exploring ways to include quantitative information as part of the composite imagery. However, limitations in local hardware processing and/or data bandwidth for users of the GOES-R data stream are challenges to overcome. This presentation will discuss the creation of these composite images as well as possible solutions to address these processing challenges. In a similar manner the Geostationary Lightning Mapper (GLM) to be launched on GOES-R presents several data management challenges. The GLM is a pioneering instrument to quantify total lightning from a geostationary platform. The expected data frequency from the GLM is to be at a sub-minute interval. Users of such a data set may have little experience in handling such a rapid update of information. To assist users, SPoRT is working with the NWS to develop tools within the user fs decision support system to allow tracking and analysis of total lightning from a storm-based perspective. This presentation will discuss the

Natural and Enhanced Attenuation of Chlorinated Solvents Using RT3D

Energy Technology Data Exchange (ETDEWEB)

Johnson, Christian D.; Truex, Michael J.; Clement, T P.

2006-07-25

RT3D (Reactive Transport in 3-Dimensions) is a reactive transport code that can be applied to model solute fate and transport for many different purposes. This document specifically addresses application of RT3D for modeling related to evaluation and implementation of Monitored Natural Attenuation (MNA). Selection of MNA as a remedy requires an evaluation process to demonstrate that MNA will meet the remediation goals. The U.S. EPA, through the Office of Solid Waste and Emergency Response (OSWER) Directive 9200.4?17P, provides the regulatory context for the evaluation and implementation of MNA. In a complementary fashion, the context for using fate and transport modeling as part of MNA evaluation is described in the EPA?s technical protocol for chlorinated solvent MNA, the Scenarios Evaluation Tool for Chlorinated Solvent MNA, and in this document. The intent of this document is to describe (1) the context for applying RT3D for chlorinated solvent MNA and (2) the attenuation processes represented in RT3D, (3) dechlorination reactions that may occur, and (4) the general approach for using RT3D reaction modules (including a summary of the RT3D reaction modules that are available) to model fate and transport of chlorinated solvents as part of MNA or for combinations of MNA and selected types of active remediation.

Selection of reference genes for quantitative RT-PCR (RT-qPCR) analysis of rat tissues under physiological and toxicological conditions

DEFF Research Database (Denmark)

Svingen, Terje; Letting, Heidi; Hadrup, Niels

2015-01-01

In biological research the analysis of gene expression levels in cells and tissues can be a powerful tool to gain insights into biological processes. For this, quantitative RT-PCR (RT-qPCR) is a popular method that often involve the use of constitutively expressed endogenous reference (or...... ‘housekeeping’) gene for normalization of data. Thus, it is essential to use reference genes that have been verified to be stably expressed within the specific experimental setting. Here, we have analysed the expression stability of 12 commonly used reference genes (Actb, B2m, Gapdh, Hprt, Pgk1, Rn18s, Rpl13a...

RT-PCR detection of Candida albicans ALS gene expression in the reconstituted human epithelium (RHE) model of oral candidiasis and in model biofilms.

Science.gov (United States)

Green, Clayton B; Cheng, Georgina; Chandra, Jyotsna; Mukherjee, Pranab; Ghannoum, Mahmoud A; Hoyer, Lois L

2004-02-01

An RT-PCR assay was developed to analyse expression patterns of genes in the Candida albicans ALS (agglutinin-like sequence) family. Inoculation of a reconstituted human buccal epithelium (RHE) model of mucocutaneous candidiasis with strain SC5314 showed destruction of the epithelial layer by C. albicans and also formation of an upper fungal layer that had characteristics similar to a biofilm. RT-PCR analysis of total RNA samples extracted from C. albicans-inoculated buccal RHE showed that ALS1, ALS2, ALS3, ALS4, ALS5 and ALS9 were consistently detected over time as destruction of the RHE progressed. Detection of transcripts from ALS7, and particularly from ALS6, was more sporadic, but not associated with a strictly temporal pattern. The expression pattern of ALS genes in C. albicans cultures used to inoculate the RHE was similar to that observed in the RHE model, suggesting that contact of C. albicans with buccal RHE does little to alter ALS gene expression. RT-PCR analysis of RNA samples extracted from model denture and catheter biofilms showed similar gene expression patterns to the buccal RHE specimens. Results from the RT-PCR analysis of biofilm RNA specimens were consistent between various C. albicans strains during biofilm development and were comparable to gene expression patterns in planktonic cells. The RT-PCR assay described here will be useful for analysis of human clinical specimens and samples from other disease models. The method will provide further insight into the role of ALS genes and their encoded proteins in the diverse interactions between C. albicans and its host.

Massively parallel sequencing of forensic STRs

DEFF Research Database (Denmark)

Parson, Walther; Ballard, David; Budowle, Bruce

2016-01-01

The DNA Commission of the International Society for Forensic Genetics (ISFG) is reviewing factors that need to be considered ahead of the adoption by the forensic community of short tandem repeat (STR) genotyping by massively parallel sequencing (MPS) technologies. MPS produces sequence data that...

Nucleotide sequence of the coat protein gene of the Skierniewice isolate of plum pox virus (PPV)

International Nuclear Information System (INIS)

Wypijewski, K.; Musial, W.; Augustyniak, J.; Malinowski, T.

1994-01-01

The coat protein (CP) gene of the Skierniewice isolate of plum pox virus (PPV-S) has been amplified using the reverse transcription - polymerase chain reaction (RT-PCR), cloned and sequenced. The nucleotide sequence of the gene and the deduced amino-acid sequences of PPV-S CP were compared with those of other PPV strains. The nucleotide sequence showed very high homology to most of the published sequences. The motif: Asp-Ala-Gly (DAG), important for the aphid transmissibility, was present in the amino-acid sequence. Our isolate did not react in ELISA with monoclonal antibodies MAb06 supposed to be specific for PPV-D. (author). 32 refs, 1 fig., 2 tabs

Milan Kundera poeediparukaga faun / Kärt Hellerma

Index Scriptorium Estoniae

Hellerma, Kärt, 1956-

1996-01-01

Arvustus: Kundera, Milan. Surematus. Tln. : Monokkel, 1995. Ilmunud ka kogumikus: Hellerma, Kärt. Kohanenud kirjandus : valik kirjanduskriitikat 1987-2006. Eesti Keele Sihtasutus : Tallinn, 2006. Lk. 176-179

libgapmis: extending short-read alignments.

Science.gov (United States)

Alachiotis, Nikolaos; Berger, Simon; Flouri, Tomáš; Pissis, Solon P; Stamatakis, Alexandros

2013-01-01

A wide variety of short-read alignment programmes have been published recently to tackle the problem of mapping millions of short reads to a reference genome, focusing on different aspects of the procedure such as time and memory efficiency, sensitivity, and accuracy. These tools allow for a small number of mismatches in the alignment; however, their ability to allow for gaps varies greatly, with many performing poorly or not allowing them at all. The seed-and-extend strategy is applied in most short-read alignment programmes. After aligning a substring of the reference sequence against the high-quality prefix of a short read--the seed--an important problem is to find the best possible alignment between a substring of the reference sequence succeeding and the remaining suffix of low quality of the read--extend. The fact that the reads are rather short and that the gap occurrence frequency observed in various studies is rather low suggest that aligning (parts of) those reads with a single gap is in fact desirable. In this article, we present libgapmis, a library for extending pairwise short-read alignments. Apart from the standard CPU version, it includes ultrafast SSE- and GPU-based implementations. libgapmis is based on an algorithm computing a modified version of the traditional dynamic-programming matrix for sequence alignment. Extensive experimental results demonstrate that the functions of the CPU version provided in this library accelerate the computations by a factor of 20 compared to other programmes. The analogous SSE- and GPU-based implementations accelerate the computations by a factor of 6 and 11, respectively, compared to the CPU version. The library also provides the user the flexibility to split the read into fragments, based on the observed gap occurrence frequency and the length of the read, thereby allowing for a variable, but bounded, number of gaps in the alignment. We present libgapmis, a library for extending pairwise short-read alignments. We

WE-H-207B-00: MRgRT

Energy Technology Data Exchange (ETDEWEB)

NONE

2016-06-15

In recent years, steady progress has been made towards the implementation of MRI in external beam radiation therapy for processes ranging from treatment simulation to in-room guidance. Novel procedures relying mostly on MR data are currently implemented in the clinic. This session will cover topics such as (a) commissioning and quality control of the MR in-room imagers and simulators specific to RT, (b) treatment planning requirements, constraints and challenges when dealing with various MR data, (c) quantification of organ motion with an emphasis on treatment delivery guidance, and (d) MR-driven strategies for adaptive RT workflows. The content of the session was chosen to address both educational and practical key aspects of MR guidance. Learning Objectives: Good understanding of MR testing recommended for in-room MR imaging as well as image data validation for RT chain (e.g. image transfer, filtering for consistency, spatial accuracy, manipulation for task specific); Familiarity with MR-based planning procedures: motivation, core workflow requirements, current status, challenges; Overview of the current methods for the quantification of organ motion; Discussion on approaches for adaptive treatment planning and delivery. T. Stanescu - License agreement with Modus Medical Devices to develop a phantom for the quantification of MR image system-related distortions.; T. Stanescu, N/A.

WE-H-207B-00: MRgRT

International Nuclear Information System (INIS)

2016-01-01

In recent years, steady progress has been made towards the implementation of MRI in external beam radiation therapy for processes ranging from treatment simulation to in-room guidance. Novel procedures relying mostly on MR data are currently implemented in the clinic. This session will cover topics such as (a) commissioning and quality control of the MR in-room imagers and simulators specific to RT, (b) treatment planning requirements, constraints and challenges when dealing with various MR data, (c) quantification of organ motion with an emphasis on treatment delivery guidance, and (d) MR-driven strategies for adaptive RT workflows. The content of the session was chosen to address both educational and practical key aspects of MR guidance. Learning Objectives: Good understanding of MR testing recommended for in-room MR imaging as well as image data validation for RT chain (e.g. image transfer, filtering for consistency, spatial accuracy, manipulation for task specific); Familiarity with MR-based planning procedures: motivation, core workflow requirements, current status, challenges; Overview of the current methods for the quantification of organ motion; Discussion on approaches for adaptive treatment planning and delivery. T. Stanescu - License agreement with Modus Medical Devices to develop a phantom for the quantification of MR image system-related distortions.; T. Stanescu, N/A

PASSIOMA: Exploring Expressed Sequence Tags during Flower Development in Passiflora spp.

Directory of Open Access Journals (Sweden)

Lucas Cutri

2012-01-01

Full Text Available The genus Passiflora provides a remarkable example of floral complexity and diversity. The extreme variation of Passiflora flower morphologies allowed a wide range of interactions with pollinators to evolve. We used the analysis of expressed sequence tags (ESTs as an approach for the characterization of genes expressed during Passiflora reproductive development. Analyzing the Passiflora floral EST database (named PASSIOMA, we found sequences showing significant sequence similarity to genes known to be involved in reproductive development such as MADS-box genes. Some of these sequences were studied using RT-PCR and in situ hybridization confirming their expression during Passiflora flower development. The detection of these novel sequences can contribute to the development of EST-based markers for important agronomic traits as well as to the establishment of genomic tools to study the naturally occurring floral diversity among Passiflora species.

On the Origin of Reverse Transcriptase-Using CRISPR-Cas Systems and Their Hyperdiverse, Enigmatic Spacer Repertoires

Directory of Open Access Journals (Sweden)

Sukrit Silas

2017-07-01

Full Text Available Cas1 integrase is the key enzyme of the clustered regularly interspaced short palindromic repeat (CRISPR-Cas adaptation module that mediates acquisition of spacers derived from foreign DNA by CRISPR arrays. In diverse bacteria, the cas1 gene is fused (or adjacent to a gene encoding a reverse transcriptase (RT related to group II intron RTs. An RT-Cas1 fusion protein has been recently shown to enable acquisition of CRISPR spacers from RNA. Phylogenetic analysis of the CRISPR-associated RTs demonstrates monophyly of the RT-Cas1 fusion, and coevolution of the RT and Cas1 domains. Nearly all such RTs are present within type III CRISPR-Cas loci, but their phylogeny does not parallel the CRISPR-Cas type classification, indicating that RT-Cas1 is an autonomous functional module that is disseminated by horizontal gene transfer and can function with diverse type III systems. To compare the sequence pools sampled by RT-Cas1-associated and RT-lacking CRISPR-Cas systems, we obtained samples of a commercially grown cyanobacterium—Arthrospira platensis. Sequencing of the CRISPR arrays uncovered a highly diverse population of spacers. Spacer diversity was particularly striking for the RT-Cas1-containing type III-B system, where no saturation was evident even with millions of sequences analyzed. In contrast, analysis of the RT-lacking type III-D system yielded a highly diverse pool but reached a point where fewer novel spacers were recovered as sequencing depth was increased. Matches could be identified for a small fraction of the non-RT-Cas1-associated spacers, and for only a single RT-Cas1-associated spacer. Thus, the principal source(s of the spacers, particularly the hypervariable spacer repertoire of the RT-associated arrays, remains unknown.

Adaptive Basis Selection for Exponential Family Smoothing Splines with Application in Joint Modeling of Multiple Sequencing Samples

OpenAIRE

Ma, Ping; Zhang, Nan; Huang, Jianhua Z.; Zhong, Wenxuan

2017-01-01

Second-generation sequencing technologies have replaced array-based technologies and become the default method for genomics and epigenomics analysis. Second-generation sequencing technologies sequence tens of millions of DNA/cDNA fragments in parallel. After the resulting sequences (short reads) are mapped to the genome, one gets a sequence of short read counts along the genome. Effective extraction of signals in these short read counts is the key to the success of sequencing technologies. No...

The First Report of miRNAs from a Thysanopteran Insect, Thrips palmi Karny Using High-Throughput Sequencing.

Directory of Open Access Journals (Sweden)

K B Rebijith

Full Text Available Thrips palmi Karny (Thysanoptera: Thripidae is the sole vector of Watermelon bud necrosis tospovirus, where the crop loss has been estimated to be around USD 50 million annually. Chemical insecticides are of limited use in the management of T. palmi due to the thigmokinetic behaviour and development of high levels of resistance to insecticides. There is an urgent need to find out an effective futuristic management strategy, where the small RNAs especially microRNAs hold great promise as a key player in the growth and development. miRNAs are a class of short non-coding RNAs involved in regulation of gene expression either by mRNA cleavage or by translational repression. We identified and characterized a total of 77 miRNAs from T. palmi using high-throughput deep sequencing. Functional classifications of the targets for these miRNAs revealed that majority of them are involved in the regulation of transcription and translation, nucleotide binding and signal transduction. We have also validated few of these miRNAs employing stem-loop RT-PCR, qRT-PCR and Northern blot. The present study not only provides an in-depth understanding of the biological and physiological roles of miRNAs in governing gene expression but may also lead as an invaluable tool for the management of thysanopteran insects in the future.

Peptomics, identification of novel cationic Arabidopsis peptides with conserved sequence motifs

DEFF Research Database (Denmark)

Olsen, Addie Nina; Mundy, John; Skriver, Karen

2002-01-01

Arabidopsis family of 34 genes. The predicted peptides are characterized by a conserved C-terminal sequence motif and additional primary structure conservation in a core region. The majority of these genes had not previously been annotated. A subset of the predicted peptides show high overall sequence...... similarity to Rapid Alkalinization Factor (RALF), a peptide isolated from tobacco. We therefore refer to this peptide family as RALFL for RALF-Like. RT-PCR analysis confirmed that several of the Arabidopsis genes are expressed and that their expression patterns vary. The identification of a large gene family...

Sequence-to-Sequence Prediction of Vehicle Trajectory via LSTM Encoder-Decoder Architecture

OpenAIRE

Park, Seong Hyeon; Kim, ByeongDo; Kang, Chang Mook; Chung, Chung Choo; Choi, Jun Won

2018-01-01

In this paper, we propose a deep learning based vehicle trajectory prediction technique which can generate the future trajectory sequence of surrounding vehicles in real time. We employ the encoder-decoder architecture which analyzes the pattern underlying in the past trajectory using the long short-term memory (LSTM) based encoder and generates the future trajectory sequence using the LSTM based decoder. This structure produces the $K$ most likely trajectory candidates over occupancy grid ma...

Quantitative real-time RT-PCR and chromogenic in situ hybridization

DEFF Research Database (Denmark)

Rosa, Fabíola E; Silveira, Sara M; Silveira, Cássia G T

2009-01-01

. METHODS: To elucidate the molecular profile of HER-2 status, mRNA and protein expression in 75 invasive breast carcinomas were analyzed by real time quantitative RT-PCR (qRT-PCR) and IHC, respectively. Amplifications were evaluated in 43 of these cases by CISH and in 11 by FISH. RESULTS: The concordance...

Direct recovery of infectious Pestivirus from a full-length RT-PCR amplicon

DEFF Research Database (Denmark)

Rasmussen, Thomas Bruun; Reimann, Ilona; Hoffmann, Bernd

2008-01-01

This study describes the use of a novel and rapid long reverse transcription (RT)-PCR for the generation of infectious full-length cDNA of pestiviruses. To produce rescued viruses, full-length RT-PCR amplicons of 12.3 kb, including a T7-promotor, were transcribed directly in vitro, and the result......This study describes the use of a novel and rapid long reverse transcription (RT)-PCR for the generation of infectious full-length cDNA of pestiviruses. To produce rescued viruses, full-length RT-PCR amplicons of 12.3 kb, including a T7-promotor, were transcribed directly in vitro......, and the resulting RNA transcripts were electroporated into ovine cells. Infectious virus was obtained after one cell culture passage. The rescued viruses had a phenotype similar to the parental Border Disease virus strain. Therefore, direct generation of infectious pestiviruses from full-length RT-PCR cDNA products...

Effect of low frequency ultrasound on combined rt-PA and eptifibatide thrombolysis in human clots.

Science.gov (United States)

Meunier, Jason M; Holland, Christy K; Pancioli, Arthur M; Lindsell, Christopher J; Shaw, George J

2009-01-01

Fibrinolytics such as recombinant tissue plasminogen activator (rt-PA) are used to treat thrombotic disease such as acute myocardial infarction (AMI) and ischemic stroke. Interest in increasing efficacy and reducing side effects has led to the study of adjuncts such as GP IIb-IIIa inhibitors and ultrasound (US) enhanced thrombolysis. Currently, GP IIb-IIIa inhibitor and fibrinolytic treatment are often used in AMI, and are under investigation for stroke treatment. However, little is known of the efficacy of combined GP IIb-IIIa inhibitor, fibrinolytic and ultrasound treatment. We measure the lytic efficacy of rt-PA, eptifibatide (Epf) and 120 kHz ultrasound treatment in an in-vitro human clot model. Blood was drawn from 15 subjects after IRB approval. Clots were made in 20 microL pipettes, and placed in a water tank for microscopic visualization during lytic treatment. Clots were exposed to control, rt-PA (rt-PA), eptifibatide (Epf), or rt-PA+eptifibatide (rt-PA + Epf), with (+US) or without (-US) ultrasound for 30 minutes at 37 degrees C in human plasma. Clot lysis was measured over time, using a microscopic imaging technique. The fractional clot loss (FCL) and initial lytic rate (LR) were used to quantify lytic efficacy. LR values for (- US) treated clots were 0.8+/-0.1(control), 1.8+/-0.3 (Epf), 1.5+/-0.2 (rt-PA), and 1.3+/-0.4 (rt-PA + Epf) (% clot width/minute) respectively. In comparison, the (+ US) group exhibited LR values of 1.6+/-0.2 (control), 4.3+/-0.4 (Epf), 6.3+/-0.4 (rt-PA), and 4.6+/-0.6 (rt-PA + Epf). For (- US) treated clots, FCL was 6.0+/-0.8 (control), 9.2+/-2.5 (Epf), 15.6+/-1.7 (rt-PA), and 28.0+/-2.2% (rt-PA + Epf) respectively. FCL for (+ US) clots was 13.5+/-2.4 (control), 20.7+/-6.4 (Epf), 44.4+/-3.6 (rt-PA) and 30.3+/-3.6% (rt-PA + Epf) respectively. Although the addition of eptifibatide enhances the in-vitro lytic efficacy of rt-PA in the absence of ultrasound, the efficacy of ultrasound and rt-PA is greater than that of combined

Quantitative RT-PCR based platform for rapid quantification of the transcripts of highly homologous multigene families and their members during grain development

DEFF Research Database (Denmark)

Kaczmarczyk, Agnieszka Ewa; Bowra, Steve; Elek, Zoltan

2012-01-01

expression combined with genetic variation in large multigene families with high homology among the alleles is very challenging. Results We designed a rapid qRT-PCR system with the aim of characterising the variation in the expression of hordein genes families. All the known D-, C-, B-, and gamma......-hordein sequences coding full length open reading frames were collected from commonly available databases. Phylogenetic analysis was performed and the members of the different hordein families were classified into subfamilies. Primer sets were designed to discriminate the gene expression level of whole families...... and its subgroups. More over the results indicate the genotypic specific gene expression. Conclusions Quantitative RT-PCR with SYBR Green labelling can be a useful technique to follow gene expression levels of large gene families with highly homologues members. We showed variation in the temporal...

Pärt, Arvo: "Litany" / Patric Wiklacz

Index Scriptorium Estoniae

Wiklacz, Patric

1996-01-01

Uuest heliplaadist "Pärt, Arvo: "Litany". Litany. Psalom. Trisagion. The Hilliard Ensemble, Orchestre de chambre de Tallinn, Choeur de chambre Philharmonique d'Estonie, Tõnu Kaljuste. Orchestre de chambre de Lituanie, Saulius Sondeckis. ECM New Series ECM 1592, distribution Polygram 449 810-2 (CD:158F)

Pärt, Arvo: "Litany" / Barry Witherden

Index Scriptorium Estoniae

Witherden, Barry

1996-01-01

Uuest heliplaadist "Pärt, Arvo: "Litany". Litany. Psalom. Trisagion. The Hilliard Ensemble, Tallinn Chamber Orchestra, Estonian Philharmonic Chamber Orchestra, Estonian Philharmonic Chamber Choir, Tõnu Kaljuste. Orchestre de chambre de Lituanie, Saulius Sondeckis". ECM New Series ECM 1592, distribution Polygram 449 810-2 (CD:158F)

Have you tried spermine? A rapid and cost-effective method to eliminate dextran sodium sulfate inhibition of PCR and RT-PCR.

Science.gov (United States)

Krych, Łukasz; Kot, Witold; Bendtsen, Katja M B; Hansen, Axel K; Vogensen, Finn K; Nielsen, Dennis S

2018-01-01

The Dextran Sulfate Sodium (DSS) induced colitis mouse model is commonly used to investigate human inflammatory bowel disease (IBD). Nucleic acid extracts originating from these animals are often contaminated with DSS, which is a strong inhibitor of many enzymatic based molecular biology reactions including PCR and reverse-transcription (RT). Methods for removing DSS from nucleic acids extracts exist for RNA, but no effective protocol for DNA or cDNA is currently available. However, spermine has previously been shown to be an effective agent for counteracting DSS inhibition of polynucleotide kinase, which led to the hypothesis, that spermine could be used to counteract DSS inhibition of PCR and RT. We investigated the means of adding spermine in an adequate concentration to PCR based protocols (including qPCR, two-step RT-qPCR, and amplicon sequencing library preparation) to remove DSS inhibition. Within the range up to 0.01g/L, spermine can be added to PCR/qPCR or RT prophylactically without a significant reduction of reaction efficiency. Addition of spermine at the concentration of 0.08g/L can be used to recover qualitative PCR signal inhibited by DSS in concentrations up to 0.32g/L. For optimal quantitative analysis, the concentration of spermine requires fine adjustment. Hence, we present here a simple fluorometric based method for adjusting the concentration of spermine ensuring an optimal efficiency of the reaction exposed to an unknown concentration of DSS. In conclusion, we demonstrate a cost effective and easy method to counteract DSS inhibition in PCR and two-step RT-qPCR. Fixed or fine-tuned concentrations of spermine can be administered depending on the qualitative or quantitative character of the analysis. Copyright © 2017 Elsevier B.V. All rights reserved.

BarraCUDA - a fast short read sequence aligner using graphics processing units

Directory of Open Access Journals (Sweden)

Klus Petr

2012-01-01

Full Text Available Abstract Background With the maturation of next-generation DNA sequencing (NGS technologies, the throughput of DNA sequencing reads has soared to over 600 gigabases from a single instrument run. General purpose computing on graphics processing units (GPGPU, extracts the computing power from hundreds of parallel stream processors within graphics processing cores and provides a cost-effective and energy efficient alternative to traditional high-performance computing (HPC clusters. In this article, we describe the implementation of BarraCUDA, a GPGPU sequence alignment software that is based on BWA, to accelerate the alignment of sequencing reads generated by these instruments to a reference DNA sequence. Findings Using the NVIDIA Compute Unified Device Architecture (CUDA software development environment, we ported the most computational-intensive alignment component of BWA to GPU to take advantage of the massive parallelism. As a result, BarraCUDA offers a magnitude of performance boost in alignment throughput when compared to a CPU core while delivering the same level of alignment fidelity. The software is also capable of supporting multiple CUDA devices in parallel to further accelerate the alignment throughput. Conclusions BarraCUDA is designed to take advantage of the parallelism of GPU to accelerate the alignment of millions of sequencing reads generated by NGS instruments. By doing this, we could, at least in part streamline the current bioinformatics pipeline such that the wider scientific community could benefit from the sequencing technology. BarraCUDA is currently available from http://seqbarracuda.sf.net

BarraCUDA - a fast short read sequence aligner using graphics processing units

LENUS (Irish Health Repository)

Klus, Petr

2012-01-13

Abstract Background With the maturation of next-generation DNA sequencing (NGS) technologies, the throughput of DNA sequencing reads has soared to over 600 gigabases from a single instrument run. General purpose computing on graphics processing units (GPGPU), extracts the computing power from hundreds of parallel stream processors within graphics processing cores and provides a cost-effective and energy efficient alternative to traditional high-performance computing (HPC) clusters. In this article, we describe the implementation of BarraCUDA, a GPGPU sequence alignment software that is based on BWA, to accelerate the alignment of sequencing reads generated by these instruments to a reference DNA sequence. Findings Using the NVIDIA Compute Unified Device Architecture (CUDA) software development environment, we ported the most computational-intensive alignment component of BWA to GPU to take advantage of the massive parallelism. As a result, BarraCUDA offers a magnitude of performance boost in alignment throughput when compared to a CPU core while delivering the same level of alignment fidelity. The software is also capable of supporting multiple CUDA devices in parallel to further accelerate the alignment throughput. Conclusions BarraCUDA is designed to take advantage of the parallelism of GPU to accelerate the alignment of millions of sequencing reads generated by NGS instruments. By doing this, we could, at least in part streamline the current bioinformatics pipeline such that the wider scientific community could benefit from the sequencing technology. BarraCUDA is currently available from http:\\/\\/seqbarracuda.sf.net

Serotype identification and VP1 coding sequence analysis of foot-and-mouth disease virus from outbreaks in Eastern and Northern Uganda in 2008/9

DEFF Research Database (Denmark)

Kasambula, L.; Belsham, Graham; Siegismund, H. R.

2012-01-01

regions, and the presence of FMDV RNA in these samples was determined using a standard diagnostic RT-PCR assay. From the total of 27 positive samples, the VP1 coding region was amplified and sequenced. Each of these sequences showed >99% identity to each other, and just five distinct sequences were...

Analysis of a new strain of Euphorbia mosaic virus with distinct replication specificity unveils a lineage of begomoviruses with short Rep sequences in the DNA-B intergenic region

Directory of Open Access Journals (Sweden)

Argüello-Astorga Gerardo R

2010-10-01

Full Text Available Abstract Background Euphorbia mosaic virus (EuMV is a member of the SLCV clade, a lineage of New World begomoviruses that display distinctive features in their replication-associated protein (Rep and virion-strand replication origin. The first entirely characterized EuMV isolate is native from Yucatan Peninsula, Mexico; subsequently, EuMV was detected in weeds and pepper plants from another region of Mexico, and partial DNA-A sequences revealed significant differences in their putative replication specificity determinants with respect to EuMV-YP. This study was aimed to investigate the replication compatibility between two EuMV isolates from the same country. Results A new isolate of EuMV was obtained from pepper plants collected at Jalisco, Mexico. Full-length clones of both genomic components of EuMV-Jal were biolistically inoculated into plants of three different species, which developed symptoms indistinguishable from those induced by EuMV-YP. Pseudorecombination experiments with EuMV-Jal and EuMV-YP genomic components demonstrated that these viruses do not form infectious reassortants in Nicotiana benthamiana, presumably because of Rep-iteron incompatibility. Sequence analysis of the EuMV-Jal DNA-B intergenic region (IR led to the unexpected discovery of a 35-nt-long sequence that is identical to a segment of the rep gene in the cognate viral DNA-A. Similar short rep sequences ranging from 35- to 51-nt in length were identified in all EuMV isolates and in three distinct viruses from South America related to EuMV. These short rep sequences in the DNA-B IR are positioned downstream to a ~160-nt non-coding domain highly similar to the CP promoter of begomoviruses belonging to the SLCV clade. Conclusions EuMV strains are not compatible in replication, indicating that this begomovirus species probably is not a replicating lineage in nature. The genomic analysis of EuMV-Jal led to the discovery of a subgroup of SLCV clade viruses that contain in

Teekond Kirjasõna Vabariigis / Märt Väljataga ; intervjueerinud Marek Tamm

Index Scriptorium Estoniae

Väljataga, Märt, 1965-

2015-01-01

Märt Väljataga tõlgitud R. Rorty artiklist "Pragmatism ja filosoofia", mis ilmus Akadeemia 1992, nr. 2, lk. 281-282, 285, tema filosoofiast ja Märt Väljataga tööst ajakirja "Vikerkaar" toimetajana ning tõlkijana

Parallel motif extraction from very long sequences

KAUST Repository

Sahli, Majed; Mansour, Essam; Kalnis, Panos

2013-01-01

Motifs are frequent patterns used to identify biological functionality in genomic sequences, periodicity in time series, or user trends in web logs. In contrast to a lot of existing work that focuses on collections of many short sequences, modern

The de Bono LAMS Sequence Series: Template Designs as Knowledge-Mobilising Strategy for 21st Century Higher Education

Science.gov (United States)

Dobozy, Eva

2012-01-01

In this paper, the five interlocking de Bono LAMS sequences are introduced as a new form of generic template designs. This transdisciplinary knowledge-mobilising strategy is based on Edward de Bono's attention-directing ideas and thinking skills, commonly known as the CoRT tools. The development of the de Bono LAMS sequence series is an important…

Factors influencing the use of RT in NSW: a qualitative study exploring consumer and health professional practices

International Nuclear Information System (INIS)

Sundaresan, Puma; Milross, Christopher G.; Stockler, Martin R.; Smith, Andrea; Evans, Alison; King, Madeleine T.

2014-01-01

Radiotherapy (RT) is an essential and cost-effective cancer treatment. It is underutilised in Australia. Bridging the gap between actual and optimal RT utilisation requires not only provision of adequate RT infrastructure but also an understanding of the factors that influence the extent to which this opportunity for RT is utilised. This study explored factors perceived to affect RT-related decision making by consumers and health professionals (HPs). Six semi-structured focus groups (FGs) and 13 interviews were conducted at three geographical locations in NSW, Australia (n=26 consumers and 30 HPs). Audio recordings of FGs and interviews were transcribed verbatim and analysed thematically. An exhaustive list of issues perceived to affect consumer and HP RT decisions was identified. There were common themes across participant groups and locations. Perceptions of RT and its benefits, as well as accurate communication of the expected benefits and risks of RT, were highlighted as important to decision making. Perceived factors relating to 'inconvenience' of RT were multifaceted and included travel, relocation, accommodation, time away from work and financial challenges. Perceived potential barriers to RT referral included knowledge of RT and RT services, availability of a local or visiting RT service, referrer bias, and the low profile of RT. Important drivers during RT decisions appear to include the perceived benefit, risks and inconvenience of RT. Underutilisation of RT may also result from multiple barriers at the referrer level. Further research into whether these factors influence actual RT decisions is needed.

Temperature affects thrombolytic efficacy using rt-PA and eptifibatide, an in vitro study.

Science.gov (United States)

Meunier, Jason M; Chang, Wan-Tsu W; Bluett, Brent; Wenker, Evan; Lindsell, Christopher J; Shaw, George J

2012-09-01

The potential for hypothermia as a neuroprotectant during stroke has led to its increase in clinical use. At the same time, combination pharmaceutical therapies for ischemic stroke using recombinant tissue plasminogen activator (rt-PA), and GP IIb-IIIa inhibitors, such as Eptifibatide (Epf ), are under study. However, there is little data on how the reactions triggered by these agents are impacted by temperature. Here, clot lysis during exposure to the combination of rt-PA and Epf is measured in an in vitro human clot model at hypothermic temperatures. The hypothesis is that lytic efficacy of rt-PA and Epf decreases with decreasing temperature. Whole blood clots from 31 volunteers were exposed to rt-PA (0.5 μg/mL) and Epf (0.63 μg/mL) in human fresh-frozen plasma (rt-PA+Epf ), rt-PA alone in plasma (rt-PA Alone), or to plasma alone (Control), at temperatures from 30°C to 37°C, for 30 minutes. Clot lysis was measured using a microscopic imaging technique; the mean fractional clot loss (FCL) at 30 minutes was used to determine lytic efficacy. Temperature had a significant impact on FCL in clots exposed to rt-PA+Epf, with the FCL being lower at 30°C to 36°C than at 37°C. The FCL remained significantly higher for rt-PA+Epf–treated clots than Controls regardless of temperature, with the exception of measurements made at 30°C when no significant differences in the FCL were observed between groups. The use of hypothermia as a neuroprotectant may negatively impact the therapeutic benefit of thrombolytic agents.

Development and implementation of the quality control panel of RT-PCR and real-time RT-PCR for avian influenza A (H5N1 surveillance network in mainland China

Directory of Open Access Journals (Sweden)

Wang Wei

2011-03-01

Full Text Available Abstract Background Reverse transcription PCR (RT-PCR and real time RT-PCR (rRT-PCR have been indispensable methods for influenza surveillance, especially for determination of avian influenza. The movement of testing beyond reference lab introduced the need of quality control, including the implementation of an evaluation system for validating personal training and sample proficiency testing. Methods We developed a panel with lysates of seasonal influenza virus (H1N1, H3N2 and B, serials of diluted H5N1 virus lysates, and in-vitro transcribed H5 hemaglutinin (HA and an artificial gene RNAs for RT-PCR and rRT-PCR quality control assessment. The validations of stability and reproducibility were performed on the panel. Additionally, the panel was implemented to assess the detection capability of Chinese human avian influenza networks. Results The panel has relatively high stability and good reproducibility demonstrated by kappa's tests. In the implementation of panel on Chinese human avian influenza networks, the results suggested that there were a relatively low number of discrepancies for both concise and reproducibility in Chinese avian influenza virus net works. Conclusions A quality control panel of RT-PCR and real-time RT-PCR for avian influenza A (H5N1 surveillance network was developed. An availably statistical data, which are used to assess the detection capability of networks on avian influenza virus (H5N1, can be obtained relatively easily through implementation of the panel on networks.

GapMis: a tool for pairwise sequence alignment with a single gap.

Science.gov (United States)

Flouri, Tomás; Frousios, Kimon; Iliopoulos, Costas S; Park, Kunsoo; Pissis, Solon P; Tischler, German

2013-08-01

Pairwise sequence alignment has received a new motivation due to the advent of recent patents in next-generation sequencing technologies, particularly so for the application of re-sequencing---the assembly of a genome directed by a reference sequence. After the fast alignment between a factor of the reference sequence and a high-quality fragment of a short read by a short-read alignment programme, an important problem is to find the alignment between a relatively short succeeding factor of the reference sequence and the remaining low-quality part of the read allowing a number of mismatches and the insertion of a single gap in the alignment. We present GapMis, a tool for pairwise sequence alignment with a single gap. It is based on a simple algorithm, which computes a different version of the traditional dynamic programming matrix. The presented experimental results demonstrate that GapMis is more suitable and efficient than most popular tools for this task.

Short rest interval lengths between sets optimally enhance body composition and performance with 8 weeks of strength resistance training in older men.

Science.gov (United States)

Villanueva, Matthew G; Lane, Christianne Joy; Schroeder, E Todd

2015-02-01

To determine if 8 weeks of periodized strength resistance training (RT) utilizing relatively short rest interval lengths (RI) in between sets (SS) would induce greater improvements in body composition and muscular performance, compared to the same RT program utilizing extended RI (SL). 22 male volunteers (SS: n = 11, 65.6 ± 3.4 years; SL: n = 11, 70.3 ± 4.9 years) were assigned to one of two strength RT groups, following 4 weeks of periodized hypertrophic RT (PHRT): strength RT with 60-s RI (SS) or strength RT with 4-min RI (SL). Prior to randomization, all 22 study participants trained 3 days/week, for 4 weeks, targeting hypertrophy; from week 4 to week 12, SS and SL followed the same periodized strength RT program for 8 weeks, with RI the only difference in their RT prescription. Following PHRT, all study participants experienced increases in lean body mass (LBM) (p body strength (p body fat (p high-intensity strength RT with shortened RI induces significantly greater enhancements in body composition, muscular performance, and functional performance, compared to the same RT prescription with extended RI, in older men. Applied professionals may optimize certain RT-induced adaptations, by incorporating shortened RI.

Phylogeny and resistance profiles of HIV-1 POL sequences from rectal biopsies and blood

DEFF Research Database (Denmark)

Katzenstein, Terese Lea; Petersen, A B; Storgaard, M

2010-01-01

The phylogeny and resistance profiles of human immunodeficiency virus type 1 (HIV-1) protease (PR) and reverse transcriptase (RT) sequences were compared among six patients with HIV-1 who had received numerous treatments. RNA and DNA fractions were obtained from concurrent blood and rectal biopsy...

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

International Nuclear Information System (INIS)

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

2005-01-01

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

Development of a novel real-time RT-PCR assay to detect Seneca Valley virus-1 associated with emerging cases of vesicular disease in pigs.

Science.gov (United States)

Fowler, Veronica L; Ransburgh, Russell H; Poulsen, Elizabeth G; Wadsworth, Jemma; King, Donald P; Mioulet, Valerie; Knowles, Nick J; Williamson, Susanna; Liu, Xuming; Anderson, Gary A; Fang, Ying; Bai, Jianfa

2017-01-01

Seneca Valley virus 1 (SVV-1) can cause vesicular disease that is clinically indistinguishable from foot-and-mouth disease, vesicular stomatitis and swine vesicular disease. SVV-1-associated disease has been identified in pigs in several countries, namely USA, Canada, Brazil and China. Diagnostic tests are required to reliably detect this emerging virus, and this report describes the development and evaluation of a novel real-time (r) reverse-transcription (RT) PCR assay (rRT-PCR), targeting the viral polymerase gene (3D) of SVV-1. This new assay detected all historical and contemporary SVV-1 isolates examined (n=8), while no cross-reactivity was observed with nucleic acid templates prepared from other vesicular disease viruses or common swine pathogens. The analytical sensitivity of the rRT-PCR was 0.79 TCID 50 /ml and the limit of detection was equivalent using two different rRT-PCR master-mixes. The performance of the test was further evaluated using pig nasal (n=25) and rectal swab samples (n=25), where concordant results compared to virus sequencing were generated for 43/50 samples. The availability of this assay, will enable laboratories to rapidly detect SVV-1 in cases of vesicular disease in pigs, negated for notifiable diseases, and could enable existing knowledge gaps to be investigated surrounding the natural epidemiology of SVV-1. Copyright © 2016 Elsevier B.V. All rights reserved.

Yeast genome sequencing:

DEFF Research Database (Denmark)

Piskur, Jure; Langkjær, Rikke Breinhold

2004-01-01

For decades, unicellular yeasts have been general models to help understand the eukaryotic cell and also our own biology. Recently, over a dozen yeast genomes have been sequenced, providing the basis to resolve several complex biological questions. Analysis of the novel sequence data has shown...... of closely related species helps in gene annotation and to answer how many genes there really are within the genomes. Analysis of non-coding regions among closely related species has provided an example of how to determine novel gene regulatory sequences, which were previously difficult to analyse because...... they are short and degenerate and occupy different positions. Comparative genomics helps to understand the origin of yeasts and points out crucial molecular events in yeast evolutionary history, such as whole-genome duplication and horizontal gene transfer(s). In addition, the accumulating sequence data provide...

A Pan-Lyssavirus Taqman Real-Time RT-PCR Assay for the Detection of Highly Variable Rabies virus and Other Lyssaviruses.

Science.gov (United States)

Wadhwa, Ashutosh; Wilkins, Kimberly; Gao, Jinxin; Condori Condori, Rene Edgar; Gigante, Crystal M; Zhao, Hui; Ma, Xiaoyue; Ellison, James A; Greenberg, Lauren; Velasco-Villa, Andres; Orciari, Lillian; Li, Yu

2017-01-01

Rabies, resulting from infection by Rabies virus (RABV) and related lyssaviruses, is one of the most deadly zoonotic diseases and is responsible for up to 70,000 estimated human deaths worldwide each year. Rapid and accurate laboratory diagnosis of rabies is essential for timely administration of post-exposure prophylaxis in humans and control of the disease in animals. Currently, only the direct fluorescent antibody (DFA) test is recommended for routine rabies diagnosis. Reverse-transcription polymerase chain reaction (RT-PCR) based diagnostic methods have been widely adapted for the diagnosis of other viral pathogens, but there is currently no widely accepted rapid real-time RT-PCR assay for the detection of all lyssaviruses. In this study, we demonstrate the validation of a newly developed multiplex real-time RT-PCR assay named LN34, which uses a combination of degenerate primers and probes along with probe modifications to achieve superior coverage of the Lyssavirus genus while maintaining sensitivity and specificity. The primers and probes of the LN34 assay target the highly conserved non-coding leader region and part of the nucleoprotein (N) coding sequence of the Lyssavirus genome to maintain assay robustness. The probes were further modified by locked nucleotides to increase their melting temperature to meet the requirements for an optimal real-time RT-PCR assay. The LN34 assay was able to detect all RABV variants and other lyssaviruses in a validation panel that included representative RABV isolates from most regions of the world as well as representatives of 13 additional Lyssavirus species. The LN34 assay was successfully used for both ante-mortem and post-mortem diagnosis of over 200 clinical samples as well as field derived surveillance samples. This assay represents a major improvement over previously published rabies specific RT-PCR and real-time RT-PCR assays because of its ability to universally detect RABV and other lyssaviruses, its high

A Pan-Lyssavirus Taqman Real-Time RT-PCR Assay for the Detection of Highly Variable Rabies virus and Other Lyssaviruses.

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Ashutosh Wadhwa

2017-01-01

Full Text Available Rabies, resulting from infection by Rabies virus (RABV and related lyssaviruses, is one of the most deadly zoonotic diseases and is responsible for up to 70,000 estimated human deaths worldwide each year. Rapid and accurate laboratory diagnosis of rabies is essential for timely administration of post-exposure prophylaxis in humans and control of the disease in animals. Currently, only the direct fluorescent antibody (DFA test is recommended for routine rabies diagnosis. Reverse-transcription polymerase chain reaction (RT-PCR based diagnostic methods have been widely adapted for the diagnosis of other viral pathogens, but there is currently no widely accepted rapid real-time RT-PCR assay for the detection of all lyssaviruses. In this study, we demonstrate the validation of a newly developed multiplex real-time RT-PCR assay named LN34, which uses a combination of degenerate primers and probes along with probe modifications to achieve superior coverage of the Lyssavirus genus while maintaining sensitivity and specificity. The primers and probes of the LN34 assay target the highly conserved non-coding leader region and part of the nucleoprotein (N coding sequence of the Lyssavirus genome to maintain assay robustness. The probes were further modified by locked nucleotides to increase their melting temperature to meet the requirements for an optimal real-time RT-PCR assay. The LN34 assay was able to detect all RABV variants and other lyssaviruses in a validation panel that included representative RABV isolates from most regions of the world as well as representatives of 13 additional Lyssavirus species. The LN34 assay was successfully used for both ante-mortem and post-mortem diagnosis of over 200 clinical samples as well as field derived surveillance samples. This assay represents a major improvement over previously published rabies specific RT-PCR and real-time RT-PCR assays because of its ability to universally detect RABV and other lyssaviruses

Evaluation of full S1 gene sequencing of classical and variant infectious bronchitis viruses extracted from allantoic fluid and FTA cards.

Science.gov (United States)

Manswr, Basim; Ball, Christopher; Forrester, Anne; Chantrey, Julian; Ganapathy, Kannan

2018-05-01

Sequence variability in the S1 gene determines the genotype of infectious bronchitis virus (IBV) strains. A single RT-PCR assay was developed to amplify and sequence the full S1 gene for six classical and variant IBVs (M41, D274, 793B, IS/885/00, IS/1494/06 and Q1) enriched in allantoic fluid (AF) or the same AF but inoculated onto Flinders Technology Association (FTA) cards. Representative strains from each genotype were grown in SPF eggs and RNA was extracted from AF. Full S1 gene amplification was achieved using primer A and primer 22.51. Products were sequenced using primer A, 1050+, 1380+ and SX3+ to obtain short sequences covering the full gene. Following serial dilutions of AF, detection limits of the partial assay were higher than those of the full S1 gene. Partial S1 sequences exhibited higher than average nucleotide similarity percentages (79%; 352bp) compared to full S1 sequences (77%; 1,756bp), suggesting that full S1 analysis allows greater strain differentiation. For IBV detection from AF inoculated FTA cards, four serotypes were incubated for up to 21 days at three temperatures; 4 o C, 24 o C and 40 o C. RNA was extracted and tested with partial and full S1 protocols. Through partial sequencing, all IBVs were successfully detected at all sampling points and storage temperatures. In contrast, using full S1 sequencing was not possible to amplify the gene beyond 14 days or when stored at 40°C. Data presented shows that for full S1 sequencing, a substantial amount of RNA is needed. Field samples collected onto FTA cards are unlikely to yield such quantity or quality.

Extension of radiotherapy (involved field vs subtotal nodal) after short-term chemotherapy in early-stage Hodgkin's disease (ESHD)

International Nuclear Information System (INIS)

Zanini, M.; Bonfante, V.; Soncini, F.; Di Russo, A.; Santoro, A.; Viviani, S.; Devizzi, L.; Gasparini, M.; Tesoro, Tess J.D.; Valagussa, P.; Bonadonna, G.

1995-01-01

Aims of this study were to avoid staging laparotomy, to improve the results of radiotherapy (RT) alone by a combined modality approach, to evaluate the optimal extent of RT after short-term chemotherapy (CT) and to reduce the long-term toxicity in patients cured for ESHD. From (2(90)) to (1(95)), 110 consecutive patients with clinical stage I (bulky and/or B), IIA, IIA bulky and IIEA entered a randomized trial comparing subtotal nodal irradiation (STNI) vs involved field radiotherapy (IFRT), both after short-term chemotherapy (ABVD for 4 cycles). The doses of RT were 30 Gy to uninvolved and 36 Gy to involved sites, respectively. With a median follow-up of 38 mos., the main results in 96 evaluable patients (stage I: 13 pts., stage II: 83 pts., median age 28 yrs., range 17-64) are as reported below. Most patients achieved complete remission (CR) after CT (92%), while 8 partial responders attained CR with RT (IFRT 5 pts., STNI 3 pts.). Four stage II patients relapsed (IFRT 2, pts., STNI 2 pts.), mainly in extranodal or not irradiated sites. Up-to-date the most remarkable toxicities include: acute myocardial infarction (1%), aspecific EKG changes (5%), reductions in pulmonary function tests (most mild and/or transient) in 45%, subclinical or clinical hypothyroidism 26% (only in pts. irradiated to the neck), 3 cases of azoospermia in 23 evaluable patients (13%) and transient amenorreha in(1(44)) patients. No second malignancies have been so far observed. The results of this study seem to indicate that 4 courses of ABVD plus RT are an effective treatment in clinical ESHD, with moderate long-term toxicities. In almost 10% of patients RT changed partial into complete remission. Should these results be confirmed on a longer follow-up, IFRT will definitely replace STNI in a combined modality treatment

[Enterovirus sequencing as a new approach to the laboratory diagnosis for clinical and epidemiological purposes].

Science.gov (United States)

Rainetová, P; Jiřincová, H; Musílek, M; Nováková, L; Vodičková, I; Štruncová, V; Švecová, M; Pazdiora, P; Piskunová, N; Trubač, P; Zajíc, T; Havlíčková, M

2015-06-01

Introducing enterovirus sequencing as an advanced approach to classify the viruses isolated according to the novel nomenclature and to characterize isolates in detail. Seventy-five specimens collected from 64 patients in two hospitals, Liberec Regional Hospital, and Plzeň University Hospital, were analyzed. The study patients' age ranged from four to 54 years, with a median of 15 years in males and 16 years in females. In most patients, the reasons for admission were intense headache, fever, vomiting, tiredness, meningeal symptoms, intestinal symptoms (in two patients), and skin symptoms (in one patient). The specimens collected were rectal and throat swabs, cerebrospinal fluid (CSF) and stool specimens. Molecular detection and typing were performed using the RT-PCR method. A segment of the 5´non-coding RNA was selected for typing. Specimens were amplified using single-step PCR with external primers and with the same primers extended to include M13 sequences (Generi-Biotech). The LASERGENE software (DIASTAR) was used in sequence editing, alignment, and quality check. The sequences obtained were checked against the central GenBank sequence database using the BLAST algorithm. The identification of the study isolates resulted in 61 ECHO viruses 30, three coxsackie viruses B1, one coxsackie virus B3, one coxsackie virus A9, one enterovirus 86, one enterovirus 71, Two ECHO viruses 13/coxsackie virus B5, one ECHO virus 7/30/coxsackie virus B4, one coxsackie virus B4/enterovirus B, one enterovirus 87/ECHO virus 30/enterovirus B, and one ECHO virus 3. All viruses isolated, except enterovirus 71 classified into group A, were of group B. The enteroviruses were identified unambigously, although the sequencing only targeted a short, conserved segment that showed considerable variability. The sequencing was an effective alternative to enterovirus identification by the neutralisation test and allowed for detailed characterization of the isolates. The predominance of ECHO 30 as

Association of sequences in the coat protein/readthrough domain of potato mop-top virus with transmission by Spongospora subterranea.

Science.gov (United States)

Reavy, B; Arif, M; Cowan, G H; Torrance, L

1998-10-01

A monofungal culture of Spongospora subterranea was unable to acquire and transmit the T isolate of potato mop-top pomovirus (PMTV-T), which has been maintained by manual transmission in the laboratory for 30 years. A recently obtained field isolate (PMTV-S) was efficiently acquired and transmitted by the same fungus culture. Sequence analysis of the readthrough (RT) protein-coding region of PMTV-S showed the presence of an additional 543 nt in the 3' half of the coding region relative to that of PMTV-T. These additional nucleotides preserved the reading frame of the RT protein and inserted 181 amino acids into the RT protein. This was confirmed by a comparison by immunoblotting of the sizes of the RT protein of PMTV-T and other recent isolates of PMTV.

Development of tailored real-time RT-PCR assays for the detection and differentiation of serotype O, A and Asia-1 foot-and-mouth disease virus lineages circulating in the Middle East

DEFF Research Database (Denmark)

Reid, Scott M.; Mioulet, Valerie; Knowles, Nick J.

2014-01-01

transcription polymerase chain reaction (rRT-PCR) assays using primer/probe sets designed from the VP1 coding region of the virus genomes were developed for the specific detection of serotype O, A and Asia-1 FMD viruses (FMDVs) circulating in the Middle East. These assays were evaluated using representative...... by the generic rRT-PCR diagnostic assays but negative by virus isolation and antigen-detection ELISA which would otherwise have to be serotyped by nucleotide sequencing. A similar approach could be used to develop serotyping assays for FMDV strains circulating in other regions of the world....

Gene discovery and transcript analyses in the corn smut pathogen Ustilago maydis: expressed sequence tag and genome sequence comparison

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Saville Barry J

2007-09-01

Full Text Available Abstract Background Ustilago maydis is the basidiomycete fungus responsible for common smut of corn and is a model organism for the study of fungal phytopathogenesis. To aid in the annotation of the genome sequence of this organism, several expressed sequence tag (EST libraries were generated from a variety of U. maydis cell types. In addition to utility in the context of gene identification and structure annotation, the ESTs were analyzed to identify differentially abundant transcripts and to detect evidence of alternative splicing and anti-sense transcription. Results Four cDNA libraries were constructed using RNA isolated from U. maydis diploid teliospores (U. maydis strains 518 × 521 and haploid cells of strain 521 grown under nutrient rich, carbon starved, and nitrogen starved conditions. Using the genome sequence as a scaffold, the 15,901 ESTs were assembled into 6,101 contiguous expressed sequences (contigs; among these, 5,482 corresponded to predicted genes in the MUMDB (MIPS Ustilago maydis database, while 619 aligned to regions of the genome not yet designated as genes in MUMDB. A comparison of EST abundance identified numerous genes that may be regulated in a cell type or starvation-specific manner. The transcriptional response to nitrogen starvation was assessed using RT-qPCR. The results of this suggest that there may be cross-talk between the nitrogen and carbon signalling pathways in U. maydis. Bioinformatic analysis identified numerous examples of alternative splicing and anti-sense transcription. While intron retention was the predominant form of alternative splicing in U. maydis, other varieties were also evident (e.g. exon skipping. Selected instances of both alternative splicing and anti-sense transcription were independently confirmed using RT-PCR. Conclusion Through this work: 1 substantial sequence information has been provided for U. maydis genome annotation; 2 new genes were identified through the discovery of 619

Sequence-specific inhibition of microRNA-130a gene by CRISPR/Cas9 system in breast cancer cell line

Science.gov (United States)

Ainina Abdollah, Nur; Das Kumitaa, Theva; Yusof Narazah, Mohd; Razak, Siti Razila Abdul

2017-05-01

MicroRNAs (miRNAs) are short stranded noncoding RNA that play important roles in apoptosis, cell survival, development and cell proliferation. However, gene expression control via small regulatory RNA, particularly miRNA in breast cancer is still less explored. Therefore, this project aims to develop an approach to target microRNA-130a using the Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR)/Cas9 system in MCF7, breast cancer cell line. The 20 bp sequences target at stem loop, 3ʹ and 5ʹ end of miR130a were cloned into pSpCas9(BB)-2A-GFP (PX458) plasmid, and the positive clones were confirmed by sequencing. A total of 5 μg of PX458-miR130a was transfected to MCF7 using Lipofectamine® 3000 according to manufacturer’s protocol. The transfected cells were maintained in the incubator at 37 °C under humidified 5% CO2. After 48 hours, cells were harvested and total RNA was extracted using miRNeasy Mini Kit (Qiagen). cDNAs were synthesised specific to miR-130a using TaqMan MicroRNA Reverse Transcription Kit (Applied Biosystems). Then, qRT-PCR was carried out using TaqMan Universal Master Mix (Applied Biosystems) to quantify the knockdown level of mature miRNAs in the cells. Result showed that miR-130a-5p was significantly downregulated in MCF7 cell line. However, no significant changes were observed for sequences targeting miR-130a-3p and stem loop. Thus, this study showed that the expression of miR-130a-5p was successfully down-regulated using CRISPR silencing system. This technique may be useful to manipulate the level of miRNA in various cell types to answer clinical questions at the molecular level.

System of programming units for the K556RT4 and K556RT5 fixed programmed memory devices

International Nuclear Information System (INIS)

Bobkov, S.G.; Ermolin, Yu.V.; Kantserov, V.A.; Strigin, V.B.

1983-01-01

The programming system of constant programmable memory devices K556RT4 and K556RT5 that consist of two units (a programming device and an electrothermotraining unit) is described. The modules are made in the KAMAK standard. The programming device takes up 2 normal places, while the electrothermotraining block takes up 1 place. As information recording is done using a computer the time for programming is reduced and the possibility of errors is limited as compared with the manual method. The computer introduces the whole word to be recorded, not the separate parts, in the programming device. The transition to a new digit of a given word in the programming device is done automatically. This reduces the expense of computer time and accelerates the programming of microdiagrams

Rapid detection of bovine coronavirus by a semi-nested RT-PCR Detecção rápida do Coronavírus Bovino (BCoV por meio de uma semi-nested RT-PCR

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Karen M. Asano

2009-11-01

Full Text Available Bovine coronavirus (BCoV is a member of the group 2 of the Coronavirus (Nidovirales: Coronaviridae and the causative agent of enteritis in both calves and adult bovine, as well as respiratory disease in calves. The present study aimed to develop a semi-nested RT-PCR for the detection of BCoV based on representative up-to-date sequences of the nucleocapsid gene, a conserved region of coronavirus genome. Three primers were designed, the first round with a 463bp and the second (semi-nested with a 306bp predicted fragment. The analytical sensitivity was determined by 10-fold serial dilutions of the BCoV Kakegawa strain (HA titre: 256 in DEPC treated ultra-pure water, in fetal bovine serum (FBS and in a BCoV-free fecal suspension, when positive results were found up to the 10-2, 10-3 and 10-7 dilutions, respectively, which suggests that the total amount of RNA in the sample influence the precipitation of pellets by the method of extraction used. When fecal samples was used, a large quantity of total RNA serves as carrier of BCoV RNA, demonstrating a high analytical sensitivity and lack of possible substances inhibiting the PCR. The final semi-nested RT-PCR protocol was applied to 25 fecal samples from adult cows, previously tested by a nested RT-PCR RdRp used as a reference test, resulting in 20 and 17 positives for the first and second tests, respectively, and a substantial agreement was found by kappa statistics (0.694. The high sensitivity and specificity of the new proposed method and the fact that primers were designed based on current BCoV sequences give basis to a more accurate diagnosis of BCoV-caused diseases, as well as to further insights on protocols for the detection of other Coronavirus representatives of both Animal and Public Health importance.O Coronavírus bovino (BCoV pertence ao grupo 2 do gênero Coronavirus (Nidovirales: Coronaviridae e é agente causador de enterites tanto em bezerros como em bovinos adultos, bem como de doen

[Disappearance of residual disease confirmed by RT-PCR following induction chemotherapy in two hypoplastic leukemia patients with t(8;21)].

Science.gov (United States)

Sawada, M; Tsurumi, H; Yamada, T; Hara, T; Oyama, M; Moriwaki, H

1999-04-01

Reverse transcriptase-polymerase chain reaction (RT-PCR) methods often detect the AML1/MTG8 fusion transcript even in acute myelogenous leukemia (AML) patients with t(8;21) who have been in long-term remission. We encountered 2 hypoplastic leukemia patients with t(8;21) who achieved cytogenetic remission with short-term conventional chemotherapy. Patient 1 was a 42-year-old woman. Chromosomal analysis detected t(8;21) (q22;q22) and PCR analysis (35 cycles PCR amplification; detection limit 1 x 10(-5) cells) detected the AML1/MTG8 fusion transcript. Complete remission was obtained with 1 course of chemotherapy consisting of low-dose cytarabine (20 mg x 14 days) and etoposide (50 mg x 14 days). After 2 courses of consolidation chemotherapy consisting of conventional-dose cytarabine and mitoxantrone, the RT-PCR findings were negative for the AML1/MTG8 fusion transcript. Patient 2 was a 67-year-old man. Cytogenetic analysis detected t(8;21) (q22;q22), and was positive for the AML1/MTG8 fusion transcript. After 2 courses of induction chemotherapy comprising low-dose cytarabine (20 mg x 14 days) and etoposide (50 mg x 14 days), and 3 courses of conventional consolidation chemotherapy, RT-PCR analysis confirmed the disappearance of the AML1/MTG8 fusion transcript.

SU-F-J-174: A Series of Computational Human Phantoms in DICOM-RT Format for Normal Tissue Dose Reconstruction in Epidemiological Studies

International Nuclear Information System (INIS)

Pyakuryal, A; Moroz, B; Lee, C; Pelletier, C; Jung, J; Lee, C

2016-01-01

Purpose: Epidemiological studies of second cancer risk in radiotherapy patients often require individualized dose estimates of normal tissues. Prior to 3D conformal radiation therapy planning, patient anatomy information was mostly limited to 2D radiological images or not even available. Generic patient CT images are often used in commercial radiotherapy treatment planning system (TPS) to reconstruct normal tissue doses. The objective of the current work was to develop a series of reference size computational human phantoms in DICOM-RT format for direct use in dose reconstruction in TPS. Methods: Contours of 93 organs and tissues were extracted from a series of pediatric and adult hybrid computational human phantoms (newborn, 1-, 5-, 10-, 15-year-old, and adult males and females) using Rhinoceros software. A MATLAB script was created to convert the contours into the DICOM-RT structure format. The simulated CT images with the resolution of 1×1×3 mm3 were also generated from the binary phantom format and coupled with the DICOM-structure files. Accurate volumes of the organs were drawn in the format using precise delineation of the contours in converted format. Due to complex geometry of organs, higher resolution (1×1×1 mm3) was found to be more efficient in the conversion of newborn and 1-year-old phantoms. Results: Contour sets were efficiently converted into DICOM-RT structures in relatively short time (about 30 minutes for each phantom). A good agreement was observed in the volumes between the original phantoms and the converted contours for large organs (NRMSD<1.0%) and small organs (NRMSD<7.7%). Conclusion: A comprehensive series of computational human phantoms in DICOM-RT format was created to support epidemiological studies of second cancer risks in radiotherapy patients. We confirmed the DICOM-RT phantoms were successfully imported into the TPS programs of major vendors.

SU-F-J-174: A Series of Computational Human Phantoms in DICOM-RT Format for Normal Tissue Dose Reconstruction in Epidemiological Studies

Energy Technology Data Exchange (ETDEWEB)

Pyakuryal, A; Moroz, B [National Cancer Institute, National Institutes of Health, Rockville, MD (United States); Lee, C [University of Michigan, Ann Arbor, MI (United States); Pelletier, C; Jung, J [East Carolina University Greenville, NC (United States); Lee, C [National Cancer Institute, Rockville, MD (United States)

2016-06-15

Purpose: Epidemiological studies of second cancer risk in radiotherapy patients often require individualized dose estimates of normal tissues. Prior to 3D conformal radiation therapy planning, patient anatomy information was mostly limited to 2D radiological images or not even available. Generic patient CT images are often used in commercial radiotherapy treatment planning system (TPS) to reconstruct normal tissue doses. The objective of the current work was to develop a series of reference size computational human phantoms in DICOM-RT format for direct use in dose reconstruction in TPS. Methods: Contours of 93 organs and tissues were extracted from a series of pediatric and adult hybrid computational human phantoms (newborn, 1-, 5-, 10-, 15-year-old, and adult males and females) using Rhinoceros software. A MATLAB script was created to convert the contours into the DICOM-RT structure format. The simulated CT images with the resolution of 1×1×3 mm3 were also generated from the binary phantom format and coupled with the DICOM-structure files. Accurate volumes of the organs were drawn in the format using precise delineation of the contours in converted format. Due to complex geometry of organs, higher resolution (1×1×1 mm3) was found to be more efficient in the conversion of newborn and 1-year-old phantoms. Results: Contour sets were efficiently converted into DICOM-RT structures in relatively short time (about 30 minutes for each phantom). A good agreement was observed in the volumes between the original phantoms and the converted contours for large organs (NRMSD<1.0%) and small organs (NRMSD<7.7%). Conclusion: A comprehensive series of computational human phantoms in DICOM-RT format was created to support epidemiological studies of second cancer risks in radiotherapy patients. We confirmed the DICOM-RT phantoms were successfully imported into the TPS programs of major vendors.

Application and evaluation of RT-PCR-ELISA for the nucleoprotein and RT-PCR for detection of low-pathogenic H5 and H7 subtypes of avian influenza virus

DEFF Research Database (Denmark)

Dybkær, Karen; Munch, Mette; Handberg, Kurt J.

2004-01-01

Three 1-tube Reverse Transcriptase Polymerase Chain Reactions (RT-PCR) directed against the genes encoding the nucleoprotein (NP) and the H5 and H7 hemagglutinin (HA) gene, respectively, were used for detection of avian influenza virus (AIV) in various specimens. A total of 1,040 samples...... originating from chickens experimentally infected with 2 different low pathogenic avian influenza viruses, from domestic ducks and from wild aquatic birds were examined. The outcome of 1) the universal AIV RT-PCR including a PCR-enzyme-linked immunosorbent assay (ELISA) procedure directed against NP (NP RT...

Polyadenylated Sequencing Primers Enable Complete Readability of PCR Amplicons Analyzed by Dideoxynucleotide Sequencing

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Martin Beránek

2012-01-01

Full Text Available Dideoxynucleotide DNA sequencing is one of the principal procedures in molecular biology. Loss of an initial part of nucleotides behind the 3' end of the sequencing primer limits the readability of sequenced amplicons. We present a method which extends the readability by using sequencing primers modified by polyadenylated tails attached to their 5' ends. Performing a polymerase chain reaction, we amplified eight amplicons of six human genes (AMELX, APOE, HFE, MBL2, SERPINA1 and TGFB1 ranging from 106 bp to 680 bp. Polyadenylation of the sequencing primers minimized the loss of bases in all amplicons. Complete sequences of shorter products (AMELX 106 bp, SERPINA1 121 bp, HFE 208 bp, APOE 244 bp, MBL2 317 bp were obtained. In addition, in the case of TGFB1 products (366 bp, 432 bp, and 680 bp, respectively, the lengths of sequencing readings were significantly longer if adenylated primers were used. Thus, single strand dideoxynucleotide sequencing with adenylated primers enables complete or near complete readability of short PCR amplicons.

Unified Deep Learning Architecture for Modeling Biology Sequence.

Science.gov (United States)

Wu, Hongjie; Cao, Chengyuan; Xia, Xiaoyan; Lu, Qiang

2017-10-09

Prediction of the spatial structure or function of biological macromolecules based on their sequence remains an important challenge in bioinformatics. When modeling biological sequences using traditional sequencing models, characteristics, such as long-range interactions between basic units, the complicated and variable output of labeled structures, and the variable length of biological sequences, usually lead to different solutions on a case-by-case basis. This study proposed the use of bidirectional recurrent neural networks based on long short-term memory or a gated recurrent unit to capture long-range interactions by designing the optional reshape operator to adapt to the diversity of the output labels and implementing a training algorithm to support the training of sequence models capable of processing variable-length sequences. Additionally, the merge and pooling operators enhanced the ability to capture short-range interactions between basic units of biological sequences. The proposed deep-learning model and its training algorithm might be capable of solving currently known biological sequence-modeling problems through the use of a unified framework. We validated our model on one of the most difficult biological sequence-modeling problems currently known, with our results indicating the ability of the model to obtain predictions of protein residue interactions that exceeded the accuracy of current popular approaches by 10% based on multiple benchmarks.

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

Directory of Open Access Journals (Sweden)

Thierry-Mieg Danielle

2009-06-01

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

Standardized comparison of the relative impacts of HIV-1 reverse transcriptase (RT) mutations on nucleoside RT inhibitor susceptibility.

Science.gov (United States)

Melikian, George L; Rhee, Soo-Yon; Taylor, Jonathan; Fessel, W Jeffrey; Kaufman, David; Towner, William; Troia-Cancio, Paolo V; Zolopa, Andrew; Robbins, Gregory K; Kagan, Ron; Israelski, Dennis; Shafer, Robert W

2012-05-01

Determining the phenotypic impacts of reverse transcriptase (RT) mutations on individual nucleoside RT inhibitors (NRTIs) has remained a statistical challenge because clinical NRTI-resistant HIV-1 isolates usually contain multiple mutations, often in complex patterns, complicating the task of determining the relative contribution of each mutation to HIV drug resistance. Furthermore, the NRTIs have highly variable dynamic susceptibility ranges, making it difficult to determine the relative effect of an RT mutation on susceptibility to different NRTIs. In this study, we analyzed 1,273 genotyped HIV-1 isolates for which phenotypic results were obtained using the PhenoSense assay (Monogram, South San Francisco, CA). We used a parsimonious feature selection algorithm, LASSO, to assess the possible contributions of 177 mutations that occurred in 10 or more isolates in our data set. We then used least-squares regression to quantify the impact of each LASSO-selected mutation on each NRTI. Our study provides a comprehensive view of the most common NRTI resistance mutations. Because our results were standardized, the study provides the first analysis that quantifies the relative phenotypic effects of NRTI resistance mutations on each of the NRTIs. In addition, the study contains new findings on the relative impacts of thymidine analog mutations (TAMs) on susceptibility to abacavir and tenofovir; the impacts of several known but incompletely characterized mutations, including E40F, V75T, Y115F, and K219R; and a tentative role in reduced NRTI susceptibility for K64H, a novel NRTI resistance mutation.

Kirjanduskriitikute suvemängud / Kärt Hellerma

Index Scriptorium Estoniae

Hellerma, Kärt, 1956-

2007-01-01

Arvustus: Lodge, David. Ühest kohast teise / tlk. Aet Varik. Tallinn : Varrak, 2005 ; Lodge, David. Väike maailm / tlk. Kersti Unt. Tallinn : Varrak, 1996 ; Lodge, David. Väärt töö / tlk. Kersti Unt. Tallinn : Varrak, 2006

Modeling qRT-PCR dynamics with application to cancer biomarker quantification.

Science.gov (United States)

Chervoneva, Inna; Freydin, Boris; Hyslop, Terry; Waldman, Scott A

2017-01-01

Quantitative reverse transcription polymerase chain reaction (qRT-PCR) is widely used for molecular diagnostics and evaluating prognosis in cancer. The utility of mRNA expression biomarkers relies heavily on the accuracy and precision of quantification, which is still challenging for low abundance transcripts. The critical step for quantification is accurate estimation of efficiency needed for computing a relative qRT-PCR expression. We propose a new approach to estimating qRT-PCR efficiency based on modeling dynamics of polymerase chain reaction amplification. In contrast, only models for fluorescence intensity as a function of polymerase chain reaction cycle have been used so far for quantification. The dynamics of qRT-PCR efficiency is modeled using an ordinary differential equation model, and the fitted ordinary differential equation model is used to obtain effective polymerase chain reaction efficiency estimates needed for efficiency-adjusted quantification. The proposed new qRT-PCR efficiency estimates were used to quantify GUCY2C (Guanylate Cyclase 2C) mRNA expression in the blood of colorectal cancer patients. Time to recurrence and GUCY2C expression ratios were analyzed in a joint model for survival and longitudinal outcomes. The joint model with GUCY2C quantified using the proposed polymerase chain reaction efficiency estimates provided clinically meaningful results for association between time to recurrence and longitudinal trends in GUCY2C expression.

Reverse transcriptase-quantitative polymerase chain reaction (RT ...

African Journals Online (AJOL)

zino

2014-02-05

Feb 5, 2014 ... ecological studies - A review ... The objective of this review is to assess the importance of RT-qPCR in soil related ... phenol extraction step with heat inactivation of the added .... Real time polymerase chain reaction (PCR).

Evaluation of NGS and RT-PCR Methods for ALK Rearrangement in European NSCLC Patients: Results from the European Thoracic Oncology Platform Lungscape Project.

Science.gov (United States)

Letovanec, Igor; Finn, Stephen; Zygoura, Panagiota; Smyth, Paul; Soltermann, Alex; Bubendorf, Lukas; Speel, Ernst-Jan; Marchetti, Antonio; Nonaka, Daisuke; Monkhorst, Kim; Hager, Henrik; Martorell, Miguel; Sejda, Aleksandra; Cheney, Richard; Hernandez-Losa, Javier; Verbeken, Eric; Weder, Walter; Savic, Spasenija; Di Lorito, Alessia; Navarro, Atilio; Felip, Enriqueta; Warth, Arne; Baas, Paul; Meldgaard, Peter; Blackhall, Fiona; Dingemans, Anne-Marie; Dienemann, Hendrik; Dziadziuszko, Rafal; Vansteenkiste, Johan; O'Brien, Cathal; Geiger, Thomas; Sherlock, Jon; Schageman, Jeoffrey; Dafni, Urania; Kammler, Roswitha; Kerr, Keith; Thunnissen, Erik; Stahel, Rolf; Peters, Solange

2018-03-01

The reported prevalence of ALK receptor tyrosine kinase gene (ALK) rearrangement in NSCLC ranges from 2% to 7%. The primary standard diagnostic method is fluorescence in situ hybridization (FISH). Recently, immunohistochemistry (IHC) has also proved to be a reproducible and sensitive technique. Reverse-transcriptase polymerase chain reaction (RT-PCR) has also been advocated, and most recently, the advent of targeted next-generation sequencing (NGS) for ALK and other fusions has become possible. This study compares anaplastic lymphoma kinase (ALK) evaluation with all four techniques in resected NSCLC from the large European Thoracic Oncology Platform Lungscape cohort. A total of 96 cases from the European Thoracic Oncology Platform Lungscape iBiobank, with any ALK immunoreactivity were examined by FISH, central RT-PCR, and NGS. An H-score higher than 120 defines IHC positivity. RNA was extracted from the same formalin-fixed, paraffin-embedded tissues. For RT-PCR, primers covered the most frequent ALK translocations. For NGS, the Oncomine Solid Tumour Fusion Transcript Kit (Thermo Fisher Scientific, Waltham, MA) was used. The concordance was assessed using the Cohen κ coefficient (two-sided α ≤ 5%). NGS provided results for 77 of the 95 cases tested (81.1%), whereas RT-PCR provided results for 77 of 96 (80.2%). Concordance occurred in 55 cases of the 60 cases tested with all four methods (43 ALK negative and 12 ALK positive). Using ALK copositivity for IHC and FISH as the criterion standard, we derived a sensitivity for RT-PCR/NGS of 70.0%/85.0%, with a specificity of 87.1%/79.0%. When either RT-PCR or NGS was combined with IHC, the sensitivity remained the same, whereas the specificity increased to 88.7% and 83.9% respectively. NGS evaluation with the Oncomine Solid Tumour Fusion transcript kit and RT-PCR proved to have high sensitivity and specificity, advocating their use in routine practice. For maximal sensitivity and specificity, ALK status should be

Phylogenetic analysis of HIV-1 reverse transcriptase sequences from 382 patients recruited in JJ Hospital of Mumbai, India, between 2002 and 2008.

Science.gov (United States)

Deshpande, Alaka; Jauvin, Valerie; Pinson, Patricia; Jeannot, Anne Cecile; Fleury, Herve J

2009-06-01

Analysis of reverse transcriptase (RT) sequences of 382 HIV-1 isolates from untreated and treated patients recruited in JJ Hospital (Mumbai, India) between 2002 and 2008 shows that subtype C is largely predominant (98%) and that non-C sequences cluster with A1, B, CRF01_AE, and CRF06_cpx.

Pärt, Arvo. Litany; Psalom; Trisagion / Andreas Obst

Index Scriptorium Estoniae

Obst, Andreas

1996-01-01

Pärt, Arvo. Litany; Psalom; Trisagion. Hilliard Ensemble, Estonian Philharmonic Chamber Choir, Tallinn Chamber Orchestra, Tõnu Kaljuste; Lithuanian Chamber Orchestra, Saulius Sondeckis. ECM 1592/449 810-2

Narcisisma saistība ar pašnovērtējumu.

OpenAIRE

Strode, Indra

2008-01-01

Pētījuma mēķis ir noskaidrot narcisisma saistību ar pašvērtējumu. Pētījuma respondentu skaits – 60 cilvēki (30 sievietes, 30vīrieši). Vecuma ierobežojums – no 20 līdz 30 gadi. Pašvērtējuma līmeņa noteikšanai tika izmantota Rozenberga pašnovērtējuma aptauja („Rosenberg Self-Esteem Scale”, Rosenberg, 1965), narcisisma noteikšanai tik izmantota Narcistiskas personības aptauja (Raskin & Terry, 1988). Vīriešu narcisisma un pašvērtējuma rādītāji uzrādīja statistiski nozīmīgu korelāciju, tāpat kā si...

Endurance Training Intensity Does Not Mediate Interference to Maximal Lower-Body Strength Gain during Short-Term Concurrent Training.

Science.gov (United States)

Fyfe, Jackson J; Bartlett, Jonathan D; Hanson, Erik D; Stepto, Nigel K; Bishop, David J

2016-01-01

We determined the effect of concurrent training incorporating either high-intensity interval training (HIT) or moderate-intensity continuous training (MICT) on maximal strength, counter-movement jump (CMJ) performance, and body composition adaptations, compared with single-mode resistance training (RT). Twenty-three recreationally-active males (mean ± SD: age, 29.6 ± 5.5 y; [Formula: see text], 44 ± 11 mL kg -1 ·min -1 ) underwent 8 weeks (3 sessions·wk -1 ) of either: (1) HIT combined with RT (HIT+RT group, n = 8), (2) work-matched MICT combined with RT (MICT+RT group, n = 7), or (3) RT performed alone (RT group, n = 8). Measures of aerobic capacity, maximal (1-RM) strength, CMJ performance and body composition (DXA) were obtained before (PRE), mid-way (MID), and after (POST) training. Maximal (one-repetition maximum [1-RM]) leg press strength was improved from PRE to POST for RT (mean change ± 90% confidence interval; 38.5 ± 8.5%; effect size [ES] ± 90% confidence interval; 1.26 ± 0.24; P body lean mass was similarly increased for RT (4.1 ± 2.0%; ES; 0.33 ± 0.16; P = 0.023) and MICT+RT (3.6 ± 2.4%; ES; 0.45 ± 0.30; P = 0.052); however, this change was attenuated for HIT+RT (1.8 ± 1.6%; ES; 0.13 ± 0.12; P = 0.069). We conclude that concurrent training incorporating either HIT or work-matched MICT similarly attenuates improvements in maximal lower-body strength and indices of CMJ performance compared with RT performed alone. This suggests endurance training intensity is not a critical mediator of interference to maximal strength gain during short-term concurrent training.

Mamma diagnostics for MTRA (medical-radiological personnel)/RT (radiologists); Mammadiagnostik fuer MTRA/RT

Energy Technology Data Exchange (ETDEWEB)

Fischer, Uwe; Baum, Friedemann

2014-07-01

The text book on mamma diagnostics for MTRA (medical-radiological personnel)/RT (radiologists) covers the following issues: Anatomy, development and physiology of mammary glands; tumor development an breast cancer risk; pathology, non-imaging diagnostics; mammography: physical-technical fundamentals; mammography: analogue technique; mammography: digital technique; mammography: quality assurance; mammography: legal questions and radiation protection; mammography: new developments; mammography: setting technique; mammography: use and appraisal; mamma-sonography: technique and methodology; mamma-sonography: assignment and appraisal, mamma-NMR: technique and methodology; mamma-NMR: assignment and appraisal lymph node diagnostics; mamma interventions; biopsy; mamma interventions: marking examination concepts; therapeutic concepts; hygienic concepts; communication and interaction.

Canine distemper virus detection by different methods of One-Step RT-qPCR

Directory of Open Access Journals (Sweden)

Claudia de Camargo Tozato

2016-01-01

Full Text Available ABSTRACT: Three commercial kits of One-Step RT-qPCR were evaluated for the molecular diagnosis of Canine Distemper Virus. Using the kit that showed better performance, two systems of Real-time RT-PCR (RT-qPCR assays were tested and compared for analytical sensitivity to Canine Distemper Virus RNA detection: a One-Step RT-qPCR (system A and a One-Step RT-qPCR combined with NESTED-qPCR (system B. Limits of detection for both systems were determined using a serial dilution of Canine Distemper Virus synthetic RNA or a positive urine sample. In addition, the same urine sample was tested using samples with prior centrifugation or ultracentrifugation. Commercial kits of One-Step RT-qPCR assays detected canine distemper virus RNA in 10 (100% urine samples from symptomatic animals tested. The One-Step RT-qPCR kit that showed better results was used to evaluate the analytical sensitivity of the A and B systems. Limit of detection using synthetic RNA for the system A was 11 RNA copies µL-1 and 110 RNA copies µl-1 for first round System B. The second round of the NESTED-qPCR for System B had a limit of detection of 11 copies µl-1. Relationship between Ct values and RNA concentration was linear. The RNA extracted from the urine dilutions was detected in dilutions of 10-3 and10-2 by System A and B respectively. Urine centrifugation increased the analytical sensitivity of the test and proved to be useful for routine diagnostics. The One-Step RT-qPCR is a fast, sensitive and specific method for canine distemper routine diagnosis and research projects that require sensitive and quantitative methodology.

"I know your name, but not your number"--Patients with verbal short-term memory deficits are impaired in learning sequences of digits.

Science.gov (United States)

Bormann, Tobias; Seyboth, Margret; Umarova, Roza; Weiller, Cornelius

2015-06-01

Studies on verbal learning in patients with impaired verbal short-term memory (vSTM) have revealed dissociations among types of verbal information. Patients with impaired vSTM are able to learn lists of known words but fail to acquire new word forms. This suggests that vSTM is involved in new word learning. The present study assessed both new word learning and the learning of digit sequences in two patients with impaired vSTM. In two experiments, participants were required to learn people's names, ages and professions, or their four digit 'phone numbers'. The STM patients were impaired on learning unknown family names and phone numbers, but managed to acquire other verbal information. In contrast, a patient with a severe verbal episodic memory impairment was impaired across information types. These results indicate verbal STM involvement in the learning of digit sequences. Copyright © 2015 Elsevier Ltd. All rights reserved.

Identification of suitable reference genes for gene expression normalization in qRT-PCR analysis in watermelon.

Directory of Open Access Journals (Sweden)

Qiusheng Kong

Full Text Available Watermelon is one of the major Cucurbitaceae crops and the recent availability of genome sequence greatly facilitates the fundamental researches on it. Quantitative real-time reverse transcriptase PCR (qRT-PCR is the preferred method for gene expression analyses, and using validated reference genes for normalization is crucial to ensure the accuracy of this method. However, a systematic validation of reference genes has not been conducted on watermelon. In this study, transcripts of 15 candidate reference genes were quantified in watermelon using qRT-PCR, and the stability of these genes was compared using geNorm and NormFinder. geNorm identified ClTUA and ClACT, ClEF1α and ClACT, and ClCAC and ClTUA as the best pairs of reference genes in watermelon organs and tissues under normal growth conditions, abiotic stress, and biotic stress, respectively. NormFinder identified ClYLS8, ClUBCP, and ClCAC as the best single reference genes under the above experimental conditions, respectively. ClYLS8 and ClPP2A were identified as the best reference genes across all samples. Two to nine reference genes were required for more reliable normalization depending on the experimental conditions. The widely used watermelon reference gene 18SrRNA was less stable than the other reference genes under the experimental conditions. Catalase family genes were identified in watermelon genome, and used to validate the reliability of the identified reference genes. ClCAT1and ClCAT2 were induced and upregulated in the first 24 h, whereas ClCAT3 was downregulated in the leaves under low temperature stress. However, the expression levels of these genes were significantly overestimated and misinterpreted when 18SrRNA was used as a reference gene. These results provide a good starting point for reference gene selection in qRT-PCR analyses involving watermelon.

Pärt, Arvo: "Miserere", "Festina lente" / Volkmar Fischer

Index Scriptorium Estoniae

Fischer, Volkmar

1991-01-01

Uuest heliplaadist Pärt, Arvo: "Miserere", "Festina lente", "Sarah Was Ninety Years Old". The Hilliard Ensemble. Orchester der Beethovenhalle Bonn. Dirigent Dennis Russel Davies (ECM Records, distr Amigo)

Localisation of Abundant and Organ-Specific Genes Expressed in Rosa hybrida Leaves and Flower Buds by Direct In Situ RT-PCR

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Agata Jedrzejuk

2012-01-01

Full Text Available In situ PCR is a technique that allows specific nucleic acid sequences to be detected in individual cells and tissues. In situ PCR and IS-RT-PCR are elegant techniques that can increase both sensitivity and throughput, but they are, at best, only semiquantitative; therefore, it is desirable first to ascertain the expression pattern by conventional means to establish the suitable conditions for each probe. In plants, in situ RT-PCR is widely used in the expression localisation of specific genes, including MADS-box and other function-specific genes or housekeeping genes in floral buds and other organs. This method is especially useful in small organs or during early developmental stages when the separation of particular parts is impossible. In this paper, we compared three different labelling and immunodetection methods by using in situ RT-PCR in Rosa hybrida flower buds and leaves. As target genes, we used the abundant β-actin and RhFUL gene, which is expressed only in the leaves and petals/sepals of flower buds. We used digoxygenin-11-dUTP, biotin-11-dUTP, and fluorescein-12-dUTP-labelled nucleotides and antidig-AP/ streptavidin-fluorescein-labelled antibodies. All of the used methods gave strong, specific signal and all of them may be used in localization of gene expression on tissue level in rose organs.

Direct Detection and Identification of Enteroviruses from Faeces of Healthy Nigerian Children Using a Cell-Culture Independent RT-Seminested PCR Assay

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Temitope Oluwasegun Cephas Faleye

2016-01-01

Full Text Available Recently, a cell-culture independent protocol for detection of enteroviruses from clinical specimen was recommended by the WHO for surveillance alongside the previously established protocols. Here, we investigated whether this new protocol will show the same enterovirus diversity landscape as the established cell-culture dependent protocols. Faecal samples were collected from sixty apparently healthy children in Ibadan, Nigeria. Samples were resuspended in phosphate buffered saline, RNA was extracted, and the VP1 gene was amplified using WHO recommended RT-snPCR protocol. Amplicons were sequenced and sequences subjected to phylogenetic analysis. Fifteen (25% of the 60 samples yielded the expected band size. Of the 15 amplicons sequenced, 12 were exploitable. The remaining 3 had electropherograms with multiple peaks and were unexploitable. Eleven of the 12 exploitable sequences were identified as Coxsackievirus A1 (CVA1, CVA3, CVA4, CVA8, CVA20, echovirus 32 (E32, enterovirus 71 (EV71, EVB80, and EVC99. Subsequently, the last exploitable sequence was identified as enterobacteriophage baseplate gene by nucleotide BLAST. The results of this study document the first description of molecular sequence data on CVA1, CVA8, and E32 strains present in Nigeria. The result further showed that species A enteroviruses were more commonly detected in the region when cell-culture bias is bypassed.

Genomic Sequence of a Ranavirus Isolated from Short-Finned Eel (Anguilla australis)

DEFF Research Database (Denmark)

Subramaniam, Kuttichantran; Toffan, Anna; Cappellozza, Elisabetta

2016-01-01

The short-finned eel ranavirus (SERV) was isolated from short-finned eel imported to Italy from New Zealand. Phylogenomic analyses revealed that SERV is a unique member of the genus Ranavirus, family Iridoviridae, branching at the base of the tree near other fish ranaviruses....

Pärt, Arvo: Seven Magnificat Antiphons / Barry Witherden

Index Scriptorium Estoniae

Witherden, Barry

1996-01-01

Uuest heliplaadist "Pärt, Arvo: Seven Magnificat Antiphons. Magnificat. Summa; Tormis, Veljo: The Curse Upon Iron. Karelian Destiny. BBC Singers, Bo Holton". Collins Classics 1472-2 (61 minutes: DDD)

Design of Long Period Pseudo-Random Sequences from the Addition of m -Sequences over 𝔽 p

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Ren Jian

2004-01-01

Full Text Available Pseudo-random sequence with good correlation property and large linear span is widely used in code division multiple access (CDMA communication systems and cryptology for reliable and secure information transmission. In this paper, sequences with long period, large complexity, balance statistics, and low cross-correlation property are constructed from the addition of m -sequences with pairwise-prime linear spans (AMPLS. Using m -sequences as building blocks, the proposed method proved to be an efficient and flexible approach to construct long period pseudo-random sequences with desirable properties from short period sequences. Applying the proposed method to 𝔽 2 , a signal set ( ( 2 n − 1 ( 2 m − 1 , ( 2 n + 1 ( 2 m + 1 , ( 2 ( n + 1 / 2 + 1 ( 2 ( m + 1 / 2 + 1 is constructed.

Red blood cells in cerebrospinal fluid as possible inhibitory factor for enterovirus RT-PCR

Directory of Open Access Journals (Sweden)

Sérgio Monteiro de Almeida

Full Text Available ABSTRACT The presence of hemoglobin in samples are considered an important inhibitory factor for polymerase chain reaction (PCR. The aim of this study was to examine the influence of red blood cells (RBCs in cerebrospinal fluid (CSF as an inhibitory factor to reverse transcription polymerase chain reaction (RT-PCR for enteroviruses (EV. Forty-four CSF samples from patients showing characteristics of viral meningitis were assessed for EV by RT-PCR. Viral RNA extracted with guanidine isothyocianate buffer and virus detection was performed by in-house nested PCR. Positivity for EV RT-PCR was higher in CSF samples without RBCs than in samples with RBCs: 13(26% and 36(9.2%, p = 0.001. In the group with positive EV RT-PCR, the mean + SD CSF RBC was 37 ± 183 cell/mm3; the group with negative results had 580 + 2,890 cell/mm3 (p = 0.007. The acceptable upper limit for CSF RBCs that could not influence RT-PCR was 108 cells/mm3. CSF samples with negative results for EV RT-PCR have more erythrocytes.

On the Origin of Reverse Transcriptase-Using CRISPR-Cas Systems and Their Hyperdiverse, Enigmatic Spacer Repertoires.

Science.gov (United States)

Silas, Sukrit; Makarova, Kira S; Shmakov, Sergey; Páez-Espino, David; Mohr, Georg; Liu, Yi; Davison, Michelle; Roux, Simon; Krishnamurthy, Siddharth R; Fu, Becky Xu Hua; Hansen, Loren L; Wang, David; Sullivan, Matthew B; Millard, Andrew; Clokie, Martha R; Bhaya, Devaki; Lambowitz, Alan M; Kyrpides, Nikos C; Koonin, Eugene V; Fire, Andrew Z

2017-07-11

Cas1 integrase is the key enzyme of the clustered regularly interspaced short palindromic repeat (CRISPR)-Cas adaptation module that mediates acquisition of spacers derived from foreign DNA by CRISPR arrays. In diverse bacteria, the cas1 gene is fused (or adjacent) to a gene encoding a reverse transcriptase (RT) related to group II intron RTs. An RT-Cas1 fusion protein has been recently shown to enable acquisition of CRISPR spacers from RNA. Phylogenetic analysis of the CRISPR-associated RTs demonstrates monophyly of the RT-Cas1 fusion, and coevolution of the RT and Cas1 domains. Nearly all such RTs are present within type III CRISPR-Cas loci, but their phylogeny does not parallel the CRISPR-Cas type classification, indicating that RT-Cas1 is an autonomous functional module that is disseminated by horizontal gene transfer and can function with diverse type III systems. To compare the sequence pools sampled by RT-Cas1-associated and RT-lacking CRISPR-Cas systems, we obtained samples of a commercially grown cyanobacterium- Arthrospira platensis Sequencing of the CRISPR arrays uncovered a highly diverse population of spacers. Spacer diversity was particularly striking for the RT-Cas1-containing type III-B system, where no saturation was evident even with millions of sequences analyzed. In contrast, analysis of the RT-lacking type III-D system yielded a highly diverse pool but reached a point where fewer novel spacers were recovered as sequencing depth was increased. Matches could be identified for a small fraction of the non-RT-Cas1-associated spacers, and for only a single RT-Cas1-associated spacer. Thus, the principal source(s) of the spacers, particularly the hypervariable spacer repertoire of the RT-associated arrays, remains unknown. IMPORTANCE While the majority of CRISPR-Cas immune systems adapt to foreign genetic elements by capturing segments of invasive DNA, some systems carry reverse transcriptases (RTs) that enable adaptation to RNA molecules. From

Complete sequence of Tvv1, a family of Ty 1 copia-like retrotransposons of Vitis vinifera L., reconstituted by chromosome walking.

Science.gov (United States)

Pelsy, F.; Merdinoglu, D.

2002-09-01

A chromosome-walking strategy was used to sequence and characterize retrotransposons in the grapevine genome. The reconstitution of a family of retroelements, named Tvv1, was achieved by six successive steps. These elements share a single, highly conserved open reading frame 4,153 nucleotides-long, putatively encoding the gag, pro, int, rt and rh proteins. Comparison of the Tvv1 open reading frame coding potential with those of drosophila copia and tobacco Tnt1, revealed that Tvv1 is closely related to Ty 1 copia-like retrotransposons. A highly variable untranslated leader region, upstream of the open reading frame, allowed us to differentiate Tvv1 variants, which represent a family of at least 28 copies, in varying sizes. This internal region is flanked by two long terminal repeats in direct orientation, sized between 149 and 157 bp. Among elements theoretically sized from 4,970 to 5,550 bp, we describe the full-length sequence of a reference element Tvv1-1, 5,343 nucleotides-long. The full-length sequence of Tvv1-1 compared to pea PDR1 shows a 53.3% identity. In addition, both elements contain long terminal repeats of nearly the same size in which the U5 region could be entirely absent. Therefore, we assume that Tvv1 and PDR1 could constitute a particular class of short LTRs retroelements.

Analysis on the arcelin expression in bruchid pest resistant wild pulses using real time RT-qPCR.

Science.gov (United States)

Sakthivelkumar, Shanmugavel; Veeramani, Velayutham; Hilda, Karuppiah; Arumugam, Munusamy; Janarthanan, Sundaram

2014-12-01

Arcelin, the antimetabolic protein from wild pulses is a known natural insecticidal molecule. Wild pulses with high arcelin content could serve as potential source to. increase the levels of insect resistance in cultivated pulse crops. In this study, arcelin (Arl) gene expression was screened in seven stored product insect pest resistant wild pulse varieties using real time RT-qPCR. Arcelin gene specific real time PCR primers were synthesized from arcelin mRNA sequence of the wild pulse variety, Lablab purpureus. The results revealed different levels of arcelin gene expression in the tested varieties. Canavalia virosa registered significantly high content indicating its suitability for utilization of arcelin gene in developing stored product insect pest resistance with other cultivated pulses.

Identification of Biomolecular Building Blocks by Recognition Tunneling: Stride towards Nanopore Sequencing of Biomolecules

Science.gov (United States)

Sen, Suman

DNA, RNA and Protein are three pivotal biomolecules in human and other organisms, playing decisive roles in functionality, appearance, diseases development and other physiological phenomena. Hence, sequencing of these biomolecules acquires the prime interest in the scientific community. Single molecular identification of their building blocks can be done by a technique called Recognition Tunneling (RT) based on Scanning Tunneling Microscope (STM). A single layer of specially designed recognition molecule is attached to the STM electrodes, which trap the targeted molecules (DNA nucleoside monophosphates, RNA nucleoside monophosphates or amino acids) inside the STM nanogap. Depending on their different binding interactions with the recognition molecules, the analyte molecules generate stochastic signal trains accommodating their "electronic fingerprints". Signal features are used to detect the molecules using a machine learning algorithm and different molecules can be identified with significantly high accuracy. This, in turn, paves the way for rapid, economical nanopore sequencing platform, overcoming the drawbacks of Next Generation Sequencing (NGS) techniques. To read DNA nucleotides with high accuracy in an STM tunnel junction a series of nitrogen-based heterocycles were designed and examined to check their capabilities to interact with naturally occurring DNA nucleotides by hydrogen bonding in the tunnel junction. These recognition molecules are Benzimidazole, Imidazole, Triazole and Pyrrole. Benzimidazole proved to be best among them showing DNA nucleotide classification accuracy close to 99%. Also, Imidazole reader can read an abasic monophosphate (AP), a product from depurination or depyrimidination that occurs 10,000 times per human cell per day. In another study, I have investigated a new universal reader, 1-(2-mercaptoethyl)pyrene (Pyrene reader) based on stacking interactions, which should be more specific to the canonical DNA nucleosides. In addition

Märt Agu: "Isa vari käib mul kogu aeg kannul" / Märt Agu ; interv. Aigi Viira

Index Scriptorium Estoniae

Agu, Märt

2006-01-01

1. okt. tähistas oma 30-aastast sünnipäeva ja oma ellukutsuja Mait Agu 55-sünniaastapäeva Tallinna Ülikooli koreograafiaosakond. Sel puhul esitati Estonias Mait Agu tantsulavastus "Kodalased ja kaasaegsed", mille seadis uuesti lavale Märt Agu oma isa loodud koreograafia põhjal

A Formal Approach for RT-DVS Algorithms Evaluation Based on Statistical Model Checking

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Shengxin Dai

2015-01-01

Full Text Available Energy saving is a crucial concern in embedded real time systems. Many RT-DVS algorithms have been proposed to save energy while preserving deadline guarantees. This paper presents a novel approach to evaluate RT-DVS algorithms using statistical model checking. A scalable framework is proposed for RT-DVS algorithms evaluation, in which the relevant components are modeled as stochastic timed automata, and the evaluation metrics including utilization bound, energy efficiency, battery awareness, and temperature awareness are expressed as statistical queries. Evaluation of these metrics is performed by verifying the corresponding queries using UPPAAL-SMC and analyzing the statistical information provided by the tool. We demonstrate the applicability of our framework via a case study of five classical RT-DVS algorithms.

Stem-Loop RT-qPCR as an Efficient Tool for the Detection and Quantification of Small RNAs in Giardia lamblia

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Jaime Marcial-Quino

2016-12-01

Full Text Available Stem-loop quantitative reverse transcription PCR (RT-qPCR is a molecular technique used for identification and quantification of individual small RNAs in cells. In this work, we used a Universal ProbeLibrary (UPL-based design to detect—in a rapid, sensitive, specific, and reproducible way—the small nucleolar RNA (snoRNA GlsR17 and its derived miRNA (miR2 of Giardia lamblia using a stem-loop RT-qPCR approach. Both small RNAs could be isolated from both total RNA and small RNA samples. Identification of the two small RNAs was carried out by sequencing the PCR-amplified small RNA products upon ligation into the pJET1.2/blunt vector. GlsR17 is constitutively expressed during the 72 h cultures of trophozoites, while the mature miR2 is present in 2-fold higher abundance during the first 48 h than at 72 h. Because it has been suggested that miRNAs in G. lamblia have an important role in the regulation of gene expression, the use of the stem-loop RT-qPCR method could be valuable for the study of miRNAs of G. lamblia. This methodology will be a powerful tool for studying gene regulation in G. lamblia, and will help to better understand the features and functions of these regulatory molecules and how they work within the RNA interference (RNAi pathway in G. lamblia.

Stem-Loop RT-qPCR as an Efficient Tool for the Detection and Quantification of Small RNAs in Giardia lamblia

Science.gov (United States)

Marcial-Quino, Jaime; Gómez-Manzo, Saúl; Fierro, Francisco; Vanoye-Carlo, America; Rufino-González, Yadira; Sierra-Palacios, Edgar; Castillo-Villanueva, Adriana; Castillo-Rodríguez, Rosa Angélica; Rodríguez-Bustamante, Eduardo; Arreguin-Espinosa, Roberto; Reyes-Vivas, Horacio

2016-01-01

Stem-loop quantitative reverse transcription PCR (RT-qPCR) is a molecular technique used for identification and quantification of individual small RNAs in cells. In this work, we used a Universal ProbeLibrary (UPL)-based design to detect—in a rapid, sensitive, specific, and reproducible way—the small nucleolar RNA (snoRNA) GlsR17 and its derived miRNA (miR2) of Giardia lamblia using a stem-loop RT-qPCR approach. Both small RNAs could be isolated from both total RNA and small RNA samples. Identification of the two small RNAs was carried out by sequencing the PCR-amplified small RNA products upon ligation into the pJET1.2/blunt vector. GlsR17 is constitutively expressed during the 72 h cultures of trophozoites, while the mature miR2 is present in 2-fold higher abundance during the first 48 h than at 72 h. Because it has been suggested that miRNAs in G. lamblia have an important role in the regulation of gene expression, the use of the stem-loop RT-qPCR method could be valuable for the study of miRNAs of G. lamblia. This methodology will be a powerful tool for studying gene regulation in G. lamblia, and will help to better understand the features and functions of these regulatory molecules and how they work within the RNA interference (RNAi) pathway in G. lamblia. PMID:27999395

Comparison of preoperative short-course radiotherapy and long-course radiochemotherapy for locally advanced rectal cancer

International Nuclear Information System (INIS)

Guckenberger, M.; Saur, G.; Wehner, D.; Sweeney, R.A.; Flentje, M.; Thalheimer, A.; Germer, C.T.

2012-01-01

Background: The purpose of this work was to perform a single institution comparison between preoperative short-course radiotherapy (SC-RT) and long-course radiochemotherapy (LC-RCHT) for locally advanced rectal cancer. Methods: A total of 225 patients with clinical stage UICC II-III rectal cancer were treated with SC-RT (29 Gy in 10 twice daily fractions followed by immediate surgery; n = 108) or LC-RCHT (54 Gy in 28 fractions with simultaneous 5-fluorouracil (5-FU) ± oxaliplatin chemotherapy followed by delayed surgery; n = 117). All patients in the LC-RCHT cohort and patients in the SC-RT with pathological UICC stage ≥ II received adjuvant chemotherapy. Before 2004, the standard of care was SC-RT with LC-RCHT reserved for patients where downstaging was considered as required for sphincter preservation or curative resection. In the later period, SC-RT was practiced only for patients unfit for radiochemotherapy. Results: Patients in the LC-RCHT cohort had a significantly higher proportion of cT4 tumors, clinical node positivity, and lower tumor location. The 5-year local control (LC) and overall survival (OS) were 91% and 66% without differences between the SC-RT and LC-RCHT groups. Acute toxicity was increased during LC-RCHT (grade ≥ II 1% vs. 33%) and there were no differences in postoperative complications. Severe late toxicity grade ≥ III was increased after SC-RT (12% vs. 3%). Of patients aged > 80 years, 7 of 7 patients and 4 of 9 patients received curative surgery after SC-RT and LC-RCHT, respectively. Conclusion: Despite the fact that patients with worse prognostic factors were treated with LC-RCHT, there were no significant differences in LC and OS between the SC-RT and LC-RCHT group. Age > 80 years was identified as a significant risk factor for LC-RCHT and these patients could be treated preferably with SC-RT. (orig.)

High-Throughput Next-Generation Sequencing of Polioviruses

Science.gov (United States)

Montmayeur, Anna M.; Schmidt, Alexander; Zhao, Kun; Magaña, Laura; Iber, Jane; Castro, Christina J.; Chen, Qi; Henderson, Elizabeth; Ramos, Edward; Shaw, Jing; Tatusov, Roman L.; Dybdahl-Sissoko, Naomi; Endegue-Zanga, Marie Claire; Adeniji, Johnson A.; Oberste, M. Steven; Burns, Cara C.

2016-01-01

ABSTRACT The poliovirus (PV) is currently targeted for worldwide eradication and containment. Sanger-based sequencing of the viral protein 1 (VP1) capsid region is currently the standard method for PV surveillance. However, the whole-genome sequence is sometimes needed for higher resolution global surveillance. In this study, we optimized whole-genome sequencing protocols for poliovirus isolates and FTA cards using next-generation sequencing (NGS), aiming for high sequence coverage, efficiency, and throughput. We found that DNase treatment of poliovirus RNA followed by random reverse transcription (RT), amplification, and the use of the Nextera XT DNA library preparation kit produced significantly better results than other preparations. The average viral reads per total reads, a measurement of efficiency, was as high as 84.2% ± 15.6%. PV genomes covering >99 to 100% of the reference length were obtained and validated with Sanger sequencing. A total of 52 PV genomes were generated, multiplexing as many as 64 samples in a single Illumina MiSeq run. This high-throughput, sequence-independent NGS approach facilitated the detection of a diverse range of PVs, especially for those in vaccine-derived polioviruses (VDPV), circulating VDPV, or immunodeficiency-related VDPV. In contrast to results from previous studies on other viruses, our results showed that filtration and nuclease treatment did not discernibly increase the sequencing efficiency of PV isolates. However, DNase treatment after nucleic acid extraction to remove host DNA significantly improved the sequencing results. This NGS method has been successfully implemented to generate PV genomes for molecular epidemiology of the most recent PV isolates. Additionally, the ability to obtain full PV genomes from FTA cards will aid in facilitating global poliovirus surveillance. PMID:27927929

Ring trial 2016 for Bluetongue virus detection by real-time RT-PCR in France.

Science.gov (United States)

Sailleau, Corinne; Viarouge, Cyril; Breard, Emmanuel; Vitour, Damien; Zientara, Stephan

2017-05-01

Since the unexpected emergence of BTV-8 in Northern Europe and the incursion of BTV-8 and 1 in France in 2006-2007, molecular diagnosis has considerably evolved. Several real-time RT-PCR (rtRT-PCR) methods have been developed and published, and are currently being used in many countries across Europe for BTV detection and typing. In France, the national reference laboratory (NRL) for orbiviruses develops and validates 'ready-to-use' kits with private companies for viral RNA detection. The regional laboratories network that was set up to deal with a heavy demand for analyses has used these available kits. From 2007, ring tests were organized to monitor the performance of the French laboratories. This study presents the results of 63 regional laboratories in the ring trial organized in 2016. Blood samples were sent to the laboratories. Participants were asked to use the rtRT-PCR methods in place in their laboratory, for detection of all BTV serotypes and specifically BTV-8. The French regional laboratories are able to detect and genotype BTV in affected animals. Despite the use of several methods (i.e. RNA extraction and different commercial rtRT-PCRs), the network is homogeneous. The ring trial demonstrated that the French regional veterinary laboratories have reliable and robust BTV diagnostic tools for BTV genome detection.

Identification of microRNAs from Amur grape (Vitis amurensis Rupr.) by deep sequencing and analysis of microRNA variations with bioinformatics.

Science.gov (United States)

Wang, Chen; Han, Jian; Liu, Chonghuai; Kibet, Korir Nicholas; Kayesh, Emrul; Shangguan, Lingfei; Li, Xiaoying; Fang, Jinggui

2012-03-29

MicroRNA (miRNA) is a class of functional non-coding small RNA with 19-25 nucleotides in length while Amur grape (Vitis amurensis Rupr.) is an important wild fruit crop with the strongest cold resistance among the Vitis species, is used as an excellent breeding parent for grapevine, and has elicited growing interest in wine production. To date, there is a relatively large number of grapevine miRNAs (vv-miRNAs) from cultivated grapevine varieties such as Vitis vinifera L. and hybrids of V. vinifera and V. labrusca, but there is no report on miRNAs from Vitis amurensis Rupr, a wild grapevine species. A small RNA library from Amur grape was constructed and Solexa technology used to perform deep sequencing of the library followed by subsequent bioinformatics analysis to identify new miRNAs. In total, 126 conserved miRNAs belonging to 27 miRNA families were identified, and 34 known but non-conserved miRNAs were also found. Significantly, 72 new potential Amur grape-specific miRNAs were discovered. The sequences of these new potential va-miRNAs were further validated through miR-RACE, and accumulation of 18 new va-miRNAs in seven tissues of grapevines confirmed by real time RT-PCR (qRT-PCR) analysis. The expression levels of va-miRNAs in flowers and berries were found to be basically consistent in identity to those from deep sequenced sRNAs libraries of combined corresponding tissues. We also describe the conservation and variation of va-miRNAs using miR-SNPs and miR-LDs during plant evolution based on comparison of orthologous sequences, and further reveal that the number and sites of miR-SNP in diverse miRNA families exhibit distinct divergence. Finally, 346 target genes for the new miRNAs were predicted and they include a number of Amur grape stress tolerance genes and many genes regulating anthocyanin synthesis and sugar metabolism. Deep sequencing of short RNAs from Amur grape flowers and berries identified 72 new potential miRNAs and 34 known but non-conserved mi

Identification of microRNAs from Amur grape (vitis amurensis Rupr. by deep sequencing and analysis of microRNA variations with bioinformatics

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Wang Chen

2012-03-01

Full Text Available Abstract Background MicroRNA (miRNA is a class of functional non-coding small RNA with 19-25 nucleotides in length while Amur grape (Vitis amurensis Rupr. is an important wild fruit crop with the strongest cold resistance among the Vitis species, is used as an excellent breeding parent for grapevine, and has elicited growing interest in wine production. To date, there is a relatively large number of grapevine miRNAs (vv-miRNAs from cultivated grapevine varieties such as Vitis vinifera L. and hybrids of V. vinifera and V. labrusca, but there is no report on miRNAs from Vitis amurensis Rupr, a wild grapevine species. Results A small RNA library from Amur grape was constructed and Solexa technology used to perform deep sequencing of the library followed by subsequent bioinformatics analysis to identify new miRNAs. In total, 126 conserved miRNAs belonging to 27 miRNA families were identified, and 34 known but non-conserved miRNAs were also found. Significantly, 72 new potential Amur grape-specific miRNAs were discovered. The sequences of these new potential va-miRNAs were further validated through miR-RACE, and accumulation of 18 new va-miRNAs in seven tissues of grapevines confirmed by real time RT-PCR (qRT-PCR analysis. The expression levels of va-miRNAs in flowers and berries were found to be basically consistent in identity to those from deep sequenced sRNAs libraries of combined corresponding tissues. We also describe the conservation and variation of va-miRNAs using miR-SNPs and miR-LDs during plant evolution based on comparison of orthologous sequences, and further reveal that the number and sites of miR-SNP in diverse miRNA families exhibit distinct divergence. Finally, 346 target genes for the new miRNAs were predicted and they include a number of Amur grape stress tolerance genes and many genes regulating anthocyanin synthesis and sugar metabolism. Conclusions Deep sequencing of short RNAs from Amur grape flowers and berries identified 72

Real-time RT-PCR high-resolution melting curve analysis and multiplex RT-PCR to detect and differentiate grapevine leafroll-associated associated virus 3 variant groups I, II, III and VI

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Bester Rachelle

2012-09-01

Full Text Available Abstract Background Grapevine leafroll-associated virus 3 (GLRaV-3 is the main contributing agent of leafroll disease worldwide. Four of the six GLRaV-3 variant groups known have been found in South Africa, but their individual contribution to leafroll disease is unknown. In order to study the pathogenesis of leafroll disease, a sensitive and accurate diagnostic assay is required that can detect different variant groups of GLRaV-3. Methods In this study, a one-step real-time RT-PCR, followed by high-resolution melting (HRM curve analysis for the simultaneous detection and identification of GLRaV-3 variants of groups I, II, III and VI, was developed. A melting point confidence interval for each variant group was calculated to include at least 90% of all melting points observed. A multiplex RT-PCR protocol was developed to these four variant groups in order to assess the efficacy of the real-time RT-PCR HRM assay. Results A universal primer set for GLRaV-3 targeting the heat shock protein 70 homologue (Hsp70h gene of GLRaV-3 was designed that is able to detect GLRaV-3 variant groups I, II, III and VI and differentiate between them with high-resolution melting curve analysis. The real-time RT-PCR HRM and the multiplex RT-PCR were optimized using 121 GLRaV-3 positive samples. Due to a considerable variation in melting profile observed within each GLRaV-3 group, a confidence interval of above 90% was calculated for each variant group, based on the range and distribution of melting points. The intervals of groups I and II could not be distinguished and a 95% joint confidence interval was calculated for simultaneous detection of group I and II variants. An additional primer pair targeting GLRaV-3 ORF1a was developed that can be used in a subsequent real-time RT-PCR HRM to differentiate between variants of groups I and II. Additionally, the multiplex RT-PCR successfully validated 94.64% of the infections detected with the real-time RT-PCR HRM

De Novo Assembly of Human Herpes Virus Type 1 (HHV-1) Genome, Mining of Non-Canonical Structures and Detection of Novel Drug-Resistance Mutations Using Short- and Long-Read Next Generation Sequencing Technologies.

Science.gov (United States)

Karamitros, Timokratis; Harrison, Ian; Piorkowska, Renata; Katzourakis, Aris; Magiorkinis, Gkikas; Mbisa, Jean Lutamyo

2016-01-01

Human herpesvirus type 1 (HHV-1) has a large double-stranded DNA genome of approximately 152 kbp that is structurally complex and GC-rich. This makes the assembly of HHV-1 whole genomes from short-read sequencing data technically challenging. To improve the assembly of HHV-1 genomes we have employed a hybrid genome assembly protocol using data from two sequencing technologies: the short-read Roche 454 and the long-read Oxford Nanopore MinION sequencers. We sequenced 18 HHV-1 cell culture-isolated clinical specimens collected from immunocompromised patients undergoing antiviral therapy. The susceptibility of the samples to several antivirals was determined by plaque reduction assay. Hybrid genome assembly resulted in a decrease in the number of contigs in 6 out of 7 samples and an increase in N(G)50 and N(G)75 of all 7 samples sequenced by both technologies. The approach also enhanced the detection of non-canonical contigs including a rearrangement between the unique (UL) and repeat (T/IRL) sequence regions of one sample that was not detectable by assembly of 454 reads alone. We detected several known and novel resistance-associated mutations in UL23 and UL30 genes. Genome-wide genetic variability ranged from genomes will be useful in determining genetic determinants of drug resistance, virulence, pathogenesis and viral evolution. The numerous, complex repeat regions of the HHV-1 genome currently remain a barrier towards this goal.

Questioning short-term memory and its measurement: Why digit span measures long-term associative learning.

Science.gov (United States)

Jones, Gary; Macken, Bill

2015-11-01

Traditional accounts of verbal short-term memory explain differences in performance for different types of verbal material by reference to inherent characteristics of the verbal items making up memory sequences. The role of previous experience with sequences of different types is ostensibly controlled for either by deliberate exclusion or by presenting multiple trials constructed from different random permutations. We cast doubt on this general approach in a detailed analysis of the basis for the robust finding that short-term memory for digit sequences is superior to that for other sequences of verbal material. Specifically, we show across four experiments that this advantage is not due to inherent characteristics of digits as verbal items, nor are individual digits within sequences better remembered than other types of individual verbal items. Rather, the advantage for digit sequences stems from the increased frequency, compared to other verbal material, with which digits appear in random sequences in natural language, and furthermore, relatively frequent digit sequences support better short-term serial recall than less frequent ones. We also provide corpus-based computational support for the argument that performance in a short-term memory setting is a function of basic associative learning processes operating on the linguistic experience of the rememberer. The experimental and computational results raise questions not only about the role played by measurement of digit span in cognition generally, but also about the way in which long-term memory processes impact on short-term memory functioning. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

A simple, flexible and efficient PCR-fusion/Gateway cloning procedure for gene fusion, site-directed mutagenesis, short sequence insertion and domain deletions and swaps

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Etchells J Peter

2009-10-01

Full Text Available Abstract Background The progress and completion of various plant genome sequencing projects has paved the way for diverse functional genomic studies that involve cloning, modification and subsequent expression of target genes. This requires flexible and efficient procedures for generating binary vectors containing: gene fusions, variants from site-directed mutagenesis, addition of protein tags together with domain swaps and deletions. Furthermore, efficient cloning procedures, ideally high throughput, are essential for pyramiding of multiple gene constructs. Results Here, we present a simple, flexible and efficient PCR-fusion/Gateway cloning procedure for construction of binary vectors for a range of gene fusions or variants with single or multiple nucleotide substitutions, short sequence insertions, domain deletions and swaps. Results from selected applications of the procedure which include ORF fusion, introduction of Cys>Ser mutations, insertion of StrepII tag sequence and domain swaps for Arabidopsis secondary cell wall AtCesA genes are demonstrated. Conclusion The PCR-fusion/Gateway cloning procedure described provides an elegant, simple and efficient solution for a wide range of diverse and complicated cloning tasks. Through streamlined cloning of sets of gene fusions and modification variants into binary vectors for systematic functional studies of gene families, our method allows for efficient utilization of the growing sequence and expression data.

Mixed Sequence Reader: A Program for Analyzing DNA Sequences with Heterozygous Base Calling

Science.gov (United States)

Chang, Chun-Tien; Tsai, Chi-Neu; Tang, Chuan Yi; Chen, Chun-Houh; Lian, Jang-Hau; Hu, Chi-Yu; Tsai, Chia-Lung; Chao, Angel; Lai, Chyong-Huey; Wang, Tzu-Hao; Lee, Yun-Shien

2012-01-01

The direct sequencing of PCR products generates heterozygous base-calling fluorescence chromatograms that are useful for identifying single-nucleotide polymorphisms (SNPs), insertion-deletions (indels), short tandem repeats (STRs), and paralogous genes. Indels and STRs can be easily detected using the currently available Indelligent or ShiftDetector programs, which do not search reference sequences. However, the detection of other genomic variants remains a challenge due to the lack of appropriate tools for heterozygous base-calling fluorescence chromatogram data analysis. In this study, we developed a free web-based program, Mixed Sequence Reader (MSR), which can directly analyze heterozygous base-calling fluorescence chromatogram data in .abi file format using comparisons with reference sequences. The heterozygous sequences are identified as two distinct sequences and aligned with reference sequences. Our results showed that MSR may be used to (i) physically locate indel and STR sequences and determine STR copy number by searching NCBI reference sequences; (ii) predict combinations of microsatellite patterns using the Federal Bureau of Investigation Combined DNA Index System (CODIS); (iii) determine human papilloma virus (HPV) genotypes by searching current viral databases in cases of double infections; (iv) estimate the copy number of paralogous genes, such as β-defensin 4 (DEFB4) and its paralog HSPDP3. PMID:22778697

GIF Video Sentiment Detection Using Semantic Sequence

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Dazhen Lin

2017-01-01

Full Text Available With the development of social media, an increasing number of people use short videos in social media applications to express their opinions and sentiments. However, sentiment detection of short videos is a very challenging task because of the semantic gap problem and sequence based sentiment understanding problem. In this context, we propose a SentiPair Sequence based GIF video sentiment detection approach with two contributions. First, we propose a Synset Forest method to extract sentiment related semantic concepts from WordNet to build a robust SentiPair label set. This approach considers the semantic gap between label words and selects a robust label subset which is related to sentiment. Secondly, we propose a SentiPair Sequence based GIF video sentiment detection approach that learns the semantic sequence to understand the sentiment from GIF videos. Our experiment results on GSO-2016 (GIF Sentiment Ontology data show that our approach not only outperforms four state-of-the-art classification methods but also shows better performance than the state-of-the-art middle level sentiment ontology features, Adjective Noun Pairs (ANPs.

Human papillomavirus detection using the Abbott RealTime high-risk HPV tests compared with conventional nested PCR coupled to high-throughput sequencing of amplification products in cervical smear specimens from a Gabonese female population.

Science.gov (United States)

Moussavou-Boundzanga, Pamela; Koumakpayi, Ismaël Hervé; Labouba, Ingrid; Leroy, Eric M; Belembaogo, Ernest; Berthet, Nicolas

2017-12-21

Cervical cancer is the fourth most common malignancy in women worldwide. However, screening with human papillomavirus (HPV) molecular tests holds promise for reducing cervical cancer incidence and mortality in low- and middle-income countries. The performance of the Abbott RealTime High-Risk HPV test (AbRT) was evaluated in 83 cervical smear specimens and compared with a conventional nested PCR coupled to high-throughput sequencing (HTS) to identify the amplicons. The AbRT assay detected at least one HPV genotype in 44.57% of women regardless of the grade of cervical abnormalities. Except for one case, good concordance was observed for the genotypes detected with the AbRT assay in the high-risk HPV category determined with HTS of the amplicon generated by conventional nested PCR. The AbRT test is an easy and reliable molecular tool and was as sensitive as conventional nested PCR in cervical smear specimens for detection HPVs associated with high-grade lesions. Moreover, sequencing amplicons using an HTS approach effectively identified the genotype of the hrHPV identified with the AbRT test.

Structural Evolution of the R-T Phase Boundary in KNN-Based Ceramics

KAUST Repository

Lv, Xiang

2017-10-04

Although a rhombohedral-tetragonal (R-T) phase boundary is known to substantially enhance the piezoelectric properties of potassium-sodium niobate ceramics, the structural evolution of the R-T phase boundary itself is still unclear. In this work, the structural evolution of R-T phase boundary from -150 °C to 200 °C is investigated in (0.99-x)K0.5Na0.5Nb1-ySbyO3-0.01CaSnO3-xBi0.5K0.5HfO3 (where x=0~0.05 with y=0.035, and y=0~0.07 with x=0.03) ceramics. Through temperature-dependent powder X-ray diffraction (XRD) patterns and Raman spectra, the structural evolution was determined to be Rhombohedral (R, <-125 °C) → Rhombohedral+Orthorhombic (R+O, -125 °C to 0 °C) → Rhombohedral+Tetragonal (R+T, 0 °C to 150 °C) → dominating Tetragonal (T, 200 °C to Curie temperature (TC)) → Cubic (C, >TC). In addition, the enhanced electrical properties (e.g., a direct piezoelectric coefficient (d33) of ~450±5 pC/N, a conversion piezoelectric coefficient (d33*) of ~580±5 pm/V, an electromechanical coupling factor (kp) of ~0.50±0.02, and TC~250 °C), fatigue-free behavior, and good thermal stability were exhibited by the ceramics possessing the R-T phase boundary. This work improves understanding of the physical mechanism behind the R-T phase boundary in KNN-based ceramics and is an important step towards their adoption in practical applications. This article is protected by copyright. All rights reserved.

Narr kui ajastu metafoor / Kärt Hellerma

Index Scriptorium Estoniae

Hellerma, Kärt, 1956-

2001-01-01

William Shakespeare'i "Kuningas Lear" (lav. Priit Pedajas) ja Pedro Calder̤n de la Barca "Elu on unenägu" (lav. Ingo Normet) Eesti Draamateatris. Ilmunud ka kogumikus : Hellerma, Kärt. Avanenud ruum. Tallinn : Eesti Keele Sihtasutus, 2006, lk. 213-216. Pealk. Narr kui meie aja kangelane

Pärt, Arvo: De profundis / Barry Witherden

Index Scriptorium Estoniae

Witherden, Barry

1997-01-01

Uuest heliplaadist "Pärt, Arvo: De profundis. Missa Sillabica. Solfeggio. And one of the Pharisees. Cantate Domino. Summa. Seven Magnificat Antiphons. The Beautitudes. Magnificat. Christopher Bowers-Broadbent (org.), Daniel Kennedy (perc.). Theatre of Voices / Paul Hillier (bass)". Harmonia Mundi HMU40 7182; HMU90 7182 (76 minutes:DDD)

Cell-type-specific responses of RT4 neural cell lines to dibutyryl-cAMP: branch determination versus maturation

International Nuclear Information System (INIS)

Droms, K.; Sueoka, N.

1987-01-01

This report describes the induction of cell-type-specific maturation, by dibutyryl-cAMP and testololactone, of neuronal and glial properties in a family of cell lines derived from a rat peripheral neurotumor, RT4. This maturation allows further understanding of the process of determination because of the close lineage relationship between the cell types of the RT4 family. The RT4 family is characterized by the spontaneous conversion of one of the cell types, RT4-AC (stem-cell type), to any of three derivative cell types, RT4-B, RT4-D, or RT4-E, with a frequency of about 10(-5). The RT4-AC cells express some properties characteristic of both neuronal and glial cells. Of these neural properties expressed by RT4-AC cells, only the neuronal properties are expressed by the RT4-B and RT4-E cells, and only the glial properties are expressed by the RT4-D cells. This in vitro cell-type conversion of RT4-AC to three derivative cell types is a branch point for the coordinate regulation of several properties and seems to resemble determination in vivo. In our standard culture conditions, several other neuronal and glial properties are not expressed by these cell types. However, addition of dibutyryl-cAMP induces expression of additional properties, in a cell-type-specific manner: formation of long cellular processes in the RT4-B8 and RT4-E5 cell lines and expression of high-affinity uptake of gamma-aminobutyric acid, by a glial-cell-specific mechanism, in the RT4-D6-2 cell line. These new properties are maximally expressed 2-3 days after addition of dibutyryl-cAMP

Aplicación de RT-Linux en el control de motores de pasos. Parte II; Appication of RT-Linux in the Control of Steps Motors. Part II

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Ernesto Duany Renté

2011-02-01

Full Text Available Este trabajo complementa al presentado anteriormente: "Aplicación de RT-Linux en el control de motoresde pasos. Primera parte", de manera que se puedan relacionar a las tareas de adquisición y control para laobtención de un sistema lo más exacto posible. Las técnicas empleadas son las de tiempo real aprovechandolas posibilidades del microkernel RT-Linux y los software libres contenidos en sistemas Unix/Linux. Lasseñales se obtienen mediante un conversor AD y mostradas en pantalla empleando el Gnuplot.  The work presented in this paper is a complement to the control and acquisition tasks which were explainedin "Application of RT-Linux in the Control of Steps Motors. First Part", so that those both real time taskscan be fully related in order to make the whole control system more accurate. The employed techniquesare those of Real Time Taking advantage of the possibilities of the micro kernel RT-Linux and the freesoftware distributed in the Unix/Linux operating systems. The signals are obtained by means of an ADconverter and are shown in screen using Gnuplot.

Short-term memory stores organized by information domain.

Science.gov (United States)

Noyce, Abigail L; Cestero, Nishmar; Shinn-Cunningham, Barbara G; Somers, David C

2016-04-01

Vision and audition have complementary affinities, with vision excelling in spatial resolution and audition excelling in temporal resolution. Here, we investigated the relationships among the visual and auditory modalities and spatial and temporal short-term memory (STM) using change detection tasks. We created short sequences of visual or auditory items, such that each item within a sequence arose at a unique spatial location at a unique time. On each trial, two successive sequences were presented; subjects attended to either space (the sequence of locations) or time (the sequence of inter item intervals) and reported whether the patterns of locations or intervals were identical. Each subject completed blocks of unimodal trials (both sequences presented in the same modality) and crossmodal trials (Sequence 1 visual, Sequence 2 auditory, or vice versa) for both spatial and temporal tasks. We found a strong interaction between modality and task: Spatial performance was best on unimodal visual trials, whereas temporal performance was best on unimodal auditory trials. The order of modalities on crossmodal trials also mattered, suggesting that perceptual fidelity at encoding is critical to STM. Critically, no cost was attributable to crossmodal comparison: In both tasks, performance on crossmodal trials was as good as or better than on the weaker unimodal trials. STM representations of space and time can guide change detection in either the visual or the auditory modality, suggesting that the temporal or spatial organization of STM may supersede sensory-specific organization.

Short-term memory stores organized by information domain

Science.gov (United States)

Noyce, Abigail L.; Cestero, Nishmar; Shinn-Cunningham, Barbara G.; Somers, David C.

2016-01-01

Vision and audition have complementary affinities, with vision excelling in spatial resolution and audition excelling in temporal resolution. Here, we investigate the relationships among visual and auditory modalities and spatial and temporal short-term memory (STM) using change detection tasks. We created short sequences of visual or auditory items, such that each item within a sequence arose at a unique spatial location at a unique time. On each trial, two successive sequences were presented; subjects attended to either space (the sequence of locations), or time (the sequence of inter-item intervals), and reported whether the patterns of locations or intervals were identical. Each subject completed blocks of unimodal trials (both sequences presented in the same modality) and crossmodal trials (sequence 1 visual and sequence 2 auditory, or vice versa) for both spatial and temporal tasks. We found a strong interaction between modality and task: spatial performance was best on unimodal visual trials, while temporal performance was best on unimodal auditory trials. The order of modalities on crossmodal trials also mattered, suggesting that perceptual fidelity at encoding is critical to STM. Critically, there was no cost attributable to crossmodal comparison: in both tasks, performance on crossmodal trials was as good or better than on the weaker unimodal trials. STM representations of space and time can guide change detection in either the visual or the auditory modality, suggesting that temporal or spatial organization of STM may supersede sensory-specific organization. PMID:26791231

RT-PCR-ELISA as a tool for diagnosis of low-pathogenicity avian influenza

DEFF Research Database (Denmark)

Dybkaer, Karen; Munch, Mette; Handberg, Kurt Jensen

2003-01-01

A one-tube reverse transcriptase/polymerase chain reaction coupled with an enzyme-linked immunosorbent assay (RT-PCR-ELISA) was developed for the rapid detection of avian influenza virus (AIV) in clinical specimens. A total of 419 swab pools were analyzed from chickens experimentally infected wit...... of the twenty-three VI-positive specimens were negative when tested by RT-PCR-ELISA. The diagnostic sensitivity and specificity of the RT-PCR-ELISA was 91% and 97%, respectively, using VI in SPF eggs as the gold reference standard....

Probabilistic topic modeling for the analysis and classification of genomic sequences

Science.gov (United States)

2015-01-01

Background Studies on genomic sequences for classification and taxonomic identification have a leading role in the biomedical field and in the analysis of biodiversity. These studies are focusing on the so-called barcode genes, representing a well defined region of the whole genome. Recently, alignment-free techniques are gaining more importance because they are able to overcome the drawbacks of sequence alignment techniques. In this paper a new alignment-free method for DNA sequences clustering and classification is proposed. The method is based on k-mers representation and text mining techniques. Methods The presented method is based on Probabilistic Topic Modeling, a statistical technique originally proposed for text documents. Probabilistic topic models are able to find in a document corpus the topics (recurrent themes) characterizing classes of documents. This technique, applied on DNA sequences representing the documents, exploits the frequency of fixed-length k-mers and builds a generative model for a training group of sequences. This generative model, obtained through the Latent Dirichlet Allocation (LDA) algorithm, is then used to classify a large set of genomic sequences. Results and conclusions We performed classification of over 7000 16S DNA barcode sequences taken from Ribosomal Database Project (RDP) repository, training probabilistic topic models. The proposed method is compared to the RDP tool and Support Vector Machine (SVM) classification algorithm in a extensive set of trials using both complete sequences and short sequence snippets (from 400 bp to 25 bp). Our method reaches very similar results to RDP classifier and SVM for complete sequences. The most interesting results are obtained when short sequence snippets are considered. In these conditions the proposed method outperforms RDP and SVM with ultra short sequences and it exhibits a smooth decrease of performance, at every taxonomic level, when the sequence length is decreased. PMID:25916734

Short segment search method for phylogenetic analysis using nested sliding windows

Science.gov (United States)

Iskandar, A. A.; Bustamam, A.; Trimarsanto, H.

2017-10-01

To analyze phylogenetics in Bioinformatics, coding DNA sequences (CDS) segment is needed for maximal accuracy. However, analysis by CDS cost a lot of time and money, so a short representative segment by CDS, which is envelope protein segment or non-structural 3 (NS3) segment is necessary. After sliding window is implemented, a better short segment than envelope protein segment and NS3 is found. This paper will discuss a mathematical method to analyze sequences using nested sliding window to find a short segment which is representative for the whole genome. The result shows that our method can find a short segment which more representative about 6.57% in topological view to CDS segment than an Envelope segment or NS3 segment.

Fast high-resolution MR imaging using the snapshot-FLASH MR sequence

International Nuclear Information System (INIS)

Matthaei, D.; Haase, A.; Henrich, D.; Duhmke, E.

1990-01-01

Snapshot, fast low-angle short (FLASH) MR imaging using an accelerated FLASH-MR sequence provides MR images with measuring times far below 1 second. The short TE of this sequence prevents susceptibility artifacts in gradient-echo imaging. In this paper variations of the sequence are shown that provide high resolution images with T1-weighted IR, T2-weighted SE, and chemical shift (CHESS) contrast sequences. METHODS AND MATERIALS: A whole-body 2-T system (Bruker-Medizintechnik) were used in combination with a 60-cm gradient system (providing gradient strength of 5 mT/m) to study healthy volunteers. The measuring time for a 256 x 256 image matrix was 800 msec. This sequence has been used in combination with T1-weighted IR, T2-weighted SE, and CHESS variations

Development of single step RT-PCR for detection of Kyasanur forest disease virus from clinical samples

Directory of Open Access Journals (Sweden)

Gouri Chaubal

2018-02-01

Discussion and conclusion: The previously published sensitive real time RT-PCR assay requires higher cost in terms of reagents and machine setup and technical expertise has been the primary reason for development of this assay. A single step RT-PCR is relatively easy to perform and more cost effective than real time RT-PCR in smaller setups in the absence of Biosafety Level-3 facility. This study reports the development and optimization of single step RT-PCR assay which is more sensitive and less time-consuming than nested RT-PCR and cost effective for rapid diagnosis of KFD viral RNA.

Complete Genome Sequence of Diaphorina citri-associated C virus, a Novel Putative RNA Virus of the Asian Citrus Psyllid, Diaphorina citri

OpenAIRE

Nouri, Shahideh; Salem, Nid?; Falk, Bryce W.

2016-01-01

We present here the complete nucleotide sequence and genome organization of a novel putative RNA virus identified in field populations of the Asian citrus psyllid, Diaphorina citri, through sequencing of the transcriptome followed by reverse transcription-PCR (RT-PCR). We tentatively named this virus Diaphorina citri-associated C virus (DcACV). DcACV is an unclassified positive-sense RNA virus.

Pärt, Arvo: Litany. Psalom. Trisagion / Robert Cowan

Index Scriptorium Estoniae

Cowan, Robert

1996-01-01

Uuest heliplaadist "Pärt, Arvo: Litany. Psalom. Trisagion. Hilliard Ensemble, Estonian Philharmonic Chamber Choir, Tallinn Chamber Orchestra, Tõnu Kaljuste; Lithuanian Chamber Orchestra, Saulius Sondeckis" ECM New Series 449 810-4; 449 810-2 (42 minutes: DDD)

Performance of nested RT-PCR on CSF for tuberculous meningitis diagnosis in HIV-infected patients.

Science.gov (United States)

Gualberto, F A S; Gonçalves, M G; Fukasawa, L O; Santos, A M Ramos Dos; Sacchi, C T; Harrison, L H; Boulware, D R; Vidal, J E

2017-10-01

Timely diagnosis of tuberculous meningitis (TBM) in patients with human immunodeficiency virus (HIV) infection remains a challenge. Despite the current scale-up of the Xpert® MTB/RIF assay, other molecular diagnostic tools are necessary, particularly in referral centres in low- and middle-income countries without Xpert testing. To determine the diagnostic performance of nested real-time polymerase chain reaction (nRT-PCR) in HIV-infected TBM patients categorised according to standardised clinical case definitions. Based on clinical, laboratory and imaging data, HIV-infected patients with suspected TBM were prospectively categorised as 'definite TBM', 'probable TBM', 'possible TBM' or 'not TBM'. We evaluated nRT-PCR sensitivity and specificity in diagnosing TBM among definite TBM cases, and among definite + probable TBM cases. Ninety-two participants were enrolled in the study. nRT-PCR sensitivity for definite TBM (n = 8) was 100% (95%CI 67-100) and 86% (95%CI 60-96) for both definite and probable TBM (n = 6). Assuming that 'not TBM' patients (n = 74) were true-negatives, nRT-PCR specificity was 100% (95%CI 95-100). The possible TBM group (n = 4) had no nRT-PCR positives. The nRT-PCR is a useful rule-in test for HIV-infected patients with TBM according to international consensus case definitions. As nRT-PCR cannot exclude TBM, studies comparing and combining nRT-PCR with other assays are necessary for a rule-out test.

A fast and robust method for full genome sequencing of Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) Type 1 and Type 2

DEFF Research Database (Denmark)

Kvisgaard, Lise Kirstine; Hjulsager, Charlotte Kristiane; Fahnøe, Ulrik

2013-01-01

. In the present study, fast and robust methods for long range RT-PCR amplification and subsequent next generation sequencing (NGS) were developed and validated on nine Type 1 and nine Type 2 PRRSV viruses. The methods generated robust and reliable sequences both on primary material and cell culture adapted...... viruses and the protocols performed well on all three NGS platforms tested (Roche 454 FLX, Illumina HiSeq2000, and Ion Torrent PGM™ Sequencer). These methods will greatly facilitate the generation of more full genome PRRSV sequences globally....

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.

SASqPCR: robust and rapid analysis of RT-qPCR data in SAS.

Directory of Open Access Journals (Sweden)

Daijun Ling

Full Text Available Reverse transcription quantitative real-time PCR (RT-qPCR is a key method for measurement of relative gene expression. Analysis of RT-qPCR data requires many iterative computations for data normalization and analytical optimization. Currently no computer program for RT-qPCR data analysis is suitable for analytical optimization and user-controllable customization based on data quality, experimental design as well as specific research aims. Here I introduce an all-in-one computer program, SASqPCR, for robust and rapid analysis of RT-qPCR data in SAS. This program has multiple macros for assessment of PCR efficiencies, validation of reference genes, optimization of data normalizers, normalization of confounding variations across samples, and statistical comparison of target gene expression in parallel samples. Users can simply change the macro variables to test various analytical strategies, optimize results and customize the analytical processes. In addition, it is highly automatic and functionally extendable. Thus users are the actual decision-makers controlling RT-qPCR data analyses. SASqPCR and its tutorial are freely available at http://code.google.com/p/sasqpcr/downloads/list.

Long-term earthquake forecasts based on the epidemic-type aftershock sequence (ETAS model for short-term clustering

Directory of Open Access Journals (Sweden)

Jiancang Zhuang

2012-07-01

Full Text Available Based on the ETAS (epidemic-type aftershock sequence model, which is used for describing the features of short-term clustering of earthquake occurrence, this paper presents some theories and techniques related to evaluating the probability distribution of the maximum magnitude in a given space-time window, where the Gutenberg-Richter law for earthquake magnitude distribution cannot be directly applied. It is seen that the distribution of the maximum magnitude in a given space-time volume is determined in the longterm by the background seismicity rate and the magnitude distribution of the largest events in each earthquake cluster. The techniques introduced were applied to the seismicity in the Japan region in the period from 1926 to 2009. It was found that the regions most likely to have big earthquakes are along the Tohoku (northeastern Japan Arc and the Kuril Arc, both with much higher probabilities than the offshore Nankai and Tokai regions.

Repeat-aware modeling and correction of short read errors.

Science.gov (United States)

Yang, Xiao; Aluru, Srinivas; Dorman, Karin S

2011-02-15

High-throughput short read sequencing is revolutionizing genomics and systems biology research by enabling cost-effective deep coverage sequencing of genomes and transcriptomes. Error detection and correction are crucial to many short read sequencing applications including de novo genome sequencing, genome resequencing, and digital gene expression analysis. Short read error detection is typically carried out by counting the observed frequencies of kmers in reads and validating those with frequencies exceeding a threshold. In case of genomes with high repeat content, an erroneous kmer may be frequently observed if it has few nucleotide differences with valid kmers with multiple occurrences in the genome. Error detection and correction were mostly applied to genomes with low repeat content and this remains a challenging problem for genomes with high repeat content. We develop a statistical model and a computational method for error detection and correction in the presence of genomic repeats. We propose a method to infer genomic frequencies of kmers from their observed frequencies by analyzing the misread relationships among observed kmers. We also propose a method to estimate the threshold useful for validating kmers whose estimated genomic frequency exceeds the threshold. We demonstrate that superior error detection is achieved using these methods. Furthermore, we break away from the common assumption of uniformly distributed errors within a read, and provide a framework to model position-dependent error occurrence frequencies common to many short read platforms. Lastly, we achieve better error correction in genomes with high repeat content. The software is implemented in C++ and is freely available under GNU GPL3 license and Boost Software V1.0 license at "http://aluru-sun.ece.iastate.edu/doku.php?id = redeem". We introduce a statistical framework to model sequencing errors in next-generation reads, which led to promising results in detecting and correcting errors

Deep Sequencing Reveals the Complete Genome and Evidence for Transcriptional Activity of the First Virus-Like Sequences Identified in Aristotelia chilensis (Maqui Berry

Directory of Open Access Journals (Sweden)

Javier Villacreses

2015-04-01

Full Text Available Here, we report the genome sequence and evidence for transcriptional activity of a virus-like element in the native Chilean berry tree Aristotelia chilensis. We propose to name the endogenous sequence as Aristotelia chilensis Virus 1 (AcV1. High-throughput sequencing of the genome of this tree uncovered an endogenous viral element, with a size of 7122 bp, corresponding to the complete genome of AcV1. Its sequence contains three open reading frames (ORFs: ORFs 1 and 2 shares 66%–73% amino acid similarity with members of the Caulimoviridae virus family, especially the Petunia vein clearing virus (PVCV, Petuvirus genus. ORF1 encodes a movement protein (MP; ORF2 a Reverse Transcriptase (RT and a Ribonuclease H (RNase H domain; and ORF3 showed no amino acid sequence similarity with any other known virus proteins. Analogous to other known endogenous pararetrovirus sequences (EPRVs, AcV1 is integrated in the genome of Maqui Berry and showed low viral transcriptional activity, which was detected by deep sequencing technology (DNA and RNA-seq. Phylogenetic analysis of AcV1 and other pararetroviruses revealed a closer resemblance with Petuvirus. Overall, our data suggests that AcV1 could be a new member of Caulimoviridae family, genus Petuvirus, and the first evidence of this kind of virus in a fruit plant.

Simultaneous detection and identification of four cherry viruses by two step multiplex RT-PCR with an internal control of plant nad5 mRNA.

Science.gov (United States)

Noorani, Md Salik; Awasthi, Prachi; Sharma, Maheshwar Prasad; Ram, Raja; Zaidi, Aijaz Asgar; Hallan, Vipin

2013-10-01

A multiplex reverse transcription-polymerase chain reaction (mRT-PCR) was developed and standardized for the simultaneous detection of four cherry viruses: Cherry virus A (CVA, Genus; Capillovirus), Cherry necrotic rusty mottle virus (CNRMV, unassigned species of the Betaflexiviridae), Little cherry virus 1 (LChV-1, Genus; Closterovirus) and Prunus necrotic ringspot virus (PNRSV, Genus; Ilarvirus) with nad5 as plant internal control. A reliable and quick method for total plant RNA extraction from pome and stone fruit trees was also developed. To minimize primer dimer formation, a single antisense primer for CVA and CNRMV was used. A mixture of random hexamer and oligo (dT) primer was used for cDNA synthesis, which was highly suited and economic for multiplexing. All four viruses were detected successfully by mRT-PCR in artificially created viral RNA mixture and field samples of sweet cherry. The identity of the viruses was confirmed by sequencing. The assay could detect above viruses in diluted cDNA (10(-4)) and RNA (10(-3), except PNRSV which was detected only till ten times lesser dilution). The developed mRT-PCR will not only be useful for the detection of viruses from single or multiple infections of sweet cherry plants but also for other stone and pome fruits. The developed method will be therefore quite helpful for virus indexing, plant quarantine and certification programs. This is the first report for the simultaneous detection of four cherry viruses by mRT-PCR. Copyright © 2013 Elsevier B.V. All rights reserved.

Characterization of Erwinia amylovora strains from different host plants using repetitive-sequences PCR analysis, and restriction fragment length polymorphism and short-sequence DNA repeats of plasmid pEA29.

Science.gov (United States)

Barionovi, D; Giorgi, S; Stoeger, A R; Ruppitsch, W; Scortichini, M

2006-05-01

The three main aims of the study were the assessment of the genetic relationship between a deviating Erwinia amylovora strain isolated from Amelanchier sp. (Maloideae) grown in Canada and other strains from Maloideae and Rosoideae, the investigation of the variability of the PstI fragment of the pEA29 plasmid using restriction fragment length polymorphism (RFLP) analysis and the determination of the number of short-sequence DNA repeats (SSR) by DNA sequence analysis in representative strains. Ninety-three strains obtained from 12 plant genera and different geographical locations were examined by repetitive-sequences PCR using Enterobacterial Repetitive Intergenic Consensus, BOX and Repetitive Extragenic Palindromic primer sets. Upon the unweighted pair group method with arithmetic mean analysis, a deviating strain from Amelanchier sp. was analysed using amplified ribosomal DNA restriction analysis (ARDRA) analysis and the sequencing of the 16S rDNA gene. This strain showed 99% similarity to other E. amylovora strains in the 16S gene and the same banding pattern with ARDRA. The RFLP analysis of pEA29 plasmid using MspI and Sau3A restriction enzymes showed a higher variability than that previously observed and no clear-cut grouping of the strains was possible. The number of SSR units reiterated two to 12 times. The strains obtained from pear orchards showing for the first time symptoms of fire blight had a low number of SSR units. The strains from Maloideae exhibit a wider genetic variability than previously thought. The RFLP analysis of a fragment of the pEA29 plasmid would not seem a reliable method for typing E. amylovora strains. A low number of SSR units was observed with first epidemics of fire blight. The current detection techniques are mainly based on the genetic similarities observed within the strains from the cultivated tree-fruit crops. For a more reliable detection of the fire blight pathogen also in wild and ornamentals Rosaceous plants the genetic

Karje kunstniku südamest / Kärt Hellerma

Index Scriptorium Estoniae

Hellerma, Kärt, 1956-

1997-01-01

Arvustus: Dinesen, Isak [Blixen, Karen]. Babette'i pidusöök / tlk. Piret Peiker. Tallinn : Varrak, 1996. Ilmunud ka kogumikus: Hellerma, Kärt. Kohanenud kirjandus : valik kirjanduskriitikat 1987-2006. Eesti Keele Sihtasutus : Tallinn, 2006. Lk. 167-169

NASA/SPoRt: GOES-R Activities in Support of Product Development, Management, and Training

Science.gov (United States)

Fuell, Kevin; Jedlovec, Gary; Molthan, Andrew; Stano, Geoffrey

2012-01-01

SPoRT is using current capabilities of MODIS and VIIRS, combined with current GOES (i.e. Hybrid Imagery) to demonstrate mesoscale capabilities of future ABI instrument. SPoRT is transitioning RGBs from EUMETSAT standard "recipes" to demonstrate a method to more efficiently handle the increase channels/frequency of ABI. Challenges for RGB production exist. Internal vs. external production, Bit depth needed, Adding quantitative information, etc. SPoRT forming group to address these issues. SPoRT is leading efforts on the application of total lightning in operations and to educate users of this new capability. Training in many forms is used to support testbed activities and is a key part to the transition process.

Short Note DNA sequences from the Little Brown Bustard Eupodotis ...

African Journals Online (AJOL)

Taxonomic classification of birds based exclusively on morphology and plumage traits has often been found to be inconsistent with true evolutionary history when tested with molecular phylogenies based on neutrally evolving markers. Here we present cytochrome-b gene sequences for the poorly known Little Brown ...

Quantitative RT-PCR for titration of replication-defective recombinant Semliki Forest virus.

Science.gov (United States)

Puglia, Ana L P; Rezende, Alexandre G; Jorge, Soraia A C; Wagner, Renaud; Pereira, Carlos A; Astray, Renato M

2013-11-01

Virus titration may constitute a drawback in the development and use of replication-defective viral vectors like Semliki Forest virus (SFV). The standardization and validation of a reverse transcription quantitative PCR (qRT-PCR) method for SFV titration is presented here. The qRT-PCR target is located within the nsp1 gene of the non-structural polyprotein SFV region (SFV RNA), which allows the strategy to be used for several different recombinant SFV constructs. Titer determinations were carried out by performing virus titration and infection assays with SFVs containing an RNA coding region for the rabies virus glycoprotein (RVGP) or green fluorescent protein (GFP). Results showed that the standardized qRT-PCR is applicable for different SFV constructs, and showed good reproducibility. To evaluate the correlation between the amount of functional SFV RNA in a virus lot and its infectivity in BHK-21 cell cultures, a temperature mediated titer decrease was performed and successfully quantitated by qRT-PCR. When used for cell infection at the same multiplicity of infection (MOI), the temperature treated SFV-RVGP samples induced the same levels of RVGP expression. Similarly, when different SFV-GFP lots with different virus titers, as accessed by qRT-PCR, were used for cell infection at the same MOI, the cultures showed comparable amounts of fluorescent cells. The data demonstrate a good correlation between the amount of virus used for infection, as measured by its SFV RNA, and the protein synthesis in the cells. In conclusion, the qRT-PCR method developed here is accurate and enables the titration of replication-defective SFV vectors, an essential aid for viral vector development as well as for establishment of production bioprocesses. Copyright © 2013 Elsevier B.V. All rights reserved.

Study of rt-PA therapy for acute stroke in older patients

International Nuclear Information System (INIS)

Ono, Yasuhiro; Toyoshima, Atsuhiko; Toyota, Yasunori; Kuramoto, Satoshi; Katsumata, Atsushi; Kawauchi, Masamitsu; Matsumoto, Yuzo

2011-01-01

Intravenous alteplase (rt-PA) therapy has been widely approved for acute ischemic stroke. We examined symptomatic intracranial hemorrhage (SICH) and outcomes in older patients treated with rt-PA therapy. We divided 130 consecutive patients treated with rt-PA into two groups: 72 patients younger than 80 years (age <80 group) and 58 patients older than 80 years (age ≥80 group). On CT and MRI scans, SICH was observed in 5 patients (7.1%) in the age <80 group and 4 patients (6.9%) in the age ≥80 group. The SICH rate did not differ significantly between the two groups (odds ratio (OR) 1.21, 95% confidence interval (CI) 0.04-2.39, P=0.6174). The patients in the age ≥80 group had a significantly lower mRS 0-1 rate (8.8 vs. 30.0%; OR 7.20, 95% CI 6.05-8.34, P=0.0009) and significantly higher mRS 6 (mortality) rate (19.3 vs. 9.0%; OR 6.52, 95% CI 5.39-7.65, P=0.0015) compared with those in the age <80 group. These data suggest that the factor of age is not related to SICH but is related to patients' outcomes in rt-PA therapy. (author)

GOKaRT: Graphical Online Search Tool for Maps

Directory of Open Access Journals (Sweden)

Mechthild Schüler

2008-09-01

Full Text Available The map department of the Staats- und Universitätsbibliothek Göttingen together with the Berlin State Library propose a project to develop a web-based graphic cataloguing and search system for maps, to be funded by the German Research Foundation. This tool shall be made available to all map holdings in archives, libraries, university departments and museums in Germany as a comfortable means for the administration of map holdings and as a search tool. Sheets belonging to map series as well as single maps (old and new will be registered cooperatively by the participants with simple tools. This cooperation in data maintenance will facilitate the work especially for understaffed map holdings. Depending on the type of map there are four different mechanisms for map reference. For map series electronic index sheets are used which will show information regarding the various issues of the map sheets. Due to the intuitive graphic search entry GOKaRT-users will easily find the required maps of a certain region available in a chosen holding. User administration modules ensure comfortable handling. GOKaRT is being developed on the basis of licence-free open source programmes. In case financing is provided by the German Research Foundation, GOKaRT can be used free of charge internationally. This would require a contract stipulating data exchange between the partners as well as permanent storage and usability of the data.

Data acquisition and real-time bolometer tomography using LabVIEW RT

International Nuclear Information System (INIS)

Giannone, L.; Eich, T.; Fuchs, J.C.; Ravindran, M.; Ruan, Q.; Wenzel, L.; Cerna, M.; Concezzi, S.

2011-01-01

The currently available multi-core PCI Express systems running LabVIEW RT (real-time), equipped with FPGA cards for data acquisition and real-time parallel signal processing, greatly shorten the design and implementation cycles of large-scale, real-time data acquisition and control systems. This paper details a data acquisition and real-time tomography system using LabVIEW RT for the bolometer diagnostic on the ASDEX Upgrade tokamak (Max Planck Institute for Plasma Physics, Garching, Germany). The transformation matrix for tomography is pre-computed based on the geometry of distributed radiation sources and sensors. A parallelized iterative algorithm is adapted to solve a constrained linear system for the reconstruction of the radiated power density. Real-time bolometer tomography is performed with LabVIEW RT. Using multi-core machines to execute the parallelized algorithm, a cycle time well below 1 ms is reached.

DaRT: A CALL System to Help Students Practice and Develop Reasoning Skills in Choosing English Articles.

Science.gov (United States)

Yoshii, Rika; Milne, Alastair

1998-01-01

Describes DaRT, a computer assisted language-learning system for helping English-as-a-Second-Language students master English articles. DaRT uses a diagrammatic reasoning tool to present communicative contexts for exercises in choosing appropriate articles. This paper describes the development of DaRT and DaRT's system components and concludes…

Short echo time, fast gradient-echo imaging

International Nuclear Information System (INIS)

Haacke, E.M.; Lenz, G.W.

1987-01-01

Present fast-gradient-echoes schemes can acquire volume data rapidly and are flexible in T1 or T1/T2 contrast behavior. However, sequences used to date employ echo time (TE) values of about 15 ms +- 5 and, because of in vivo field inhomogeneities (short T2), they suffer badly from signal loss near sinuses and tissue boundaries. The authors implemented sequences with TE = 4-6 ms and found significant improvement in image quality, especially at high fields. Examples with long TEs vs. short TEs are given in the knee, spine, head, and orbits. Further advantages include (1) faster repetition times (15 ms), (2) higher-quality spin-density or T1-weighted images, and (3) reduction of blood motion artifacts

Recent evolutionary history of human immunodeficiency virus type 1 subtype B: Reconstruction of epidemic onset based on sequence distances to the common ancestor

NARCIS (Netherlands)

Lukashov, Vladimir V.; Goudsmit, Jaap

2002-01-01

We obtained and studied HIV-1 sequences with a known sampling year from three outbreaks of the HIV-1 epidemic: 141 env V3 (270 nt) sampled between 1984 and 1992 and 117 pol prot/RT (804 nt) sequences sampled between 1986 and 1999 from Dutch homosexual men and injecting drug users (IDUs), as well as

Thermal regulation 2000: one year later; RT 2000: un an apres

Energy Technology Data Exchange (ETDEWEB)

NONE

2002-05-01

This meeting, organized by the research center of Gaz de France (Cegibat), aims at drawing up a progress status of the French thermal regulation (RT 2000), one year after its enforcement on June 1, 2000: status, experience feedback and recommendations; Gaz de France assistance and solutions; tools at the disposition of designers (labelling of equipments, simulation softwares..); practical examples of implementation of the RT 2000 in office buildings, residential buildings and sport complexes. (J.S.)

L. Harrison: Mass (to St. Anthony); A. Pärt: Berliner Messe / Robert Cowan

Index Scriptorium Estoniae

Cowan, Robert

1994-01-01

Uuest heliplaadist "L. Harrison: Mass (to St. Anthony); A. Pärt: Berliner Messe. Oregon Repertory Singers / Gilbert Seeley. Koch International Classics CD 37 177-2; Pärt - comparative version: Estonian Phil. Chbr. Ch., Tallinn CO / Kaljuste" (11/93)(ECM) 439 162-2

Investigating the Decision-Making of Response to Intervention (RtI) Teams within the School Setting

Science.gov (United States)

Thur, Scott M.

2015-01-01

The purpose of this study was to measure decision-making influences within RtI teams. The study examined the factors that influence school personnel involved in three areas of RtI: determining which RtI measures and tools teams select and implement (i.e. Measures and Tools), evaluating the data-driven decisions that are made based on the…

Recent Upgrades to NASA SPoRT Initialization Datasets for the Environmental Modeling System

Science.gov (United States)

Case, Jonathan L.; LaFontaine, Frank J.; Molthan, Andrew L.; Zavodsky, Bradley T.; Rozumalski, Robert A.

2012-01-01

The NASA Short-term Prediction Research and Transition (SPoRT) Center has developed several products for its National Weather Service (NWS) partners that can initialize specific fields for local model runs within the NOAA/NWS Science and Training Resource Center (STRC) Environmental Modeling System (EMS). In last year's NWA abstract on this topic, the suite of SPoRT products supported in the STRC EMS was presented, which includes a Sea Surface Temperature (SST) composite, a Great Lakes sea-ice extent, a Green Vegetation Fraction (GVF) composite, and NASA Land Information System (LIS) gridded output. This abstract and companion presentation describes recent upgrades made to the SST and GVF composites, as well as the real-time LIS runs. The Great Lakes sea-ice product is unchanged from 2011. The SPoRT SST composite product has been expanded geographically and as a result, the resolution has been coarsened from 1 km to 2 km to accommodate the larger domain. The expanded domain covers much of the northern hemisphere from eastern Asia to western Europe (0 N to 80 N latitude and 150 E to 10 E longitude). In addition, the NESDIS POES-GOES product was added to fill in gaps caused by the Moderate Resolution Imaging Spectroradiometer (MODIS) being unable to sense in cloudy regions, replacing the recently-lost Advanced Microwave Scanning Radiometer for EOS with negligible change to product fidelity. The SST product now runs twice per day for Terra and Aqua combined data collections from 0000 to 1200 UTC and from 1200 to 0000 UTC, with valid analysis times at 0600 and 1800 UTC. The twice-daily compositing technique reduces the overall latency of the previous version while still representing the diurnal cycle characteristics. The SST composites are available at approximately four hours after the end of each collection period (i.e. 1600 UTC for the nighttime analysis and 0400 UTC for the daytime analysis). The real-time MODIS GVF composite has only received minor updates in the

WE-H-207B-04: Strategies for Adaptive RT

Energy Technology Data Exchange (ETDEWEB)

Green, O. [Washington University School of Medicine (United States)

2016-06-15

In recent years, steady progress has been made towards the implementation of MRI in external beam radiation therapy for processes ranging from treatment simulation to in-room guidance. Novel procedures relying mostly on MR data are currently implemented in the clinic. This session will cover topics such as (a) commissioning and quality control of the MR in-room imagers and simulators specific to RT, (b) treatment planning requirements, constraints and challenges when dealing with various MR data, (c) quantification of organ motion with an emphasis on treatment delivery guidance, and (d) MR-driven strategies for adaptive RT workflows. The content of the session was chosen to address both educational and practical key aspects of MR guidance. Learning Objectives: Good understanding of MR testing recommended for in-room MR imaging as well as image data validation for RT chain (e.g. image transfer, filtering for consistency, spatial accuracy, manipulation for task specific); Familiarity with MR-based planning procedures: motivation, core workflow requirements, current status, challenges; Overview of the current methods for the quantification of organ motion; Discussion on approaches for adaptive treatment planning and delivery. T. Stanescu - License agreement with Modus Medical Devices to develop a phantom for the quantification of MR image system-related distortions.; T. Stanescu, N/A.

WE-H-207B-02: MR-Driven RT Planning

Energy Technology Data Exchange (ETDEWEB)

Cao, M. [UCLA School of Medicine (United States)

2016-06-15

In recent years, steady progress has been made towards the implementation of MRI in external beam radiation therapy for processes ranging from treatment simulation to in-room guidance. Novel procedures relying mostly on MR data are currently implemented in the clinic. This session will cover topics such as (a) commissioning and quality control of the MR in-room imagers and simulators specific to RT, (b) treatment planning requirements, constraints and challenges when dealing with various MR data, (c) quantification of organ motion with an emphasis on treatment delivery guidance, and (d) MR-driven strategies for adaptive RT workflows. The content of the session was chosen to address both educational and practical key aspects of MR guidance. Learning Objectives: Good understanding of MR testing recommended for in-room MR imaging as well as image data validation for RT chain (e.g. image transfer, filtering for consistency, spatial accuracy, manipulation for task specific); Familiarity with MR-based planning procedures: motivation, core workflow requirements, current status, challenges; Overview of the current methods for the quantification of organ motion; Discussion on approaches for adaptive treatment planning and delivery. T. Stanescu - License agreement with Modus Medical Devices to develop a phantom for the quantification of MR image system-related distortions.; T. Stanescu, N/A.

WE-H-207B-04: Strategies for Adaptive RT

International Nuclear Information System (INIS)

Green, O.

2016-01-01

In recent years, steady progress has been made towards the implementation of MRI in external beam radiation therapy for processes ranging from treatment simulation to in-room guidance. Novel procedures relying mostly on MR data are currently implemented in the clinic. This session will cover topics such as (a) commissioning and quality control of the MR in-room imagers and simulators specific to RT, (b) treatment planning requirements, constraints and challenges when dealing with various MR data, (c) quantification of organ motion with an emphasis on treatment delivery guidance, and (d) MR-driven strategies for adaptive RT workflows. The content of the session was chosen to address both educational and practical key aspects of MR guidance. Learning Objectives: Good understanding of MR testing recommended for in-room MR imaging as well as image data validation for RT chain (e.g. image transfer, filtering for consistency, spatial accuracy, manipulation for task specific); Familiarity with MR-based planning procedures: motivation, core workflow requirements, current status, challenges; Overview of the current methods for the quantification of organ motion; Discussion on approaches for adaptive treatment planning and delivery. T. Stanescu - License agreement with Modus Medical Devices to develop a phantom for the quantification of MR image system-related distortions.; T. Stanescu, N/A

WE-H-207B-02: MR-Driven RT Planning

International Nuclear Information System (INIS)

Cao, M.

2016-01-01

In recent years, steady progress has been made towards the implementation of MRI in external beam radiation therapy for processes ranging from treatment simulation to in-room guidance. Novel procedures relying mostly on MR data are currently implemented in the clinic. This session will cover topics such as (a) commissioning and quality control of the MR in-room imagers and simulators specific to RT, (b) treatment planning requirements, constraints and challenges when dealing with various MR data, (c) quantification of organ motion with an emphasis on treatment delivery guidance, and (d) MR-driven strategies for adaptive RT workflows. The content of the session was chosen to address both educational and practical key aspects of MR guidance. Learning Objectives: Good understanding of MR testing recommended for in-room MR imaging as well as image data validation for RT chain (e.g. image transfer, filtering for consistency, spatial accuracy, manipulation for task specific); Familiarity with MR-based planning procedures: motivation, core workflow requirements, current status, challenges; Overview of the current methods for the quantification of organ motion; Discussion on approaches for adaptive treatment planning and delivery. T. Stanescu - License agreement with Modus Medical Devices to develop a phantom for the quantification of MR image system-related distortions.; T. Stanescu, N/A

Simultaneous detection of enteropathogenic viruses in buffalos faeces using multiplex reverse transcription-polymerase chain reaction (mRT-PCR

Directory of Open Access Journals (Sweden)

U. Pagnini

2010-02-01

Full Text Available A multiplex reverse transcription- polymerase chain reaction (mRT-PCR assay that detects Bovine Viral Diarrhoea Virus, Bovine Coronavirus, and Group A Rotaviruses in infected cell-culture fluids and clinical faecal samples is described. One hundred twenty faecal samples from buffalo calves with acute gastroenteritis were tested. The mRT-PCR was validated against simplex RT-PCR with published primers for Pestivirus, Coronavirus and Rotavirus. The multiplex RT-PCR was equally sensitive and specific in detecting viral infections compared with simplex RT-PCR. The mRT-PCR readily identified viruses by discriminating the size of their amplified gene products. This mRT-PCR may be a sensitive and rapid assay for surveillance of buffalo enteric viruses in field specimens. This novel multiplex RT-PCR is an attractive technique for the rapid, specific, and cost-effective laboratory diagnosis of acute gastroenteritis.

SU-F-J-219: Predicting Ventilation Change Due to Radiation Therapy: Dependency On Pre-RT Ventilation and Effort Correction

Energy Technology Data Exchange (ETDEWEB)

Patton, T; Du, K; Bayouth, J [University of Wisconsin, Madison, WI (United States); Christensen, G; Reinhardt, J [University of Iowa, Iowa City, IA (United States)

2016-06-15

Purpose: Ventilation change caused by radiation therapy (RT) can be predicted using four-dimensional computed tomography (4DCT) and image registration. This study tested the dependency of predicted post-RT ventilation on effort correction and pre-RT lung function. Methods: Pre-RT and 3 month post-RT 4DCT images were obtained for 13 patients. The 4DCT images were used to create ventilation maps using a deformable image registration based Jacobian expansion calculation. The post-RT ventilation maps were predicted in four different ways using the dose delivered, pre-RT ventilation, and effort correction. The pre-RT ventilation and effort correction were toggled to determine dependency. The four different predicted ventilation maps were compared to the post-RT ventilation map calculated from image registration to establish the best prediction method. Gamma pass rates were used to compare the different maps with the criteria of 2mm distance-to-agreement and 6% ventilation difference. Paired t-tests of gamma pass rates were used to determine significant differences between the maps. Additional gamma pass rates were calculated using only voxels receiving over 20 Gy. Results: The predicted post-RT ventilation maps were in agreement with the actual post-RT maps in the following percentage of voxels averaged over all subjects: 71% with pre-RT ventilation and effort correction, 69% with no pre-RT ventilation and effort correction, 60% with pre-RT ventilation and no effort correction, and 58% with no pre-RT ventilation and no effort correction. When analyzing only voxels receiving over 20 Gy, the gamma pass rates were respectively 74%, 69%, 65%, and 55%. The prediction including both pre- RT ventilation and effort correction was the only prediction with significant improvement over using no prediction (p<0.02). Conclusion: Post-RT ventilation is best predicted using both pre-RT ventilation and effort correction. This is the only prediction that provided a significant

Identification of optimum sequencing depth especially for de novo genome assembly of small genomes using next generation sequencing data.

Science.gov (United States)

Desai, Aarti; Marwah, Veer Singh; Yadav, Akshay; Jha, Vineet; Dhaygude, Kishor; Bangar, Ujwala; Kulkarni, Vivek; Jere, Abhay

2013-01-01

Next Generation Sequencing (NGS) is a disruptive technology that has found widespread acceptance in the life sciences research community. The high throughput and low cost of sequencing has encouraged researchers to undertake ambitious genomic projects, especially in de novo genome sequencing. Currently, NGS systems generate sequence data as short reads and de novo genome assembly using these short reads is computationally very intensive. Due to lower cost of sequencing and higher throughput, NGS systems now provide the ability to sequence genomes at high depth. However, currently no report is available highlighting the impact of high sequence depth on genome assembly using real data sets and multiple assembly algorithms. Recently, some studies have evaluated the impact of sequence coverage, error rate and average read length on genome assembly using multiple assembly algorithms, however, these evaluations were performed using simulated datasets. One limitation of using simulated datasets is that variables such as error rates, read length and coverage which are known to impact genome assembly are carefully controlled. Hence, this study was undertaken to identify the minimum depth of sequencing required for de novo assembly for different sized genomes using graph based assembly algorithms and real datasets. Illumina reads for E.coli (4.6 MB) S.kudriavzevii (11.18 MB) and C.elegans (100 MB) were assembled using SOAPdenovo, Velvet, ABySS, Meraculous and IDBA-UD. Our analysis shows that 50X is the optimum read depth for assembling these genomes using all assemblers except Meraculous which requires 100X read depth. Moreover, our analysis shows that de novo assembly from 50X read data requires only 6-40 GB RAM depending on the genome size and assembly algorithm used. We believe that this information can be extremely valuable for researchers in designing experiments and multiplexing which will enable optimum utilization of sequencing as well as analysis resources.

Kaks vaadet Ameerikast Jacques Derridale / Märt Väljataga

Index Scriptorium Estoniae

Väljataga, Märt, 1965-

1998-01-01

Kommentaariks Vikerkaares 1998, nr. 7-8 ilmunud artiklitele: Mark Lilla 'Jacques Derrida poliitika' ja Richard Rorty 'Ironistlikust teooriast privaatsete allusioonideni' (mõlemad tõlkinud Märt Väljataga)

Long-read sequencing and de novo assembly of a Chinese genome

Science.gov (United States)

Short-read sequencing has enabled the de novo assembly of several individual human genomes, but with inherent limitations in characterizing repeat elements. Here we sequence a Chinese individual HX1 by single-molecule real-time (SMRT) long-read sequencing, construct a physical map by NanoChannel arr...

Complete Genome Sequence of Diaphorina citri-associated C virus, a Novel Putative RNA Virus of the Asian Citrus Psyllid, Diaphorina citri.

Science.gov (United States)

Nouri, Shahideh; Salem, Nidà; Falk, Bryce W

2016-07-21

We present here the complete nucleotide sequence and genome organization of a novel putative RNA virus identified in field populations of the Asian citrus psyllid, Diaphorina citri, through sequencing of the transcriptome followed by reverse transcription-PCR (RT-PCR). We tentatively named this virus Diaphorina citri-associated C virus (DcACV). DcACV is an unclassified positive-sense RNA virus. Copyright © 2016 Nouri et al.

Effects of aging and dopamine genotypes on the emergence of explicit memory during sequence learning.

Science.gov (United States)

Schuck, Nicolas W; Frensch, Peter A; Schjeide, Brit-Maren M; Schröder, Julia; Bertram, Lars; Li, Shu-Chen

2013-11-01

The striatum and medial temporal lobe play important roles in implicit and explicit memory, respectively. Furthermore, recent studies have linked striatal dopamine modulation to both implicit as well as explicit sequence learning and suggested a potential role of the striatum in the emergence of explicit memory during sequence learning. With respect to aging, previous findings indicated that implicit memory is less impaired than explicit memory in older adults and that genetic effects on cognition are magnified by aging. To understand the links between these findings, we investigated effects of aging and genotypes relevant for striatal dopamine on the implicit and explicit components of sequence learning. Reaction time (RT) and error data from 80 younger (20-30 years) and 70 older adults (60-71 years) during a serial reaction time task showed that age differences in learning-related reduction of RTs emerged gradually over the course of learning. Verbal recall and measures derived from the process-dissociation procedure revealed that younger adults acquired more explicit memory about the sequence than older adults, potentially causing age differences in RT gains in later stages of learning. Of specific interest, polymorphisms of the dopamine- and cAMP-regulated neuronal phosphoprotein (DARPP-32, rs907094) and dopamine transporter (DAT, VNTR) genes showed interactive effects on overall RTs and verbal recall of the sequence in older but not in younger adults. Together our findings show that variations in genotypes relevant for dopamine functions are associated more with aging-related impairments in the explicit than the implicit component of sequence learning, providing support for theories emphasizing the role of dopaminergic modulation in cognitive aging and the magnification of genetic effects in human aging. © 2013 Elsevier Ltd. All rights reserved.

Resolving the Complexity of Human Skin Metagenomes Using Single-Molecule Sequencing

Directory of Open Access Journals (Sweden)

Yu-Chih Tsai

2016-02-01

Full Text Available Deep metagenomic shotgun sequencing has emerged as a powerful tool to interrogate composition and function of complex microbial communities. Computational approaches to assemble genome fragments have been demonstrated to be an effective tool for de novo reconstruction of genomes from these communities. However, the resultant “genomes” are typically fragmented and incomplete due to the limited ability of short-read sequence data to assemble complex or low-coverage regions. Here, we use single-molecule, real-time (SMRT sequencing to reconstruct a high-quality, closed genome of a previously uncharacterized Corynebacterium simulans and its companion bacteriophage from a skin metagenomic sample. Considerable improvement in assembly quality occurs in hybrid approaches incorporating short-read data, with even relatively small amounts of long-read data being sufficient to improve metagenome reconstruction. Using short-read data to evaluate strain variation of this C. simulans in its skin community at single-nucleotide resolution, we observed a dominant C. simulans strain with moderate allelic heterozygosity throughout the population. We demonstrate the utility of SMRT sequencing and hybrid approaches in metagenome quantitation, reconstruction, and annotation.

Resolving the Complexity of Human Skin Metagenomes Using Single-Molecule Sequencing

Science.gov (United States)

Tsai, Yu-Chih; Deming, Clayton; Segre, Julia A.; Kong, Heidi H.; Korlach, Jonas

2016-01-01

ABSTRACT Deep metagenomic shotgun sequencing has emerged as a powerful tool to interrogate composition and function of complex microbial communities. Computational approaches to assemble genome fragments have been demonstrated to be an effective tool for de novo reconstruction of genomes from these communities. However, the resultant “genomes” are typically fragmented and incomplete due to the limited ability of short-read sequence data to assemble complex or low-coverage regions. Here, we use single-molecule, real-time (SMRT) sequencing to reconstruct a high-quality, closed genome of a previously uncharacterized Corynebacterium simulans and its companion bacteriophage from a skin metagenomic sample. Considerable improvement in assembly quality occurs in hybrid approaches incorporating short-read data, with even relatively small amounts of long-read data being sufficient to improve metagenome reconstruction. Using short-read data to evaluate strain variation of this C. simulans in its skin community at single-nucleotide resolution, we observed a dominant C. simulans strain with moderate allelic heterozygosity throughout the population. We demonstrate the utility of SMRT sequencing and hybrid approaches in metagenome quantitation, reconstruction, and annotation. PMID:26861018

One-step multiplex real-time RT-PCR assay for detecting and genotyping wild-type group A rotavirus strains and vaccine strains (Rotarix® and RotaTeq®) in stool samples

Science.gov (United States)

Mijatovic-Rustempasic, Slavica; Esona, Mathew D.; Tam, Ka Ian; Quaye, Osbourne; Bowen, Michael D.

2016-01-01

Background. Group A rotavirus (RVA) infection is the major cause of acute gastroenteritis (AGE) in young children worldwide. Introduction of two live-attenuated rotavirus vaccines, RotaTeq® and Rotarix®, has dramatically reduced RVA associated AGE and mortality in developed as well as in many developing countries. High-throughput methods are needed to genotype rotavirus wild-type strains and to identify vaccine strains in stool samples. Quantitative RT-PCR assays (qRT-PCR) offer several advantages including increased sensitivity, higher throughput, and faster turnaround time. Methods. In this study, a one-step multiplex qRT-PCR assay was developed to detect and genotype wild-type strains and vaccine (Rotarix® and RotaTeq®) rotavirus strains along with an internal processing control (Xeno or MS2 RNA). Real-time RT-PCR assays were designed for VP7 (G1, G2, G3, G4, G9, G12) and VP4 (P[4], P[6] and P[8]) genotypes. The multiplex qRT-PCR assay also included previously published NSP3 qRT-PCR for rotavirus detection and Rotarix® NSP2 and RotaTeq® VP6 qRT-PCRs for detection of Rotarix® and RotaTeq® vaccine strains respectively. The multiplex qRT-PCR assay was validated using 853 sequence confirmed stool samples and 24 lab cultured strains of different rotavirus genotypes. By using thermostable rTth polymerase enzyme, dsRNA denaturation, reverse transcription (RT) and amplification (PCR) steps were performed in single tube by uninterrupted thermocycling profile to reduce chances of sample cross contamination and for rapid generation of results. For quantification, standard curves were generated using dsRNA transcripts derived from RVA gene segments. Results. The VP7 qRT-PCRs exhibited 98.8–100% sensitivity, 99.7–100% specificity, 85–95% efficiency and a limit of detection of 4–60 copies per singleplex reaction. The VP7 qRT-PCRs exhibited 81–92% efficiency and limit of detection of 150–600 copies in multiplex reactions. The VP4 qRT-PCRs exhibited 98.8�

A voxel-based investigation for MRI-only radiotherapy of the brain using ultra short echo times

Science.gov (United States)

Edmund, Jens M.; Kjer, Hans M.; Van Leemput, Koen; Hansen, Rasmus H.; Andersen, Jon AL; Andreasen, Daniel

2014-12-01

Radiotherapy (RT) based on magnetic resonance imaging (MRI) as the only modality, so-called MRI-only RT, would remove the systematic registration error between MR and computed tomography (CT), and provide co-registered MRI for assessment of treatment response and adaptive RT. Electron densities, however, need to be assigned to the MRI images for dose calculation and patient setup based on digitally reconstructed radiographs (DRRs). Here, we investigate the geometric and dosimetric performance for a number of popular voxel-based methods to generate a so-called pseudo CT (pCT). Five patients receiving cranial irradiation, each containing a co-registered MRI and CT scan, were included. An ultra short echo time MRI sequence for bone visualization was used. Six methods were investigated for three popular types of voxel-based approaches; (1) threshold-based segmentation, (2) Bayesian segmentation and (3) statistical regression. Each approach contained two methods. Approach 1 used bulk density assignment of MRI voxels into air, soft tissue and bone based on logical masks and the transverse relaxation time T2 of the bone. Approach 2 used similar bulk density assignments with Bayesian statistics including or excluding additional spatial information. Approach 3 used a statistical regression correlating MRI voxels with their corresponding CT voxels. A similar photon and proton treatment plan was generated for a target positioned between the nasal cavity and the brainstem for all patients. The CT agreement with the pCT of each method was quantified and compared with the other methods geometrically and dosimetrically using both a number of reported metrics and introducing some novel metrics. The best geometrical agreement with CT was obtained with the statistical regression methods which performed significantly better than the threshold and Bayesian segmentation methods (excluding spatial information). All methods agreed significantly better with CT than a reference water MRI

Coval: improving alignment quality and variant calling accuracy for next-generation sequencing data.

Directory of Open Access Journals (Sweden)

Shunichi Kosugi

Full Text Available Accurate identification of DNA polymorphisms using next-generation sequencing technology is challenging because of a high rate of sequencing error and incorrect mapping of reads to reference genomes. Currently available short read aligners and DNA variant callers suffer from these problems. We developed the Coval software to improve the quality of short read alignments. Coval is designed to minimize the incidence of spurious alignment of short reads, by filtering mismatched reads that remained in alignments after local realignment and error correction of mismatched reads. The error correction is executed based on the base quality and allele frequency at the non-reference positions for an individual or pooled sample. We demonstrated the utility of Coval by applying it to simulated genomes and experimentally obtained short-read data of rice, nematode, and mouse. Moreover, we found an unexpectedly large number of incorrectly mapped reads in 'targeted' alignments, where the whole genome sequencing reads had been aligned to a local genomic segment, and showed that Coval effectively eliminated such spurious alignments. We conclude that Coval significantly improves the quality of short-read sequence alignments, thereby increasing the calling accuracy of currently available tools for SNP and indel identification. Coval is available at http://sourceforge.net/projects/coval105/.

Real-time RT-PCR, a necessary tool to support the diagnosis and surveillance of rotavirus in Mexico.

Science.gov (United States)

De La Cruz Hernández, Sergio Isaac; Anaya Molina, Yazmin; Gómez Santiago, Fabián; Terán Vega, Heidi Lizbeth; Monroy Leyva, Elda; Méndez Pérez, Héctor; García Lozano, Herlinda

2018-04-01

Rotavirus produces diarrhea in children under 5 years old. Most of those conventional methods such as polyacrylamide gel electrophoresis (PAGE) and reverse transcription-polymerase chain reaction (RT-PCR) have been used for rotavirus detection. However, these techniques need a multi-step process to get the results. In comparison with conventional methods, the real-time RT-PCR is a highly sensitive method, which allows getting the results in only one day. In this study a real-time RT-PCR assay was tested using a panel of 440 samples from patients with acute gastroenteritis, and characterized by PAGE and RT-PCR. The results show that the real-time RT-PCR detected rotavirus from 73% of rotavirus-negative samples analyzed by PAGE and RT-PCR; thus, the percentage of rotavirus-positive samples increased to 81%. The results indicate that this real-time RT-PCR should be part of a routine analysis, and as a support of the diagnosis of rotavirus in Mexico. Copyright © 2017 Elsevier Inc. All rights reserved.

Parallel motif extraction from very long sequences

KAUST Repository

Sahli, Majed

2013-01-01

Motifs are frequent patterns used to identify biological functionality in genomic sequences, periodicity in time series, or user trends in web logs. In contrast to a lot of existing work that focuses on collections of many short sequences, modern applications require mining of motifs in one very long sequence (i.e., in the order of several gigabytes). For this case, there exist statistical approaches that are fast but inaccurate; or combinatorial methods that are sound and complete. Unfortunately, existing combinatorial methods are serial and very slow. Consequently, they are limited to very short sequences (i.e., a few megabytes), small alphabets (typically 4 symbols for DNA sequences), and restricted types of motifs. This paper presents ACME, a combinatorial method for extracting motifs from a single very long sequence. ACME arranges the search space in contiguous blocks that take advantage of the cache hierarchy in modern architectures, and achieves almost an order of magnitude performance gain in serial execution. It also decomposes the search space in a smart way that allows scalability to thousands of processors with more than 90% speedup. ACME is the only method that: (i) scales to gigabyte-long sequences; (ii) handles large alphabets; (iii) supports interesting types of motifs with minimal additional cost; and (iv) is optimized for a variety of architectures such as multi-core systems, clusters in the cloud, and supercomputers. ACME reduces the extraction time for an exact-length query from 4 hours to 7 minutes on a typical workstation; handles 3 orders of magnitude longer sequences; and scales up to 16, 384 cores on a supercomputer. Copyright is held by the owner/author(s).

A MapReduce Framework for DNA Sequencing Data Processing

Directory of Open Access Journals (Sweden)

Samy Ghoneimy

2016-12-01

Full Text Available Genomics and Next Generation Sequencers (NGS like Illumina Hiseq produce data in the order of ‎‎200 billion base pairs in a single one-week run for a 60x human genome coverage, which ‎requires modern high-throughput experimental technologies that can ‎only be tackled with high performance computing (HPC and specialized software algorithms called ‎‎“short read aligners”. This paper focuses on the implementation of the DNA sequencing as a set of MapReduce programs that will accept a DNA data set as a FASTQ file and finally generate a VCF (variant call format file, which has variants for a given DNA data set. In this paper MapReduce/Hadoop along with Burrows-Wheeler Aligner (BWA, Sequence Alignment/Map (SAM ‎tools, are fully utilized to provide various utilities for manipulating alignments, including sorting, merging, indexing, ‎and generating alignments. The Map-Sort-Reduce process is designed to be suited for a Hadoop framework in ‎which each cluster is a traditional N-node Hadoop cluster to utilize all of the Hadoop features like HDFS, program ‎management and fault tolerance. The Map step performs multiple instances of the short read alignment algorithm ‎‎(BoWTie that run in parallel in Hadoop. The ordered list of the sequence reads are used as input tuples and the ‎output tuples are the alignments of the short reads. In the Reduce step many parallel instances of the Short ‎Oligonucleotide Analysis Package for SNP (SOAPsnp algorithm run in the cluster. Input tuples are sorted ‎alignments for a partition and the output tuples are SNP calls. Results are stored via HDFS, and then archived in ‎SOAPsnp format. ‎ The proposed framework enables extremely fast discovering somatic mutations, inferring population genetical ‎parameters, and performing association tests directly based on sequencing data without explicit genotyping or ‎linkage-based imputation. It also demonstrate that this method achieves comparable

Differentiating Visual from Response Sequencing during Long-term Skill Learning.

Science.gov (United States)

Lynch, Brighid; Beukema, Patrick; Verstynen, Timothy

2017-01-01

The dual-system model of sequence learning posits that during early learning there is an advantage for encoding sequences in sensory frames; however, it remains unclear whether this advantage extends to long-term consolidation. Using the serial RT task, we set out to distinguish the dynamics of learning sequential orders of visual cues from learning sequential responses. On each day, most participants learned a new mapping between a set of symbolic cues and responses made with one of four fingers, after which they were exposed to trial blocks of either randomly ordered cues or deterministic ordered cues (12-item sequence). Participants were randomly assigned to one of four groups (n = 15 per group): Visual sequences (same sequence of visual cues across training days), Response sequences (same order of key presses across training days), Combined (same serial order of cues and responses on all training days), and a Control group (a novel sequence each training day). Across 5 days of training, sequence-specific measures of response speed and accuracy improved faster in the Visual group than any of the other three groups, despite no group differences in explicit awareness of the sequence. The two groups that were exposed to the same visual sequence across days showed a marginal improvement in response binding that was not found in the other groups. These results indicate that there is an advantage, in terms of rate of consolidation across multiple days of training, for learning sequences of actions in a sensory representational space, rather than as motoric representations.

Complete Genome Sequence of the Largest Known Flavi-Like Virus, Diaphorina citri flavi-like virus, a Novel Virus of the Asian Citrus Psyllid, Diaphorina citri

OpenAIRE

Matsumura, Emilyn E.; Nerva, Luca; Nigg, Jared C.; Falk, Bryce W.; Nouri, Shahideh

2016-01-01

A novel flavi-like virus tentatively named Diaphorina citri flavi-like virus (DcFLV) was identified in field populations of Diaphorina citri through small RNA and transcriptome sequencing followed by reverse transcription (RT)-PCR. We report here the complete nucleotide sequence and genome organization of DcFLV, the largest flavi-like virus identified to date.

Distinct electrophysiological indices of maintenance in auditory and visual short-term memory.

Science.gov (United States)

Lefebvre, Christine; Vachon, François; Grimault, Stephan; Thibault, Jennifer; Guimond, Synthia; Peretz, Isabelle; Zatorre, Robert J; Jolicœur, Pierre

2013-11-01

We compared the electrophysiological correlates for the maintenance of non-musical tones sequences in auditory short-term memory (ASTM) to those for the short-term maintenance of sequences of coloured disks held in visual short-term memory (VSTM). The visual stimuli yielded a sustained posterior contralateral negativity (SPCN), suggesting that the maintenance of sequences of coloured stimuli engaged structures similar to those involved in the maintenance of simultaneous visual displays. On the other hand, maintenance of acoustic sequences produced a sustained negativity at fronto-central sites. This component is named the Sustained Anterior Negativity (SAN). The amplitude of the SAN increased with increasing load in ASTM and predicted individual differences in the performance. There was no SAN in a control condition with the same auditory stimuli but no memory task, nor one associated with visual memory. These results suggest that the SAN is an index of brain activity related to the maintenance of representations in ASTM that is distinct from the maintenance of representations in VSTM. © 2013 Elsevier Ltd. All rights reserved.

Endurance training intensity does not mediate interference to maximal lower-body strength gain during short-term concurrent training.

Directory of Open Access Journals (Sweden)

Jackson J Fyfe

2016-11-01

-matched MICT similarly attenuates improvements in maximal lower-body strength and indices of CMJ performance compared with RT performed alone. This suggests endurance training intensity is not a critical mediator of interference to maximal strength gain during short-term concurrent training.

JVM: Java Visual Mapping tool for next generation sequencing read.

Science.gov (United States)

Yang, Ye; Liu, Juan

2015-01-01

We developed a program JVM (Java Visual Mapping) for mapping next generation sequencing read to reference sequence. The program is implemented in Java and is designed to deal with millions of short read generated by sequence alignment using the Illumina sequencing technology. It employs seed index strategy and octal encoding operations for sequence alignments. JVM is useful for DNA-Seq, RNA-Seq when dealing with single-end resequencing. JVM is a desktop application, which supports reads capacity from 1 MB to 10 GB.

Similarity as an organising principle in short-term memory.

Science.gov (United States)

LeCompte, D C; Watkins, M J

1993-03-01

The role of stimulus similarity as an organising principle in short-term memory was explored in a series of seven experiments. Each experiment involved the presentation of a short sequence of items that were drawn from two distinct physical classes and arranged such that item class changed after every second item. Following presentation, one item was re-presented as a probe for the 'target' item that had directly followed it in the sequence. Memory for the sequence was considered organised by class if probability of recall was higher when the probe and target were from the same class than when they were from different classes. Such organisation was found when one class was auditory and the other was visual (spoken vs. written words, and sounds vs. pictures). It was also found when both classes were auditory (words spoken in a male voice vs. words spoken in a female voice) and when both classes were visual (digits shown in one location vs. digits shown in another). It is concluded that short-term memory can be organised on the basis of sensory modality and on the basis of certain features within both the auditory and visual modalities.

Phase I study of neoadjuvant accelerated short course radiation therapy with photons and capecitabine for resectable pancreatic cancer

International Nuclear Information System (INIS)

Wo, Jennifer Y.; Mamon, Harvey J.; Ferrone, Cristina R.; Ryan, David P.; Blaszkowsky, Lawrence S.; Kwak, Eunice L.; Tseng, Yolanda D.; Napolitano, Brian N.; Ancukiewicz, Marek; Swanson, Richard S.; Lillemoe, Keith D.; Castillo, Carlos Fernandez-del; Hong, Theodore S.

2014-01-01

Purpose: In this phase I study, we sought to determine the feasibility and tolerability of neoadjuvant short course radiotherapy (SC-CRT) delivered with photon RT with concurrent capecitabine for resectable pancreatic adenocarcinoma. Materials and methods: Ten patients with localized, resectable pancreatic adenocarcinoma were enrolled from December 2009 to August 2011. In dose level I, patients received 3 Gy × 10. In dose level 2, patients received 5 Gy × 5 (every other day). In dose level 3, patients received 5 Gy × 5 (consecutive days). Capecitabine was given during weeks 1 and 2. Surgery was performed 1–3 weeks after completion of chemotherapy. Results: With an intended accrual of 12 patients, the study was closed early due to unexpected intraoperative complications. Compared to the companion phase I proton study, patients treated with photons had increased intraoperative RT fibrosis reported by surgeons (27% vs. 63%). Among those undergoing a Whipple resection, increased RT fibrosis translated to an increased mean OR time of 69 min. Dosimetric comparison revealed significantly increased low dose exposure to organs at risk for patients treated with photon RT. Conclusions: This phase I experience evaluating the tolerability of neoadjuvant SC-CRT with photon RT closed early due to unexpected intraoperative complications

Feasibility of small animal cranial irradiation with the microRT system

International Nuclear Information System (INIS)

Kiehl, Erich L.; Stojadinovic, Strahinja; Malinowski, Kathleen T.; Limbrick, David; Jost, Sarah C.; Garbow, Joel R.; Rubin, Joshua B.; Deasy, Joseph O.; Khullar, Divya; Izaguirre, Enrique W.; Parikh, Parag J.; Low, Daniel A.; Hope, Andrew J.

2008-01-01

Purpose: To develop and validate methods for small-animal CNS radiotherapy using the microRT system. Materials and Methods: A custom head immobilizer was designed and built to integrate with a pre-existing microRT animal couch. The Delrin couch-immobilizer assembly, compatible with multiple imaging modalities (CT, microCT, microMR, microPET, microSPECT, optical), was first imaged via CT in order to verify the safety and reproducibility of the immobilization method. Once verified, the subject animals were CT-scanned while positioned within the couch-immobilizer assembly for treatment planning purposes. The resultant images were then imported into CERR, an in-house-developed research treatment planning system, and registered to the microRTP treatment planning space using rigid registration. The targeted brain was then contoured and conformal radiotherapy plans were constructed for two separate studies: (1) a whole-brain irradiation comprised of two lateral beams at the 90 degree sign and 270 degree sign microRT treatment positions and (2) a hemispheric (left-brain) irradiation comprised of a single A-P vertex beam at the 0 degree sign microRT treatment position. During treatment, subject animals (n=48) were positioned to the CERR-generated treatment coordinates using the three-axis microRT motor positioning system and were irradiated using a clinical Ir-192 high-dose-rate remote after-loading system. The radiation treatment course consisted of 5 Gy fractions, 3 days per week. 90% of the subjects received a total dose of 30 Gy and 10% received a dose of 60 Gy. Results: Image analysis verified the safety and reproducibility of the immobilizer. CT scans generated from repeated reloading and repositioning of the same subject animal in the couch-immobilizer assembly were fused to a baseline CT. The resultant analysis revealed a 0.09 mm average, center-of-mass translocation and negligible volumetric error in the contoured, murine brain. The experimental use of the head

A RADIATION TRANSFER SOLVER FOR ATHENA USING SHORT CHARACTERISTICS

International Nuclear Information System (INIS)

Davis, Shane W.; Stone, James M.; Jiang Yanfei

2012-01-01

We describe the implementation of a module for the Athena magnetohydrodynamics (MHD) code that solves the time-independent, multi-frequency radiative transfer (RT) equation on multidimensional Cartesian simulation domains, including scattering and non-local thermodynamic equilibrium (LTE) effects. The module is based on well known and well tested algorithms developed for modeling stellar atmospheres, including the method of short characteristics to solve the RT equation, accelerated Lambda iteration to handle scattering and non-LTE effects, and parallelization via domain decomposition. The module serves several purposes: it can be used to generate spectra and images, to compute a variable Eddington tensor (VET) for full radiation MHD simulations, and to calculate the heating and cooling source terms in the MHD equations in flows where radiation pressure is small compared with gas pressure. For the latter case, the module is combined with the standard MHD integrators using operator splitting: we describe this approach in detail, including a new constraint on the time step for stability due to radiation diffusion modes. Implementation of the VET method for radiation pressure dominated flows is described in a companion paper. We present results from a suite of test problems for both the RT solver itself and for dynamical problems that include radiative heating and cooling. These tests demonstrate that the radiative transfer solution is accurate and confirm that the operator split method is stable, convergent, and efficient for problems of interest. We demonstrate there is no need to adopt ad hoc assumptions of questionable accuracy to solve RT problems in concert with MHD: the computational cost for our general-purpose module for simple (e.g., LTE gray) problems can be comparable to or less than a single time step of Athena's MHD integrators, and only few times more expensive than that for more general (non-LTE) problems.

Association Between RT-Induced Changes in Lung Tissue Density and Global Lung Function

International Nuclear Information System (INIS)

Ma Jinli; Zhang Junan; Zhou Sumin; Hubbs, Jessica L.; Foltz, Rodney J.; Hollis, Donna R.; Light, Kim L.; Wong, Terence Z.; Kelsey, Christopher R.; Marks, Lawrence B.

2009-01-01

Purpose: To assess the association between radiotherapy (RT)-induced changes in computed tomography (CT)-defined lung tissue density and pulmonary function tests (PFTs). Methods and Materials: Patients undergoing incidental partial lung RT were prospectively assessed for global (PFTs) and regional (CT and single photon emission CT [SPECT]) lung function before and, serially, after RT. The percent reductions in the PFT and the average changes in lung density were compared (Pearson correlations) in the overall group and subgroups stratified according to various clinical factors. Comparisons were also made between the CT- and SPECT-based computations using the Mann-Whitney U test. Results: Between 1991 and 2004, 343 patients were enrolled in this study. Of these, 111 patients had a total of 203 concurrent post-RT evaluations of changes in lung density and PFTs available for the analyses, and 81 patients had a total of 141 concurrent post-RT SPECT images. The average increases in lung density were related to the percent reductions in the PFTs, albeit with modest correlation coefficients (range, 0.20-0.43). The analyses also indicated that the association between lung density and PFT changes is essentially equivalent to the corresponding association with SPECT-defined lung perfusion. Conclusion: We found a weak quantitative association between the degree of increase in lung density as defined by CT and the percent reduction in the PFTs.

SU-E-J-42: Customized Deformable Image Registration Using Open-Source Software SlicerRT

Energy Technology Data Exchange (ETDEWEB)

Gaitan, J Cifuentes; Chin, L; Pignol, J [Sunnybrook Health Sciences Centre, Toronto, Ontario (Canada); Kirby, N; Pouliot, J [UC San Francisco, San Francisco, CA (United States); Lasso, A; Pinter, C; Fichtinger, G [Queen' s University, Kingston, Ontario (Canada)

2014-06-01

Purpose: SlicerRT is a flexible platform that allows the user to incorporate the necessary images registration and processing tools to improve clinical workflow. This work validates the accuracy and the versatility of the deformable image registration algorithm of the free open-source software SlicerRT using a deformable physical pelvic phantom versus available commercial image fusion algorithms. Methods: Optical camera images of nonradiopaque markers implanted in an anatomical pelvic phantom were used to measure the ground-truth deformation and evaluate the theoretical deformations for several DIR algorithms. To perform the registration, full and empty bladder computed tomography (CT) images of the phantom were obtained and used as fixed and moving images, respectively. The DIR module, found in SlicerRT, used a B-spline deformable image registration with multiple optimization parameters that allowed customization of the registration including a regularization term that controlled the amount of local voxel displacement. The virtual deformation field at the center of the phantom was obtained and compared to the experimental ground-truth values. The parameters of SlicerRT were then varied to improve spatial accuracy. To quantify image similarity, the mean absolute difference (MAD) parameter using Hounsfield units was calculated. In addition, the Dice coefficient of the contoured rectum was evaluated to validate the strength of the algorithm to transfer anatomical contours. Results: Overall, SlicerRT achieved one of the lowest MAD values across the algorithm spectrum, but slightly smaller mean spatial errors in comparison to MIM software (MIM). On the other hand, SlicerRT created higher mean spatial errors than Velocity Medical Solutions (VEL), although obtaining an improvement on the DICE to 0.91. The large spatial errors were attributed to the poor contrast in the prostate bladder interface of the phantom. Conclusion: Based phantom validation, SlicerRT is capable of