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Sample records for based rna extraction

  1. Analysis of current and alternative phenol based RNA extraction methodologies for cyanobacteria

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    Lindblad Peter

    2009-08-01

    Full Text Available Abstract Background The validity and reproducibility of gene expression studies depend on the quality of extracted RNA and the degree of genomic DNA contamination. Cyanobacteria are gram-negative prokaryotes that synthesize chlorophyll a and carry out photosynthetic water oxidation. These organisms possess an extended array of secondary metabolites that impair cell lysis, presenting particular challenges when it comes to nucleic acid isolation. Therefore, we used the NHM5 strain of Nostoc punctiforme ATCC 29133 to compare and improve existing phenol based chemistry and procedures for RNA extraction. Results With this work we identify and explore strategies for improved and lower cost high quality RNA isolation from cyanobacteria. All the methods studied are suitable for RNA isolation and its use for downstream applications. We analyse different Trizol based protocols, introduce procedural changes and describe an alternative RNA extraction solution. Conclusion It was possible to improve purity of isolated RNA by modifying protocol procedures. Further improvements, both in RNA purity and experimental cost, were achieved by using a new extraction solution, PGTX.

  2. Novel extraction strategy of ribosomal RNA and genomic DNA from cheese for PCR-based investigations.

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    Bonaïti, Catherine; Parayre, Sandrine; Irlinger, Françoise

    2006-03-15

    Cheese microorganisms, such as bacteria and fungi, constitute a complex ecosystem that plays a central role in cheeses ripening. The molecular study of cheese microbial diversity and activity is essential but the extraction of high quality nucleic acid may be problematic: the cheese samples are characterised by a strong buffering capacity which negatively influenced the yield of the extracted rRNA. The objective of this study is to develop an effective method for the direct and simultaneous isolation of yeast and bacterial ribosomal RNA and genomic DNA from the same cheese samples. DNA isolation was based on a protocol used for nucleic acids isolation from anaerobic digestor, without preliminary washing step with the combined use of the action of chaotropic agent (acid guanidinium thiocyanate), detergents (SDS, N-lauroylsarcosine), chelating agent (EDTA) and a mechanical method (bead beating system). The DNA purification was carried out by two washing steps of phenol-chloroform. RNA was isolated successfully after the second acid extraction step by recovering it from the phenolic phase of the first acid extraction. The novel method yielded pure preparation of undegraded RNA accessible for reverse transcription-PCR. The extraction protocol of genomic DNA and rRNA was applicable to complex ecosystem of different cheese matrices.

  3. The use of carrier RNA to enhance DNA extraction from microfluidic-based silica monoliths.

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    Shaw, Kirsty J; Thain, Lauren; Docker, Peter T; Dyer, Charlotte E; Greenman, John; Greenway, Gillian M; Haswell, Stephen J

    2009-10-12

    DNA extraction was carried out on silica-based monoliths within a microfluidic device. Solid-phase DNA extraction methodology was applied in which the DNA binds to silica in the presence of a chaotropic salt, such as guanidine hydrochloride, and is eluted in a low ionic strength solution, such as water. The addition of poly-A carrier RNA to the chaotropic salt solution resulted in a marked increase in the effective amount of DNA that could be recovered (25ng) compared to the absence of RNA (5ng) using the silica-based monolith. These findings confirm that techniques utilising nucleic acid carrier molecules can enhance DNA extraction methodologies in microfluidic applications.

  4. Protocol: A simple phenol-based method for 96-well extraction of high quality RNA from Arabidopsis

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    Coustham Vincent

    2011-03-01

    Full Text Available Abstract Background Many experiments in modern plant molecular biology require the processing of large numbers of samples for a variety of applications from mutant screens to the analysis of natural variants. A severe bottleneck to many such analyses is the acquisition of good yields of high quality RNA suitable for use in sensitive downstream applications such as real time quantitative reverse-transcription-polymerase chain reaction (real time qRT-PCR. Although several commercial kits are available for high-throughput RNA extraction in 96-well format, only one non-kit method has been described in the literature using the commercial reagent TRIZOL. Results We describe an unusual phenomenon when using TRIZOL reagent with young Arabidopsis seedlings. This prompted us to develop a high-throughput RNA extraction protocol (HTP96 adapted from a well established phenol:chloroform-LiCl method (P:C-L that is cheap, reliable and requires no specialist equipment. With this protocol 192 high quality RNA samples can be prepared in 96-well format in three hours (less than 1 minute per sample with less than 1% loss of samples. We demonstrate that the RNA derived from this protocol is of high quality and suitable for use in real time qRT-PCR assays. Conclusion The development of the HTP96 protocol has vastly increased our sample throughput, allowing us to fully exploit the large sample capacity of modern real time qRT-PCR thermocyclers, now commonplace in many labs, and develop an effective high-throughput gene expression platform. We propose that the HTP96 protocol will significantly benefit any plant scientist with the task of obtaining hundreds of high quality RNA extractions.

  5. A rapid silica spin column-based method of RNA extraction from fruit trees for RT-PCR detection of viruses.

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    Yang, Fan; Wang, Guoping; Xu, Wenxing; Hong, Ni

    2017-09-01

    Efficient recovery of high quality RNA is very important for successful RT-PCR detection of plant RNA viruses. High levels of polyphenols and polysaccharides in plant tissues can irreversibly bind to and/or co-precipitate with RNA, which influences RNA isolation. In this study, a silica spin column-based RNA isolation method was developed by using commercially available silica columns combined with the application of a tissue lysis solution, and binding and washing buffers with high concentration guanidinium thiocyanate (GuSCN, 50% w/v), which helps remove plant proteins, polysaccharides and polyphenolic compounds. The method was successfully used to extract high quality RNA from citrus (Citrus aurantifolia), grapevine (Vitis vinifera), peach (Prunus persica), pear (Pyrus spp.), taro (Colocosia esculenta) and tobacco (Nicotiana benthamiana) samples. The method was comparable to conventional CTAB method in RNA isolation efficiency, but it was more sample-adaptable and cost-effective than commercial kits. High quality RNA isolated using silica spin column-based method was successfully used for the RT-PCR and/or multiplex RT-PCR amplification of woody fruit tree viruses and a viroid. The study provided a useful tool for the detection and characterization of plant viruses. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Graphene Oxide Conjugated Magnetic Beads for RNA Extraction.

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    Pham, Xuan-Hung; Baek, Ahruem; Kim, Tae Han; Lee, Sang Hun; Rho, Won-Yeop; Chung, Woo-Jae; Kim, Dong-Eun; Jun, Bong-Hyun

    2017-08-04

    A magnetic material that consists of silica-coated magnetic beads conjugated with graphene oxide (GO) was successfully prepared for facile ribonucleic acid (RNA) extraction. When the GO-modified magnetic beads were applied to separate the RNA from the lysed cell, the cellular RNAs were readily adsorbed to and readily desorbed from the surface of the GO-modified magnetic beads by urea. The amount of RNA extracted by the GO-modified magnetic beads was ≈170 % as much as those of the control extracted by a conventional phenol-based chaotropic solution. These results demonstrate that the facile method of RNA separation by using GO-modified magnetic beads as an adsorbent is an efficient and simple way to purify intact cellular RNAs and/or microRNA from cell lysates. © 2017 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. Parallel RNA extraction using magnetic beads and a droplet array.

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    Shi, Xu; Chen, Chun-Hong; Gao, Weimin; Chao, Shih-Hui; Meldrum, Deirdre R

    2015-02-21

    Nucleic acid extraction is a necessary step for most genomic/transcriptomic analyses, but it often requires complicated mechanisms to be integrated into a lab-on-a-chip device. Here, we present a simple, effective configuration for rapidly obtaining purified RNA from low concentration cell medium. This Total RNA Extraction Droplet Array (TREDA) utilizes an array of surface-adhering droplets to facilitate the transportation of magnetic purification beads seamlessly through individual buffer solutions without solid structures. The fabrication of TREDA chips is rapid and does not require a microfabrication facility or expertise. The process takes less than 5 minutes. When purifying mRNA from bulk marine diatom samples, its repeatability and extraction efficiency are comparable to conventional tube-based operations. We demonstrate that TREDA can extract the total mRNA of about 10 marine diatom cells, indicating that the sensitivity of TREDA approaches single-digit cell numbers.

  8. Extraction of total nucleic acid based on silica-coated magnetic particles for RT-qPCR detection of plant RNA virus/viroid.

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    Sun, Ning; Deng, Congliang; Zhao, Xiaoli; Zhou, Qi; Ge, Guanglu; Liu, Yi; Yan, Wenlong; Xia, Qiang

    2014-02-01

    In this study, a nucleic acid extraction method based on silica-coated magnetic particles (SMPs) and RT-qPCR assay was developed to detect Arabis mosaic virus (ArMV), Lily symptomless virus (LSV), Hop stunt viroid (HSVd) and grape yellow speckle viroid 1 (GYSVd-1). The amplification sequences of RT-qPCR were reversely transcribed in vitro as RNA standard templates. The standard curves covered six or seven orders of magnitude with a detection limit of 100 copies per each assay. Extraction efficiency of the SMPs method was evaluated by recovering spiked ssRNAs from plant samples and compared to two commercial kits (TRIzol and RNeasy Plant mini kit). Results showed that the recovery rate of SMPs method was comparable to the commercial kits when spiked ssRNAs were extracted from lily leaves, whereas it was two or three times higher than commercial kits when spiked ssRNAs were extracted from grapevine leaves. SMPs method was also used to extract viral nucleic acid from15 ArMV-positive lily leaf samples and 15 LSV-positive lily leaf samples. SMPs method did not show statistically significant difference from other methods on detecting ArMV, but LSV. The SMPs method has the same level of virus load as the TRIzol, and its mean virus load of was 0.5log10 lower than the RNeasy Plant mini kit. Nucleic acid was extracted from 19 grapevine-leaf samples with SMPs and the two commercial kits and subsequently screened for HSVd and GYSVd-1 by RT-qPCR. Regardless of HSVd or GYSVd-1, SMPs method outperforms other methods on both positive rate and the viroid load. In conclusion, SMPs method was able to efficiently extract the nucleic acid of RNA viruses or viroids, especially grapevine viroids, from lily-leaf or grapevine-leaf samples for RT-qPCR detection. Copyright © 2013 Elsevier B.V. All rights reserved.

  9. Optimization of RNA Extraction from Rat Pancreatic Tissue

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    Sanaz Dastgheib

    2014-05-01

    Full Text Available Background: Optimized RNA extraction from tissues and cell lines consists of four main stages regardless of the method of extraction: 1 homogenizing, 2 effective denaturation of proteins from RNA, 3 inactivation of ribonuclease, and 4 removal of any DNA, protein, and carbohydrate contamination. Isolation of undamaged intact RNA is challenging when the related tissue contains high levels of RNase. Various technical difficulties occur during extraction of RNA from pancreatic tissue due to spontaneous autolysis. Since standard routine protocols yield unacceptable results in pancrease, we have designed a simple method for RNA extraction by comparing different protocols. Methods: We obtained 20-30 mg pancreatic tissues in less than 2 min from 30 rats. Several methods were performed to extract RNA from pancreatic tissue and evaluate its integrity. All methods were performed three times to obtain reproducible results. Results: Immersing pancreatic tissue in RNA-later for 24 h at -80ºC yielded high quality RNA by using the TriPure reagent which was comparable to the commercial RNeasy Micro Kit. The quality of RNA was evaluated by spectrophotometer, electrophoresis and RT-PCR. We separated intact 28S and 18S ribosomal RNA (rRNA when our procedure was compared with the RNeasy Micro Kit. Finally, full length of the actin gene was amplified by RT-PCR. Conclusion: We designed a simple, fast, cost-effective method for complete RNA extraction from the least amount of quantitatively intact pancreatic tissue

  10. A Simple and Efficient RNA Extraction Method from Deep-Sea Hydrothermal Vent Chimney Structures.

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    Muto, Hisashi; Takaki, Yoshihiro; Hirai, Miho; Mino, Sayaka; Sawayama, Shigeki; Takai, Ken; Nakagawa, Satoshi

    2017-12-27

    RNA-based microbiological analyses, e.g., transcriptome and reverse transcription-quantitative PCR, require a relatively large amount of high quality RNA. RNA-based analyses on microbial communities in deep-sea hydrothermal environments often encounter methodological difficulties with RNA extraction due to the presence of unique minerals in and the low biomass of samples. In the present study, we assessed RNA extraction methods for deep-sea vent chimneys that had complex mineral compositions. Mineral-RNA adsorption experiments were conducted using mock chimney minerals and Escherichia coli total RNA solution, and showed that detectable RNA significantly decreased possibly due to adsorption onto minerals. This decrease in RNA was prevented by the addition of sodium tripolyphosphate (STPP), deoxynucleotide triphosphates (dNTPs), salmon sperm DNA, and NaOH. The addition of STPP was also effective for RNA extraction from the mixture of E. coli cells and mock chimney minerals when TRIzol reagent and the RNeasy column were used, but not when the RNeasy PowerSoil total RNA kit was used. A combination of STPP, TRIzol reagent, the RNeasy column, and sonication resulted in the highest RNA yield from a natural chimney. This indirect extraction procedure is simple, rapid, inexpensive, and may be used for large-scale RNA extraction.

  11. Extraction of low molecular weight RNA from Citrus trifolita tissues ...

    African Journals Online (AJOL)

    We employed a simple and quick method involving trizol for total RNA extraction from citrus tissues, then generation of LMW RNA using 4M LiCl, which have been successfully utilized in studies in our laboratory. Compared with traditional methods, this method is less expensive and produced high RNA yields while avoiding ...

  12. Urinary extracellular vesicles for RNA extraction: optimization of a protocol devoid of prokaryote contamination.

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    Tataruch-Weinert, Dorota; Musante, Luca; Kretz, Oliver; Holthofer, Harry

    2016-01-01

    Urinary extracellular vesicles (UEVs) represent an ideal platform for biomarker discovery. They carry different types of RNA species, and reported profile discrepancies related to the presence/absence of 18s and 28s rRNA remain controversial. Moreover, sufficient urinary RNA yields and respective quality RNA profiles are still to be fully established. UEVs were enriched by hydrostatic filtration dialysis, and RNA content was extracted using 7 different commercially available techniques. RNA quantity was assessed using spectrophotometry and fluorometry, whilst RNA quality was determined by capillary electrophoresis. The presence of prokaryotic transcriptome was stressed when cellular RNA, as a control, was spiked into the UEVs samples before RNA extraction. The presence of bacteria in hydrostatic filtration dialysis above 1,000 kDa molecular weight cut-off and in crude urine was confirmed with growth media plates. The efficiency in removing urinary bacteria was evaluated by differential centrifugation, filtration (0.22 µm filters) and chemical pretreatment (water purification tablet). For volumes of urine >200 ml, the chemical treatment provides ease of handling without affecting vesicle integrity, protein and RNA profiles. This protocol was selected to enrich RNA with 7 methods, and its respective quality and quantity were assessed. The results were given as follows: (a) Fluorometry gave more repeatability and reproducibility than spectrophotometry to assess the RNA yields, (b) UEVs were enriched with small RNA, (c) Ribosomal RNA peaks were not observed for any RNA extraction method used and (d) RNA yield was higher for column-based method designed for urinary exosome, whilst the highest relative microRNA presence was obtained using TRIzol method. Our results show that the presence of bacteria can lead to misidentification in the electrophoresis peaks. Fluorometry is more reliable than spectrophotometry. RNA isolation method must be selected in conjunction with

  13. Comparison of RNA extraction methods from biofilm samples of Staphylococcus epidermidis

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    França Angela

    2011-12-01

    Full Text Available Abstract Background Microbial biofilms are communities of bacteria adhered to a surface and surrounded by an extracellular polymeric matrix. Biofilms have been associated with increased antibiotic resistance and tolerance to the immune system. Staphylococcus epidermidis is the major bacterial species found in biofilm-related infections on indwelling medical devices. Obtaining high quality mRNA from biofilms is crucial to validate the transcriptional measurements associated with the switching to the biofilm mode of growth. Therefore, we selected three commercially available RNA extraction kits with distinct characteristics, including those using silica membrane or organic extraction methods, and enzymatic or mechanical cell lysis, and evaluated the RNA quality obtained from two distinct S. epidermidis bacterial biofilms. Results RNA extracted using the different kits was evaluated for quantity, purity, integrity, and functionally. All kits were able to extract intact and functional total RNA from the biofilms generated from each S. epidermidis strain. The results demonstrated that the kit based on mechanical lysis and organic extraction (FastRNA® Pro Blue was the only one that was able to isolate pure and large quantities of RNA. Normalized expression of the icaA virulence gene showed that RNA extracted with PureLink™ had a significant lower concentration of icaA mRNA transcripts than the other kits tested. Conclusions When working with complex samples, such as biofilms, that contain a high content extracellular polysaccharide and proteins, special care should be taken when selecting the appropriate RNA extraction system, in order to obtain accurate, reproducible, and biologically significant results. Among the RNA extraction kits tested, FastRNA® Pro Blue was the best option for both S. epidermidis biofilms used.

  14. Fragment-based modelling of single stranded RNA bound to RNA recognition motif containing proteins

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    de Beauchene, Isaure Chauvot; de Vries, Sjoerd J.; Zacharias, Martin

    2016-01-01

    Abstract Protein-RNA complexes are important for many biological processes. However, structural modeling of such complexes is hampered by the high flexibility of RNA. Particularly challenging is the docking of single-stranded RNA (ssRNA). We have developed a fragment-based approach to model the structure of ssRNA bound to a protein, based on only the protein structure, the RNA sequence and conserved contacts. The conformational diversity of each RNA fragment is sampled by an exhaustive library of trinucleotides extracted from all known experimental protein–RNA complexes. The method was applied to ssRNA with up to 12 nucleotides which bind to dimers of the RNA recognition motifs (RRMs), a highly abundant eukaryotic RNA-binding domain. The fragment based docking allows a precise de novo atomic modeling of protein-bound ssRNA chains. On a benchmark of seven experimental ssRNA–RRM complexes, near-native models (with a mean heavy-atom deviation of <3 Å from experiment) were generated for six out of seven bound RNA chains, and even more precise models (deviation < 2 Å) were obtained for five out of seven cases, a significant improvement compared to the state of the art. The method is not restricted to RRMs but was also successfully applied to Pumilio RNA binding proteins. PMID:27131381

  15. 16S rRNA Gene Sequence Analysis of Drinking Water Using RNA and DNA Extracts as Targets for Clone Library Development

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    The bacterial composition of chlorinated drinking water was analyzed using 16S rRNA gene clone libraries derived from DNA extracts of 12 samples and compared to clone libraries previously generated using RNA extracts from the same samples. Phylogenetic analysis of 761 DNA-based ...

  16. Automated extraction protocol for quantification of SARS-Coronavirus RNA in serum: an evaluation study

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    Lui Wing-bong

    2006-02-01

    Full Text Available Abstract Background We have previously developed a test for the diagnosis and prognostic assessment of the severe acute respiratory syndrome (SARS based on the detection of the SARS-coronavirus RNA in serum by real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR. In this study, we evaluated the feasibility of automating the serum RNA extraction procedure in order to increase the throughput of the assay. Methods An automated nucleic acid extraction platform using the MagNA Pure LC instrument (Roche Diagnostics was evaluated. We developed a modified protocol in compliance with the recommended biosafety guidelines from the World Health Organization based on the use of the MagNA Pure total nucleic acid large volume isolation kit for the extraction of SARS-coronavirus RNA. The modified protocol was compared with a column-based extraction kit (QIAamp viral RNA mini kit, Qiagen for quantitative performance, analytical sensitivity and precision. Results The newly developed automated protocol was shown to be free from carry-over contamination and have comparable performance with other standard protocols and kits designed for the MagNA Pure LC instrument. However, the automated method was found to be less sensitive, less precise and led to consistently lower serum SARS-coronavirus concentrations when compared with the column-based extraction method. Conclusion As the diagnostic efficiency and prognostic value of the serum SARS-CoV RNA RT-PCR test is critically associated with the analytical sensitivity and quantitative performance contributed both by the RNA extraction and RT-PCR components of the test, we recommend the use of the column-based manual RNA extraction method.

  17. Comparison of RNA extraction methods in Thai aromatic coconut water

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    Nopporn Jaroonchon

    2015-10-01

    Full Text Available Many researches have reported that nucleic acid in coconut water is in free form and at very low yields which makes it difficult to process in molecular studies. Our research attempted to compare two extraction methods to obtain a higher yield of total RNA in aromatic coconut water and monitor its change at various fruit stages. The first method used ethanol and sodium acetate as reagents; the second method used lithium chloride. We found that extraction using only lithium chloride gave a higher total RNA yield than the method using ethanol to precipitate nucleic acid. In addition, the total RNA from both methods could be used in amplification of betaine aldehyde dehydrogenase2 (Badh2 genes, which is involved in coconut aroma biosynthesis, and could be used to perform further study as we expected. From the molecular study, the nucleic acid found in coconut water increased with fruit age.

  18. Urinary extracellular vesicles for RNA extraction: optimization of a protocol devoid of prokaryote contamination

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    Dorota Tataruch-Weinert

    2016-06-01

    Full Text Available Background: Urinary extracellular vesicles (UEVs represent an ideal platform for biomarker discovery. They carry different types of RNA species, and reported profile discrepancies related to the presence/absence of 18s and 28s rRNA remain controversial. Moreover, sufficient urinary RNA yields and respective quality RNA profiles are still to be fully established. Methods: UEVs were enriched by hydrostatic filtration dialysis, and RNA content was extracted using 7 different commercially available techniques. RNA quantity was assessed using spectrophotometry and fluorometry, whilst RNA quality was determined by capillary electrophoresis. Results: The presence of prokaryotic transcriptome was stressed when cellular RNA, as a control, was spiked into the UEVs samples before RNA extraction. The presence of bacteria in hydrostatic filtration dialysis above 1,000 kDa molecular weight cut-off and in crude urine was confirmed with growth media plates. The efficiency in removing urinary bacteria was evaluated by differential centrifugation, filtration (0.22 µm filters and chemical pretreatment (water purification tablet. For volumes of urine >200 ml, the chemical treatment provides ease of handling without affecting vesicle integrity, protein and RNA profiles. This protocol was selected to enrich RNA with 7 methods, and its respective quality and quantity were assessed. The results were given as follows: (a Fluorometry gave more repeatability and reproducibility than spectrophotometry to assess the RNA yields, (b UEVs were enriched with small RNA, (c Ribosomal RNA peaks were not observed for any RNA extraction method used and (d RNA yield was higher for column-based method designed for urinary exosome, whilst the highest relative microRNA presence was obtained using TRIzol method. Conclusion: Our results show that the presence of bacteria can lead to misidentification in the electrophoresis peaks. Fluorometry is more reliable than spectrophotometry. RNA

  19. Improving Griffith's protocol for co-extraction of microbial DNA and RNA in adsorptive soils

    DEFF Research Database (Denmark)

    Paulin, Mélanie Marie; Nicolaisen, Mette Haubjerg; Jacobsen, Carsten Suhr

    2013-01-01

    Quantification of microbial gene expression is increasingly being used to study key functions in soil microbial communities, yet major limitations still exist for efficient extraction of nucleic acids, especially RNA for transcript analysis, from this complex matrix. We present an improved......-time PCR on both the RNA (after conversion to cDNA) and the DNA fraction of the extracts. Non-adsorptive soils were characterized by low clay content and/or high phosphate content, whereas adsorptive soils had clay contents above 20% and/or a strong presence of divalent Ca in combination with high p......H. Modifications to the co-extraction protocol improved nucleic acid extraction efficiency from all adsorptive soils and were successfully validated by DGGE analysis of the indigenous community based on 16S rRNA gene and transcripts in soils representing low biomass and/or high clay content. This new approach...

  20. Molecular Techniques for Dicistrovirus Detection without RNA Extraction or Purification

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    Jailson F. B. Querido

    2013-01-01

    Full Text Available Dicistroviridae is a new family of small, nonenveloped, and +ssRNA viruses pathogenic to both beneficial arthropods and insect pests as well. Triatoma virus (TrV, a dicistrovirus, is a pathogen of Triatoma infestans (Hemiptera: Reduviidae, one of the main vectors of Chagas disease. In this work, we report a single-step method to identify TrV, a dicistrovirus, isolated from fecal samples of triatomines. The identification method proved to be quite sensitive, even without the extraction and purification of RNA virus.

  1. Extraction of total RNA in the developing chicken forebrain

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    Sayed Rasoul Zaker

    2014-01-01

    Full Text Available Background: Gene expression of Gama-Aminobutyric acid (GABA A receptor subunits may change during development. Procedures in molecular biology are required to understand the gene expression profile GABA A R in chicken. The outcome of the results depends on good-quality high-molecular-weight RNA. Several procedures can be used to isolate RNA from the brain of chicken; however, most of them are time-consuming and require disruption of cells or freeze and thaw in the presence of RNase inhibitors. The aim of this experiment was isolation of RNA from chicken embryonic brain tissues using appropriate RNA extraction kit. Materials and Methods: Fertilized eggs from Ross breed (Gallus gallus were incubated at 38°C and 60% relative humidity in a forced-draft incubator and were turned every 3 h. After 3, 7, 14 and 20 days of incubation, eggs were cooled on ice to induce deep anesthesia. Then whole brains were dissected out. As brains could not be excised in a reproducible way from earlier embryos (embryonic days 4 and 6, whole heads were collected. Chicken embryos between day 7 to 20 and 1 day after birth were decapitated, and their brains removed. Samples were immediately inserted into lysis buffer and stored at −70°C. Total RNA was isolated and a contaminating genomic deoxyribonucleic acid (DNA was digested. RNA quality was checked using gel electrophoresis. Results: We obtained 52 mg/ml to 745 mg/ml with A260/280 1.7-2.2. Only high-quality RNA, with no signs of degradation, was used for further experiments. Conclusion: In conclusion, protocol was found to be suitable for the isolation of total RNA from embryonic chicken cells.

  2. RNA extraction from decaying wood for (meta)transcriptomic analyses.

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    Adamo, Martino; Voyron, Samuele; Girlanda, Mariangela; Marmeisse, Roland

    2017-10-01

    Wood decomposition is a key step of the terrestrial carbon cycle and is of economic importance. It is essentially a microbiological process performed by fungi and to an unknown extent by bacteria. To gain access to the genes expressed by the diverse microbial communities participating in wood decay, we developed an RNA extraction protocol from this recalcitrant material rich in polysaccharides and phenolic compounds. This protocol was implemented on 22 wood samples representing as many tree species from 11 plant families in the Angiosperms and Gymnosperms. RNA was successfully extracted from all samples and converted into cDNAs from which were amplified both fungal and bacterial protein coding genes, including genes encoding hydrolytic enzymes participating in lignocellulose hydrolysis. This protocol applicable to a wide range of decomposing wood types represents a first step towards a metatranscriptomic analysis of wood degradation under natural conditions.

  3. antaRNA: ant colony-based RNA sequence design.

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    Kleinkauf, Robert; Mann, Martin; Backofen, Rolf

    2015-10-01

    RNA sequence design is studied at least as long as the classical folding problem. Although for the latter the functional fold of an RNA molecule is to be found ,: inverse folding tries to identify RNA sequences that fold into a function-specific target structure. In combination with RNA-based biotechnology and synthetic biology ,: reliable RNA sequence design becomes a crucial step to generate novel biochemical components. In this article ,: the computational tool antaRNA is presented. It is capable of compiling RNA sequences for a given structure that comply in addition with an adjustable full range objective GC-content distribution ,: specific sequence constraints and additional fuzzy structure constraints. antaRNA applies ant colony optimization meta-heuristics and its superior performance is shown on a biological datasets. http://www.bioinf.uni-freiburg.de/Software/antaRNA CONTACT: backofen@informatik.uni-freiburg.de Supplementary data are available at Bioinformatics online. © The Author 2015. Published by Oxford University Press.

  4. Sample preservation, transport and processing strategies for honeybee RNA extraction: Influence on RNA yield, quality, target quantification and data normalization.

    Science.gov (United States)

    Forsgren, Eva; Locke, Barbara; Semberg, Emilia; Laugen, Ane T; Miranda, Joachim R de

    2017-08-01

    Viral infections in managed honey bees are numerous, and most of them are caused by viruses with an RNA genome. Since RNA degrades rapidly, appropriate sample management and RNA extraction methods are imperative to get high quality RNA for downstream assays. This study evaluated the effect of various sampling-transport scenarios (combinations of temperature, RNA stabilizers, and duration) of transport on six RNA quality parameters; yield, purity, integrity, cDNA synthesis efficiency, target detection and quantification. The use of water and extraction buffer were also compared for a primary bee tissue homogenate prior to RNA extraction. The strategy least affected by time was preservation of samples at -80°C. All other regimens turned out to be poor alternatives unless the samples were frozen or processed within 24h. Chemical stabilizers have the greatest impact on RNA quality and adding an extra homogenization step (a QIAshredder™ homogenizer) to the extraction protocol significantly improves the RNA yield and chemical purity. This study confirms that RIN values (RNA Integrity Number), should be used cautiously with bee RNA. Using water for the primary homogenate has no negative effect on RNA quality as long as this step is no longer than 15min. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. ncRNA-class Web Tool: Non-coding RNA feature extraction and pre-miRNA classification web tool

    KAUST Repository

    Kleftogiannis, Dimitrios A.; Theofilatos, Konstantinos A.; Papadimitriou, Stergios; Tsakalidis, Athanasios K.; Likothanassis, Spiridon D.; Mavroudi, Seferina P.

    2012-01-01

    Until recently, it was commonly accepted that most genetic information is transacted by proteins. Recent evidence suggests that the majority of the genomes of mammals and other complex organisms are in fact transcribed into non-coding RNAs (ncRNAs), many of which are alternatively spliced and/or processed into smaller products. Non coding RNA genes analysis requires the calculation of several sequential, thermodynamical and structural features. Many independent tools have already been developed for the efficient calculation of such features but to the best of our knowledge there does not exist any integrative approach for this task. The most significant amount of existing work is related to the miRNA class of non-coding RNAs. MicroRNAs (miRNAs) are small non-coding RNAs that play a significant role in gene regulation and their prediction is a challenging bioinformatics problem. Non-coding RNA feature extraction and pre-miRNA classification Web Tool (ncRNA-class Web Tool) is a publicly available web tool ( http://150.140.142.24:82/Default.aspx ) which provides a user friendly and efficient environment for the effective calculation of a set of 58 sequential, thermodynamical and structural features of non-coding RNAs, plus a tool for the accurate prediction of miRNAs. © 2012 IFIP International Federation for Information Processing.

  6. Extracting microRNA-gene relations from biomedical literature using distant supervision.

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    Andre Lamurias

    Full Text Available Many biomedical relation extraction approaches are based on supervised machine learning, requiring an annotated corpus. Distant supervision aims at training a classifier by combining a knowledge base with a corpus, reducing the amount of manual effort necessary. This is particularly useful for biomedicine because many databases and ontologies have been made available for many biological processes, while the availability of annotated corpora is still limited. We studied the extraction of microRNA-gene relations from text. MicroRNA regulation is an important biological process due to its close association with human diseases. The proposed method, IBRel, is based on distantly supervised multi-instance learning. We evaluated IBRel on three datasets, and the results were compared with a co-occurrence approach as well as a supervised machine learning algorithm. While supervised learning outperformed on two of those datasets, IBRel obtained an F-score 28.3 percentage points higher on the dataset for which there was no training set developed specifically. To demonstrate the applicability of IBRel, we used it to extract 27 miRNA-gene relations from recently published papers about cystic fibrosis. Our results demonstrate that our method can be successfully used to extract relations from literature about a biological process without an annotated corpus. The source code and data used in this study are available at https://github.com/AndreLamurias/IBRel.

  7. Comparison of methods for miRNA extraction from plasma and quantitative recovery of RNA from plasma and cerebrospinal fluid

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    Melissa A McAlexander

    2013-05-01

    Full Text Available Interest in extracellular RNA has intensified as evidence accumulates that these molecules may be useful as indicators of a wide variety of biological conditions. To establish specific extracellular RNA molecules as clinically relevant biomarkers, reproducible recovery from biological samples and reliable measurements of the isolated RNA are paramount. Towards these ends, careful and rigorous comparisons of technical procedures are needed at all steps from sample handling to RNA isolation to RNA measurement protocols. In the investigations described in this methods paper, RT-qPCR was used to examine the apparent recovery of specific endogenous miRNAs and a spiked-in synthetic RNA from blood plasma samples. RNA was isolated using several widely used RNA isolation kits, with or without the addition of glycogen as a carrier. Kits examined included total RNA isolation systems that have been commercially available for several years and commonly adapted for extraction of biofluid RNA, as well as more recently introduced biofluids-specific RNA methods. Our conclusions include the following: some RNA isolation methods appear to be superior to others for the recovery of RNA from biological fluids; addition of a carrier molecule seems to be beneficial for some but not all isolation methods; and partially or fully quantitative recovery of RNA is observed from increasing volumes of plasma and cerebrospinal fluid.

  8. Evaluation of a low-cost procedure for sampling, long-term storage, and extraction of RNA from blood for qPCR analyses

    DEFF Research Database (Denmark)

    Mærkedahl, Rasmus Baadsgaard; Frøkiær, Hanne; Lauritzen, Lotte

    2015-01-01

    Abstract Background: In large clinical trials, where RNA cannot be extracted immediately after sampling, preserving RNA in whole blood is a crucial initial step in obtaining robust qPCR data. The current golden standard for RNA preservation is costly and designed for time-consuming column-based RNA....../binding solution over time and between samples stored and extracted by the two systems. Conclusions: The MagMAX system can be used for storage of human blood for up to 4 months and is equivalent to the PAXgene system for RNA extraction. It furthermore, provides a means for significant cost reduction in large...

  9. Efficiency of RNA extraction from selected bacteria in the context of biogas production and metatranscriptomics.

    Science.gov (United States)

    Stark, Lucy; Giersch, Tina; Wünschiers, Röbbe

    2014-10-01

    Understanding the microbial population in anaerobic digestion is an essential task to increase efficient substrate use and process stability. The metabolic state, represented e.g. by the transcriptome, of a fermenting system can help to find markers for monitoring industrial biogas production to prevent failures or to model the whole process. Advances in next-generation sequencing make transcriptomes accessible for large-scale analyses. In order to analyze the metatranscriptome of a mixed-species sample, isolation of high-quality RNA is the first step. However, different extraction methods may yield different efficiencies in different species. Especially in mixed-species environmental samples, unbiased isolation of transcripts is important for meaningful conclusions. We applied five different RNA-extraction protocols to nine taxonomic diverse bacterial species. Chosen methods are based on various lysis and extraction principles. We found that the extraction efficiency of different methods depends strongly on the target organism. RNA isolation of gram-positive bacteria was characterized by low yield whilst from gram-negative species higher concentrations can be obtained. Transferring our results to mixed-species investigations, such as metatranscriptomics with biofilms or biogas plants, leads to the conclusion that particular microorganisms might be over- or underrepresented depending on the method applied. Special care must be taken when using such metatranscriptomics data for, e.g. process modeling. Copyright © 2013 Elsevier Ltd. All rights reserved.

  10. Extraction of low molecular weight RNA from Citrus trifolita tissues ...

    African Journals Online (AJOL)

    Jane

    2010-12-20

    Dec 20, 2010 ... A critical prerequisite in miRNA studies is acquisition of high quality LMW RNA. LMW RNA is ..... air-dried for a few minutes and then exposed to BIOMAX X-ray film for 48 h using an .... Approaches to microRNA discovery. Nat.

  11. Feature-Based and String-Based Models for Predicting RNA-Protein Interaction

    Directory of Open Access Journals (Sweden)

    Donald Adjeroh

    2018-03-01

    Full Text Available In this work, we study two approaches for the problem of RNA-Protein Interaction (RPI. In the first approach, we use a feature-based technique by combining extracted features from both sequences and secondary structures. The feature-based approach enhanced the prediction accuracy as it included much more available information about the RNA-protein pairs. In the second approach, we apply search algorithms and data structures to extract effective string patterns for prediction of RPI, using both sequence information (protein and RNA sequences, and structure information (protein and RNA secondary structures. This led to different string-based models for predicting interacting RNA-protein pairs. We show results that demonstrate the effectiveness of the proposed approaches, including comparative results against leading state-of-the-art methods.

  12. RNA-based therapies for genodermatoses

    NARCIS (Netherlands)

    Bornert, Olivier; Peking, Patricia; Bremer, Jeroen; Koller, Ulrich; van den Akker, Peter C.; Aartsma-Rus, Annemieke; Pasmooij, Anna M. G.; Murauer, Eva M.; Nystroem, Alexander

    Genetic disorders affecting the skin, genodermatoses, constitute a large and heterogeneous group of diseases, for which treatment is generally limited to management of symptoms. RNA-based therapies are emerging as a powerful tool to treat genodermatoses. In this review, we discuss in detail RNA

  13. Entropy-based model for miRNA isoform analysis.

    Directory of Open Access Journals (Sweden)

    Shengqin Wang

    Full Text Available MiRNAs have been widely studied due to their important post-transcriptional regulatory roles in gene expression. Many reports have demonstrated the evidence of miRNA isoform products (isomiRs in high-throughput small RNA sequencing data. However, the biological function involved in these molecules is still not well investigated. Here, we developed a Shannon entropy-based model to estimate isomiR expression profiles of high-throughput small RNA sequencing data extracted from miRBase webserver. By using the Kolmogorov-Smirnov statistical test (KS test, we demonstrated that the 5p and 3p miRNAs present more variants than the single arm miRNAs. We also found that the isomiR variant, except the 3' isomiR variant, is strongly correlated with Minimum Free Energy (MFE of pre-miRNA, suggesting the intrinsic feature of pre-miRNA should be one of the important factors for the miRNA regulation. The functional enrichment analysis showed that the miRNAs with high variation, particularly the 5' end variation, are enriched in a set of critical functions, supporting these molecules should not be randomly produced. Our results provide a probabilistic framework for miRNA isoforms analysis, and give functional insights into pre-miRNA processing.

  14. Establishment of the total RNA extraction system for lily bulbs with ...

    African Journals Online (AJOL)

    USER

    2011-12-07

    Dec 7, 2011 ... The brightness of. CTAB was higher than that of SDS, which indicated that only improved CTAB method can extract biologically active. RNA from lily bulbs. DISCUSSION. The presence of RNase, which can be classified into endogenous and exogenous, is the major cause for the failure of RNA extraction.

  15. Extraction of low molecular weight RNA from Citrus trifolita tissues ...

    African Journals Online (AJOL)

    Jane

    2010-12-20

    Dec 20, 2010 ... many biological and metabolic processes, including tissue ... water before getting LMW RNA. ... 1 cm) were collected from five-year-old trifoliate orange (C. ... using 700 µl 75% ethanol and total RNA was precipitated by.

  16. Evaluation of RNA extraction methods in rice and their application in expression analysis of resistance genes against Magnaporthe oryzae

    Directory of Open Access Journals (Sweden)

    Parisa Azizi

    2017-01-01

    Full Text Available Extraction of RNA of high quality and integrity is essential for gene expression studies and all downstream RNA-based techniques. The leaves of 16 merit Malaysian rice varieties were used to isolate total RNA using five different methods. The quantity, quality and integrity of extracted RNA were confirmed using three different means. The ratios of A260/280 ranged from 2.12 to 2.20. Electrophoresis (1.5% agarose gel was performed, illustrating intact and sharp bands representing the 28S, 18S, 5.8S and 5S ribosomal subunits of RNA, presenting intact RNA. RNA quality was verified using semi-quantitative polymerase chain reaction (sqPCR. The objective of this study was to identify different genes involved in the resistance of rice plants using high-quality RNA extracted 31 h after inoculation of Magnaporthe oryzae pathotype P7.2. The expression levels of eight blast resistance genes, Pikh, Pib, Pita, Pi21, Pi9, Os11gRGA8, OsWRKY22 and OsWRKY45, were evaluated by real-time PCR (RT-PCR. Real-time PCR was performed to identify candidate genes using RNA extracted by the TRIzol method, which showed the highest score compared with other methods in terms of RNA quantity, purity and integrity. In addition, the results of real-time PCR confirmed that the up-regulation of seven blast resistance genes may confer stronger resistance for the MR 276 variety against M. oryzae pathotype P7.2.

  17. RNA preservation agents and nucleic acid extraction method bias perceived bacterial community composition.

    Directory of Open Access Journals (Sweden)

    Ann McCarthy

    Full Text Available Bias is a pervasive problem when characterizing microbial communities. An important source is the difference in lysis efficiencies of different populations, which vary depending on the extraction protocol used. To avoid such biases impacting comparisons between gene and transcript abundances in the environment, the use of one protocol that simultaneously extracts both types of nucleic acids from microbial community samples has gained popularity. However, knowledge regarding tradeoffs to combined nucleic acid extraction protocols is limited, particularly regarding yield and biases in the observed community composition. Here, we evaluated a commercially available protocol for simultaneous extraction of DNA and RNA, which we adapted for freshwater microbial community samples that were collected on filters. DNA and RNA yields were comparable to other commonly used, but independent DNA and RNA extraction protocols. RNA protection agents benefited RNA quality, but decreased DNA yields significantly. Choice of extraction protocol influenced the perceived bacterial community composition, with strong method-dependent biases observed for specific phyla such as the Verrucomicrobia. The combined DNA/RNA extraction protocol detected significantly higher levels of Verrucomicrobia than the other protocols, and those higher numbers were confirmed by microscopic analysis. Use of RNA protection agents as well as independent sequencing runs caused a significant shift in community composition as well, albeit smaller than the shift caused by using different extraction protocols. Despite methodological biases, sample origin was the strongest determinant of community composition. However, when the abundance of specific phylogenetic groups is of interest, researchers need to be aware of the biases their methods introduce. This is particularly relevant if different methods are used for DNA and RNA extraction, in addition to using RNA protection agents only for RNA

  18. Comparative evaluation of total RNA extraction methods in Theobroma cacao using shoot apical meristems.

    Science.gov (United States)

    Silva, D V; Branco, S M J; Holanda, I S A; Royaert, S; Motamayor, J C; Marelli, J P; Corrêa, R X

    2016-03-04

    Theobroma cacao is a species of great economic importance with its beans used for chocolate production. The tree has been a target of various molecular studies. It contains many polyphenols, which complicate the extraction of nucleic acids with the extraction protocols requiring a large amount of plant material. These issues, therefore, necessitate the optimization of the protocols. The aim of the present study was to evaluate different methods for extraction of total RNA from shoot apical meristems of T. cacao 'CCN 51' and to assess the influence of storage conditions for the meristems on the extraction. The study also aimed to identify the most efficient protocol for RNA extraction using a small amount of plant material. Four different protocols were evaluated for RNA extraction using one shoot apical meristem per sample. Among these protocols, one that was more efficient was then tested to extract RNA using four different numbers of shoot apical meristems, subjected to three different storage conditions. The best protocol was tested for cDNA amplification using reverse transcription-polymerase chain reaction; the cDNA quality was determined to be satisfactory for molecular analyses. The study revealed that with the best RNA extraction protocol, one shoot apical meristem was sufficient for extraction of high-quality total RNA. The results obtained might enable advances in genetic analyses and molecular studies using reduced amount of plant material.

  19. SELEX-Based Screening of Exosome-Tropic RNA.

    Science.gov (United States)

    Yamashita, Takuma; Shinotsuka, Haruka; Takahashi, Yuki; Kato, Kana; Nishikawa, Makiya; Takakura, Yoshinobu

    2017-01-01

    Cell-derived nanosized vesicles or exosomes are expected to become delivery carriers for functional RNAs, such as small interfering RNA (siRNA). A method to efficiently load functional RNAs into exosomes is required for the development of exosome-based delivery carriers of functional RNAs. However, there is no method to find exosome-tropic exogenous RNA sequences. In this study, we used a systematic evolution of ligands by exponential enrichment (SELEX) method to screen exosome-tropic RNAs that can be used to load functional RNAs into exosomes by conjugation. Pooled single stranded 80-base RNAs, each of which contains a randomized 40-base sequence, were transfected into B16-BL6 murine melanoma cells and exosomes were collected from the cells. RNAs extracted from the exosomes were subjected to next round of SELEX. Cloning and sequencing of RNAs in SELEX-screened RNA pools showed that 29 of 56 clones had a typical RNA sequence. The sequence found by SELEX was enriched in exosomes after transfection to B16-BL6 cells. The results show that the SELEX-based method can be used for screening of exosome-tropic RNAs.

  20. Evidence for an RNA polymerization activity in axolotl and Xenopus egg extracts.

    Directory of Open Access Journals (Sweden)

    Hélène Pelczar

    2010-12-01

    Full Text Available We have previously reported a post-transcriptional RNA amplification observed in vivo following injection of in vitro synthesized transcripts into axolotl oocytes, unfertilized (UFE or fertilized eggs. To further characterize this phenomenon, low speed extracts (LSE from axolotl and Xenopus UFE were prepared and tested in an RNA polymerization assay. The major conclusions are: i the amphibian extracts catalyze the incorporation of radioactive ribonucleotide in RNase but not DNase sensitive products showing that these products correspond to RNA; ii the phenomenon is resistant to α-amanitin, an inhibitor of RNA polymerases II and III and to cordycepin (3'dAMP, but sensitive to cordycepin 5'-triphosphate, an RNA elongation inhibitor, which supports the existence of an RNA polymerase activity different from polymerases II and III; the detection of radiolabelled RNA comigrating at the same length as the exogenous transcript added to the extracts allowed us to show that iii the RNA polymerization is not a 3' end labelling and that iv the radiolabelled RNA is single rather than double stranded. In vitro cell-free systems derived from amphibian UFE therefore validate our previous in vivo results hypothesizing the existence of an evolutionary conserved enzymatic activity with the properties of an RNA dependent RNA polymerase (RdRp.

  1. Optimization of phenol extraction procedures for preparation of RNA from mammalian lymphoid organs

    Energy Technology Data Exchange (ETDEWEB)

    Griffin, G.D.; Sellin, H.G.; Novelli, G.D.

    1978-07-01

    Methods have been developed to optimize the extraction of RNA from mammalian lymphoid organs (spleen) with respect to both quantity and quality of RNA and with minimal DNA contamination. Nuclease inhibitors, including diethyl pyrocarbonate, polyvinyl sulfate, and bentonite were used in the initial disruption of the tissue, which was accomplished by blender, Dounce homogenizer, or preparation of a cell suspension. Seven buffer systems, varying with respect to pH, detergent, and NaCl concentration, were used in the initial extraction with phenol, and the temperature of extraction was also varied. Protocols involving the selective use of naphthalene 1.5-disulfonic acid and sodium dodecyl sulfate were developed to provide an initial RNA extract with minimal DNA content. Dounce homogenization, followed by separate treatment of nuclear and cytosol fractions, was found to be the most effective technique, both in terms of RNA yield (averaging 76%) and the quality of RNA recovered (as judged by gel electrophoresis). RNA from blender preparations contained larger amounts of DNA and RNA yield was decreased to 54%. RNA extracted from spleen cell suspensions was of poor quality and gave very poor yield (27%).

  2. Literature-based condition-specific miRNA-mRNA target prediction.

    Directory of Open Access Journals (Sweden)

    Minsik Oh

    Full Text Available miRNAs are small non-coding RNAs that regulate gene expression by binding to the 3'-UTR of genes. Many recent studies have reported that miRNAs play important biological roles by regulating specific mRNAs or genes. Many sequence-based target prediction algorithms have been developed to predict miRNA targets. However, these methods are not designed for condition-specific target predictions and produce many false positives; thus, expression-based target prediction algorithms have been developed for condition-specific target predictions. A typical strategy to utilize expression data is to leverage the negative control roles of miRNAs on genes. To control false positives, a stringent cutoff value is typically set, but in this case, these methods tend to reject many true target relationships, i.e., false negatives. To overcome these limitations, additional information should be utilized. The literature is probably the best resource that we can utilize. Recent literature mining systems compile millions of articles with experiments designed for specific biological questions, and the systems provide a function to search for specific information. To utilize the literature information, we used a literature mining system, BEST, that automatically extracts information from the literature in PubMed and that allows the user to perform searches of the literature with any English words. By integrating omics data analysis methods and BEST, we developed Context-MMIA, a miRNA-mRNA target prediction method that combines expression data analysis results and the literature information extracted based on the user-specified context. In the pathway enrichment analysis using genes included in the top 200 miRNA-targets, Context-MMIA outperformed the four existing target prediction methods that we tested. In another test on whether prediction methods can re-produce experimentally validated target relationships, Context-MMIA outperformed the four existing target prediction

  3. A discontinuous RNA platform mediates RNA virus replication: building an integrated model for RNA-based regulation of viral processes.

    Directory of Open Access Journals (Sweden)

    Baodong Wu

    2009-03-01

    Full Text Available Plus-strand RNA viruses contain RNA elements within their genomes that mediate a variety of fundamental viral processes. The traditional view of these elements is that of local RNA structures. This perspective, however, is changing due to increasing discoveries of functional viral RNA elements that are formed by long-range RNA-RNA interactions, often spanning thousands of nucleotides. The plus-strand RNA genomes of tombusviruses exemplify this concept by possessing different long-range RNA-RNA interactions that regulate both viral translation and transcription. Here we report that a third fundamental tombusvirus process, viral genome replication, requires a long-range RNA-based interaction spanning approximately 3000 nts. In vivo and in vitro analyses suggest that the discontinuous RNA platform formed by the interaction facilitates efficient assembly of the viral RNA replicase. This finding has allowed us to build an integrated model for the role of global RNA structure in regulating the reproduction of a eukaryotic RNA virus, and the insights gained have extended our understanding of the multifunctional nature of viral RNA genomes.

  4. Comparison of RNA Extraction Methods for the Identification of Grapevine fan leaf virus

    Directory of Open Access Journals (Sweden)

    Z. Gholampour

    2016-06-01

    Full Text Available Introduction: To now, more than 70 viral diseases have been reported from grapevine. Serological methods are regular diagnostic tools of grapevine viruses, however, their sensitivity has affected by seasonal fluctuations of the virus. Reverse transcription polymerase chain reaction provides significant improvement in detection of grapevine viruses. Extraction of high-quality RNA is essential for the successful application of many molecular techniques, such as RT-PCR. Extraction of high-quality RNA from the leaves of woody plants, such as grapevine, is particularly challenging because of high concentrations of polysaccharides, polyphenols, and other secondary metabolites. Some RNA extraction methods yield pellets that are poorly soluble, indicating the presence of unknown contaminants, whereas others are gelatinous, indicating the presence of polysaccharides. RNA can make complexes with polysaccharides and phenolic compounds render the RNA unusable for applications such as reverse transcription. Grapevine fanleaf virus is a member of the genus Nepovirus in the family Secoviridae. The GFLV genome consists of two positive-sense single stranded RNAs. The genome has a poly (A tail at the 3´ terminus and a covalently linked VPG protein at the 5´ terminus. Several extraction methods had been reported to be used for identification of GFLV in grapevine. Some of them require harmful chemical material; disadvantages of other are high costs. Immunocapture-RT-PCR requires preparation of specific antibody and direct binding RT-PCR (DB-RT-PCR has a high contamination risk. In this study, four RNA extraction protocols were compared with a commercial isolation kit to explore the most efficient RNA isolation method for grapevines. Material and Methods: 40 leaf samples were randomly collected during the growing season of 2011-2012. GFLV was detected in leaf samples by enzyme linked immunosorbent assay (ELISA Using specific antibodies raised against Iranian

  5. Efficient RNA extraction protocol for the wood mangrove species Laguncularia racemosa suited for next-generation RNA sequencing

    International Nuclear Information System (INIS)

    Wilwerth, M. W.; Rossetto, P.

    2016-01-01

    Mangrove flora and habitat have immeasurable importance in marine and coastal ecology as well as in the economy. Despite their importance, they are constantly threatened by oil spill accidents and environmental contamination; therefore, it is crucial to understand the changes in gene expression to better predict toxicity in these plants. Among the species of Atlantic coast mangrove (Americas and Africa), Laguncularia racemosa, or white mangrove, is a conspicuous species. The wide distribution of L. racemosa in areas where marine oil exploration is rapidly increasing make it a candidate mangrove species model to uncover the impact of oil spills at the molecular level with the use of massive transcriptome sequencing. However, for this purpose, the RNA extraction protocol should ensure low levels of contaminants and structure integrity. In this study, eight RNA extraction methods were tested and analysed using downstream applications. The InviTrap Spin Plant RNA Mini Kit performed best with regard to purity and integrity. Moreover, the obtained RNA was submitted to cDNA synthesis and RT-PCR, successfully generating amplification products of the expected size. These Results show the applicability of the RNA obtained here for downstream methodologies, such as the construction of cDNA libraries for the Illumina Hi-seq platform. (author)

  6. 64 original article optimization of rna extraction in mycobacterium

    African Journals Online (AJOL)

    Dr Oboro VO

    ... on thick 7H10 agar plates supplemented with 10% oleic acid- ... RNA lysis solution (0.15 M sucrose, 10 mM sodium acetate [pH 5.2] .... medium was discarded and the infected macrophage ..... Clin Invest 1995, 96(1):245-249. 18. Buchmeier ...

  7. Shielding the messenger (RNA): microRNA-based anticancer therapies

    Science.gov (United States)

    Sotillo, Elena; Thomas-Tikhonenko, Andrei

    2011-01-01

    It has been a decade since scientists realized that microRNAs (miRNAs) are not an oddity invented by worms to regulate gene expression at post-transcriptional levels. Rather, many of these 21–22-nucleotide-short RNAs exist in invertebrates and vertebrates alike and some of them are in fact highly conserved. miRNAs are now recognized as an important class of non-coding small RNAs that inhibit gene expression by targeting mRNA stability and translation. In the last ten years, our knowledge of the miRNAs world was expanding at vertiginous speed, propelled by the development of computational engines for miRNA identification and target prediction, biochemical tools and techniques to modulate miRNA activity, and last but not least, the emergence of miRNA-centric animal models. One important conclusion that has emerged from this effort is that many microRNAs and their cognate targets are strongly implicated in cancer, either as oncogenes or tumor and metastasis suppressors. In this review we will discuss the diverse role that miRNAs play in cancer initiation and progression and also the tools with which miRNA expression could be corrected in vivo. While the idea of targeting microRNAs towards therapeutic ends is getting considerable traction, basic, translational, and clinical research done in the next few years will tell whether this promise is well-founded. PMID:21514318

  8. iDoRNA: An Interacting Domain-based Tool for Designing RNA-RNA Interaction Systems

    Directory of Open Access Journals (Sweden)

    Jittrawan Thaiprasit

    2016-03-01

    Full Text Available RNA-RNA interactions play a crucial role in gene regulation in living organisms. They have gained increasing interest in the field of synthetic biology because of their potential applications in medicine and biotechnology. However, few novel regulators based on RNA-RNA interactions with desired structures and functions have been developed due to the challenges of developing design tools. Recently, we proposed a novel tool, called iDoDe, for designing RNA-RNA interacting sequences by first decomposing RNA structures into interacting domains and then designing each domain using a stochastic algorithm. However, iDoDe did not provide an optimal solution because it still lacks a mechanism to optimize the design. In this work, we have further developed the tool by incorporating a genetic algorithm (GA to find an RNA solution with maximized structural similarity and minimized hybridized RNA energy, and renamed the tool iDoRNA. A set of suitable parameters for the genetic algorithm were determined and found to be a weighting factor of 0.7, a crossover rate of 0.9, a mutation rate of 0.1, and the number of individuals per population set to 8. We demonstrated the performance of iDoRNA in comparison with iDoDe by using six RNA-RNA interaction models. It was found that iDoRNA could efficiently generate all models of interacting RNAs with far more accuracy and required far less computational time than iDoDe. Moreover, we compared the design performance of our tool against existing design tools using forty-four RNA-RNA interaction models. The results showed that the performance of iDoRNA is better than RiboMaker when considering the ensemble defect, the fitness score and computation time usage. However, it appears that iDoRNA is outperformed by NUPACK and RNAiFold 2.0 when considering the ensemble defect. Nevertheless, iDoRNA can still be an useful alternative tool for designing novel RNA-RNA interactions in synthetic biology research. The source code of iDoRNA

  9. RNA-Based Vaccines in Cancer Immunotherapy

    Directory of Open Access Journals (Sweden)

    Megan A. McNamara

    2015-01-01

    Full Text Available RNA vaccines traditionally consist of messenger RNA synthesized by in vitro transcription using a bacteriophage RNA polymerase and template DNA that encodes the antigen(s of interest. Once administered and internalized by host cells, the mRNA transcripts are translated directly in the cytoplasm and then the resulting antigens are presented to antigen presenting cells to stimulate an immune response. Alternatively, dendritic cells can be loaded with either tumor associated antigen mRNA or total tumor RNA and delivered to the host to elicit a specific immune response. In this review, we will explain why RNA vaccines represent an attractive platform for cancer immunotherapy, discuss modifications to RNA structure that have been developed to optimize mRNA vaccine stability and translational efficiency, and describe strategies for nonviral delivery of mRNA vaccines, highlighting key preclinical and clinical data related to cancer immunotherapy.

  10. How Severely Is DNA Quantification Hampered by RNA Co-extraction?

    Science.gov (United States)

    Sanchez, Ignacio; Remm, Matthieu; Frasquilho, Sonia; Betsou, Fay; Mathieson, William

    2015-10-01

    The optional RNase digest that is part of many DNA extraction protocols is often omitted, either because RNase is not provided in the kit or because users do not want to risk contaminating their laboratory. Consequently, co-eluting RNA can become a "contaminant" of unknown magnitude in a DNA extraction. We extracted DNA from liver, lung, kidney, and heart tissues and established that 28-52% of the "DNA" as assessed by spectrophotometry is actually RNA (depending on tissue type). Including an RNase digest in the extraction protocol reduced 260:280 purity ratios. Co-eluting RNA drives an overestimation of DNA yield when quantification is carried out using OD 260 nm spectrophotometry, or becomes an unquantified contaminant when spectrofluorometry is used for DNA quantification. This situation is potentially incompatible with the best practice guidelines for biobanks issued by organizations such as the International Society for Biological and Environmental Repositories, which state that biospecimens should be accurately characterized in terms of their identity, purity, concentration, and integrity. Consequently, we conclude that an RNase digest must be included in DNA extractions if pure DNA is required. We also discuss the implications of unquantified RNA contamination in DNA samples in the context of laboratory accreditation schemes.

  11. Comparison of RNA Extraction Methods for Molecular Analysis of Oral Cytology

    Directory of Open Access Journals (Sweden)

    Mônica Ghislaine Oliveira Alves

    2016-01-01

    Full Text Available Objective of work: The aim of this study was to compare three methods of RNA extraction for molecular analysis of oral cytology to establish the best technique, considering its concentration and purity for molecular tests of oral lesions such as real-time reverse transcriptase reaction. Material and methods: The sample included exfoliative cytology from the oral cavity mucosa of patients with no visible clinical changes, using Orcellex Rovers Brush®. The extraction of total RNA was performed using the following three techniques: 30 samples were extracted by Trizol® technique, 30 by the DirectzolTM RNA Miniprep system and 30 by the RNeasy mini Kit. The absorbance was measured by spectrophotometer to estimate the purity. The estimated RNA concentration was obtained by multiplying the value of A260 (ng/mL by 40. Statistical analysis of the obtained data was performed using GraphPad Prism 5.03 software with Student t, analysis of variance and Bonferroni tests, considering p ≤0.05. Results: Trizol® group revealed higher average concentration, followed by Direct-zolTM and Rneasy group. It was observed that the RNA Direct-zolTM group had the highest purity, followed by RNeasy and Trizol® groups, allowing for the two ratios. Conclusion: Considering all aspects, concentration, purity and time spent in the procedures, the Direct-zolTM group showed the best results.

  12. Extraction of High Quality RNA from Cannabis sativa Bast Fibres: A Vademecum for Molecular Biologists

    Directory of Open Access Journals (Sweden)

    Gea Guerriero

    2016-07-01

    Full Text Available In plants there is no universal protocol for RNA extraction, since optimizations are required depending on the species, tissues and developmental stages. Some plants/tissues are rich in secondary metabolites or synthesize thick cell walls, which hinder an efficient RNA extraction. One such example is bast fibres, long extraxylary cells characterized by a thick cellulosic cell wall. Given the economic importance of bast fibres, which are used in the textile sector, as well as in biocomposites as green substitutes of glass fibres, it is desirable to better understand their development from a molecular point of view. This knowledge favours the development of biotechnological strategies aimed at improving specific properties of bast fibres. To be able to perform high-throughput analyses, such as, for instance, transcriptomics of bast fibres, RNA extraction is a crucial and limiting step. We here detail a protocol enabling the rapid extraction of high quality RNA from the bast fibres of textile hemp, Cannabis sativa L., a multi-purpose fibre crop standing in the spotlight of research.

  13. siRNA delivery with lipid-based systems

    DEFF Research Database (Denmark)

    Foged, Camilla

    2012-01-01

    A key hurdle for the further development of RNA interference (RNAi) therapeutics like small interfering RNA (siRNA) is their safe and effective delivery. Lipids are promising and versatile carriers because they are based on Nature's own building blocks and can be provided with properties which......RNA into more hydrophobic lipoplexes, which promote passage of the siRNA across cellular membrane barriers, especially when lipids are added that facilitate membrane fusion. Despite these attractive features, siRNA delivery vehicles are facing a number of challenges such as the limited delivery efficiency...

  14. Extraction of mRNA from coagulated horse blood and analysis of inflammation-related cytokine responses to coagulation

    DEFF Research Database (Denmark)

    Bovbjerg, Kirsten Katrine Lindegaard; Heegaard, Peter M. H.; Skovgaard, Kerstin

    2010-01-01

    the peripheral blood mononuclear cells present in the blood clot, homogenizing the clot by rotating knife homogenization (GentleMACS, Miltenyi Biotec) in the presence of QIAzol extraction buffer (Qiagen). The RNA extracted yielded high concentrations of total RNA (50-265 ng/μl) and quality measures (RIN=8...

  15. The European Regulatory Environment of RNA-Based Vaccines.

    Science.gov (United States)

    Hinz, Thomas; Kallen, Kajo; Britten, Cedrik M; Flamion, Bruno; Granzer, Ulrich; Hoos, Axel; Huber, Christoph; Khleif, Samir; Kreiter, Sebastian; Rammensee, Hans-Georg; Sahin, Ugur; Singh-Jasuja, Harpreet; Türeci, Özlem; Kalinke, Ulrich

    2017-01-01

    A variety of different mRNA-based drugs are currently in development. This became possible, since major breakthroughs in RNA research during the last decades allowed impressive improvements of translation, stability and delivery of mRNA. This article focuses on antigen-encoding RNA-based vaccines that are either directed against tumors or pathogens. mRNA-encoded vaccines are developed both for preventive or therapeutic purposes. Most mRNA-based vaccines are directly administered to patients. Alternatively, primary autologous cells from cancer patients are modified ex vivo by the use of mRNA and then are adoptively transferred to patients. In the EU no regulatory guidelines presently exist that specifically address mRNA-based vaccines. The existing regulatory framework, however, clearly defines that mRNA-based vaccines in most cases have to be centrally approved. Interestingly, depending on whether RNA-based vaccines are directed against tumors or infectious disease, they are formally considered gene therapy products or not, respectively. Besides an overview on the current clinical use of mRNA vaccines in various therapeutic areas a detailed discussion of the current regulatory situation is provided and regulatory perspectives are discussed.

  16. 16S rRNA Gene Sequence Analysis of Drinking Water Using RNA and DNA Extracts as Targets for Clone Library Development - Poster

    Science.gov (United States)

    We examined the bacterial composition of chlorinated drinking water using 16S rRNA gene clone libraries derived from RNA and DNA extracted from twelve water samples collected in three different months (June, August, and September of 2007). Phylogenetic analysis of 1234 and 1117 ...

  17. PIG-B: a homemade monophasic cocktail for the extraction of RNA.

    Science.gov (United States)

    Weber, K; Bolander, M E; Sarkar, G

    1998-02-01

    An inexpensive monophasic reagent has been developed for the extraction of total RNA from cells or tissues. The main ingredients of the reagent are Phenol, Isoamyl alcohol, Guanidinium isothiocyanate, and Beta-mercaptoethanol (PIG-B). The quality and yield of RNA obtained by this reagent is at par with that obtained by TRIzol, an expensive but widely used monophasic reagent available commercially. The complete composition and method of preparation of PIG-B is provided to aid preparation of the reagent in the laboratory.

  18. Protocol: high throughput silica-based purification of RNA from Arabidopsis seedlings in a 96-well format.

    Science.gov (United States)

    Salvo-Chirnside, Eliane; Kane, Steven; Kerr, Lorraine E

    2011-12-02

    The increasing popularity of systems-based approaches to plant research has resulted in a demand for high throughput (HTP) methods to be developed. RNA extraction from multiple samples in an experiment is a significant bottleneck in performing systems-level genomic studies. Therefore we have established a high throughput method of RNA extraction from Arabidopsis thaliana to facilitate gene expression studies in this widely used plant model. We present optimised manual and automated protocols for the extraction of total RNA from 9-day-old Arabidopsis seedlings in a 96 well plate format using silica membrane-based methodology. Consistent and reproducible yields of high quality RNA are isolated averaging 8.9 μg total RNA per sample (~20 mg plant tissue). The purified RNA is suitable for subsequent qPCR analysis of the expression of over 500 genes in triplicate from each sample. Using the automated procedure, 192 samples (2 × 96 well plates) can easily be fully processed (samples homogenised, RNA purified and quantified) in less than half a day. Additionally we demonstrate that plant samples can be stored in RNAlater at -20°C (but not 4°C) for 10 months prior to extraction with no significant effect on RNA yield or quality. Additionally, disrupted samples can be stored in the lysis buffer at -20°C for at least 6 months prior to completion of the extraction procedure providing a flexible sampling and storage scheme to facilitate complex time series experiments.

  19. Efficient recovery of whole blood RNA - a comparison of commercial RNA extraction protocols for high-throughput applications in wildlife species

    Directory of Open Access Journals (Sweden)

    Schwochow Doreen

    2012-06-01

    Full Text Available Abstract Background Since the emergence of next generation sequencing platforms, unprecedented opportunities have arisen in the study of natural vertebrate populations. In particular, insights into the genetic and epigenetic mechanisms of adaptation can be revealed through study of the expression profiles of genes. However, as a pre-requisite to expression profiling, care must be taken in RNA preparation as factors like DNA contamination, RNA integrity or transcript abundance can affect downstream applications. Here, we evaluated five commonly used RNA extraction methods using whole blood sampled under varying conditions from 20 wild carnivores. Results Despite the use of minute starting volumes, all methods produced quantifiable RNA extracts (1.4 – 18.4 μg with varying integrity (RIN 4.6 - 7.7, the latter being significantly affected by the storage and extraction method used. We observed a significant overall effect of the extraction method on DNA contamination. One particular extraction method, the LeukoLOCK™ filter system, yielded high RNA integrity along with low DNA contamination and efficient depletion of hemoglobin transcripts highly abundant in whole blood. In a proof of concept sequencing experiment, we found globin RNA transcripts to occupy up to ¼ of all sequencing reads if libraries were not depleted of hemoglobin prior to sequencing. Conclusion By carefully choosing the appropriate RNA extraction method, whole blood can become a valuable source for high-throughput applications like expression arrays or transcriptome sequencing from natural populations. Additionally, candidate genes showing signs of selection could subsequently be genotyped in large population samples using whole blood as a source for RNA without harming individuals from rare or endangered species.

  20. [In silico CRISPR-based sgRNA design].

    Science.gov (United States)

    Wang, Yuanli; Chuai, Guohui; Yan, Jifang; Shi, Lei; Liu, Qi

    2017-10-25

    CRISPR-based genome editing has been widely implemented in various cell types. In-silico single guide RNA (sgRNA) design is a key step for successful gene editing using CRISPR system. Continuing efforts are made to refine in-silico sgRNA design with high on-target efficacy and reduced off-target effects. In this paper, we summarize the present sgRNA design tools, and show that efficient in-silico models can be built that integrate current heterogeneous genome-editing data to derive unbiased sgRNA design rules and identify key features for improving sgRNA design. Our review shows that systematic comparisons and evaluation of on-target and off-target effects of sgRNA will allow more precise genome editing and gene therapies using the CRISPR system.

  1. Protocol: high throughput silica-based purification of RNA from Arabidopsis seedlings in a 96-well format

    OpenAIRE

    Salvo-Chirnside, Eliane; Kane, Steven; Kerr, Lorraine E

    2011-01-01

    Abstract The increasing popularity of systems-based approaches to plant research has resulted in a demand for high throughput (HTP) methods to be developed. RNA extraction from multiple samples in an experiment is a significant bottleneck in performing systems-level genomic studies. Therefore we have established a high throughput method of RNA extraction from Arabidopsis thaliana to facilitate gene expression studies in this widely used plant model. We present optimised manual and automated p...

  2. Soil DNA extraction procedure influences protist 18S rRNA gene community profiling outcome

    DEFF Research Database (Denmark)

    Santos, Susana S.; Nunes, Ines Marques; Nielsen, Tue K.

    2017-01-01

    Advances in sequencing technologies allow deeper studies of the soil protist diversity and function. However, little attention has been given to the impact of the chosen soil DNA extraction procedure to the overall results. We examined the effect of three acknowledged DNA recovery methods, two...... manual methods (ISOm-11063, GnS-GII) and one commercial kit (MoBio), on soil protist community structures obtained from different sites with different land uses. Results from 18S rRNA gene amplicon sequencing suggest that DNA extraction method significantly affect the replicate homogeneity, the total...... number of operational taxonomic units (OTUs) recovered and the overall taxonomic structure and diversity of soil protist communities. However, DNA extraction effects did not overwhelm the natural variation among samples, as the community data still strongly grouped by geographical location...

  3. [Effects of lipopolysaccharides extracted from Porphyromonas endodontalis on the expression of IL-1beta mRNA and IL-6 mRNA in osteoblasts].

    Science.gov (United States)

    Yang, Di; Li, Ren; Qiu, Li-Hong; Li, Chen

    2009-04-01

    To quantify the IL-1 beta mRNA and IL-6 mRNA expression induced by lipopolysaccharides (LPS)extracted from Porphyromonas endodontalis(P.e) in osteoblasts, and to relate P.e-LPS to bone absorption pathogenesis in lesions of chronical apical periodontitis. MG63 was treated with different concentrations of P.e-LPS(0-50 microg/mL) for different hours(0-24h). The expression of IL-1 beta mRNA and IL-6 mRNA was detected by reverse transcription polymerase chain reaction (RT-PCR).Statistical analysis was performed using one- way ANOVA and Dunnett t test with SPSS11.0 software package. The level of IL-1 beta mRNA and IL-6 mRNA increased significantly after treatment with P.e-LPS at more than 5 microg/mL (P<0.01)and for more than 1 hour (P<0.01), which indicated that P.e-LPS induced osteoblasts to express IL-1 beta mRNA and IL-6 mRNA in dose and time dependent manners. P.e-LPS may promote bone resorption in lesions of chronical apical periodontitis by inducing IL-1 beta mRNA and IL-6 mRNA expression in osteoblasts.

  4. RNA.

    Science.gov (United States)

    Darnell, James E., Jr.

    1985-01-01

    Ribonucleic acid (RNA) converts genetic information into protein and usually must be processed to serve its function. RNA types, chemical structure, protein synthesis, translation, manufacture, and processing are discussed. Concludes that the first genes might have been spliced RNA and that humans might be closer than bacteria to primitive…

  5. In situ, real-time catabolic gene expression: Extraction and characterization of naphthalene dioxygenase mRNA transcripts from groundwater

    International Nuclear Information System (INIS)

    Wilson, M.S.; Bakermans, C.; Madsen, E.L.

    1999-01-01

    The authors developed procedures for isolating and characterizing in situ-transcribed mRNA from groundwater microorganisms catabolizing naphthalene at a coal tar waste-contaminated site. Groundwater was pumped through 0.22-microm-pore-size filters, which were then frozen to dry ice-ethanol. RNA was extracted from the frozen filters by boiling sodium dodecyl sulfate lysis and acidic phenol-chloroform extraction. Transcript characterization was performed with a series of PCR primers designed to amplify nahAc homologs. Several primer pairs were found to amplify nahAc homologs representing the entire diversity of the naphthalene-degrading genes. The environmental RNA extract was reverse transcribed, and the resultant mixture of cDNAs was amplified by PCR. A digoxigenin-labeled probe mixture was produced by PCR amplification of groundwater cDNA. This probe mixture hybridized under stringent conditions with the corresponding PCR products from naphthalene-degrading bacteria carrying a variety of nahAc homologs, indicating that diverse dioxygenase transcripts had been retrieved from groundwater. Diluted and undiluted cDNA preparations were independently amplified, and 28 of the resulting PCR products were cloned and sequenced. Sequence comparisons revealed two major groups related to the dioxygenase genes ndoB and dntAc, previously cloned from Pseudomonas putida NCIB 9816-4 and Burkholderia sp. strain DNT, respectively. A distinctive subgroup of sequences was found only in experiments performed with the undiluted cDNA preparation. To the authors' knowledge, these results are the first to directly document in situ transcription of genes encoding naphthalene catabolism at a contaminated site by indigenous microorganisms. The retrieved sequences represent greater diversity than has been detected at the study site by culture-based approaches

  6. nRC: non-coding RNA Classifier based on structural features.

    Science.gov (United States)

    Fiannaca, Antonino; La Rosa, Massimo; La Paglia, Laura; Rizzo, Riccardo; Urso, Alfonso

    2017-01-01

    Non-coding RNA (ncRNA) are small non-coding sequences involved in gene expression regulation of many biological processes and diseases. The recent discovery of a large set of different ncRNAs with biologically relevant roles has opened the way to develop methods able to discriminate between the different ncRNA classes. Moreover, the lack of knowledge about the complete mechanisms in regulative processes, together with the development of high-throughput technologies, has required the help of bioinformatics tools in addressing biologists and clinicians with a deeper comprehension of the functional roles of ncRNAs. In this work, we introduce a new ncRNA classification tool, nRC (non-coding RNA Classifier). Our approach is based on features extraction from the ncRNA secondary structure together with a supervised classification algorithm implementing a deep learning architecture based on convolutional neural networks. We tested our approach for the classification of 13 different ncRNA classes. We obtained classification scores, using the most common statistical measures. In particular, we reach an accuracy and sensitivity score of about 74%. The proposed method outperforms other similar classification methods based on secondary structure features and machine learning algorithms, including the RNAcon tool that, to date, is the reference classifier. nRC tool is freely available as a docker image at https://hub.docker.com/r/tblab/nrc/. The source code of nRC tool is also available at https://github.com/IcarPA-TBlab/nrc.

  7. General enumeration of RNA secondary structures based on new ...

    African Journals Online (AJOL)

    akpobome

    coding, transferring and retrieving genetic information, and in directing cell metabolism. The nucleic acid includes DNA and RNA molecule. RNA molecule is a single-stranded nucleic acid of four different kinds of nucleotides. The four nucleotides only differ by one part, called bases. Hence, one usually identifies nucleotides.

  8. Molecular Beacon-Based MicroRNA Imaging During Neurogenesis.

    Science.gov (United States)

    Lee, Jonghwan; Kim, Soonhag

    2016-01-01

    The fluorescence monitoring system for examining endogenous microRNA (miRNA) activity in cellular level provides crucial information on not only understanding a critical role of miRNA involving a variety of biological processes, but also evaluating miRNA expression patterns in a noninvasive manner. In this protocol, we report the details of a new procedure for a molecular beacon-based miRNA monitoring system, which includes the illustration scheme for miRNA detection strategy, exogenous miRNA detection, and measurement of endogenous miRNA expression level during neurogenesis. The fluorescence signal of miR-124a beacon quenched by BHQ2 was gradually recovered as increasing concentration of the miR-124a in tube. The functional work of miR-124a beacon was examined in intracellular environment, allowing for the internalization of the miR-124a beacon by lipofectamine, which resulted in activated fluorescent signals of the miR-124a beacon in the HeLa cells after the addition of synthetic miR-124a. The endogenous miR-124a expression level was detected by miR-124a beacon system during neurogenesis, showing brighter fluorescence intensity in cytoplasmic area of P19 cells after induction of neuronal differentiation by retinoic acid. The molecular beacon based-miRNA detection technique could be applicable to the simultaneous visualization of a variety of miRNA expression patterns using different fluorescence dyes. For the study of examining endogenous miRNA expression level using miRNA-beacon system, if cellular differentiation step is already prepared, transfection step of miR-124a beacon into P19 cells, and acquisition of activated fluorescence signal measured by confocal microscope can be conducted approximately within 6 h.

  9. Data Extraction Based on Page Structure Analysis

    Directory of Open Access Journals (Sweden)

    Ren Yichao

    2017-01-01

    Full Text Available The information we need has some confusing problems such as dispersion and different organizational structure. In addition, because of the existence of unstructured data like natural language and images, extracting local content pages is extremely difficult. In the light of of the problems above, this article will apply a method combined with page structure analysis algorithm and page data extraction algorithm to accomplish the gathering of network data. In this way, the problem that traditional complex extraction model behave poorly when dealing with large-scale data is perfectly solved and the page data extraction efficiency is also boosted to a new level. In the meantime, the article will also make a comparison about pages and content of different types between the methods of DOM structure based on the page and HTML regularities of distribution. After all of those, we may find a more efficient extract method.

  10. A simple and robust vector-based shRNA expression system used for RNA interference.

    Science.gov (United States)

    Wang, Xue-jun; Li, Ying; Huang, Hai; Zhang, Xiu-juan; Xie, Pei-wen; Hu, Wei; Li, Dan-dan; Wang, Sheng-qi

    2013-01-01

    RNA interference (RNAi) mediated by small interfering RNAs (siRNAs) or short hairpin RNAs (shRNAs) has become a powerful genetic tool for conducting functional studies. Previously, vector-based shRNA-expression strategies capable of inducing RNAi in viable cells have been developed, however, these vector systems have some disadvantages, either because they were error-prone or cost prohibitive. In this report we described the development of a simple, robust shRNA expression system utilizing 1 long oligonucleotide or 2 short oligonucleotides for half the cost of conventional shRNA construction methods and with a >95% cloning success rate. The shRNA loop sequence and stem structure were also compared and carefully selected for better RNAi efficiency. Furthermore, an easier strategy was developed based on isocaudomers which permit rapid combination of the most efficient promoter-shRNA cassettes. Finally, using this method, the conservative target sites for hepatitis B virus (HBV) knockdown were systemically screened and HBV antigen expression shown to be successfully suppressed in the presence of connected multiple shRNAs both in vitro and in vivo. This novel design describes an inexpensive and effective way to clone and express single or multiple shRNAs from the same vector with the capacity for potent and effective silencing of target genes.

  11. A simple and robust vector-based shRNA expression system used for RNA interference.

    Directory of Open Access Journals (Sweden)

    Xue-jun Wang

    Full Text Available BACKGROUND: RNA interference (RNAi mediated by small interfering RNAs (siRNAs or short hairpin RNAs (shRNAs has become a powerful genetic tool for conducting functional studies. Previously, vector-based shRNA-expression strategies capable of inducing RNAi in viable cells have been developed, however, these vector systems have some disadvantages, either because they were error-prone or cost prohibitive. RESULTS: In this report we described the development of a simple, robust shRNA expression system utilizing 1 long oligonucleotide or 2 short oligonucleotides for half the cost of conventional shRNA construction methods and with a >95% cloning success rate. The shRNA loop sequence and stem structure were also compared and carefully selected for better RNAi efficiency. Furthermore, an easier strategy was developed based on isocaudomers which permit rapid combination of the most efficient promoter-shRNA cassettes. Finally, using this method, the conservative target sites for hepatitis B virus (HBV knockdown were systemically screened and HBV antigen expression shown to be successfully suppressed in the presence of connected multiple shRNAs both in vitro and in vivo. CONCLUSION: This novel design describes an inexpensive and effective way to clone and express single or multiple shRNAs from the same vector with the capacity for potent and effective silencing of target genes.

  12. Paper-Based MicroRNA Expression Profiling from Plasma and Circulating Tumor Cells.

    Science.gov (United States)

    Leong, Sai Mun; Tan, Karen Mei-Ling; Chua, Hui Wen; Huang, Mo-Chao; Cheong, Wai Chye; Li, Mo-Huang; Tucker, Steven; Koay, Evelyn Siew-Chuan

    2017-03-01

    Molecular characterization of circulating tumor cells (CTCs) holds great promise for monitoring metastatic progression and characterizing metastatic disease. However, leukocyte and red blood cell contamination of routinely isolated CTCs makes CTC-specific molecular characterization extremely challenging. Here we report the use of a paper-based medium for efficient extraction of microRNAs (miRNAs) from limited amounts of biological samples such as rare CTCs harvested from cancer patient blood. Specifically, we devised a workflow involving the use of Flinders Technology Associates (FTA) ® Elute Card with a digital PCR-inspired "partitioning" method to extract and purify miRNAs from plasma and CTCs. We demonstrated the sensitivity of this method to detect miRNA expression from as few as 3 cancer cells spiked into human blood. Using this method, background miRNA expression was excluded from contaminating blood cells, and CTC-specific miRNA expression profiles were derived from breast and colorectal cancer patients. Plasma separated out during purification of CTCs could likewise be processed using the same paper-based method for miRNA detection, thereby maximizing the amount of patient-specific information that can be derived from a single blood draw. Overall, this paper-based extraction method enables an efficient, cost-effective workflow for maximized recovery of small RNAs from limited biological samples for downstream molecular analyses. © 2016 American Association for Clinical Chemistry.

  13. Ensemble-based prediction of RNA secondary structures.

    Science.gov (United States)

    Aghaeepour, Nima; Hoos, Holger H

    2013-04-24

    Accurate structure prediction methods play an important role for the understanding of RNA function. Energy-based, pseudoknot-free secondary structure prediction is one of the most widely used and versatile approaches, and improved methods for this task have received much attention over the past five years. Despite the impressive progress that as been achieved in this area, existing evaluations of the prediction accuracy achieved by various algorithms do not provide a comprehensive, statistically sound assessment. Furthermore, while there is increasing evidence that no prediction algorithm consistently outperforms all others, no work has been done to exploit the complementary strengths of multiple approaches. In this work, we present two contributions to the area of RNA secondary structure prediction. Firstly, we use state-of-the-art, resampling-based statistical methods together with a previously published and increasingly widely used dataset of high-quality RNA structures to conduct a comprehensive evaluation of existing RNA secondary structure prediction procedures. The results from this evaluation clarify the performance relationship between ten well-known existing energy-based pseudoknot-free RNA secondary structure prediction methods and clearly demonstrate the progress that has been achieved in recent years. Secondly, we introduce AveRNA, a generic and powerful method for combining a set of existing secondary structure prediction procedures into an ensemble-based method that achieves significantly higher prediction accuracies than obtained from any of its component procedures. Our new, ensemble-based method, AveRNA, improves the state of the art for energy-based, pseudoknot-free RNA secondary structure prediction by exploiting the complementary strengths of multiple existing prediction procedures, as demonstrated using a state-of-the-art statistical resampling approach. In addition, AveRNA allows an intuitive and effective control of the trade-off between

  14. Polyethylene glycol and polyvinylpirrolidone effect on bacterial rRNA extraction and hybridization from cells exposed to tannins.

    OpenAIRE

    ARCURI, P.B.; THONNEY, M.L.; SCHOFIELD, P.; PELL, A.N.

    2003-01-01

    In order to detect fluctuations in ruminal microbial populations due to forage tannins using 16S ribosomal RNA (rRNA) probes, recovery of intact rRNA is required. The objective of this work was to evaluate the effect of polyethylene glycol (PEG) and polyvinylpirrolidone (PVP) on extraction of bacterial rRNA, in the presence of tannins from tropical legume forages and other sources, that hybridize with oligonucleotide probes. Ruminococcus albus 8 cells were exposed to 8 g/L tannic acid or 1 g/...

  15. Refining Automatically Extracted Knowledge Bases Using Crowdsourcing

    OpenAIRE

    Li, Chunhua; Zhao, Pengpeng; Sheng, Victor S.; Xian, Xuefeng; Wu, Jian; Cui, Zhiming

    2017-01-01

    Machine-constructed knowledge bases often contain noisy and inaccurate facts. There exists significant work in developing automated algorithms for knowledge base refinement. Automated approaches improve the quality of knowledge bases but are far from perfect. In this paper, we leverage crowdsourcing to improve the quality of automatically extracted knowledge bases. As human labelling is costly, an important research challenge is how we can use limited human resources to maximize the quality i...

  16. Cytochrome c oxidase subunit 1-based human RNA quantification to enhance mRNA profiling in forensic biology

    Directory of Open Access Journals (Sweden)

    Dong Zhao

    2017-01-01

    Full Text Available RNA analysis offers many potential applications in forensic science, and molecular identification of body fluids by analysis of cell-specific RNA markers represents a new technique for use in forensic cases. However, due to the nature of forensic materials that often admixed with nonhuman cellular components, human-specific RNA quantification is required for the forensic RNA assays. Quantification assay for human RNA has been developed in the present study with respect to body fluid samples in forensic biology. The quantitative assay is based on real-time reverse transcription-polymerase chain reaction of mitochondrial RNA cytochrome c oxidase subunit I and capable of RNA quantification with high reproducibility and a wide dynamic range. The human RNA quantification improves the quality of mRNA profiling in the identification of body fluids of saliva and semen because the quantification assay can exclude the influence of nonhuman components and reduce the adverse affection from degraded RNA fragments.

  17. Evaluation of different pulverisation methods for RNA extraction in squash fruit: lyophilisation, cryogenic mill and mortar grinding.

    Science.gov (United States)

    Román, Belén; González-Verdejo, Clara I; Peña, Francisco; Nadal, Salvador; Gómez, Pedro

    2012-01-01

    Quality and integrity of RNA are critical for transcription studies in plant molecular biology. In squash fruit and other high water content crops, the grinding of tissue with mortar and pestle in liquid nitrogen fails to produce a homogeneous and fine powered sample desirable to ensure a good penetration of the extraction reagent. To develop an improved pulverisation method to facilitate the homogenisation process of squash fruit tissue prior to RNA extraction without reducing quality and yield of the extracted RNA. Three methods of pulverisation, each followed by the same extraction protocol, were compared. The first approach consisted of the lyophilisation of the sample in order to remove the excess of water before grinding, the second one used a cryogenic mill and the control one a mortar grinding of frozen tissue. The quality of the isolated RNA was tested by carrying out a quantitative real time downstream amplification. In the three situations considered, mean values for A(260) /A(280) indicated minimal interference by proteins and RNA quality indicator (RQI) values were considered appropriate for quantitative real-time polymerase chain reaction (qRT-PCR) amplification. Successful qRT-PCR amplifications were obtained with cDNA isolated with the three protocols. Both apparatus can improve and facilitate the grinding step in the RNA extraction process in zucchini, resulting in isolated RNA of high quality and integrity as revealed by qRT-PCR downstream application. This is apparently the first time that a cryogenic mill has been used to prepare fruit samples for RNA extraction, thereby improving the sampling strategy because the fine powder obtained represents a homogeneous mix of the organ tissue. Copyright © 2012 John Wiley & Sons, Ltd.

  18. A novel method for RNA extraction from FFPE samples reveals significant differences in biomarker expression between orthotopic and subcutaneous pancreatic cancer patient-derived xenografts.

    Science.gov (United States)

    Hoover, Malachia; Adamian, Yvess; Brown, Mark; Maawy, Ali; Chang, Alexander; Lee, Jacqueline; Gharibi, Armen; Katz, Matthew H; Fleming, Jason; Hoffman, Robert M; Bouvet, Michael; Doebler, Robert; Kelber, Jonathan A

    2017-01-24

    Next-generation sequencing (NGS) can identify and validate new biomarkers of cancer onset, progression and therapy resistance. Substantial archives of formalin-fixed, paraffin-embedded (FFPE) cancer samples from patients represent a rich resource for linking molecular signatures to clinical data. However, performing NGS on FFPE samples is limited by poor RNA purification methods. To address this hurdle, we developed an improved methodology for extracting high-quality RNA from FFPE samples. By briefly integrating a newly-designed micro-homogenizing (mH) tool with commercially available FFPE RNA extraction protocols, RNA recovery is increased by approximately 3-fold while maintaining standard A260/A280 ratios and RNA quality index (RQI) values. Furthermore, we demonstrate that the mH-purified FFPE RNAs are longer and of higher integrity. Previous studies have suggested that pancreatic ductal adenocarcinoma (PDAC) gene expression signatures vary significantly under in vitro versus in vivo and in vivo subcutaneous versus orthotopic conditions. By using our improved mH-based method, we were able to preserve established expression patterns of KRas-dependency genes within these three unique microenvironments. Finally, expression analysis of novel biomarkers in KRas mutant PDAC samples revealed that PEAK1 decreases and MST1R increases by over 100-fold in orthotopic versus subcutaneous microenvironments. Interestingly, however, only PEAK1 levels remain elevated in orthotopically grown KRas wild-type PDAC cells. These results demonstrate the critical nature of the orthotopic tumor microenvironment when evaluating the clinical relevance of new biomarkers in cells or patient-derived samples. Furthermore, this new mH-based FFPE RNA extraction method has the potential to enhance and expand future FFPE-RNA-NGS cancer biomarker studies.

  19. Reconstruction and analysis of the lncRNA-miRNA-mRNA network based on competitive endogenous RNA reveal functional lncRNAs in rheumatoid arthritis.

    Science.gov (United States)

    Jiang, Hui; Ma, Rong; Zou, Shubiao; Wang, Yongzhong; Li, Zhuqing; Li, Weiping

    2017-06-01

    Rheumatoid arthritis (RA) is an autoimmune disease with an unknown etiology, occurring in approximately 1.0% of general population. More and more studies have suggested that long non-coding RNAs (lncRNAs) could play important roles in various biological processes and be associated with the pathogenesis of different kinds of diseases including RA. Although a large number of lncRNAs have been found, our knowledge of their function and physiological/pathological significance is still in its infancy. In order to reveal functional lncRNAs and identify the key lncRNAs in RA, we reconstructed a global triple network based on the competitive endogenous RNA (ceRNA) theory using the data from National Center for Biotechnology Information Gene Expression Omnibus and our previous paper. Meanwhile, Gene Ontology (GO) and pathway analysis were performed using Cytoscape plug-in BinGO and Database for Annotation, Visualization, and Integration Discovery (DAVID), respectively. We found that the lncRNA-miRNA-mRNA network was composed of 7 lncRNA nodes, 90 mRNA nodes, 24 miRNA nodes, and 301 edges. The functional assay showed that 147 GO terms and 23 pathways were enriched. In addition, three lncRNAs (S5645.1, XR_006437.1, J01878) were highly related to RA, and therefore, were selected as key lncRNAs. This study suggests that specific lncRNAs are associated with the development of RA, and three lncRNAs (S5645.1, XR_006437.1, J01878) could be used as potential diagnostic biomarkers and therapeutic targets.

  20. Sample preparation for avian and porcine influenza virus cDNA amplification simplified: Boilign vs. conventional RNA extraction

    International Nuclear Information System (INIS)

    Fereidouni, S.R.; Starick, E.; Ziller, M.; Harder, T.C.; Unger, H.; Hamilton, K.; Globig, A.

    2016-01-01

    Full text: RNA extraction and purification is a fundamental step that allows for highly sensitive amplification of specific RNA targets in PCR applications. However, commercial extraction kits that are broadly used because of their robustness and high yield of purified RNA are expensive and labor-intensive. In this study, boiling in distilled water or a commerical lysis buffer of different sample matrices containing avian or porcine influenza viruses was tested as an alternative. Real-time PCR (RTaPCR) for nucleoprotein gene fragment was used as read out. Results were compared with freshly extracted RNA by use of a commercial extraction kit. Different batches of virus containing material materials, including diluted virus positive allontoic fluid or cell culture supernatnat, and avian faecal, cloacal or oropharyngeal swab samples were used in this study. Simple boiling of samples without any additional purification steps can be used as an alternative RNA preparation method to detect influenza A virus nucleoprotein RNA in oropharyngeal swab samples, allantoic fluid or cell-culture supernatant. The boiling method is not applicable for sample matrices containing faecal material. (author)

  1. Digestion of chrysanthemum stunt viroid by leaf extracts of Capsicum chinense indicates strong RNA-digesting activity.

    Science.gov (United States)

    Iraklis, Boubourakas; Kanda, Hiroko; Nabeshima, Tomoyuki; Onda, Mayu; Ota, Nao; Koeda, Sota; Hosokawa, Munetaka

    2016-08-01

    CSVd could not infect Nicotiana benthamiana when the plants were pretreated with crude leaf extract of Capsicum chinense 'Sy-2'. C. chinense leaves were revealed to contain strong RNA-digesting activity. Several studies have identified active antiviral and antiviroid agents in plants. Capsicum plants are known to contain antiviral agents, but the mechanism of their activity has not been determined. We aimed to elucidate the mechanism of Capsicum extract's antiviroid activity. Chrysanthemum stunt viroid (CSVd) was inoculated into Nicotiana benthamiana plants before or after treating the plants with a leaf extract of Capsicum chinense 'Sy-2'. CSVd infection was determined using quantitative reverse transcription-polymerase chain reaction (qRT-PCR) 3 weeks after inoculation. When Capsicum extract was sprayed or painted onto N. benthamiana before inoculation, it was effective in preventing infection by CSVd. To evaluate CSVd digestion activity in leaf extracts, CSVd was mixed with leaf extracts of Mirabilis, Phytolacca, Pelargonium and Capsicum. CSVd-digesting activities were examined by quantifying undigested CSVd using qRT-PCR, and RNA gel blotting permitted visualization of the digested CSVd. Only Capsicum leaf extract digested CSVd, and in the Capsicum treatment, small digested CSVd products were detected by RNA gel blot analysis. When the digesting experiment was performed for various cultivars and species of Capsicum, only cultivars of C. chinense showed strong CSVd-digesting activity. Our observations indicated that Capsicum extract contains strong RNA-digesting activity, leading to the conclusion that this activity is the main mechanism for protection from infection by CSVd through spraying or painting before inoculation. To our knowledge, this is the first report of a strong RNA-digesting activity by a plant extract.

  2. Identification of the miRNA-mRNA regulatory network of small cell osteosarcoma based on RNA-seq.

    Science.gov (United States)

    Xie, Lin; Liao, Yedan; Shen, Lida; Hu, Fengdi; Yu, Sunlin; Zhou, Yonghong; Zhang, Ya; Yang, Yihao; Li, Dongqi; Ren, Minyan; Yuan, Zhongqin; Yang, Zuozhang

    2017-06-27

    Small cell osteosarcoma (SCO) is a rare subtype of osteosarcoma characterized by highly aggressive progression and a poor prognosis. The miRNA and mRNA expression profiles of peripheral blood mononuclear cells (PBMCs) were obtained in 3 patients with SCO and 10 healthy individuals using high-throughput RNA-sequencing. We identified 37 dysregulated miRNAs and 1636 dysregulated mRNAs in patients with SCO compared to the healthy controls. Specifically, the 37 dysregulated miRNAs consisted of 27 up-regulated miRNAs and 10 down-regulated miRNAs; the 1636 dysregulated mRNAs consisted of 555 up-regulated mRNAs and 1081 down-regulated mRNAs. The target-genes of miRNAs were predicted, and 1334 negative correlations between miRNAs and mRNAs were used to construct an miRNA-mRNA regulatory network. Dysregulated genes were significantly enriched in pathways related to cancer, mTOR signaling and cell cycle signaling. Specifically, hsa-miR-26b-5p, hsa-miR-221-3p and hsa-miR-125b-2-3p were significantly dysregulated miRNAs and exhibited a high degree of connectivity with target genes. Overall, the expression of dysregulated genes in tumor tissues and peripheral blood samples of patients with SCO measured by quantitative real-time polymerase chain reaction corroborated with our bioinformatics analyses based on the expression profiles of PBMCs from patients with SCO. Thus, hsa-miR-26b-5p, hsa-miR-221-3p and hsa-miR-125b-2-3p may be involved in SCO tumorigenesis.

  3. A path-based measurement for human miRNA functional similarities using miRNA-disease associations

    Science.gov (United States)

    Ding, Pingjian; Luo, Jiawei; Xiao, Qiu; Chen, Xiangtao

    2016-09-01

    Compared with the sequence and expression similarity, miRNA functional similarity is so important for biology researches and many applications such as miRNA clustering, miRNA function prediction, miRNA synergism identification and disease miRNA prioritization. However, the existing methods always utilized the predicted miRNA target which has high false positive and false negative to calculate the miRNA functional similarity. Meanwhile, it is difficult to achieve high reliability of miRNA functional similarity with miRNA-disease associations. Therefore, it is increasingly needed to improve the measurement of miRNA functional similarity. In this study, we develop a novel path-based calculation method of miRNA functional similarity based on miRNA-disease associations, called MFSP. Compared with other methods, our method obtains higher average functional similarity of intra-family and intra-cluster selected groups. Meanwhile, the lower average functional similarity of inter-family and inter-cluster miRNA pair is obtained. In addition, the smaller p-value is achieved, while applying Wilcoxon rank-sum test and Kruskal-Wallis test to different miRNA groups. The relationship between miRNA functional similarity and other information sources is exhibited. Furthermore, the constructed miRNA functional network based on MFSP is a scale-free and small-world network. Moreover, the higher AUC for miRNA-disease prediction indicates the ability of MFSP uncovering miRNA functional similarity.

  4. Simultaneous DNA-RNA Extraction from Coastal Sediments and Quantification of 16S rRNA Genes and Transcripts by Real-time PCR.

    Science.gov (United States)

    Tatti, Enrico; McKew, Boyd A; Whitby, Corrine; Smith, Cindy J

    2016-06-11

    Real Time Polymerase Chain Reaction also known as quantitative PCR (q-PCR) is a widely used tool in microbial ecology to quantify gene abundances of taxonomic and functional groups in environmental samples. Used in combination with a reverse transcriptase reaction (RT-q-PCR), it can also be employed to quantify gene transcripts. q-PCR makes use of highly sensitive fluorescent detection chemistries that allow quantification of PCR amplicons during the exponential phase of the reaction. Therefore, the biases associated with 'end-point' PCR detected in the plateau phase of the PCR reaction are avoided. A protocol to quantify bacterial 16S rRNA genes and transcripts from coastal sediments via real-time PCR is provided. First, a method for the co-extraction of DNA and RNA from coastal sediments, including the additional steps required for the preparation of DNA-free RNA, is outlined. Second, a step-by-step guide for the quantification of 16S rRNA genes and transcripts from the extracted nucleic acids via q-PCR and RT-q-PCR is outlined. This includes details for the construction of DNA and RNA standard curves. Key considerations for the use of RT-q-PCR assays in microbial ecology are included.

  5. In vitro base modification of Escherichia coli glutamate 2 transfer-RNA and phenylalanine transfer-RNA gene transcripts

    International Nuclear Information System (INIS)

    Shahan, M.N.

    1989-01-01

    Plasmids were constructed that contain an E. Coli tRNA 2 Glu or tRNA Phe gene in a system transcribable by T7 or SP6 RNA polymerase. Selectively 32 P-labeled transcripts of these plasmids were used to study tRNA base modification in vitro in crude extracts by nearest neighbor analysis. The synthesis of 5-methyl-aminomethyl-2-thiouridine (mnm 5 s 2 U) was studied. Complete synthesis of mnm 5 s 2 2U is not observed. Instead, 2-thiouridine (s 2 U) is synthesized. Synthesis requires ATP, cysteine, Mg + , and monovalent cation concentrations below 50 mM. The reaction has a pH optimum above 7.0. Sulfide ion will substitute for cysteine in the reaction but sulfate, sulfite, methionine, homocysteine, and β-mercaptopyruvate will not. Extracts from E. coli strains carrying either the asuE or asuF mutations have reduced s 2 U synthetic activity which supports in vivo evidence that the wild type genes are involved in 2-thiolation of uridine. The enzyme is shown to be unstable both upon storage at -80 degree C and during the modification reaction. A method was developed to study the synthesis of any one of four pseudouridines ψ found at different positions of the tRNA cloverleaf. Synthesis of ψ is observed at three of the four positions-positions 32, 39, and 55. The asuC mutation is shown to affect ψ synthesis only at position 39 confirming that it is an allele of hisT and that the hisT mutations do not affect ψ synthesis at position 32 in E. coli. Synthesis of ψ32, ψ39, and ψ55 does not require any prior modification. Synthesis of dihydrouridine, 7-methylguanosine, and 3(3-amino-3-carboxypropyl)uridine is also observed. Synthesis of 2-methyladenosine and ψ 13 is not seen. Removal of part of the aminoacyl stem reduces synthesis of all modifications examined by 3' fold or more

  6. Cryptographic Protocols Based on Root Extracting

    DEFF Research Database (Denmark)

    Koprowski, Maciej

    In this thesis we design new cryptographic protocols, whose security is based on the hardness of root extracting or more speci cally the RSA problem. First we study the problem of root extraction in nite Abelian groups, where the group order is unknown. This is a natural generalization of the...... complexity of root extraction, even if the algorithm can choose the "public exponent'' itself. In other words, both the standard and the strong RSA assumption are provably true w.r.t. generic algorithms. The results hold for arbitrary groups, so security w.r.t. generic attacks follows for any cryptographic...... groups. In all cases, security follows from a well de ned complexity assumption (the strong root assumption), without relying on random oracles. A smooth natural number has no big prime factors. The probability, that a random natural number not greater than x has all prime factors smaller than x1/u...

  7. Feasibility of RNA and DNA Extraction from Fresh Pipelle and Archival Endometrial Tissues for Use in Gene Expression and SNP Arrays

    Directory of Open Access Journals (Sweden)

    Heather D. Kissel

    2013-01-01

    Full Text Available Identifying molecular markers of endometrial hyperplasia (neoplasia progression is critical to cancer prevention. To assess RNA and DNA quantity and quality from routinely collected endometrial samples and evaluate the performance of RNA- and DNA-based arrays across endometrial tissue types, we collected fresh frozen (FF Pipelle, FF curettage, and formalin-fixed paraffin-embedded (FFPE hysterectomy specimens (benign indications from eight women. Additionally, neoplastic and uninvolved tissues from 24 FFPE archival hysterectomy specimens with endometrial hyperplasias and carcinomas were assessed. RNA was extracted from 15 of 16 FF and 51 of 51 FFPE samples, with yields >1.2 μg for 13/15 (87% FF and 50/51 (98% FFPE samples. Extracted RNA was of high quality; all samples performed successfully on the Illumina whole-genome cDNA-mediated annealing, selection, extension, and ligation (WG-DASL array and performance did not vary by tissue type. While DNA quantity from FFPE samples was excellent, quality was not sufficient for successful performance on the Affymetrix SNP Array 6.0. In conclusion, FF Pipelle samples, which are minimally invasive, yielded excellent quantity and quality of RNA for gene expression arrays (similar to FF curettage and should be considered for use in genomic studies. FFPE-derived DNA should be evaluated on new rapidly evolving sequencing platforms.

  8. Protocol: high throughput silica-based purification of RNA from Arabidopsis seedlings in a 96-well format

    Directory of Open Access Journals (Sweden)

    Salvo-Chirnside Eliane

    2011-12-01

    Full Text Available Abstract The increasing popularity of systems-based approaches to plant research has resulted in a demand for high throughput (HTP methods to be developed. RNA extraction from multiple samples in an experiment is a significant bottleneck in performing systems-level genomic studies. Therefore we have established a high throughput method of RNA extraction from Arabidopsis thaliana to facilitate gene expression studies in this widely used plant model. We present optimised manual and automated protocols for the extraction of total RNA from 9-day-old Arabidopsis seedlings in a 96 well plate format using silica membrane-based methodology. Consistent and reproducible yields of high quality RNA are isolated averaging 8.9 μg total RNA per sample (~20 mg plant tissue. The purified RNA is suitable for subsequent qPCR analysis of the expression of over 500 genes in triplicate from each sample. Using the automated procedure, 192 samples (2 × 96 well plates can easily be fully processed (samples homogenised, RNA purified and quantified in less than half a day. Additionally we demonstrate that plant samples can be stored in RNAlater at -20°C (but not 4°C for 10 months prior to extraction with no significant effect on RNA yield or quality. Additionally, disrupted samples can be stored in the lysis buffer at -20°C for at least 6 months prior to completion of the extraction procedure providing a flexible sampling and storage scheme to facilitate complex time series experiments.

  9. SPIDIA-RNA: second external quality assessment for the pre-analytical phase of blood samples used for RNA based analyses.

    Directory of Open Access Journals (Sweden)

    Francesca Malentacchi

    Full Text Available One purpose of the EC funded project, SPIDIA, is to develop evidence-based quality guidelines for the pre-analytical handling of blood samples for RNA molecular testing. To this end, two pan-European External Quality Assessments (EQAs were implemented. Here we report the results of the second SPIDIA-RNA EQA. This second study included modifications in the protocol related to the blood collection process, the shipping conditions and pre-analytical specimen handling for participants. Participating laboratories received two identical proficiency blood specimens collected in tubes with or without an RNA stabilizer. For pre-defined specimen storage times and temperatures, laboratories were asked to perform RNA extraction from whole blood according to their usual procedure and to return extracted RNA to the SPIDIA facility for further analysis. These RNA samples were evaluated for purity, yield, integrity, stability, presence of interfering substances, and gene expression levels for the validated markers of RNA stability: FOS, IL1B, IL8, GAPDH, FOSB and TNFRSF10c. Analysis of the gene expression results of FOS, IL8, FOSB, and TNFRSF10c, however, indicated that the levels of these transcripts were significantly affected by blood collection tube type and storage temperature. These results demonstrated that only blood collection tubes containing a cellular RNA stabilizer allowed reliable gene expression analysis within 48 h from blood collection for all the genes investigated. The results of these two EQAs have been proposed for use in the development of a Technical Specification by the European Committee for Standardization.

  10. Size and Base Composition of RNA in Supercoiled Plasmid DNA

    Science.gov (United States)

    Williams, Peter H.; Boyer, Herbert W.; Helinski, Donald R.

    1973-01-01

    The average size and base composition of the covalently integrated RNA segment in supercoiled ColE1 DNA synthesized in Escherichia coli in the presence of chloramphenicol (CM-ColE1 DNA) have been determined by two independent methods. The two approaches yielded similar results, indicating that the RNA segment in CM-ColE1 DNA contains GMP at the 5′ end and comprises on the average 25 to 26 ribonucleotides with a base composition of 10-11 G, 3 A, 5-6 C, and 6-7 U. PMID:4359488

  11. Automated, simple, and efficient influenza RNA extraction from clinical respiratory swabs using TruTip and epMotion.

    Science.gov (United States)

    Griesemer, Sara B; Holmberg, Rebecca; Cooney, Christopher G; Thakore, Nitu; Gindlesperger, Alissa; Knickerbocker, Christopher; Chandler, Darrell P; St George, Kirsten

    2013-09-01

    Rapid, simple and efficient influenza RNA purification from clinical samples is essential for sensitive molecular detection of influenza infection. Automation of the TruTip extraction method can increase sample throughput while maintaining performance. To automate TruTip influenza RNA extraction using an Eppendorf epMotion robotic liquid handler, and to compare its performance to the bioMerieux easyMAG and Qiagen QIAcube instruments. Extraction efficacy and reproducibility of the automated TruTip/epMotion protocol was assessed from influenza-negative respiratory samples spiked with influenza A and B viruses. Clinical extraction performance from 170 influenza A and B-positive respiratory swabs was also evaluated and compared using influenza A and B real-time RT-PCR assays. TruTip/epMotion extraction efficacy was 100% in influenza virus-spiked samples with at least 745 influenza A and 370 influenza B input gene copies per extraction, and exhibited high reproducibility over four log10 concentrations of virus (extraction were also positive following TruTip extraction. Overall Ct value differences obtained between TruTip/epMotion and easyMAG/QIAcube clinical extracts ranged from 1.24 to 1.91. Pairwise comparisons of Ct values showed a high correlation of the TruTip/epMotion protocol to the other methods (R2>0.90). The automated TruTip/epMotion protocol is a simple and rapid extraction method that reproducibly purifies influenza RNA from respiratory swabs, with comparable efficacy and efficiency to both the easyMAG and QIAcube instruments. Copyright © 2013 Elsevier B.V. All rights reserved.

  12. RNA-Based Assessment of Diversity and Composition of Active Archaeal Communities in the German Bight

    Directory of Open Access Journals (Sweden)

    Bernd Wemheuer

    2012-01-01

    Full Text Available Archaea play an important role in various biogeochemical cycles. They are known extremophiles inhabiting environments such as thermal springs or hydrothermal vents. Recent studies have revealed a significant abundance of Archaea in moderate environments, for example, temperate sea water. Nevertheless, the composition and ecosystem function of these marine archaeal communities is largely unknown. To assess diversity and composition of active archaeal communities in the German Bight, seven marine water samples were taken and studied by RNA-based analysis of ribosomal 16S rRNA. For this purpose, total RNA was extracted from the samples and converted to cDNA. Archaeal community structures were investigated by pyrosequencing-based analysis of 16S rRNA amplicons generated from cDNA. To our knowledge, this is the first study combining next-generation sequencing and metatranscriptomics to study archaeal communities in marine habitats. The pyrosequencing-derived dataset comprised 62,045 archaeal 16S rRNA sequences. We identified Halobacteria as the predominant archaeal group across all samples with increased abundance in algal blooms. Thermoplasmatales (Euryarchaeota and the Marine Group I (Thaumarchaeota were identified in minor abundances. It is indicated that archaeal community patterns were influenced by environmental conditions.

  13. Aqueous biphasic systems containing PEG-based deep eutectic solvents for high-performance partitioning of RNA.

    Science.gov (United States)

    Zhang, Hongmei; Wang, Yuzhi; Zhou, Yigang; Xu, Kaijia; Li, Na; Wen, Qian; Yang, Qin

    2017-08-01

    In this work, 16 kinds of novel deep eutectic solvents (DESs) composed of polyethylene glycol (PEG) and quaternary ammonium salts, were coupled with Aqueous Biphasic Systems (ABSs) to extract RNA. The phase forming ability of ABSs were comprehensively evaluated, involving the effects of various proportions of DESs' components, carbon chain length and anions species of quaternary ammonium salts, average molecular weights of PEG and inorganic salts nature. Then the systems were applied in RNA extraction, and the results revealed that the extraction efficiency values were distinctly enhanced by relatively lower PEG content in DESs, smaller PEG molecular weights, longer carbon chain of quaternary ammonium salts and more hydrophobic inorganic salts. Then the systems composed of [TBAB][PEG600] and Na 2 SO 4 were utilized in the influence factor experiments, proving that the electrostatic interaction was the dominant force for RNA extraction. Therefore, back-extraction efficiency values ranging between 85.19% and 90.78% were obtained by adjusting the ionic strength. Besides, the selective separation of RNA and tryptophane (Trp) was successfully accomplished. It was found that 86.19% RNA was distributed in the bottom phase, while 72.02% Trp was enriched in the top phase in the novel ABSs. Finally, dynamic light scattering (DLS) and transmission electron microscope (TEM) were used to further investigate the extraction mechanism. The proposed method reveals the outstanding feasibility of the newly developed ABSs formed by PEG-based DESs and inorganic salts for the green extraction of RNA. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. EFFECTS OF STORAGE, RNA EXTRACTION, GENECHIP TYPE, AND DONOR SEX ON GENE EXPRESSION PROFILING OF HUMAN WHOLE BLOOD

    Science.gov (United States)

    Background: Gene expression profiling of whole blood may be useful for monitoring toxicological exposure and for diagnosis and monitoring of various diseases. Several methods are available that can be used to transport, store, and extract RNA from whole blood, but it is not clear...

  15. DNA/RNA-based formulations for treatment of breast cancer.

    Science.gov (United States)

    Xie, Zhaolu; Zeng, Xianghui

    2017-12-01

    To develop a successful formulation for the gene therapy of breast cancer, an effective therapeutic nucleic acid and a proper delivery system are essential. Increased understanding of breast cancer, and developments in biotechnology, material science and nanotechnology have provided a major impetus in the development of effective formulations for the gene therapy of breast cancer. Areas covered: We discuss DNA/RNA-based formulations that can inhibit the growth of breast cancer cells and control the progress of breast cancer. Targets for the gene therapy of breast cancer, DNA/RNA-based therapeutics and delivery systems are summarized. And examples of successful DNA/RNA-based formulations for breast cancer gene therapy are reviewed. Expert opinion: Several challenges remain in developing effective DNA/RNA-based formulations for treatment of breast cancer. Firstly, most of the currently utilized targets are not effective enough as monotherapy for breast cancer. Secondly, the requirements for co-delivery system make the preparation of formulation more complicated. Thirdly, nanoparticles with the modification of tumor-targeting ligands could be more unstable in circulation and normal tissues. Lastly, immune responses against the viral vectors are unfavorable for the gene therapy of breast cancer because of the damage to the host and the impaired therapeutic ability.

  16. Proto-ribosome: a theoretical approach based on RNA relics

    OpenAIRE

    Demongeot, Jacques

    2017-01-01

    We describe in this paper, based on already published articles, a contribution to the theory postulating the existence of a proto-ribosome, which could have appeared early at the origin of life and we discuss the interest of this notion in an evolutionary perspective, taking into account the existence of possible RNA relics of this proto-ribosome.

  17. Short Communication An efficient method for simultaneous extraction of high-quality RNA and DNA from various plant tissues.

    Science.gov (United States)

    Oliveira, R R; Viana, A J C; Reátegui, A C E; Vincentz, M G A

    2015-12-29

    Determination of gene expression is an important tool to study biological processes and relies on the quality of the extracted RNA. Changes in gene expression profiles may be directly related to mutations in regulatory DNA sequences or alterations in DNA cytosine methylation, which is an epigenetic mark. Correlation of gene expression with DNA sequence or epigenetic mark polymorphism is often desirable; for this, a robust protocol to isolate high-quality RNA and DNA simultaneously from the same sample is required. Although commercial kits and protocols are available, they are mainly optimized for animal tissues and, in general, restricted to RNA or DNA extraction, not both. In the present study, we describe an efficient and accessible method to extract both RNA and DNA simultaneously from the same sample of various plant tissues, using small amounts of starting material. The protocol was efficient in the extraction of high-quality nucleic acids from several Arabidopsis thaliana tissues (e.g., leaf, inflorescence stem, flower, fruit, cotyledon, seedlings, root, and embryo) and from other tissues of non-model plants, such as Avicennia schaueriana (Acanthaceae), Theobroma cacao (Malvaceae), Paspalum notatum (Poaceae), and Sorghum bicolor (Poaceae). The obtained nucleic acids were used as templates for downstream analyses, such as mRNA sequencing, quantitative real time-polymerase chain reaction, bisulfite treatment, and others; the results were comparable to those obtained with commercial kits. We believe that this protocol could be applied to a broad range of plant species, help avoid technical and sampling biases, and facilitate several RNA- and DNA-dependent analyses.

  18. Crataegus Special Extract WS 1442 Effects on eNOS and microRNA 155.

    Science.gov (United States)

    Wang, Xinwen; Liang, Yan; Shi, Jian; Zhu, Hao-Jie; Bleske, Barry E

    2018-04-16

    Increased expression of microRNA 155 (miR-155) results in a decrease in endothelial nitric oxide synthase (eNOS) expression and impaired endothelial function. Factors that have been shown to increase expression of miR-155 may be mitigated by WS 1442, an extract of hawthorn leaves and flowers ( Crataegus special extract) that contains a range of pharmacologically active substances including oligomeric proanthocyanidins and flavonoids. The purpose of this study is to determine the effect of WS 1442 on the expression of miR-155 and eNOS in the presence of tumor necrosis factor (TNF- α ). Human umbilical vein endothelial cells (HUVECs) were studied after the exposure to TNF- α , with or without simvastatin (positive control) and WS 1442. The expression levels of eNOS, phosphorylated eNOS, and miR-155 in the different HUVEC treatment groups were determined by western blot and quantitative real-time polymerase chain reaction, respectively. To evaluate the effect of WS 1442 on the eNOS activity, the medium and intracellular nitrate/nitrite (NO) concentrations were also analyzed using a colorimetric Griess assay kit. The results demonstrated that TNF- α upregulated miR-155 expression and decreased eNOS expression and NO concentrations. WS 1442 also increased miR-155 expression and decreased eNOS expression but, unlike TNF- α , increased phosphorylated eNOS expression and NO concentrations. Surprisingly, WS 1442 increased miR-155 expression; however, WS 1442 mitigated the overall negative effect of miR-155 on decreasing eNOS expression by increasing expression of phosphorylated eNOS and resulting in an increase in NO concentrations. In the setting where miR-155 may be expressed, WS 1442 may offer vascular protection by increasing the expression of phosphorylated eNOS. Georg Thieme Verlag KG Stuttgart · New York.

  19. Refining Automatically Extracted Knowledge Bases Using Crowdsourcing

    Directory of Open Access Journals (Sweden)

    Chunhua Li

    2017-01-01

    Full Text Available Machine-constructed knowledge bases often contain noisy and inaccurate facts. There exists significant work in developing automated algorithms for knowledge base refinement. Automated approaches improve the quality of knowledge bases but are far from perfect. In this paper, we leverage crowdsourcing to improve the quality of automatically extracted knowledge bases. As human labelling is costly, an important research challenge is how we can use limited human resources to maximize the quality improvement for a knowledge base. To address this problem, we first introduce a concept of semantic constraints that can be used to detect potential errors and do inference among candidate facts. Then, based on semantic constraints, we propose rank-based and graph-based algorithms for crowdsourced knowledge refining, which judiciously select the most beneficial candidate facts to conduct crowdsourcing and prune unnecessary questions. Our experiments show that our method improves the quality of knowledge bases significantly and outperforms state-of-the-art automatic methods under a reasonable crowdsourcing cost.

  20. Refining Automatically Extracted Knowledge Bases Using Crowdsourcing.

    Science.gov (United States)

    Li, Chunhua; Zhao, Pengpeng; Sheng, Victor S; Xian, Xuefeng; Wu, Jian; Cui, Zhiming

    2017-01-01

    Machine-constructed knowledge bases often contain noisy and inaccurate facts. There exists significant work in developing automated algorithms for knowledge base refinement. Automated approaches improve the quality of knowledge bases but are far from perfect. In this paper, we leverage crowdsourcing to improve the quality of automatically extracted knowledge bases. As human labelling is costly, an important research challenge is how we can use limited human resources to maximize the quality improvement for a knowledge base. To address this problem, we first introduce a concept of semantic constraints that can be used to detect potential errors and do inference among candidate facts. Then, based on semantic constraints, we propose rank-based and graph-based algorithms for crowdsourced knowledge refining, which judiciously select the most beneficial candidate facts to conduct crowdsourcing and prune unnecessary questions. Our experiments show that our method improves the quality of knowledge bases significantly and outperforms state-of-the-art automatic methods under a reasonable crowdsourcing cost.

  1. RNA-dependent RNA polymerase: Addressing Zika outbreak by a phylogeny-based drug target study.

    Science.gov (United States)

    Stephen, Preyesh; Lin, Sheng-Xiang

    2018-01-01

    Since the first major outbreak of Zika virus (ZIKV) in 2007, ZIKV is spreading explosively through South and Central America, and recent reports in highly populated developing countries alarm the possibility of a more catastrophic outbreak. ZIKV infection in pregnant women leads to embryonic microcephaly and Guillain-Barré syndrome in adults. At present, there is limited understanding of the infectious mechanism, and no approved therapy has been reported. Despite the withdrawal of public health emergency, the WHO still considers the ZIKV as a highly significant and long-term public health challenge that the situation has to be addressed rapidly. Non-structural protein 5 is essential for capping and replication of viral RNA and comprises a methyltransferase and RNA-dependent RNA polymerase (RdRp) domain. We used molecular modeling to obtain the structure of ZIKV RdRp, and by molecular docking and phylogeny analysis, we here demonstrate the potential sites for drug screening. Two metal binding sites and an NS3-interacting region in ZIKV RdRp are demonstrated as potential drug screening sites. The docked structures reveal a remarkable degree of conservation at the substrate binding site and the potential drug screening sites. A phylogeny-based approach is provided for an emergency preparedness, where similar class of ligands could target phylogenetically related proteins. © 2017 John Wiley & Sons A/S.

  2. Determination of an effective scoring function for RNA-RNA interactions with a physics-based double-iterative method.

    Science.gov (United States)

    Yan, Yumeng; Wen, Zeyu; Zhang, Di; Huang, Sheng-You

    2018-05-18

    RNA-RNA interactions play fundamental roles in gene and cell regulation. Therefore, accurate prediction of RNA-RNA interactions is critical to determine their complex structures and understand the molecular mechanism of the interactions. Here, we have developed a physics-based double-iterative strategy to determine the effective potentials for RNA-RNA interactions based on a training set of 97 diverse RNA-RNA complexes. The double-iterative strategy circumvented the reference state problem in knowledge-based scoring functions by updating the potentials through iteration and also overcame the decoy-dependent limitation in previous iterative methods by constructing the decoys iteratively. The derived scoring function, which is referred to as DITScoreRR, was evaluated on an RNA-RNA docking benchmark of 60 test cases and compared with three other scoring functions. It was shown that for bound docking, our scoring function DITScoreRR obtained the excellent success rates of 90% and 98.3% in binding mode predictions when the top 1 and 10 predictions were considered, compared to 63.3% and 71.7% for van der Waals interactions, 45.0% and 65.0% for ITScorePP, and 11.7% and 26.7% for ZDOCK 2.1, respectively. For unbound docking, DITScoreRR achieved the good success rates of 53.3% and 71.7% in binding mode predictions when the top 1 and 10 predictions were considered, compared to 13.3% and 28.3% for van der Waals interactions, 11.7% and 26.7% for our ITScorePP, and 3.3% and 6.7% for ZDOCK 2.1, respectively. DITScoreRR also performed significantly better in ranking decoys and obtained significantly higher score-RMSD correlations than the other three scoring functions. DITScoreRR will be of great value for the prediction and design of RNA structures and RNA-RNA complexes.

  3. Cleavage of influenza RNA by using a human PUF-based artificial RNA-binding protein–staphylococcal nuclease hybrid

    International Nuclear Information System (INIS)

    Mori, Tomoaki; Nakamura, Kento; Masaoka, Keisuke; Fujita, Yusuke; Morisada, Ryosuke; Mori, Koichi; Tobimatsu, Takamasa; Sera, Takashi

    2016-01-01

    Various viruses infect animals and humans and cause a variety of diseases, including cancer. However, effective methodologies to prevent virus infection have not yet been established. Therefore, development of technologies to inactivate viruses is highly desired. We have already demonstrated that cleavage of a DNA virus genome was effective to prevent its replication. Here, we expanded this methodology to RNA viruses. In the present study, we used staphylococcal nuclease (SNase) instead of the PIN domain (PilT N-terminus) of human SMG6 as an RNA-cleavage domain and fused the SNase to a human Pumilio/fem-3 binding factor (PUF)-based artificial RNA-binding protein to construct an artificial RNA restriction enzyme with enhanced RNA-cleavage rates for influenzavirus. The resulting SNase-fusion nuclease cleaved influenza RNA at rates 120-fold greater than the corresponding PIN-fusion nuclease. The cleaving ability of the PIN-fusion nuclease was not improved even though the linker moiety between the PUF and RNA-cleavage domain was changed. Gel shift assays revealed that the RNA-binding properties of the PUF derivative used was not as good as wild type PUF. Improvement of the binding properties or the design method will allow the SNase-fusion nuclease to cleave an RNA target in mammalian animal cells and/or organisms. - Highlights: • A novel RNA restriction enzyme using SNase was developed tor cleave viral RNA. • Our enzyme cleaved influenza RNA with rates >120-fold higher rates a PIN-fusion one. • Our artificial enzyme with the L5 linker showed the highest RNA cleavage rate. • Our artificial enzyme site-selectively cleaved influenza RNA in vitro.

  4. Co-LncRNA: investigating the lncRNA combinatorial effects in GO annotations and KEGG pathways based on human RNA-Seq data.

    Science.gov (United States)

    Zhao, Zheng; Bai, Jing; Wu, Aiwei; Wang, Yuan; Zhang, Jinwen; Wang, Zishan; Li, Yongsheng; Xu, Juan; Li, Xia

    2015-01-01

    Long non-coding RNAs (lncRNAs) are emerging as key regulators of diverse biological processes and diseases. However, the combinatorial effects of these molecules in a specific biological function are poorly understood. Identifying co-expressed protein-coding genes of lncRNAs would provide ample insight into lncRNA functions. To facilitate such an effort, we have developed Co-LncRNA, which is a web-based computational tool that allows users to identify GO annotations and KEGG pathways that may be affected by co-expressed protein-coding genes of a single or multiple lncRNAs. LncRNA co-expressed protein-coding genes were first identified in publicly available human RNA-Seq datasets, including 241 datasets across 6560 total individuals representing 28 tissue types/cell lines. Then, the lncRNA combinatorial effects in a given GO annotations or KEGG pathways are taken into account by the simultaneous analysis of multiple lncRNAs in user-selected individual or multiple datasets, which is realized by enrichment analysis. In addition, this software provides a graphical overview of pathways that are modulated by lncRNAs, as well as a specific tool to display the relevant networks between lncRNAs and their co-expressed protein-coding genes. Co-LncRNA also supports users in uploading their own lncRNA and protein-coding gene expression profiles to investigate the lncRNA combinatorial effects. It will be continuously updated with more human RNA-Seq datasets on an annual basis. Taken together, Co-LncRNA provides a web-based application for investigating lncRNA combinatorial effects, which could shed light on their biological roles and could be a valuable resource for this community. Database URL: http://www.bio-bigdata.com/Co-LncRNA/. © The Author(s) 2015. Published by Oxford University Press.

  5. Modeling and prediction of extraction profile for microwave-assisted extraction based on absorbed microwave energy.

    Science.gov (United States)

    Chan, Chung-Hung; Yusoff, Rozita; Ngoh, Gek-Cheng

    2013-09-01

    A modeling technique based on absorbed microwave energy was proposed to model microwave-assisted extraction (MAE) of antioxidant compounds from cocoa (Theobroma cacao L.) leaves. By adapting suitable extraction model at the basis of microwave energy absorbed during extraction, the model can be developed to predict extraction profile of MAE at various microwave irradiation power (100-600 W) and solvent loading (100-300 ml). Verification with experimental data confirmed that the prediction was accurate in capturing the extraction profile of MAE (R-square value greater than 0.87). Besides, the predicted yields from the model showed good agreement with the experimental results with less than 10% deviation observed. Furthermore, suitable extraction times to ensure high extraction yield at various MAE conditions can be estimated based on absorbed microwave energy. The estimation is feasible as more than 85% of active compounds can be extracted when compared with the conventional extraction technique. Copyright © 2013 Elsevier Ltd. All rights reserved.

  6. Extractions of High Quality RNA from the Seeds of Jerusalem Artichoke and Other Plant Species with High Levels of Starch and Lipid.

    Science.gov (United States)

    Mornkham, Tanupat; Wangsomnuk, Preeya Puangsomlee; Fu, Yong-Bi; Wangsomnuk, Pinich; Jogloy, Sanun; Patanothai, Aran

    2013-04-29

    Jerusalem artichoke (Helianthus tuberosus L.) is an important tuber crop. However, Jerusalem artichoke seeds contain high levels of starch and lipid, making the extraction of high-quality RNA extremely difficult and the gene expression analysis challenging. This study was aimed to improve existing methods for extracting total RNA from Jerusalem artichoke dry seeds and to assess the applicability of the improved method in other plant species. Five RNA extraction methods were evaluated on Jerusalem artichoke seeds and two were modified. One modified method with the significant improvement was applied to assay seeds of diverse Jerusalem artichoke accessions, sunflower, rice, maize, peanut and marigold. The effectiveness of the improved method to extract total RNA from seeds was assessed using qPCR analysis of four selected genes. The improved method of Ma and Yang (2011) yielded a maximum RNA solubility and removed most interfering substances. The improved protocol generated 29 to 41 µg RNA/30 mg fresh weight. An A260/A280 ratio of 1.79 to 2.22 showed their RNA purity. Extracted RNA was effective for downstream applications such as first-stranded cDNA synthesis, cDNA cloning and qPCR. The improved method was also effective to extract total RNA from seeds of sunflower, rice, maize and peanut that are rich in polyphenols, lipids and polysaccharides.

  7. Extractions of High Quality RNA from the Seeds of Jerusalem Artichoke and Other Plant Species with High Levels of Starch and Lipid

    Directory of Open Access Journals (Sweden)

    Tanupat Mornkham

    2013-04-01

    Full Text Available Jerusalem artichoke (Helianthus tuberosus L. is an important tuber crop. However, Jerusalem artichoke seeds contain high levels of starch and lipid, making the extraction of high-quality RNA extremely difficult and the gene expression analysis challenging. This study was aimed to improve existing methods for extracting total RNA from Jerusalem artichoke dry seeds and to assess the applicability of the improved method in other plant species. Five RNA extraction methods were evaluated on Jerusalem artichoke seeds and two were modified. One modified method with the significant improvement was applied to assay seeds of diverse Jerusalem artichoke accessions, sunflower, rice, maize, peanut and marigold. The effectiveness of the improved method to extract total RNA from seeds was assessed using qPCR analysis of four selected genes. The improved method of Ma and Yang (2011 yielded a maximum RNA solubility and removed most interfering substances. The improved protocol generated 29 to 41 µg RNA/30 mg fresh weight. An A260/A280 ratio of 1.79 to 2.22 showed their RNA purity. Extracted RNA was effective for downstream applications such as first-stranded cDNA synthesis, cDNA cloning and qPCR. The improved method was also effective to extract total RNA from seeds of sunflower, rice, maize and peanut that are rich in polyphenols, lipids and polysaccharides.

  8. 21 CFR 172.585 - Sugar beet extract flavor base.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Sugar beet extract flavor base. 172.585 Section 172... CONSUMPTION Flavoring Agents and Related Substances § 172.585 Sugar beet extract flavor base. Sugar beet extract flavor base may be safely used in food in accordance with the provisions of this section. (a...

  9. FEATURE EXTRACTION FOR EMG BASED PROSTHESES CONTROL

    Directory of Open Access Journals (Sweden)

    R. Aishwarya

    2013-01-01

    Full Text Available The control of prosthetic limb would be more effective if it is based on Surface Electromyogram (SEMG signals from remnant muscles. The analysis of SEMG signals depend on a number of factors, such as amplitude as well as time- and frequency-domain properties. Time series analysis using Auto Regressive (AR model and Mean frequency which is tolerant to white Gaussian noise are used as feature extraction techniques. EMG Histogram is used as another feature vector that was seen to give more distinct classification. The work was done with SEMG dataset obtained from the NINAPRO DATABASE, a resource for bio robotics community. Eight classes of hand movements hand open, hand close, Wrist extension, Wrist flexion, Pointing index, Ulnar deviation, Thumbs up, Thumb opposite to little finger are taken into consideration and feature vectors are extracted. The feature vectors can be given to an artificial neural network for further classification in controlling the prosthetic arm which is not dealt in this paper.

  10. Targeting miRNA-based medicines to cystic fibrosis airway epithelial cells using nanotechnology

    Directory of Open Access Journals (Sweden)

    McKiernan PJ

    2013-10-01

    Full Text Available Paul J McKiernan,2 Orla Cunninghamm,1,2 Catherine M Greenem,2 Sally-Ann Cryan1,31School of Pharmacy, Royal College of Surgeons in Ireland, 2Respiratory Research Division, Department of Medicine, Royal College of Surgeons in Ireland Education and Research Centre, Beaumont Hospital, 3Trinity Centre for Bioengineering, Trinity College Dublin, Dublin, IrelandAbstract: Cystic fibrosis (CF is an inherited disorder characterized by chronic airway inflammation. microRNAs (miRNAs are endogenous small RNAs which act on messenger (mRNA at a post transcriptional level, and there is a growing understanding that altered expression of miRNA is involved in the CF phenotype. Modulation of miRNA by replacement using miRNA mimics (premiRs presents a new therapeutic paradigm for CF, but effective and safe methods of delivery to the CF epithelium are limiting clinical translation. Herein, polymeric nanoparticles are investigated for delivery of miRNA mimics into CF airway epithelial cells, using miR-126 as a proof-of-concept premiR cargo to determine efficiency. Two polymers, polyethyleneimine (PEI and chitosan, were used to prepare miRNA nanomedicines, characterized for their size, surface (zeta potential, and RNA complexation efficiency, and screened for delivery and cytotoxicity in CFBE41o- (human F508del cystic fibrosis transmembrane conductance regulator bronchial epithelial cells using a novel high content analysis method. RNA extraction was carried out 24 hours post transfection, and miR-126 and TOM1 (target of Myb1 expression (a validated miR-126 target was assessed. Manufacture was optimized to produce small nanoparticles that effectively complexed miRNA. Using high content analysis, PEI-based nanoparticles were more effective than chitosan-based nanoparticles in facilitating uptake of miRNA into CFBE41o- cells and this was confirmed in miR-126 assays. PEI-premiR-126 nanoparticles at low nitrogen/phosphate (N/P ratios resulted in significant knockdown of

  11. A stable RNA virus-based vector for citrus trees

    International Nuclear Information System (INIS)

    Folimonov, Alexey S.; Folimonova, Svetlana Y.; Bar-Joseph, Moshe; Dawson, William O.

    2007-01-01

    Virus-based vectors are important tools in plant molecular biology and plant genomics. A number of vectors based on viruses that infect herbaceous plants are in use for expression or silencing of genes in plants as well as screening unknown sequences for function. Yet there is a need for useful virus-based vectors for woody plants, which demand much greater stability because of the longer time required for systemic infection and analysis. We examined several strategies to develop a Citrus tristeza virus (CTV)-based vector for transient expression of foreign genes in citrus trees using a green fluorescent protein (GFP) as a reporter. These strategies included substitution of the p13 open reading frame (ORF) by the ORF of GFP, construction of a self-processing fusion of GFP in-frame with the major coat protein (CP), or expression of the GFP ORF as an extra gene from a subgenomic (sg) mRNA controlled either by a duplicated CTV CP sgRNA controller element (CE) or an introduced heterologous CE of Beet yellows virus. Engineered vector constructs were examined for replication, encapsidation, GFP expression during multiple passages in protoplasts, and for their ability to infect, move, express GFP, and be maintained in citrus plants. The most successful vectors based on the 'add-a-gene' strategy have been unusually stable, continuing to produce GFP fluorescence after more than 4 years in citrus trees

  12. [Characterization of Black and Dichothrix Cyanobacteria Based on the 16S Ribosomal RNA Gene Sequence

    Science.gov (United States)

    Ortega, Maya

    2010-01-01

    My project focuses on characterizing different cyanobacteria in thrombolitic mats found on the island of Highborn Cay, Bahamas. Thrombolites are interesting ecosystems because of the ability of bacteria in these mats to remove carbon dioxide from the atmosphere and mineralize it as calcium carbonate. In the future they may be used as models to develop carbon sequestration technologies, which could be used as part of regenerative life systems in space. These thrombolitic communities are also significant because of their similarities to early communities of life on Earth. I targeted two cyanobacteria in my research, Dichothrix spp. and whatever black is, since they are believed to be important to carbon sequestration in these thrombolitic mats. The goal of my summer research project was to molecularly identify these two cyanobacteria. DNA was isolated from each organism through mat dissections and DNA extractions. I ran Polymerase Chain Reactions (PCR) to amplify the 16S ribosomal RNA (rRNA) gene in each cyanobacteria. This specific gene is found in almost all bacteria and is highly conserved, meaning any changes in the sequence are most likely due to evolution. As a result, the 16S rRNA gene can be used for bacterial identification of different species based on the sequence of their 16S rRNA gene. Since the exact sequence of the Dichothrix gene was unknown, I designed different primers that flanked the gene based on the known sequences from other taxonomically similar cyanobacteria. Once the 16S rRNA gene was amplified, I cloned the gene into specialized Escherichia coli cells and sent the gene products for sequencing. Once the sequence is obtained, it will be added to a genetic database for future reference to and classification of other Dichothrix sp.

  13. RED WINE EXTRACT OBTAINED BY MEMBRANE-BASED SUPERCRITICAL FLUID EXTRACTION: PRELIMINARY CHARACTERIZATION OF CHEMICAL PROPERTIES.

    Directory of Open Access Journals (Sweden)

    W. Silva

    Full Text Available ABSTRACT This study aims to obtain an extract from red wine by using membrane-based supercritical fluid extraction. This technique involves the use of porous membranes as contactors during the dense gas extraction process from liquid matrices. In this work, a Cabernet Sauvignon wine extract was obtained from supercritical fluid extraction using pressurized carbon dioxide as solvent and a hollow fiber contactor as extraction setup. The process was continuously conducted at pressures between 12 and 18 MPa and temperatures ranged from 30 to 50ºC. Meanwhile, flow rates of feed wine and supercritical CO2 varied from 0.1 to 0.5 mL min-1 and from 60 to 80 mL min-1 (NCPT, respectively. From extraction assays, the highest extraction percentage value obtained from the total amount of phenolic compounds was 14% in only one extraction step at 18MPa and 35ºC. A summarized chemical characterization of the obtained extract is reported in this work; one of the main compounds in this extract could be a low molecular weight organic acid with aromatic structure and methyl and carboxyl groups. Finally, this preliminary characterization of this extract shows a remarkable ORAC value equal to 101737 ± 5324 µmol Trolox equivalents (TE per 100 g of extract.

  14. Morpholino spin-labeling for base-pair sequencing of a 3'-terminal RNA stem by proton homonuclear Overhauser enhancements: yeast ribosomal 5S RNA

    International Nuclear Information System (INIS)

    Lee, K.M.; Marshall, A.G.

    1987-01-01

    Base-pair sequences for 5S and 5.8S RNAs are not readily extracted from proton homonuclear nuclear Overhauser enhancement (NOE) connectivity experiments alone, due to extensive peak overlap in the downfield (11-15 ppm) proton NMR spectrum. In this paper, we introduce a new method for base-pair proton peak assignment for ribosomal RNAs, based upon the distance-dependent broadening of the resonances of base-pair protons spatially proximal to a paramagnetic group. Introduction of a nitroxide spin-label covalently attached to the 3'-terminal ribose provides an unequivocal starting point for base-pair hydrogen-bond proton NMR assignment. Subsequent NOE connectivities then establish the base-pair sequence for the terminal stem of a 5S RNA. Periodate oxidation of yeast 5S RNA, followed by reaction with 4-amino-2,2,6,6-tetramethylpiperidinyl-1-oxy (TEMPO-NH2) and sodium borohydride reduction, produces yeast 5S RNA specifically labeled with a paramagnetic nitroxide group at the 3'-terminal ribose. Comparison of the 500-MHz 1H NMR spectra of native and 3'-terminal spin-labeled yeast 5S RNA serves to identify the terminal base pair (G1 . C120) and its adjacent base pair (G2 . U119) on the basis of their proximity to the 3'-terminal spin-label. From that starting point, we have then identified (G . C, A . U, or G . U) and sequenced eight of the nine base pairs in the terminal helix via primary and secondary NOE's

  15. DincRNA: a comprehensive web-based bioinformatics toolkit for exploring disease associations and ncRNA function.

    Science.gov (United States)

    Cheng, Liang; Hu, Yang; Sun, Jie; Zhou, Meng; Jiang, Qinghua

    2018-06-01

    DincRNA aims to provide a comprehensive web-based bioinformatics toolkit to elucidate the entangled relationships among diseases and non-coding RNAs (ncRNAs) from the perspective of disease similarity. The quantitative way to illustrate relationships of pair-wise diseases always depends on their molecular mechanisms, and structures of the directed acyclic graph of Disease Ontology (DO). Corresponding methods for calculating similarity of pair-wise diseases involve Resnik's, Lin's, Wang's, PSB and SemFunSim methods. Recently, disease similarity was validated suitable for calculating functional similarities of ncRNAs and prioritizing ncRNA-disease pairs, and it has been widely applied for predicting the ncRNA function due to the limited biological knowledge from wet lab experiments of these RNAs. For this purpose, a large number of algorithms and priori knowledge need to be integrated. e.g. 'pair-wise best, pairs-average' (PBPA) and 'pair-wise all, pairs-maximum' (PAPM) methods for calculating functional similarities of ncRNAs, and random walk with restart (RWR) method for prioritizing ncRNA-disease pairs. To facilitate the exploration of disease associations and ncRNA function, DincRNA implemented all of the above eight algorithms based on DO and disease-related genes. Currently, it provides the function to query disease similarity scores, miRNA and lncRNA functional similarity scores, and the prioritization scores of lncRNA-disease and miRNA-disease pairs. http://bio-annotation.cn:18080/DincRNAClient/. biofomeng@hotmail.com or qhjiang@hit.edu.cn. Supplementary data are available at Bioinformatics online.

  16. Idioms-based Business Rule Extraction

    NARCIS (Netherlands)

    R Smit (Rob)

    2011-01-01

    htmlabstractThis thesis studies the extraction of embedded business rules, using the idioms of the used framework to identify them. Embedded business rules exist as source code in the software system and knowledge about them may get lost. Extraction of those business rules could make them accessible

  17. A Novel Microfluidic Device for Fully Automated Extraction of RNA from Cell Cultures, Phase I

    Data.gov (United States)

    National Aeronautics and Space Administration — Differential gene expression by RNA profiling is a universal and critical step in space biology experiments, which seek to link specific molecular events with...

  18. A Novel Microfluidic Device for Fully Automated Extraction of RNA from Cell Cultures, Phase II

    Data.gov (United States)

    National Aeronautics and Space Administration — Obtaining high quality, intact RNA from cells is an ubiquitous need in the pursuit of space biology. Our overall objective is to develop and commercialize a...

  19. Optimization-based Method for Automated Road Network Extraction

    International Nuclear Information System (INIS)

    Xiong, D

    2001-01-01

    Automated road information extraction has significant applicability in transportation. It provides a means for creating, maintaining, and updating transportation network databases that are needed for purposes ranging from traffic management to automated vehicle navigation and guidance. This paper is to review literature on the subject of road extraction and to describe a study of an optimization-based method for automated road network extraction

  20. COMPARISON OF TWO TOTAL RNA EXTRACTION PROTOCOLS FROM CHO-K1 CELLS FOR RT-PCR: CUT-OFF COST FOR RESEARCHERS

    Directory of Open Access Journals (Sweden)

    Vasila Packeer Mohamed

    2014-05-01

    Full Text Available ABSTRACT: Various methods have been described to extract RNA from adherent mammalian cells. RNA isolation in conjunction with reverse transcription polymerase chain reaction (RT-PCR is a valuable tool used to study gene expression profiling. This approach is now being used in mammalian cell bioprocessing to help understand and improve the system. The objective of this study was to compare and determine the most suitable RNA extraction method for CHO-K1 cells in a setting where a relatively large amount of samples was involved. Total RNA was extracted using Total RNA purification kit (without DNase treatment; Norgen, Canada and RNeasy mini kit (with DNase treatment; Qiagen, USA respectively. The extracted RNA was then reverse transcribed, and the cDNA was subjected to PCR-amplifying 18S. Yield from RNeasy kit was significantly higher (0.316 ± 0.033 µg/µl; p=0.004 than Total RNA purification kit (0.177 ± 0.0243 µg/µl. However, RNA purity for both methods was close to 2.0 and there was no significant difference between the methods. Total RNA purification kit is less expensive than RNeasy kit. Since there is no DNase treatment step in the former, extraction time for RNA is shorter. When the extracted RNA was subjected to RT-PCR, both methods were able to show detection of 18S at 219 bp.   Therefore, this study demonstrates that both protocols are suitable for RNA extraction for CHO-K1 cells. RNeasy mini kit (Qiagen is recommended if higher yields is the primary concern and Total RNA Purification kit (Norgen is recommended if time and cost are concerned. ABSTRAK: Pelbagai kaedah telah digunakan untuk mengekstrak RNA daripada sel mamalia lekat.  Pemencilan RNA dengan menggunakan reaksi rantai polimerase transkripsi berbalik (RT-PCR merupakan kaedah penting yang digunakan dalam mengkaji pernyataan gen berprofil.  Pendekatan ini kini digunakan dalam pemprosesan bio sel mamalia untuk memahami dan menambah baik sistem.  Tujuan kajian dijalankan

  1. Evaluation of FTA(®) card for the rescue of infectious foot-and-mouth disease virus by chemical transfection of extracted RNA in cultured cells.

    Science.gov (United States)

    Biswal, Jitendra K; Subramaniam, Saravanan; Ranjan, Rajeev; Pattnaik, Bramhadev

    2016-08-01

    Foot-and-mouth disease (FMD) is a highly contagious epidemic disease of transboundary importance. Inadequate storage and shipment of suspected clinical samples can compromise the ability to detect and characterise FMD virus (FMDV) in endemic countries, thereby, leading to the loss of valuable virological and epidemiological data. This study, investigates the potential of using FTA(®) cards for dry transportation of clinical samples and subsequent recovery of infectious FMDV by chemical transfection of FTA(®) card fixed RNA as an alternative to the conventional cell culture based virus isolation method. A higher proportion of infectious FMDV was rescued from clinical samples (cell culture isolates, tongue epithelial suspension and impression smears) by the FTA(®) card fixed RNA transfection method (76%) compared to the conventional cell culture based virus isolation (56%), suggesting a better performance of the current RNA transfection procedure. Furthermore, it was possible to rescue live virus by the transfection of RNA extracted from FTA(®) card impregnated with clinical samples that had been stored at varying temperature (4-37 °C) up to a period of six weeks. The VP1 sequence data and antigenic relationships with the vaccine strains, between viruses rescued by FTA(®) card fixed RNA transfection and conventional cell culture, were comparable. Therefore, these results support the use of the FTA(®) card for the economic, dry, non-hazardous transport of FMD suspected clinical samples from the site of collection to national/international reference laboratories. Copyright © 2016 Elsevier Ltd. All rights reserved.

  2. siRNA-like double-stranded RNAs are specifically protected against degradation in human cell extract.

    Directory of Open Access Journals (Sweden)

    John A H Hoerter

    Full Text Available RNA interference (RNAi is a set of intracellular pathways in eukaryotes that controls both exogenous and endogenous gene expression. The power of RNAi to knock down (silence any gene of interest by the introduction of synthetic small-interfering (siRNAs has afforded powerful insight into biological function through reverse genetic approaches and has borne a new field of gene therapeutics. A number of questions are outstanding concerning the potency of siRNAs, necessitating an understanding of how short double-stranded RNAs are processed by the cell. Recent work suggests unmodified siRNAs are protected in the intracellular environment, although the mechanism of protection still remains unclear. We have developed a set of doubly-fluorophore labeled RNAs (more precisely, RNA/DNA chimeras to probe in real-time the stability of siRNAs and related molecules by fluorescence resonance energy transfer (FRET. We find that these RNA probes are substrates for relevant cellular degradative processes, including the RNase H1 mediated degradation of an DNA/RNA hybrid and Dicer-mediated cleavage of a 24-nucleotide (per strand double-stranded RNA. In addition, we find that 21- and 24-nucleotide double-stranded RNAs are relatively protected in human cytosolic cell extract, but less so in blood serum, whereas an 18-nucleotide double-stranded RNA is less protected in both fluids. These results suggest that RNAi effector RNAs are specifically protected in the cellular environment and may provide an explanation for recent results showing that unmodified siRNAs in cells persist intact for extended periods of time.

  3. Optical Aperture Synthesis Object's Information Extracting Based on Wavelet Denoising

    International Nuclear Information System (INIS)

    Fan, W J; Lu, Y

    2006-01-01

    Wavelet denoising is studied to improve OAS(optical aperture synthesis) object's Fourier information extracting. Translation invariance wavelet denoising based on Donoho wavelet soft threshold denoising is researched to remove Pseudo-Gibbs in wavelet soft threshold image. OAS object's information extracting based on translation invariance wavelet denoising is studied. The study shows that wavelet threshold denoising can improve the precision and the repetition of object's information extracting from interferogram, and the translation invariance wavelet denoising information extracting is better than soft threshold wavelet denoising information extracting

  4. Enhancement of single guide RNA transcription for efficient CRISPR/Cas-based genomic engineering.

    Science.gov (United States)

    Ui-Tei, Kumiko; Maruyama, Shohei; Nakano, Yuko

    2017-06-01

    Genomic engineering using clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (Cas) protein is a promising approach for targeting the genomic DNA of virtually any organism in a sequence-specific manner. Recent remarkable advances in CRISPR/Cas technology have made it a feasible system for use in therapeutic applications and biotechnology. In the CRISPR/Cas system, a guide RNA (gRNA), interacting with the Cas protein, recognizes a genomic region with sequence complementarity, and the double-stranded DNA at the target site is cleaved by the Cas protein. A widely used gRNA is an RNA polymerase III (pol III)-driven single gRNA (sgRNA), which is produced by artificial fusion of CRISPR RNA (crRNA) and trans-activation crRNA (tracrRNA). However, we identified a TTTT stretch, known as a termination signal of RNA pol III, in the scaffold region of the sgRNA. Here, we revealed that sgRNA carrying a TTTT stretch reduces the efficiency of sgRNA transcription due to premature transcriptional termination, and decreases the efficiency of genome editing. Unexpectedly, it was also shown that the premature terminated sgRNA may have an adverse effect of inducing RNA interference. Such disadvantageous effects were avoided by substituting one base in the TTTT stretch.

  5. Heterogeneity to Homogeneity: Synthesis, Base Pairing, and Ligation Studies of 4',3'-XyluloNA/RNA and TNA/RNA Chimeric Sequences

    Science.gov (United States)

    Bhowmik, S.; Stoop, M.; Krishnamurthy, R.

    2017-07-01

    Based on the reality of "prebiotic clutter," we herein present an alternate model for pre-RNA to RNA transition, which starts, not with homogeneous-backbone system, but rather with mixtures of heterogeneous-backbone of chimeric "pre-RNA/RNA."

  6. RNA interference-based therapeutics for human immunodeficiency virus HIV-1 treatment: synthetic siRNA or vector-based shRNA?

    Science.gov (United States)

    Subramanya, Sandesh; Kim, Sang-Soo; Manjunath, N; Shankar, Premlata

    2010-02-01

    Despite the clinical benefits of highly active antiretroviral therapy (HAART), the prospect of life-long antiretroviral treatment poses significant problems, which has spurred interest in developing new drugs and strategies to treat HIV infection and eliminate persistent viral reservoirs. RNAi has emerged as a therapeutic possibility for HIV. We discuss progress in overcoming hurdles to translating transient and stable RNAi enabling technologies to clinical application for HIV; covering the past 2 - 3 years. HIV inhibition can be achieved by transfection of chemically or enzymatically synthesized siRNAs or by DNA-based vector systems expressing short hairpin RNAs (shRNAs) that are processed intracellularly into siRNA. We compare these approaches, focusing on technical and safety issues that will guide the choice of strategy for clinical use. Introduction of synthetic siRNA into cells or its stable endogenous production using vector-driven shRNA have been shown to suppress HIV replication in vitro and, in some instances, in vivo. Each method has advantages and limitations in terms of ease of delivery, duration of silencing, emergence of escape mutants and potential toxicity. Both appear to have potential as future therapeutics for HIV, once the technical and safety issues of each approach are overcome.

  7. RNA Polymerase II Second Largest Subunit Molecular Identification of Boletus griseipurpureus Corner From Thailand and Antibacterial Activity of Basidiocarp Extracts.

    Science.gov (United States)

    Aung-Aud-Chariya, Amornrat; Bangrak, Phuwadol; Lumyong, Saisamorn; Phupong, Worrapong; Aggangan, Nelly Siababa; Kamlangdee, Niyom

    2015-03-01

    Boletus griseipurpureus Corner, an edible mushroom, is a putative ectomycorrhizal fungus. Currently, the taxonomic boundary of this mushroom is unclear and its bitter taste makes it interesting for evaluating its antibacterial properties. The purpose of this study was to identify the genetic variation of this mushroom and also to evaluate any antibacterial activities. Basidiocarps were collected from 2 north-eastern provinces, Roi Et and Ubon Ratchathani, and from 2 southern provinces, Songkhla and Surat Thani, in Thailand. Genomic DNA was extracted and molecular structure was examined using the RNA polymerase II (RPB2) analysis. Antibacterial activities of basidiocarp extracts were conducted with Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 29523 and methicillin-resistant Staphylococcus aureus (MRSA) 189 using the agar-well diffusion method. All the samples collected for this study constituted a monophyletic clade, which was closely related with the Boletus group of polypore fungi. For the antibacterial study, it was found that the crude methanol extract of basidiomes inhibited the growth of all bacteria in vitro more than the crude ethyl acetate extract. Basidomes collected from four locations in Thailand had low genetic variation and their extracts inhibited the growth of all tested bacteria. The health benefits of this edible species should be evaluated further.

  8. Phylogenetic relationships of Salmonella based on rRNA sequences

    DEFF Research Database (Denmark)

    Christensen, H.; Nordentoft, Steen; Olsen, J.E.

    1998-01-01

    separated by 16S rRNA analysis and found to be closely related to the Escherichia coli and Shigella complex by both 16S and 23S rRNA analyses. The diphasic serotypes S. enterica subspp. I and VI were separated from the monophasic serotypes subspp. IIIa and IV, including S. bongori, by 23S rRNA sequence...

  9. Analytical Performances of Human Immunodeficiency Virus Type 1 RNA-Based Amplix® Real-Time PCR Platform for HIV-1 RNA Quantification

    Directory of Open Access Journals (Sweden)

    Christian Diamant Mossoro-Kpinde

    2016-01-01

    Full Text Available Objectives. We evaluated the performances of Amplix real-time PCR platform developed by Biosynex (Strasbourg, France, combining automated station extraction (Amplix station 16 Dx and real-time PCR (Amplix NG, for quantifying plasma HIV-1 RNA by lyophilized HIV-1 RNA-based Amplix reagents targeting gag and LTR, using samples from HIV-1-infected adults from Central African Republic. Results. Amplix real-time PCR assay showed low limit of detection (28 copies/mL, across wide dynamic range (1.4–10 log copies/mL, 100% sensitivity and 99% specificity, high reproducibility, and accuracy with mean bias < 5%. The assay showed excellent correlations and concordance of 95.3% with the reference HIV-1 RNA load assay (Roche, with mean absolute bias of +0.097 log copies/mL by Bland-Altman analysis. The assay was able to detect and quantify the most prevalent HIV-1 subtype strains and the majority of non-B subtypes, CRFs of HIV-1 group M, and HIV-1 groups N and O circulating in Central Africa. The Amplix assay showed 100% sensitivity and 99.6% specificity to diagnose virological failure in clinical samples from antiretroviral drug-experienced patients. Conclusions. The HIV-1 RNA-based Amplix real-time PCR platform constitutes sensitive and reliable system for clinical monitoring of HIV-1 RNA load in HIV-1-infected children and adults, particularly adapted to intermediate laboratory facilities in sub-Saharan Africa.

  10. A thermostable messenger RNA based vaccine against rabies.

    Science.gov (United States)

    Stitz, Lothar; Vogel, Annette; Schnee, Margit; Voss, Daniel; Rauch, Susanne; Mutzke, Thorsten; Ketterer, Thomas; Kramps, Thomas; Petsch, Benjamin

    2017-12-01

    Although effective rabies virus vaccines have been existing for decades, each year, rabies virus infections still cause around 50.000 fatalities worldwide. Most of these cases occur in developing countries, where these vaccines are not available. The reasons for this are the prohibitive high costs of cell culture or egg grown rabies virus vaccines and the lack of a functional cold chain in many regions in which rabies virus is endemic. Here, we describe the excellent temperature resistance of a non-replicating mRNA based rabies virus vaccine encoding the rabies virus glycoprotein (RABV-G). Prolonged storage of the vaccine from -80°C to up to +70°C for several months did not impact the protective capacity of the mRNA vaccine. Efficacy after storage was demonstrated by the induction of rabies specific virus neutralizing antibodies and protection in mice against lethal rabies infection. Moreover, storing the vaccine at oscillating temperatures between +4° and +56°C for 20 cycles in order to simulate interruptions of the cold chain during vaccine transport, did not affect the vaccine's immunogenicity and protective characteristics, indicating that maintenance of a cold chain is not essential for this vaccine.

  11. A method for high-quality RNA extraction from tall fescue

    African Journals Online (AJOL)

    Jane

    2011-07-20

    Jul 20, 2011 ... but most of those methods are time-consuming (Li et al.,. 2008). Since RNA ... forage and seed (Wang and Ge, 2004; Huang et al.,. 2008). ..... Malnoy M, Reynoird JP, Mourgues F, Chevreau E, Simoneau P (2001). A method ...

  12. Sponge-associated actinobacterial diversity: validation of the methods of actinobacterial DNA extraction and optimization of 16S rRNA gene amplification.

    Science.gov (United States)

    Yang, Qi; Franco, Christopher M M; Zhang, Wei

    2015-10-01

    Experiments were designed to validate the two common DNA extraction protocols (CTAB-based method and DNeasy Blood & Tissue Kit) used to effectively recover actinobacterial DNA from sponge samples in order to study the sponge-associated actinobacterial diversity. This was done by artificially spiking sponge samples with actinobacteria (spores, mycelia and a combination of the two). Our results demonstrated that both DNA extraction methods were effective in obtaining DNA from the sponge samples as well as the sponge samples spiked with different amounts of actinobacteria. However, it was noted that in the presence of the sponge, the bacterial 16S rRNA gene could not be amplified unless the combined DNA template was diluted. To test the hypothesis that the extracted sponge DNA contained inhibitors, dilutions of the DNA extracts were tested for six sponge species representing five orders. The results suggested that the inhibitors were co-extracted with the sponge DNA, and a high dilution of this DNA was required for the successful PCR amplification for most of the samples. The optimized PCR conditions, including primer selection, PCR reaction system and program optimization, further improved the PCR performance. However, no single PCR condition was found to be suitable for the diverse sponge samples using various primer sets. These results highlight for the first time that the DNA extraction methods used are effective in obtaining actinobacterial DNA and that the presence of inhibitors in the sponge DNA requires high dilution coupled with fine tuning of the PCR conditions to achieve success in the study of sponge-associated actinobacterial diversity.

  13. Areca nut extract up-regulates prostaglandin production, cyclooxygenase-2 mRNA and protein expression of human oral keratinocytes.

    Science.gov (United States)

    Jeng, J H; Ho, Y S; Chan, C P; Wang, Y J; Hahn, L J; Lei, D; Hsu, C C; Chang, M C

    2000-07-01

    There are about 600 million betel quid (BQ) chewers in the world. BQ chewing is associated with increased incidence of oral cancer and submucous fibrosis. In this study, areca nut (AN) extract (200-800 microg/ml) induced the prostaglandin E(2) (PGE(2)) production by 1. 4-3.4-fold and 6-keto-PGF(1 alpha) production by 1.1-1.7-fold of gingival keratinocytes (GK), respectively, following 24 h of exposure. Exposure of GK to AN extract (>400 microg/ml) led to cell retraction and intracellular vacuoles formation. At concentrations of 800 and 1200 microg/ml, AN extract induced cell death at 21-24 and 32-52% as detected by MTT assay and cellular lactate dehydrogenase release, respectively. Interestingly, AN-induced morphological changes of GK are reversible. GK can still proliferate following exposure to AN extract. Cytotoxicity of AN extract cannot be inhibited by indomethacin (1 microM) and aspirin (50 microM), indicating that prostaglandin (PG) production is not the major factor responsible for AN cytotoxicity. PGE(2) exhibited little effect on the growth of GK at concentrations ranging from 100-1000 pg/ml. Stimulating GK production of PGs by AN extract could be due to induction of cyclooxygenase-2 (COX-2) mRNA expression and protein production. These results suggest that AN ingredients are critical in the pathogenesis of oral submucous fibrosis and oral cancer via their stimulatory effects on the PGs, COX-2 production and associated tissue inflammatory responses. AN cytotoxicity to GK is not directly mediated by COX-2 stimulation and PG production.

  14. RNA Detection Based on Graphene Field-Effect Transistor Biosensor

    Directory of Open Access Journals (Sweden)

    Meng Tian

    2018-01-01

    Full Text Available Graphene has attracted much attention in biosensing applications due to its unique properties. In this paper, the monolayer graphene was grown by chemical vapor deposition (CVD method. Using the graphene as the electric channel, we have fabricated a graphene field-effect transistor (G-FET biosensor that can be used for label-free detection of RNA. Compared with conventional method, the G-FET RNA biosensor can be run in low cost, be time-saving, and be miniaturized for RNA measurement. The sensors show high performance and achieve the RNA detection sensitivity as low as 0.1 fM, which is two orders of magnitude lower than the previously reports. Moreover, the G-FET biosensor can readily distinguish target RNA from noncomplementary RNA, showing high selectivity for RNA detection. The developed G-FET RNA biosensor with high sensitivity, fast analysis speed, and simple operation may provide a new feasible direction for RNA research and biosensing.

  15. Mechanism of action of mRNA-based vaccines.

    Science.gov (United States)

    Iavarone, Carlo; O'hagan, Derek T; Yu, Dong; Delahaye, Nicolas F; Ulmer, Jeffrey B

    2017-09-01

    The present review summarizes the growing body of work defining the mechanisms of action of this exciting new vaccine technology that should allow rational approaches in the design of next generation mRNA vaccines. Areas covered: Bio-distribution of mRNA, localization of antigen production, role of the innate immunity, priming of the adaptive immune response, route of administration and effects of mRNA delivery systems. Expert commentary: In the last few years, the development of RNA vaccines had a fast growth, the rising number of proof will enable rational approaches to improving the effectiveness and safety of this modern class of medicine.

  16. RNA interference-based resistance against a legume mastrevirus

    Directory of Open Access Journals (Sweden)

    Mansoor Shahid

    2011-11-01

    Full Text Available Abstract Background RNA interference (RNAi is a homology-dependant gene silencing mechanism and has been widely used to engineer resistance in plants against RNA viruses. However, its usefulness in delivering resistance against plant DNA viruses belonging to family Geminiviridae is still being debated. Although the RNAi approach has been shown, using a transient assay, to be useful in countering monocotyledonous plant-infecting geminiviruses of the genus Mastrevirus, it has yet to be investigated as a means of delivering resistance to dicot-infecting mastreviruses. Chickpea chlorotic dwarf Pakistan virus (CpCDPKV is a legume-infecting mastrevirus that affects chickpea and other leguminous crops in Pakistan. Results Here a hairpin (hpRNAi construct containing sequences encompassing part of replication-associated protein gene, intergenic region and part of the movement protein gene of CpCDPKV under the control of the Cauliflower mosaic virus 35S promoter has been produced and stably transformed into Nicotiana benthamiana. Plants harboring the hairpin construct were challenged with CpCDPKV. All non-transgenic N. benthamiana plants developed symptoms of CpCDPKV infection within two weeks post-inoculation. In contrast, none of the inoculated transgenic plants showed symptoms of infection and no viral DNA could be detected by Southern hybridization. A real-time quantitative PCR analysis identified very low-level accumulation of viral DNA in the inoculated transgenic plants. Conclusions The results presented show that the RNAi-based resistance strategy is useful in protecting plants from a dicot-infecting mastrevirus. The very low levels of virus detected in plant tissue of transgenic plants distal to the inoculation site suggest that virus movement and/or viral replication was impaired leading to plants that showed no discernible signs of virus infection.

  17. Enhancement of Twins Fetal ECG Signal Extraction Based on Hybrid Blind Extraction Techniques

    Directory of Open Access Journals (Sweden)

    Ahmed Kareem Abdullah

    2017-07-01

    Full Text Available ECG machines are noninvasive system used to measure the heartbeat signal. It’s very important to monitor the fetus ECG signals during pregnancy to check the heat activity and to detect any problem early before born, therefore the monitoring of ECG signals have clinical significance and importance. For multi-fetal pregnancy case the classical filtering algorithms are not sufficient to separate the ECG signals between mother and fetal. In this paper the mixture consists of mixing from three ECG signals, the first signal is the mother ECG (M-ECG signal, second signal the Fetal-1 ECG (F1-ECG, and third signal is the Fetal-2 ECG (F2-ECG, these signals are extracted based on modified blind source extraction (BSE techniques. The proposed work based on hybridization between two BSE techniques to ensure that the extracted signals separated well. The results demonstrate that the proposed work very efficiently to extract the useful ECG signals

  18. Extract of Stellerachamaejasme L(ESC) inhibits growth and metastasis of human hepatocellular carcinoma via regulating microRNA expression.

    Science.gov (United States)

    Liu, Xiaoni; Wang, Shuang; Xu, Jianji; Kou, Buxin; Chen, Dexi; Wang, Yajie; Zhu, Xiaoxin

    2018-03-20

    MicroRNAs(miRNAs)are involved in the initiation and progression of hepatocellular carcinoma. ESC, an extract of Stellerachamaejasme L, had been confirmed as a potential anti-tumor extract of Traditional Chinese Medicine. In light of the important role of miRNAs in hepatocellular carcinoma, we questioned whether the inhibitory effects of ESC on hepatocellular carcinoma (HCC) were associated with miRNAs. The proliferation inhibition of ESC on HCC cells was measured with MTT assay. The migration inhibition of ESC on HCC cells was measured with transwell assay. The influences of ESC on growth and metastasis inhibition were evaluated with xenograft tumor model of HCC. Protein expressions were measured with western blot and immunofluorescence methods and miRNA profiles were detected with miRNA array. Differential miRNA and target mRNAs were verified with real-time PCR. The results showed that ESC could inhibit proliferation and epithelial mesenchymal transition (EMT) in HCC cells in vitro and tumor growth and metastasis in xenograft models in vivo. miRNA array results showed that 69 differential miRNAs in total of 429 ones were obtained in MHCC97H cells treated by ESC. hsa-miR-107, hsa-miR-638, hsa-miR-106b-5p were selected to be validated with real-time PCR method in HepG2 and MHCC97H cells. Expressions of hsa-miR-107 and hsa-miR-638 increased obviously in HCC cells treated by ESC. Target genes of three miRNAs were also validated with real-time PCR. Interestingly, only target genes of hsa-miR-107 changed greatly. ESC downregulated the MCL1, SALL4 and BCL2 gene expressions significantly but did not influence the expression of CACNA2D1. The findings suggested ESC regressed growth and metastasis of human hepatocellular carcinoma via regulating microRNAs expression and their corresponding target genes.

  19. Shortcomings of short hairpin RNA-based transgenic RNA interference in mouse oocytes

    Czech Academy of Sciences Publication Activity Database

    Sarnová, Lenka; Malík, Radek; Sedláček, Radislav; Svoboda, Petr

    2010-01-01

    Roč. 9, č. 8 (2010), s. 1-10 ISSN 1477-5751 R&D Project s: GA MŠk ME09039 Grant - others:EMBO SDIG(DE) project 1483 Institutional research plan: CEZ:AV0Z50520514 Keywords : transgenic RNAi * shRNA * oocyte Subject RIV: EB - Genetics ; Molecular Biology http://www.jnrbm.com/content/9/1/8

  20. Shortcomings of short hairpin RNA-based transgenic RNA interference in mouse oocytes

    Czech Academy of Sciences Publication Activity Database

    Sarnová, Lenka; Malík, Radek; Sedláček, Radislav; Svoboda, Petr

    2010-01-01

    Roč. 9, č. 8 (2010), s. 1-10 ISSN 1477-5751 R&D Projects: GA MŠk ME09039 Grant - others:EMBO SDIG(DE) project 1483 Institutional research plan: CEZ:AV0Z50520514 Keywords : transgenic RNAi * shRNA * oocyte Subject RIV: EB - Genetics ; Molecular Biology http://www.jnrbm.com/content/9/1/8

  1. High-Throughput Sequencing Based Methods of RNA Structure Investigation

    DEFF Research Database (Denmark)

    Kielpinski, Lukasz Jan

    In this thesis we describe the development of four related methods for RNA structure probing that utilize massive parallel sequencing. Using them, we were able to gather structural data for multiple, long molecules simultaneously. First, we have established an easy to follow experimental...... and computational protocol for detecting the reverse transcription termination sites (RTTS-Seq). This protocol was subsequently applied to hydroxyl radical footprinting of three dimensional RNA structures to give a probing signal that correlates well with the RNA backbone solvent accessibility. Moreover, we applied...

  2. PseudoBase: a database with RNA pseudoknots.

    Science.gov (United States)

    van Batenburg, F H; Gultyaev, A P; Pleij, C W; Ng, J; Oliehoek, J

    2000-01-01

    PseudoBase is a database containing structural, functional and sequence data related to RNA pseudo-knots. It can be reached at http://wwwbio. Leiden Univ.nl/ approximately Batenburg/PKB.html. This page will direct the user to a retrieval page from where a particular pseudoknot can be chosen, or to a submission page which enables the user to add pseudoknot information to the database or to an informative page that elaborates on the various aspects of the database. For each pseudoknot, 12 items are stored, e.g. the nucleotides of the region that contains the pseudoknot, the stem positions of the pseudoknot, the EMBL accession number of the sequence that contains this pseudoknot and the support that can be given regarding the reliability of the pseudoknot. Access is via a small number of steps, using 16 different categories. The development process was done by applying the evolutionary methodology for software development rather than by applying the methodology of the classical waterfall model or the more modern spiral model.

  3. Academic Activities Transaction Extraction Based on Deep Belief Network

    Directory of Open Access Journals (Sweden)

    Xiangqian Wang

    2017-01-01

    Full Text Available Extracting information about academic activity transactions from unstructured documents is a key problem in the analysis of academic behaviors of researchers. The academic activities transaction includes five elements: person, activities, objects, attributes, and time phrases. The traditional method of information extraction is to extract shallow text features and then to recognize advanced features from text with supervision. Since the information processing of different levels is completed in steps, the error generated from various steps will be accumulated and affect the accuracy of final results. However, because Deep Belief Network (DBN model has the ability to automatically unsupervise learning of the advanced features from shallow text features, the model is employed to extract the academic activities transaction. In addition, we use character-based feature to describe the raw features of named entities of academic activity, so as to improve the accuracy of named entity recognition. In this paper, the accuracy of the academic activities extraction is compared by using character-based feature vector and word-based feature vector to express the text features, respectively, and with the traditional text information extraction based on Conditional Random Fields. The results show that DBN model is more effective for the extraction of academic activities transaction information.

  4. Effect of organic bases on extraction of gadolinium carboxylates

    International Nuclear Information System (INIS)

    Sukhan, V.V.; Frankovskij, V.A.

    1982-01-01

    The effect of pyridine, 2-aminopyridine, benzylamine, antipyrine and o-phenanthroline on the extraction of capronates and bromocapronates of gadolinium with chloroform is studied. Out of the studied organic bases benzylamine produces the highest synergetic effect. In the absence of organic bases gadolinium carboxylates, solvated by three molecules of carbonic acids, are extracted into organic phase. A possihility of extractional separation of gadolinium from comparable amounts of iron with the mixture of 1 M solutions of caproic or bromocaproic acids with 1 M benzylamine from 0.1 M solution of tartaric acids is shown [ru

  5. In Silico Meets In Vivo: Towards Computational CRISPR-Based sgRNA Design.

    Science.gov (United States)

    Chuai, Guo-Hui; Wang, Qi-Long; Liu, Qi

    2017-01-01

    CRISPR-based genome editing has been widely implemented in various cell types. In silico single guide RNA (sgRNA) design is a key step for successful gene editing using the CRISPR system, and continuing efforts are aimed at refining in silico sgRNA design with high on-target efficacy and reduced off-target effects. Many sgRNA design tools are available, but careful assessments of their application scenarios and performance benchmarks across different types of genome-editing data are needed. Efficient in silico models can be built that integrate current heterogeneous genome-editing data to derive unbiased sgRNA design rules and identify key features for improving sgRNA design. Comprehensive evaluation of on-target and off-target effects of sgRNA will allow more precise genome editing and gene therapies using the CRISPR system. Copyright © 2016 Elsevier Ltd. All rights reserved.

  6. Antioxidant properties of cumin (Bunium persicum Boiss. extract and its protective role against abiotic stress tested by microRNA markers

    Directory of Open Access Journals (Sweden)

    Katarína Ražná

    2018-02-01

    Full Text Available Bunium persicum Boiss. seeds have been used for medicinal and nutritional properties such as antioxidant, antihelmetic and antimicrobial activity. The aim of this study was to to tested protective role of cumin extract against abiotic stress by microRNA markers. Secondary also was to evaluate antioxidant activity as well as total polyphenol, flavonoid and phenolic acid content of cumin extract. We observed that cumin DNA itself has not been damaged by sonication teratment. This protective impact indicates that cumin antioxidant properties can efficiently quench free radicals induced by sonication. On the other side, ultrasound-mediated formation of reactive oxygen species did induce the DNA polymorphism of lettuce samples which was detected by miRNAs-based markers. The range of sonication impact was time-dependent. Markers based of miRNA-DNA sequences has proven to be an effective tool. We have confirmed statistically significant differences (p ≤0.01 in miRNAs markers ability to detect the polymorphism due to sonication treatment.  The antioxidant activity was determined by a method using DPPH radical and phosphomolybdenum method, total polyphenol content with Folin - Ciocalteu reagent, total flavonoid with aluminium-chloride mehod and total phenolic acid with Arnova reagent. Results showed that cumin is rich for biologically active substances and can be used more in different kind of industry as a cheap source of these substances. Antioxidant activity with DPPH method was 1.18 mg TEAC.g-1 (TEAC - Trolox equivalent antioxidant capacity per g of sample and by phosphomolybdenum method 45.23 mg TEAC.g-1. Total polyphenol content achieved value 4.22 mg GAE.g-1 (GAE - gallic acid equivalent per g of sample, total flavonoid content value 10.91 mg QE.g-1 (QE - quercetin equivalent per g of sample and total phenolic acid content value 5.07 mg CAE.g-1 (CAE - caffeic acid equivalent per g of sample.

  7. Extraction of Total DNA and RNA from Marine Filter Samples and Generation of a cDNA as Universal Template for Marker Gene Studies.

    Science.gov (United States)

    Schneider, Dominik; Wemheuer, Franziska; Pfeiffer, Birgit; Wemheuer, Bernd

    2017-01-01

    Microbial communities play an important role in marine ecosystem processes. Although the number of studies targeting marker genes such as the 16S rRNA gene has been increased in the last few years, the vast majority of marine diversity is rather unexplored. Moreover, most studies focused on the entire bacterial community and thus disregarded active microbial community players. Here, we describe a detailed protocol for the simultaneous extraction of DNA and RNA from marine water samples and for the generation of cDNA from the isolated RNA which can be used as a universal template in various marker gene studies.

  8. Statistical feature extraction based iris recognition system

    Indian Academy of Sciences (India)

    Atul Bansal

    1 Department of Electronics and Communication, G.L.A. University, 17-km stone, NH#2, Delhi-Mathura Road, .... Based upon these range of values, a decision is taken about the ...... triplet half-band filter bank and flexible k-out-of-n: A post.

  9. Tracking fungal community responses to maize plants by DNA- and RNA-based pyrosequencing.

    Directory of Open Access Journals (Sweden)

    Eiko E Kuramae

    Full Text Available We assessed soil fungal diversity and community structure at two sampling times (t1 = 47 days and t2 = 104 days of plant age in pots associated with four maize cultivars, including two genetically modified (GM cultivars by high-throughput pyrosequencing of the 18S rRNA gene using DNA and RNA templates. We detected no significant differences in soil fungal diversity and community structure associated with different plant cultivars. However, DNA-based analyses yielded lower fungal OTU richness as compared to RNA-based analyses. Clear differences in fungal community structure were also observed in relation to sampling time and the nucleic acid pool targeted (DNA versus RNA. The most abundant soil fungi, as recovered by DNA-based methods, did not necessary represent the most "active" fungi (as recovered via RNA. Interestingly, RNA-derived community compositions at t1 were highly similar to DNA-derived communities at t2, based on presence/absence measures of OTUs. We recovered large proportions of fungal sequences belonging to arbuscular mycorrhizal fungi and Basidiomycota, especially at the RNA level, suggesting that these important and potentially beneficial fungi are not affected by the plant cultivars nor by GM traits (Bt toxin production. Our results suggest that even though DNA- and RNA-derived soil fungal communities can be very different at a given time, RNA composition may have a predictive power of fungal community development through time.

  10. Synthetic biology devices and circuits for RNA-based 'smart vaccines': a propositional review.

    Science.gov (United States)

    Andries, Oliwia; Kitada, Tasuku; Bodner, Katie; Sanders, Niek N; Weiss, Ron

    2015-02-01

    Nucleic acid vaccines have been gaining attention as an alternative to the standard attenuated pathogen or protein based vaccine. However, an unrealized advantage of using such DNA or RNA based vaccination modalities is the ability to program within these nucleic acids regulatory devices that would provide an immunologist with the power to control the production of antigens and adjuvants in a desirable manner by administering small molecule drugs as chemical triggers. Advances in synthetic biology have resulted in the creation of highly predictable and modular genetic parts and devices that can be composed into synthetic gene circuits with complex behaviors. With the recent advent of modified RNA gene delivery methods and developments in the RNA replicon platform, we foresee a future in which mammalian synthetic biologists will create genetic circuits encoded exclusively on RNA. Here, we review the current repertoire of devices used in RNA synthetic biology and propose how programmable 'smart vaccines' will revolutionize the field of RNA vaccination.

  11. GARN: Sampling RNA 3D Structure Space with Game Theory and Knowledge-Based Scoring Strategies.

    Science.gov (United States)

    Boudard, Mélanie; Bernauer, Julie; Barth, Dominique; Cohen, Johanne; Denise, Alain

    2015-01-01

    Cellular processes involve large numbers of RNA molecules. The functions of these RNA molecules and their binding to molecular machines are highly dependent on their 3D structures. One of the key challenges in RNA structure prediction and modeling is predicting the spatial arrangement of the various structural elements of RNA. As RNA folding is generally hierarchical, methods involving coarse-grained models hold great promise for this purpose. We present here a novel coarse-grained method for sampling, based on game theory and knowledge-based potentials. This strategy, GARN (Game Algorithm for RNa sampling), is often much faster than previously described techniques and generates large sets of solutions closely resembling the native structure. GARN is thus a suitable starting point for the molecular modeling of large RNAs, particularly those with experimental constraints. GARN is available from: http://garn.lri.fr/.

  12. Novel thiosalicylate-based ionic liquids for heavy metal extractions

    Energy Technology Data Exchange (ETDEWEB)

    Leyma, Raphlin; Platzer, Sonja [Institute of Inorganic Chemistry, Faculty of Chemistry, University of Vienna, Waehringer Str. 42, A-1090 Vienna (Austria); Jirsa, Franz [Institute of Inorganic Chemistry, Faculty of Chemistry, University of Vienna, Waehringer Str. 42, A-1090 Vienna (Austria); Department of Zoology, University of Johannesburg, PO Box 524, Auckland Park, 2006, Johannesburg (South Africa); Kandioller, Wolfgang, E-mail: wolfgang.kandioller@univie.ac.at [Institute of Inorganic Chemistry, Faculty of Chemistry, University of Vienna, Waehringer Str. 42, A-1090 Vienna (Austria); Krachler, Regina; Keppler, Bernhard K. [Institute of Inorganic Chemistry, Faculty of Chemistry, University of Vienna, Waehringer Str. 42, A-1090 Vienna (Austria)

    2016-08-15

    Highlights: • Six thiosalicylate-based ammonium and phosphonium ionic liquids (ILs) were newly synthesized. • ILs showed good extraction of cadmium, copper, and zinc. • Phosphonium ILs showed better extraction efficiencies than their ammonium counterparts. - Abstract: This study aims to develop novel ammonium and phosphonium ionic liquids (ILs) with thiosalicylate (TS) derivatives as anions and evaluate their extracting efficiencies towards heavy metals in aqueous solutions. Six ILs were synthesized, characterized, and investigated for their extracting efficacies for cadmium, copper, and zinc. Liquid-liquid extractions of Cu, Zn, or Cd with ILs after 1–24 h using model solutions (pH 7; 0.1 M CaCl{sub 2}) were assessed using flame atomic absorption spectroscopy (F-AAS). Phosphonium-based ILs trihexyltetradecylphosphonium 2-(propylthio)benzoate [P{sub 66614}][PTB] and 2-(benzylthio)benzoate [P{sub 66614}][BTB] showed best extraction efficiency for copper and cadmium, respectively and zinc was extracted to a high degree by [P{sub 66614}][BTB] exclusively.

  13. RNA-TVcurve: a Web server for RNA secondary structure comparison based on a multi-scale similarity of its triple vector curve representation.

    Science.gov (United States)

    Li, Ying; Shi, Xiaohu; Liang, Yanchun; Xie, Juan; Zhang, Yu; Ma, Qin

    2017-01-21

    RNAs have been found to carry diverse functionalities in nature. Inferring the similarity between two given RNAs is a fundamental step to understand and interpret their functional relationship. The majority of functional RNAs show conserved secondary structures, rather than sequence conservation. Those algorithms relying on sequence-based features usually have limitations in their prediction performance. Hence, integrating RNA structure features is very critical for RNA analysis. Existing algorithms mainly fall into two categories: alignment-based and alignment-free. The alignment-free algorithms of RNA comparison usually have lower time complexity than alignment-based algorithms. An alignment-free RNA comparison algorithm was proposed, in which novel numerical representations RNA-TVcurve (triple vector curve representation) of RNA sequence and corresponding secondary structure features are provided. Then a multi-scale similarity score of two given RNAs was designed based on wavelet decomposition of their numerical representation. In support of RNA mutation and phylogenetic analysis, a web server (RNA-TVcurve) was designed based on this alignment-free RNA comparison algorithm. It provides three functional modules: 1) visualization of numerical representation of RNA secondary structure; 2) detection of single-point mutation based on secondary structure; and 3) comparison of pairwise and multiple RNA secondary structures. The inputs of the web server require RNA primary sequences, while corresponding secondary structures are optional. For the primary sequences alone, the web server can compute the secondary structures using free energy minimization algorithm in terms of RNAfold tool from Vienna RNA package. RNA-TVcurve is the first integrated web server, based on an alignment-free method, to deliver a suite of RNA analysis functions, including visualization, mutation analysis and multiple RNAs structure comparison. The comparison results with two popular RNA

  14. A semi-supervised learning framework for biomedical event extraction based on hidden topics.

    Science.gov (United States)

    Zhou, Deyu; Zhong, Dayou

    2015-05-01

    Scientists have devoted decades of efforts to understanding the interaction between proteins or RNA production. The information might empower the current knowledge on drug reactions or the development of certain diseases. Nevertheless, due to the lack of explicit structure, literature in life science, one of the most important sources of this information, prevents computer-based systems from accessing. Therefore, biomedical event extraction, automatically acquiring knowledge of molecular events in research articles, has attracted community-wide efforts recently. Most approaches are based on statistical models, requiring large-scale annotated corpora to precisely estimate models' parameters. However, it is usually difficult to obtain in practice. Therefore, employing un-annotated data based on semi-supervised learning for biomedical event extraction is a feasible solution and attracts more interests. In this paper, a semi-supervised learning framework based on hidden topics for biomedical event extraction is presented. In this framework, sentences in the un-annotated corpus are elaborately and automatically assigned with event annotations based on their distances to these sentences in the annotated corpus. More specifically, not only the structures of the sentences, but also the hidden topics embedded in the sentences are used for describing the distance. The sentences and newly assigned event annotations, together with the annotated corpus, are employed for training. Experiments were conducted on the multi-level event extraction corpus, a golden standard corpus. Experimental results show that more than 2.2% improvement on F-score on biomedical event extraction is achieved by the proposed framework when compared to the state-of-the-art approach. The results suggest that by incorporating un-annotated data, the proposed framework indeed improves the performance of the state-of-the-art event extraction system and the similarity between sentences might be precisely

  15. Antioxidant properties of cumin (Bunium persicum Boiss.) extract and its protective role against abiotic stress tested by microRNA markers

    OpenAIRE

    Katarína Ražná; Nishonoy Khasanova; Eva Ivanišová; Davranov Qahramon; Miroslav Habán

    2018-01-01

    Bunium persicum Boiss. seeds have been used for medicinal and nutritional properties such as antioxidant, antihelmetic and antimicrobial activity. The aim of this study was to to tested protective role of cumin extract against abiotic stress by microRNA markers. Secondary also was to evaluate antioxidant activity as well as total polyphenol, flavonoid and phenolic acid content of cumin extract. We observed that cumin DNA itself has not been damaged by sonication teratment. This protective im...

  16. Splicing of a C. elegans myosin pre-mRNA in a human nuclear extract

    Energy Technology Data Exchange (ETDEWEB)

    Ogg, S C; Anderson, P; Wickens, M P [Univ. of Wisconsin, Madison (USA)

    1990-01-11

    Splicing of mammalian introns requires that the intron possess at least 80 nucleotides. This length requirement presumably reflects the constraints of accommodating multiple snRNPs simultaneously in the same intro. In the free-living nematode, C. elegans, introns typically are 45 to 55 nucleotides in length. In this report, the authors determine whether C. elegans introns can obviate the mammalian length requirement by virtue of their structure or sequence. They demonstrate that a 53 nucleotide intron from the unc-54 gene of C. elegans does not undergo splicing in a mammalian (HeLa) nuclear extract. However, insertion of 31 nucleotides of foreign, prokaryotic sequence into the same intron results in efficient splicing. The observed splicing proceeds by the same two-step mechanism observed with mammalian introns, and exploits the same 3{prime} and 5{prime} sites as are used in C. elegans. The branch point used lies in the inserted sequences. They conclude that C. elegans splicing components are either fewer in number or smaller than their mammalian counterparts.

  17. Noninvasive extraction of fetal electrocardiogram based on Support Vector Machine

    Science.gov (United States)

    Fu, Yumei; Xiang, Shihan; Chen, Tianyi; Zhou, Ping; Huang, Weiyan

    2015-10-01

    The fetal electrocardiogram (FECG) signal has important clinical value for diagnosing the fetal heart diseases and choosing suitable therapeutics schemes to doctors. So, the noninvasive extraction of FECG from electrocardiogram (ECG) signals becomes a hot research point. A new method, the Support Vector Machine (SVM) is utilized for the extraction of FECG with limited size of data. Firstly, the theory of the SVM and the principle of the extraction based on the SVM are studied. Secondly, the transformation of maternal electrocardiogram (MECG) component in abdominal composite signal is verified to be nonlinear and fitted with the SVM. Then, the SVM is trained, and the training results are compared with the real data to ensure the effect of the training. Meanwhile, the parameters of the SVM are optimized to achieve the best performance so that the learning machine can be utilized to fit the unknown samples. Finally, the FECG is extracted by removing the optimal estimation of MECG component from the abdominal composite signal. In order to evaluate the performance of FECG extraction based on the SVM, the Signal-to-Noise Ratio (SNR) and the visual test are used. The experimental results show that the FECG with good quality can be extracted, its SNR ratio is significantly increased as high as 9.2349 dB and the time cost is significantly decreased as short as 0.802 seconds. Compared with the traditional method, the noninvasive extraction method based on the SVM has a simple realization, the shorter treatment time and the better extraction quality under the same conditions.

  18. Level Sets and Voronoi based Feature Extraction from any Imagery

    DEFF Research Database (Denmark)

    Sharma, O.; Anton, François; Mioc, Darka

    2012-01-01

    Polygon features are of interest in many GEOProcessing applications like shoreline mapping, boundary delineation, change detection, etc. This paper presents a unique new GPU-based methodology to automate feature extraction combining level sets, or mean shift based segmentation together with Voron...

  19. Design of guanidinium ionic liquid based microwave-assisted extraction for the efficient extraction of Praeruptorin A from Radix peucedani.

    Science.gov (United States)

    Ding, Xueqin; Li, Li; Wang, Yuzhi; Chen, Jing; Huang, Yanhua; Xu, Kaijia

    2014-12-01

    A series of novel tetramethylguanidinium ionic liquids and hexaalkylguanidinium ionic liquids have been synthesized based on 1,1,3,3-tetramethylguanidine. The structures of the ionic liquids were confirmed by (1)H NMR spectroscopy and mass spectrometry. A green guanidinium ionic liquid based microwave-assisted extraction method has been developed with these guanidinium ionic liquids for the effective extraction of Praeruptorin A from Radix peucedani. After extraction, reversed-phase high-performance liquid chromatography with UV detection was employed for the analysis of Praeruptorin A. Several significant operating parameters were systematically optimized by single-factor and L9 (3(4)) orthogonal array experiments. The amount of Praeruptorin A extracted by [1,1,3,3-tetramethylguanidine]CH2CH(OH)COOH is the highest, reaching 11.05 ± 0.13 mg/g. Guanidinium ionic liquid based microwave-assisted extraction presents unique advantages in Praeruptorin A extraction compared with guanidinium ionic liquid based maceration extraction, guanidinium ionic liquid based heat reflux extraction and guanidinium ionic liquid based ultrasound-assisted extraction. The precision, stability, and repeatability of the process were investigated. The mechanisms of guanidinium ionic liquid based microwave-assisted extraction were researched by scanning electron microscopy and IR spectroscopy. All the results show that guanidinium ionic liquid based microwave-assisted extraction has a huge potential in the extraction of bioactive compounds from complex samples. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. Effect of Green Tea Extract on Systemic Metabolic Homeostasis in Diet-Induced Obese Mice Determined via RNA-Seq Transcriptome Profiles

    Directory of Open Access Journals (Sweden)

    Ji-Young Choi

    2016-10-01

    Full Text Available Green tea (GT has various health effects, including anti-obesity properties. However, the multiple molecular mechanisms of the effects have not been fully determined. The aim of this study was to elucidate the anti-obesity effects of GT via the analysis of its metabolic and transcriptional responses based on RNA-seq profiles. C57BL/6J mice were fed a normal, high-fat (60% energy as fat, or high-fat + 0.25% (w/w GT diet for 12 weeks. The GT extract ameliorated obesity, hepatic steatosis, dyslipidemia, and insulin resistance in diet-induced obesity (DIO mice. GT supplementation resulted in body weight gain reduction than mice fed high-fat through enhanced energy expenditure, and reduced adiposity. The transcriptome profiles of epididymal white adipose tissue (eWAT suggested that GT augments transcriptional responses to the degradation of branched chain amino acids (BCAAs, as well as AMP-activated protein kinase (AMPK signaling, which suggests enhanced energy homeostasis. Our findings provide some significant insights into the effects of GT for the prevention of obesity and its comorbidities. We demonstrated that the GT extract contributed to the regulation of systemic metabolic homeostasis via transcriptional responses to not only lipid and glucose metabolism, but also amino acid metabolism via BCAA degradation in the adipose tissue of DIO mice.

  1. A need for standardization in drinking water analysis – an investigation of DNA extraction procedure, primer choice and detection limit of 16S rRNA amplicon sequencing

    DEFF Research Database (Denmark)

    Brandt, Jakob; Nielsen, Per Halkjær; Albertsen, Mads

    have been made to illuminate the effects specifically related to bacterial communities in drinking water. In this study, we investigated the impact of the DNA extraction and primer choice on the observed community structure, and we also estimated the detection limit of the 16S rRNA amplicon sequencing...

  2. Learning-based meta-algorithm for MRI brain extraction.

    Science.gov (United States)

    Shi, Feng; Wang, Li; Gilmore, John H; Lin, Weili; Shen, Dinggang

    2011-01-01

    Multiple-segmentation-and-fusion method has been widely used for brain extraction, tissue segmentation, and region of interest (ROI) localization. However, such studies are hindered in practice by their computational complexity, mainly coming from the steps of template selection and template-to-subject nonlinear registration. In this study, we address these two issues and propose a novel learning-based meta-algorithm for MRI brain extraction. Specifically, we first use exemplars to represent the entire template library, and assign the most similar exemplar to the test subject. Second, a meta-algorithm combining two existing brain extraction algorithms (BET and BSE) is proposed to conduct multiple extractions directly on test subject. Effective parameter settings for the meta-algorithm are learned from the training data and propagated to subject through exemplars. We further develop a level-set based fusion method to combine multiple candidate extractions together with a closed smooth surface, for obtaining the final result. Experimental results show that, with only a small portion of subjects for training, the proposed method is able to produce more accurate and robust brain extraction results, at Jaccard Index of 0.956 +/- 0.010 on total 340 subjects under 6-fold cross validation, compared to those by the BET and BSE even using their best parameter combinations.

  3. Lipid-Based Liquid Crystalline Nanoparticles Facilitate Cytosolic Delivery of siRNA via Structural Transformation.

    Science.gov (United States)

    He, Shufang; Fan, Weiwei; Wu, Na; Zhu, Jingjing; Miao, Yunqiu; Miao, Xiaran; Li, Feifei; Zhang, Xinxin; Gan, Yong

    2018-04-11

    RNA interference (RNAi) technology has shown great promise for the treatment of cancer and other genetic disorders. Despite the efforts to increase the target tissue distribution, the safe and effective delivery of siRNA to the diseased cells with sufficient cytosolic transport is another critical factor for successful RNAi clinical application. Here, the constructed lipid-based liquid crystalline nanoparticles, called nano-Transformers, can transform thestructure in the intracellular acidic environment and perform high-efficient siRNA delivery for cancer treatment. The developed nano-Transformers have satisfactory siRNA loading efficiency and low cytotoxicity. Different from the traditional cationic nanocarriers, the endosomal membrane fusion induced by the conformational transition of lipids contributes to the easy dissociation of siRNA from nanocarriers and direct release of free siRNA into cytoplasm. We show that transfection with cyclin-dependent kinase 1 (CDK1)-siRNA-loaded nano-Transformers causes up to 95% reduction of relevant mRNA in vitro and greatly inhibits the tumor growth without causing any immunogenic response in vivo. This work highlights that the lipid-based nano-Transformers may become the next generation of siRNA delivery system with higher efficacy and improved safety profiles.

  4. Skeleton extraction based on the topology and Snakes model

    Directory of Open Access Journals (Sweden)

    Yuanxue Cai

    Full Text Available A new skeleton line extraction method based on topology and flux is proposed by analyzing the distribution characteristics of the gradient vector field in the Snakes model. The distribution characteristics of the skeleton line are accurately obtained by calculating the eigenvalues of the critical points and the flux of the gradient vector field. Then the skeleton lines can be effectively extracted. The results also show that there is no need for the pretreatment or binarization of the target image. The skeleton lines of complex gray images such as optical interference patterns can be effectively extracted by using this method. Compared to traditional methods, this method has many advantages, such as high extraction accuracy and fast processing speed. Keywords: Skeleton, Snakes model, Topology, Photoelasticity image

  5. Olea europaea leaf extract improves the treatment response of GBM stem cells by modulating miRNA expression.

    Science.gov (United States)

    Tezcan, Gulcin; Tunca, Berrin; Bekar, Ahmet; Budak, Ferah; Sahin, Saliha; Cecener, Gulsah; Egeli, Unal; Taskapılıoglu, Mevlut Ozgur; Kocaeli, Hasan; Tolunay, Sahsine; Malyer, Hulusi; Demir, Cevdet; Tumen, Gulendam

    2014-01-01

    The stem-like cells of Glioblastoma multiforme (GBM) tumors (GSCs) are one of the important determinants of recurrence and drug resistance. The aims of the current study were to evaluate the anticancer effect of Olea europaea leaf extract (OLE) on GBM cell lines, the association between OLE and TMZ responses, and the effect of OLE and the OLE-TMZ combination in GSCs and to clarify the molecular mechanism of this effect on the expression of miRNAs related to cell death. The anti-proliferative activity of OLE and the effect of the OLE-TMZ combination were tested in the T98G, U-138MG and U-87MG GBM cell lines using WST-1 assay. The mechanism of cell death was analyzed with Annexin V/FITC and TUNEL assays. The effects of OLE on the expression levels of miR-181b, miR-153, miR-145 and miR-137 and potential mRNA targets were analyzed in GSCs using RT-qPCR. OLE exhibited anti-proliferative effects via apoptosis and necrosis in the GBM cell lines. In addition, OLE significantly induced the expression of miR-153, miR-145, and miR-137 and decreased the expression of the target genes of these miRNAs in GSCs (p GBM cells with different TMZ responses, and this effect is synergistically increased when the cells are treated with a combination of OLE and TMZ. This is the first study to indicate that OLE may interfere with the pluripotency of GSCs by modulating miRNA expression. Further studies are required, but we suggest that OLE may have a potential for advanced therapeutic cancer drug studies in GBM.

  6. Internal control for real-time polymerase chain reaction based on MS2 bacteriophage for RNA viruses diagnostics.

    Science.gov (United States)

    Zambenedetti, Miriam Ribas; Pavoni, Daniela Parada; Dallabona, Andreia Cristine; Dominguez, Alejandro Correa; Poersch, Celina de Oliveira; Fragoso, Stenio Perdigão; Krieger, Marco Aurélio

    2017-05-01

    Real-time reverse transcription polymerase chain reaction (RT-PCR) is routinely used to detect viral infections. In Brazil, it is mandatory the use of nucleic acid tests to detect hepatitis C virus (HCV), hepatitis B virus and human immunodeficiency virus in blood banks because of the immunological window. The use of an internal control (IC) is necessary to differentiate the true negative results from those consequent from a failure in some step of the nucleic acid test. The aim of this study was the construction of virus-modified particles, based on MS2 bacteriophage, to be used as IC for the diagnosis of RNA viruses. The MS2 genome was cloned into the pET47b(+) plasmid, generating pET47b(+)-MS2. MS2-like particles were produced through the synthesis of MS2 RNA genome by T7 RNA polymerase. These particles were used as non-competitive IC in assays for RNA virus diagnostics. In addition, a competitive control for HCV diagnosis was developed by cloning a mutated HCV sequence into the MS2 replicase gene of pET47b(+)-MS2, which produces a non-propagating MS2 particle. The utility of MS2-like particles as IC was evaluated in a one-step format multiplex real-time RT-PCR for HCV detection. We demonstrated that both competitive and non-competitive IC could be successfully used to monitor the HCV amplification performance, including the extraction, reverse transcription, amplification and detection steps, without compromising the detection of samples with low target concentrations. In conclusion, MS2-like particles generated by this strategy proved to be useful IC for RNA virus diagnosis, with advantage that they are produced by a low cost protocol. An attractive feature of this system is that it allows the construction of a multicontrol by the insertion of sequences from more than one pathogen, increasing its applicability for diagnosing different RNA viruses.

  7. Comparison of RNA-seq and microarray-based models for clinical endpoint prediction.

    Science.gov (United States)

    Zhang, Wenqian; Yu, Ying; Hertwig, Falk; Thierry-Mieg, Jean; Zhang, Wenwei; Thierry-Mieg, Danielle; Wang, Jian; Furlanello, Cesare; Devanarayan, Viswanath; Cheng, Jie; Deng, Youping; Hero, Barbara; Hong, Huixiao; Jia, Meiwen; Li, Li; Lin, Simon M; Nikolsky, Yuri; Oberthuer, André; Qing, Tao; Su, Zhenqiang; Volland, Ruth; Wang, Charles; Wang, May D; Ai, Junmei; Albanese, Davide; Asgharzadeh, Shahab; Avigad, Smadar; Bao, Wenjun; Bessarabova, Marina; Brilliant, Murray H; Brors, Benedikt; Chierici, Marco; Chu, Tzu-Ming; Zhang, Jibin; Grundy, Richard G; He, Min Max; Hebbring, Scott; Kaufman, Howard L; Lababidi, Samir; Lancashire, Lee J; Li, Yan; Lu, Xin X; Luo, Heng; Ma, Xiwen; Ning, Baitang; Noguera, Rosa; Peifer, Martin; Phan, John H; Roels, Frederik; Rosswog, Carolina; Shao, Susan; Shen, Jie; Theissen, Jessica; Tonini, Gian Paolo; Vandesompele, Jo; Wu, Po-Yen; Xiao, Wenzhong; Xu, Joshua; Xu, Weihong; Xuan, Jiekun; Yang, Yong; Ye, Zhan; Dong, Zirui; Zhang, Ke K; Yin, Ye; Zhao, Chen; Zheng, Yuanting; Wolfinger, Russell D; Shi, Tieliu; Malkas, Linda H; Berthold, Frank; Wang, Jun; Tong, Weida; Shi, Leming; Peng, Zhiyu; Fischer, Matthias

    2015-06-25

    Gene expression profiling is being widely applied in cancer research to identify biomarkers for clinical endpoint prediction. Since RNA-seq provides a powerful tool for transcriptome-based applications beyond the limitations of microarrays, we sought to systematically evaluate the performance of RNA-seq-based and microarray-based classifiers in this MAQC-III/SEQC study for clinical endpoint prediction using neuroblastoma as a model. We generate gene expression profiles from 498 primary neuroblastomas using both RNA-seq and 44 k microarrays. Characterization of the neuroblastoma transcriptome by RNA-seq reveals that more than 48,000 genes and 200,000 transcripts are being expressed in this malignancy. We also find that RNA-seq provides much more detailed information on specific transcript expression patterns in clinico-genetic neuroblastoma subgroups than microarrays. To systematically compare the power of RNA-seq and microarray-based models in predicting clinical endpoints, we divide the cohort randomly into training and validation sets and develop 360 predictive models on six clinical endpoints of varying predictability. Evaluation of factors potentially affecting model performances reveals that prediction accuracies are most strongly influenced by the nature of the clinical endpoint, whereas technological platforms (RNA-seq vs. microarrays), RNA-seq data analysis pipelines, and feature levels (gene vs. transcript vs. exon-junction level) do not significantly affect performances of the models. We demonstrate that RNA-seq outperforms microarrays in determining the transcriptomic characteristics of cancer, while RNA-seq and microarray-based models perform similarly in clinical endpoint prediction. Our findings may be valuable to guide future studies on the development of gene expression-based predictive models and their implementation in clinical practice.

  8. Quantitation of base substitutions in eukaryotic 5S rRNA: selection for the maintenance of RNA secondary structure.

    Science.gov (United States)

    Curtiss, W C; Vournakis, J N

    1984-01-01

    Eukaryotic 5S rRNA sequences from 34 diverse species were compared by the following method: (1) The sequences were aligned; (2) the positions of substitutions were located by comparison of all possible pairs of sequences; (3) the substitution sites were mapped to an assumed general base pairing model; and (4) the R-Y model of base stacking was used to study stacking pattern relationships in the structure. An analysis of the sequence and structure variability in each region of the molecule is presented. It was found that the degree of base substitution varies over a wide range, from absolute conservation to occurrence of over 90% of the possible observable substitutions. The substitutions are located primarily in stem regions of the 5S rRNA secondary structure. More than 88% of the substitutions in helical regions maintain base pairing. The disruptive substitutions are primarily located at the edges of helical regions, resulting in shortening of the helical regions and lengthening of the adjacent nonpaired regions. Base stacking patterns determined by the R-Y model are mapped onto the general secondary structure. Intrastrand and interstrand stacking could stabilize alternative coaxial structures and limit the conformational flexibility of nonpaired regions. Two short contiguous regions are 100% conserved in all species. This may reflect evolutionary constraints imposed at the DNA level by the requirement for binding of a 5S gene transcription initiation factor during gene expression.

  9. Determination of Resistant Starch Assimilating Bacteria in Fecal Samples of Mice by In vitro RNA-Based Stable Isotope Probing

    Science.gov (United States)

    Herrmann, Elena; Young, Wayne; Rosendale, Douglas; Conrad, Ralf; Riedel, Christian U.; Egert, Markus

    2017-01-01

    The impact of the intestinal microbiota on human health is becoming increasingly appreciated in recent years. In consequence, and fueled by major technological advances, the composition of the intestinal microbiota in health and disease has been intensively studied by high throughput sequencing approaches. Observations linking dysbiosis of the intestinal microbiota with a number of serious medical conditions including chronic inflammatory disorders and allergic diseases suggest that restoration of the composition and activity of the intestinal microbiota may be a treatment option at least for some of these diseases. One possibility to shape the intestinal microbiota is the administration of prebiotic carbohydrates such as resistant starch (RS). In the present study, we aim at establishing RNA-based stable isotope probing (RNA-SIP) to identify bacterial populations that are involved in the assimilation of RS using anaerobic in vitro fermentation of murine fecal material with stable [U13C] isotope-labeled potato starch. Total RNA from these incubations was extracted, processed by gradient ultracentrifugation and fractionated by density. 16S rRNA gene sequences were amplified from reverse transcribed RNA of high and low density fractions suspected to contain labeled and unlabeled RNA, respectively. Phylogenetic analysis of the obtained sequences revealed a distinct subset of the intestinal microbiota involved in starch metabolism. The results suggest Bacteroidetes, in particular genera affiliated with Prevotellaceae, as well as members of the Ruminococcacea family to be primary assimilators of resistant starch due to a significantly higher relative abundance in higher density fractions in RNA samples isolated after 2 h of incubation. Using high performance liquid chromatography coupled to isotope ratio mass spectrometry (HPLC-IRMS) analysis, some stable isotope label was recovered from acetate, propionate and butyrate. Here, we demonstrate the suitability of RNA

  10. Determination of Resistant Starch Assimilating Bacteria in Fecal Samples of Mice by In vitro RNA-Based Stable Isotope Probing

    Directory of Open Access Journals (Sweden)

    Elena Herrmann

    2017-07-01

    Full Text Available The impact of the intestinal microbiota on human health is becoming increasingly appreciated in recent years. In consequence, and fueled by major technological advances, the composition of the intestinal microbiota in health and disease has been intensively studied by high throughput sequencing approaches. Observations linking dysbiosis of the intestinal microbiota with a number of serious medical conditions including chronic inflammatory disorders and allergic diseases suggest that restoration of the composition and activity of the intestinal microbiota may be a treatment option at least for some of these diseases. One possibility to shape the intestinal microbiota is the administration of prebiotic carbohydrates such as resistant starch (RS. In the present study, we aim at establishing RNA-based stable isotope probing (RNA-SIP to identify bacterial populations that are involved in the assimilation of RS using anaerobic in vitro fermentation of murine fecal material with stable [U13C] isotope-labeled potato starch. Total RNA from these incubations was extracted, processed by gradient ultracentrifugation and fractionated by density. 16S rRNA gene sequences were amplified from reverse transcribed RNA of high and low density fractions suspected to contain labeled and unlabeled RNA, respectively. Phylogenetic analysis of the obtained sequences revealed a distinct subset of the intestinal microbiota involved in starch metabolism. The results suggest Bacteroidetes, in particular genera affiliated with Prevotellaceae, as well as members of the Ruminococcacea family to be primary assimilators of resistant starch due to a significantly higher relative abundance in higher density fractions in RNA samples isolated after 2 h of incubation. Using high performance liquid chromatography coupled to isotope ratio mass spectrometry (HPLC-IRMS analysis, some stable isotope label was recovered from acetate, propionate and butyrate. Here, we demonstrate the

  11. Investigation of RNA Structure by High-Throughput SHAPE-Based Probing Methods

    DEFF Research Database (Denmark)

    Poulsen, Line Dahl

    of highthroughput SHAPE-based approaches to investigate RNA structure based on novel SHAPE reagents that permit selection of full-length cDNAs. The SHAPE Selection (SHAPES) method is applied to the foot-and-mouth disease virus (FMDV) plus strand RNA genome, and the data is used to construct a genome-wide structural...... that they are functional. The SHAPES method is further applied to the hepatitis C virus (HCV), where the data is used to refine known and predicted structures. Over the past years, the interest of studying RNA structure in their native environment has been increased, and to allow studying RNA structure inside living cells...... using the SHAPE Selection approach, I introduce a biotinylated probing reagent. This chemical can cross cell membranes and reacts with RNA inside the cells, allowing the structural conformations to be studied in the context of physiological relevant conditions in living cells. The methods and results...

  12. Smart Inulin-Based Polycationic Nanodevices for siRNA Delivery.

    Science.gov (United States)

    Cavallaro, G; Sardo, C; Scialabba, C; Licciardi, M; Giammona, G

    2017-01-01

    The advances of short interfering RNA (siRNA) mediated therapy provide a powerful option for the treatment of many diseases by silencing the expression of targeted genes including cancer development and progression. Inulin is a very simple and biocompatible polysaccharide proposed by our groups to produce interesting delivery systems for Nucleic Acid Based Drugs (NABDs), such as siRNA, either as polycations able to give polyplexes and polymeric coatings for nanosystems having a metallic core. In this research field, different functionalizing groups were linked to the inulin backbone with specific aims including oligoamine such as Ethylendiammine (EDA), Diethylediamine (DETA), Spermine, (SPM) etc. In this contribution the main Inulin-based nanodevices for the delivery of siRNA have been reported, analysed and compared with particular reference to their chemical design and structure, biocompatibility, siRNA complexing ability, silencing ability. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  13. An optimised method for the extraction of bacterial mRNA from plant roots infected with Escherichia coli O157:H7

    Directory of Open Access Journals (Sweden)

    Ashleigh eHolmes

    2014-06-01

    Full Text Available Analysis of microbial gene expression during host colonisation provides valuable information on the nature of interaction, beneficial or pathogenic, and the adaptive processes involved. Isolation of bacterial mRNA for in planta analysis can be challenging where host nucleic acid may dominate the preparation, or inhibitory compounds affect downstream analysis, e.g. qPCR, microarray or RNA-seq. The goal of this work was to optimise the isolation of bacterial mRNA of food-borne pathogens from living plants. Reported methods for recovery of phytopathogen-infected plant material, using hot phenol extraction and high concentration of bacterial inoculation or large amounts of infected tissues, were found to be inappropriate for plant roots inoculated with Escherichia coli O157:H7. The bacterial RNA yields were too low and increased plant material resulted in a dominance of plant RNA in the sample. To improve the yield of bacterial RNA and reduce the number of plants required, an optimised method was developed which combines bead beating with directed bacterial lysis using SDS and lysozyme. Inhibitory plant compounds, such as phenolics and polysaccharides, were counteracted with the addition of HMW-PEG and CTAB. The new method increased the total yield of bacterial mRNA substantially and allowed assessment of gene expression by qPCR. This method can be applied to other bacterial species associated with plant roots, and also in the wider context of food safety.

  14. Isolation of Microarray-Grade Total RNA, MicroRNA, and DNA from a Single PAXgene Blood RNA Tube

    DEFF Research Database (Denmark)

    Kruhøffer, Mogens; Andersen, Lars Dyrskjøt; Voss, Thorsten

    2007-01-01

    We have developed a procedure for isolation of microRNA and genomic DNA in addition to total RNA from whole blood stabilized in PAXgene Blood RNA tubes. The procedure is based on automatic extraction on a BioRobot MDx and includes isolation of DNA from a fraction of the stabilized blood...... and recovery of small RNA species that are otherwise lost. The procedure presented here is suitable for large-scale experiments and is amenable to further automation. Procured total RNA and DNA was tested using Affymetrix Expression and single-nucleotide polymorphism GeneChips, respectively, and isolated micro......RNA was tested using spotted locked nucleic acid-based microarrays. We conclude that the yield and quality of total RNA, microRNA, and DNA from a single PAXgene blood RNA tube is sufficient for downstream microarray analysis....

  15. Remaining useful life estimation based on discriminating shapelet extraction

    International Nuclear Information System (INIS)

    Malinowski, Simon; Chebel-Morello, Brigitte; Zerhouni, Noureddine

    2015-01-01

    In the Prognostics and Health Management domain, estimating the remaining useful life (RUL) of critical machinery is a challenging task. Various research topics including data acquisition, fusion, diagnostics and prognostics are involved in this domain. This paper presents an approach, based on shapelet extraction, to estimate the RUL of equipment. This approach extracts, in an offline step, discriminative rul-shapelets from an history of run-to-failure data. These rul-shapelets are patterns that are selected for their correlation with the remaining useful life of the equipment. In other words, every selected rul-shapelet conveys its own information about the RUL of the equipment. In an online step, these rul-shapelets are compared to testing units and the ones that match these units are used to estimate their RULs. Therefore, RUL estimation is based on patterns that have been selected for their high correlation with the RUL. This approach is different from classical similarity-based approaches that attempt to match complete testing units (or only late instants of testing units) with training ones to estimate the RUL. The performance of our approach is evaluated on a case study on the remaining useful life estimation of turbofan engines and performance is compared with other similarity-based approaches. - Highlights: • A data-driven RUL estimation technique based on pattern extraction is proposed. • Patterns are extracted for their correlation with the RUL. • The proposed method shows good performance compared to other techniques

  16. Model-based Bayesian signal extraction algorithm for peripheral nerves

    Science.gov (United States)

    Eggers, Thomas E.; Dweiri, Yazan M.; McCallum, Grant A.; Durand, Dominique M.

    2017-10-01

    Objective. Multi-channel cuff electrodes have recently been investigated for extracting fascicular-level motor commands from mixed neural recordings. Such signals could provide volitional, intuitive control over a robotic prosthesis for amputee patients. Recent work has demonstrated success in extracting these signals in acute and chronic preparations using spatial filtering techniques. These extracted signals, however, had low signal-to-noise ratios and thus limited their utility to binary classification. In this work a new algorithm is proposed which combines previous source localization approaches to create a model based method which operates in real time. Approach. To validate this algorithm, a saline benchtop setup was created to allow the precise placement of artificial sources within a cuff and interference sources outside the cuff. The artificial source was taken from five seconds of chronic neural activity to replicate realistic recordings. The proposed algorithm, hybrid Bayesian signal extraction (HBSE), is then compared to previous algorithms, beamforming and a Bayesian spatial filtering method, on this test data. An example chronic neural recording is also analyzed with all three algorithms. Main results. The proposed algorithm improved the signal to noise and signal to interference ratio of extracted test signals two to three fold, as well as increased the correlation coefficient between the original and recovered signals by 10-20%. These improvements translated to the chronic recording example and increased the calculated bit rate between the recovered signals and the recorded motor activity. Significance. HBSE significantly outperforms previous algorithms in extracting realistic neural signals, even in the presence of external noise sources. These results demonstrate the feasibility of extracting dynamic motor signals from a multi-fascicled intact nerve trunk, which in turn could extract motor command signals from an amputee for the end goal of

  17. JNSViewer-A JavaScript-based Nucleotide Sequence Viewer for DNA/RNA secondary structures.

    Science.gov (United States)

    Shi, Jieming; Li, Xi; Dong, Min; Graham, Mitchell; Yadav, Nehul; Liang, Chun

    2017-01-01

    Many tools are available for visualizing RNA or DNA secondary structures, but there is scarce implementation in JavaScript that provides seamless integration with the increasingly popular web computational platforms. We have developed JNSViewer, a highly interactive web service, which is bundled with several popular tools for DNA/RNA secondary structure prediction and can provide precise and interactive correspondence among nucleotides, dot-bracket data, secondary structure graphs, and genic annotations. In JNSViewer, users can perform RNA secondary structure predictions with different programs and settings, add customized genic annotations in GFF format to structure graphs, search for specific linear motifs, and extract relevant structure graphs of sub-sequences. JNSViewer also allows users to choose a transcript or specific segment of Arabidopsis thaliana genome sequences and predict the corresponding secondary structure. Popular genome browsers (i.e., JBrowse and BrowserGenome) were integrated into JNSViewer to provide powerful visualizations of chromosomal locations, genic annotations, and secondary structures. In addition, we used StructureFold with default settings to predict some RNA structures for Arabidopsis by incorporating in vivo high-throughput RNA structure profiling data and stored the results in our web server, which might be a useful resource for RNA secondary structure studies in plants. JNSViewer is available at http://bioinfolab.miamioh.edu/jnsviewer/index.html.

  18. JNSViewer—A JavaScript-based Nucleotide Sequence Viewer for DNA/RNA secondary structures

    Science.gov (United States)

    Dong, Min; Graham, Mitchell; Yadav, Nehul

    2017-01-01

    Many tools are available for visualizing RNA or DNA secondary structures, but there is scarce implementation in JavaScript that provides seamless integration with the increasingly popular web computational platforms. We have developed JNSViewer, a highly interactive web service, which is bundled with several popular tools for DNA/RNA secondary structure prediction and can provide precise and interactive correspondence among nucleotides, dot-bracket data, secondary structure graphs, and genic annotations. In JNSViewer, users can perform RNA secondary structure predictions with different programs and settings, add customized genic annotations in GFF format to structure graphs, search for specific linear motifs, and extract relevant structure graphs of sub-sequences. JNSViewer also allows users to choose a transcript or specific segment of Arabidopsis thaliana genome sequences and predict the corresponding secondary structure. Popular genome browsers (i.e., JBrowse and BrowserGenome) were integrated into JNSViewer to provide powerful visualizations of chromosomal locations, genic annotations, and secondary structures. In addition, we used StructureFold with default settings to predict some RNA structures for Arabidopsis by incorporating in vivo high-throughput RNA structure profiling data and stored the results in our web server, which might be a useful resource for RNA secondary structure studies in plants. JNSViewer is available at http://bioinfolab.miamioh.edu/jnsviewer/index.html. PMID:28582416

  19. JNSViewer-A JavaScript-based Nucleotide Sequence Viewer for DNA/RNA secondary structures.

    Directory of Open Access Journals (Sweden)

    Jieming Shi

    Full Text Available Many tools are available for visualizing RNA or DNA secondary structures, but there is scarce implementation in JavaScript that provides seamless integration with the increasingly popular web computational platforms. We have developed JNSViewer, a highly interactive web service, which is bundled with several popular tools for DNA/RNA secondary structure prediction and can provide precise and interactive correspondence among nucleotides, dot-bracket data, secondary structure graphs, and genic annotations. In JNSViewer, users can perform RNA secondary structure predictions with different programs and settings, add customized genic annotations in GFF format to structure graphs, search for specific linear motifs, and extract relevant structure graphs of sub-sequences. JNSViewer also allows users to choose a transcript or specific segment of Arabidopsis thaliana genome sequences and predict the corresponding secondary structure. Popular genome browsers (i.e., JBrowse and BrowserGenome were integrated into JNSViewer to provide powerful visualizations of chromosomal locations, genic annotations, and secondary structures. In addition, we used StructureFold with default settings to predict some RNA structures for Arabidopsis by incorporating in vivo high-throughput RNA structure profiling data and stored the results in our web server, which might be a useful resource for RNA secondary structure studies in plants. JNSViewer is available at http://bioinfolab.miamioh.edu/jnsviewer/index.html.

  20. An improved and reproducible protocol for the extraction of high quality fungal RNA from plant biomass substrates

    NARCIS (Netherlands)

    Patyshakuliyeva, Aleksandrina; Mäkelä, Miia R; Sietiö, Outi-Maaria; de Vries, Ronald P; Hildén, Kristiina S; van den Brink, J.

    2014-01-01

    Isolation of high quantity and quality RNA is a crucial step in the detection of meaningful gene expression data. Obtaining intact fungal RNA from complex lignocellulosic substrates is often difficult, producing low integrity RNA which perform poorly in downstream applications. In this study we

  1. Image feature extraction based on the camouflage effectiveness evaluation

    Science.gov (United States)

    Yuan, Xin; Lv, Xuliang; Li, Ling; Wang, Xinzhu; Zhang, Zhi

    2018-04-01

    The key step of camouflage effectiveness evaluation is how to combine the human visual physiological features, psychological features to select effectively evaluation indexes. Based on the predecessors' camo comprehensive evaluation method, this paper chooses the suitable indexes combining with the image quality awareness, and optimizes those indexes combining with human subjective perception. Thus, it perfects the theory of index extraction.

  2. Effect of cellulose-based fibers extracted from pineapple (Ananas ...

    African Journals Online (AJOL)

    New polyurethane foams were fabricated utilizing cellulose-based fibers extracted from pineapple (Ananas comosus) leaf as raw material. The pineapple leaf fibers (PALF) were treated with alkali and subsequently bleached to enhance its fiber-matrix adhesion. Polyurethane composites have been prepared by ...

  3. Chess games: a model for RNA based computation.

    Science.gov (United States)

    Cukras, A R; Faulhammer, D; Lipton, R J; Landweber, L F

    1999-10-01

    Here we develop the theory of RNA computing and a method for solving the 'knight problem' as an instance of a satisfiability (SAT) problem. Using only biological molecules and enzymes as tools, we developed an algorithm for solving the knight problem (3 x 3 chess board) using a 10-bit combinatorial pool and sequential RNase H digestions. The results of preliminary experiments presented here reveal that the protocol recovers far more correct solutions than expected at random, but the persistence of errors still presents the greatest challenge.

  4. New Kids on the Block: RNA-Based Influenza Virus Vaccines.

    Science.gov (United States)

    Scorza, Francesco Berlanda; Pardi, Norbert

    2018-04-01

    RNA-based immunization strategies have emerged as promising alternatives to conventional vaccine approaches. A substantial body of published work demonstrates that RNA vaccines can elicit potent, protective immune responses against various pathogens. Consonant with its huge impact on public health, influenza virus is one of the best studied targets of RNA vaccine research. Currently licensed influenza vaccines show variable levels of protection against seasonal influenza virus strains but are inadequate against drifted and pandemic viruses. In recent years, several types of RNA vaccines demonstrated efficacy against influenza virus infections in preclinical models. Additionally, comparative studies demonstrated the superiority of some RNA vaccines over the currently used inactivated influenza virus vaccines in animal models. Based on these promising preclinical results, clinical trials have been initiated and should provide valuable information about the translatability of the impressive preclinical data to humans. This review briefly describes RNA-based vaccination strategies, summarizes published preclinical and clinical data, highlights the roadblocks that need to be overcome for clinical applications, discusses the landscape of industrial development, and shares the authors' personal perspectives about the future of RNA-based influenza virus vaccines.

  5. Non-Watson Crick base pairs might stabilize RNA structural motifs in ...

    Indian Academy of Sciences (India)

    Watson Crick base pairs, internal loops and pseudoknots have been the highlighting feature of recent structural determination of RNAs. The recent crystal structure of group-I introns has demonstrated that these might constitute RNA structural ...

  6. Hydration sites of unpaired RNA bases: a statistical analysis of the PDB structures

    Directory of Open Access Journals (Sweden)

    Carugo Oliviero

    2011-10-01

    Full Text Available Abstract Background Hydration is crucial for RNA structure and function. X-ray crystallography is the most commonly used method to determine RNA structures and hydration and, therefore, statistical surveys are based on crystallographic results, the number of which is quickly increasing. Results A statistical analysis of the water molecule distribution in high-resolution X-ray structures of unpaired RNA nucleotides showed that: different bases have the same penchant to be surrounded by water molecules; clusters of water molecules indicate possible hydration sites, which, in some cases, match those of the major and minor grooves of RNA and DNA double helices; complex hydrogen bond networks characterize the solvation of the nucleotides, resulting in a significant rigidity of the base and its surrounding water molecules. Interestingly, the hydration sites around unpaired RNA bases do not match, in general, the positions that are occupied by the second nucleotide when the base-pair is formed. Conclusions The hydration sites around unpaired RNA bases were found. They do not replicate the atom positions of complementary bases in the Watson-Crick pairs.

  7. RNACompress: Grammar-based compression and informational complexity measurement of RNA secondary structure

    Directory of Open Access Journals (Sweden)

    Chen Chun

    2008-03-01

    Full Text Available Abstract Background With the rapid emergence of RNA databases and newly identified non-coding RNAs, an efficient compression algorithm for RNA sequence and structural information is needed for the storage and analysis of such data. Although several algorithms for compressing DNA sequences have been proposed, none of them are suitable for the compression of RNA sequences with their secondary structures simultaneously. This kind of compression not only facilitates the maintenance of RNA data, but also supplies a novel way to measure the informational complexity of RNA structural data, raising the possibility of studying the relationship between the functional activities of RNA structures and their complexities, as well as various structural properties of RNA based on compression. Results RNACompress employs an efficient grammar-based model to compress RNA sequences and their secondary structures. The main goals of this algorithm are two fold: (1 present a robust and effective way for RNA structural data compression; (2 design a suitable model to represent RNA secondary structure as well as derive the informational complexity of the structural data based on compression. Our extensive tests have shown that RNACompress achieves a universally better compression ratio compared with other sequence-specific or common text-specific compression algorithms, such as Gencompress, winrar and gzip. Moreover, a test of the activities of distinct GTP-binding RNAs (aptamers compared with their structural complexity shows that our defined informational complexity can be used to describe how complexity varies with activity. These results lead to an objective means of comparing the functional properties of heteropolymers from the information perspective. Conclusion A universal algorithm for the compression of RNA secondary structure as well as the evaluation of its informational complexity is discussed in this paper. We have developed RNACompress, as a useful tool

  8. Preclinical and clinical development of siRNA-based therapeutics.

    Science.gov (United States)

    Ozcan, Gulnihal; Ozpolat, Bulent; Coleman, Robert L; Sood, Anil K; Lopez-Berestein, Gabriel

    2015-06-29

    The discovery of RNA interference, first in plants and Caenorhabditis elegans and later in mammalian cells, led to the emergence of a transformative view in biomedical research. Knowledge of the multiple actions of non-coding RNAs has truly allowed viewing DNA, RNA and proteins in novel ways. Small interfering RNAs (siRNAs) can be used as tools to study single gene function both in vitro and in vivo and are an attractive new class of therapeutics, especially against undruggable targets for the treatment of cancer and other diseases. Despite the potential of siRNAs in cancer therapy, many challenges remain, including rapid degradation, poor cellular uptake and off-target effects. Rational design strategies, selection algorithms, chemical modifications and nanocarriers offer significant opportunities to overcome these challenges. Here, we review the development of siRNAs as therapeutic agents from early design to clinical trial, with special emphasis on the development of EphA2-targeting siRNAs for ovarian cancer treatment. Copyright © 2015 Elsevier B.V. All rights reserved.

  9. Solvent extraction of gold using ionic liquid based process

    Science.gov (United States)

    Makertihartha, I. G. B. N.; Zunita, Megawati; Rizki, Z.; Dharmawijaya, P. T.

    2017-01-01

    In decades, many research and mineral processing industries are using solvent extraction technology for metal ions separation. Solvent extraction technique has been used for the purification of precious metals such as Au and Pd, and base metals such as Cu, Zn and Cd. This process uses organic compounds as solvent. Organic solvents have some undesired properties i.e. toxic, volatile, excessive used, flammable, difficult to recycle, low reusability, low Au recovery, together with the problems related to the disposal of spent extractants and diluents, even the costs associated with these processes are relatively expensive. Therefore, a lot of research have boosted into the development of safe and environmentally friendly process for Au separation. Ionic liquids (ILs) are the potential alternative for gold extraction because they possess several desirable properties, such as a the ability to expanse temperature process up to 300°C, good solvent properties for a wide range of metal ions, high selectivity, low vapor pressures, stability up to 200°C, easy preparation, environmentally friendly (commonly called as "green solvent"), and relatively low cost. This review paper is focused in investigate of some ILs that have the potentials as solvent in extraction of Au from mineral/metal alloy at various conditions (pH, temperature, and pressure). Performances of ILs extraction of Au are studied in depth, i.e. structural relationship of ILs with capability to separate Au from metal ions aggregate. Optimal extraction conditon in order to gain high percent of Au in mineral processing is also investigated.

  10. Building Contour Extraction Based on LiDAR Point Cloud

    Directory of Open Access Journals (Sweden)

    Zhang Xu-Qing

    2017-01-01

    Full Text Available This paper presents a new method for solving the problem of utilizing the LiDAR data to extract the building contour line. For detection of the edge points between the building test points by using the least squares fitting to get the edge line of buildings and give the weight determining of the building of edge line slope depend on the length of the edge line. And then get the weighted mean of the positive and negative slope of the building edge line. Based on the structure of the adjacent edge perpendicular hypothesis, regularization processing to extract the edge of the skeleton line perpendicular. The experiments show that the extracted building edges have the good accuracy and have the good applicability in complex urban areas.

  11. Distributed Parallel Endmember Extraction of Hyperspectral Data Based on Spark

    Directory of Open Access Journals (Sweden)

    Zebin Wu

    2016-01-01

    Full Text Available Due to the increasing dimensionality and volume of remotely sensed hyperspectral data, the development of acceleration techniques for massive hyperspectral image analysis approaches is a very important challenge. Cloud computing offers many possibilities of distributed processing of hyperspectral datasets. This paper proposes a novel distributed parallel endmember extraction method based on iterative error analysis that utilizes cloud computing principles to efficiently process massive hyperspectral data. The proposed method takes advantage of technologies including MapReduce programming model, Hadoop Distributed File System (HDFS, and Apache Spark to realize distributed parallel implementation for hyperspectral endmember extraction, which significantly accelerates the computation of hyperspectral processing and provides high throughput access to large hyperspectral data. The experimental results, which are obtained by extracting endmembers of hyperspectral datasets on a cloud computing platform built on a cluster, demonstrate the effectiveness and computational efficiency of the proposed method.

  12. Feature extraction algorithm for space targets based on fractal theory

    Science.gov (United States)

    Tian, Balin; Yuan, Jianping; Yue, Xiaokui; Ning, Xin

    2007-11-01

    In order to offer a potential for extending the life of satellites and reducing the launch and operating costs, satellite servicing including conducting repairs, upgrading and refueling spacecraft on-orbit become much more frequently. Future space operations can be more economically and reliably executed using machine vision systems, which can meet real time and tracking reliability requirements for image tracking of space surveillance system. Machine vision was applied to the research of relative pose for spacecrafts, the feature extraction algorithm was the basis of relative pose. In this paper fractal geometry based edge extraction algorithm which can be used in determining and tracking the relative pose of an observed satellite during proximity operations in machine vision system was presented. The method gets the gray-level image distributed by fractal dimension used the Differential Box-Counting (DBC) approach of the fractal theory to restrain the noise. After this, we detect the consecutive edge using Mathematical Morphology. The validity of the proposed method is examined by processing and analyzing images of space targets. The edge extraction method not only extracts the outline of the target, but also keeps the inner details. Meanwhile, edge extraction is only processed in moving area to reduce computation greatly. Simulation results compared edge detection using the method which presented by us with other detection methods. The results indicate that the presented algorithm is a valid method to solve the problems of relative pose for spacecrafts.

  13. A collaborative European exercise on mRNA-based body fluid/skin typing and interpretation of DNA and RNA results

    DEFF Research Database (Denmark)

    van den Berge, M; Carracedo, A; Gomes, I

    2014-01-01

    The European Forensic Genetics Network of Excellence (EUROFORGEN-NoE) undertook a collaborative project on mRNA-based body fluid/skin typing and the interpretation of the resulting RNA and DNA data. Although both body fluids and skin are composed of a variety of cell types with different function...

  14. Base pairing and structural insights into the 5-formylcytosine in RNA duplex

    Science.gov (United States)

    Wang, Rui; Luo, Zhipu; He, Kaizhang; Delaney, Michael O.; Chen, Doris; Sheng, Jia

    2016-01-01

    Abstract 5-Formylcytidine (f5C), a previously discovered natural nucleotide in the mitochondrial tRNA of many species including human, has been recently detected as the oxidative product of 5-methylcytidine (m5C) through 5-hydroxymethylcytidine (hm5C) in total RNA of mammalian cells. The discovery indicated that these cytosine derivatives in RNA might also play important epigenetic roles similar as in DNA, which has been intensively investigated in the past few years. In this paper, we studied the base pairing specificity of f5C in different RNA duplex contexts. We found that the 5-formyl group could increase duplex thermal stability and enhance base pairing specificity. We present three high-resolution crystal structures of an octamer RNA duplex [5′-GUA(f5C)GUAC-3′]2 that have been solved under three crystallization conditions with different buffers and pH values. Our results showed that the 5-formyl group is located in the same plane as the cytosine base and forms an intra-residue hydrogen bond with the amino group in the N4 position. In addition, this modification increases the base stacking between the f5C and the neighboring bases while not causing significant global and local structure perturbations. This work provides insights into the effects of 5-formylcytosine on RNA duplex. PMID:27079978

  15. Base pair probability estimates improve the prediction accuracy of RNA non-canonical base pairs.

    Directory of Open Access Journals (Sweden)

    Michael F Sloma

    2017-11-01

    Full Text Available Prediction of RNA tertiary structure from sequence is an important problem, but generating accurate structure models for even short sequences remains difficult. Predictions of RNA tertiary structure tend to be least accurate in loop regions, where non-canonical pairs are important for determining the details of structure. Non-canonical pairs can be predicted using a knowledge-based model of structure that scores nucleotide cyclic motifs, or NCMs. In this work, a partition function algorithm is introduced that allows the estimation of base pairing probabilities for both canonical and non-canonical interactions. Pairs that are predicted to be probable are more likely to be found in the true structure than pairs of lower probability. Pair probability estimates can be further improved by predicting the structure conserved across multiple homologous sequences using the TurboFold algorithm. These pairing probabilities, used in concert with prior knowledge of the canonical secondary structure, allow accurate inference of non-canonical pairs, an important step towards accurate prediction of the full tertiary structure. Software to predict non-canonical base pairs and pairing probabilities is now provided as part of the RNAstructure software package.

  16. Base pair probability estimates improve the prediction accuracy of RNA non-canonical base pairs.

    Science.gov (United States)

    Sloma, Michael F; Mathews, David H

    2017-11-01

    Prediction of RNA tertiary structure from sequence is an important problem, but generating accurate structure models for even short sequences remains difficult. Predictions of RNA tertiary structure tend to be least accurate in loop regions, where non-canonical pairs are important for determining the details of structure. Non-canonical pairs can be predicted using a knowledge-based model of structure that scores nucleotide cyclic motifs, or NCMs. In this work, a partition function algorithm is introduced that allows the estimation of base pairing probabilities for both canonical and non-canonical interactions. Pairs that are predicted to be probable are more likely to be found in the true structure than pairs of lower probability. Pair probability estimates can be further improved by predicting the structure conserved across multiple homologous sequences using the TurboFold algorithm. These pairing probabilities, used in concert with prior knowledge of the canonical secondary structure, allow accurate inference of non-canonical pairs, an important step towards accurate prediction of the full tertiary structure. Software to predict non-canonical base pairs and pairing probabilities is now provided as part of the RNAstructure software package.

  17. Mapping Optimal Charge Density and Length of ROMP-Based PTDMs for siRNA Internalization.

    Science.gov (United States)

    Caffrey, Leah M; deRonde, Brittany M; Minter, Lisa M; Tew, Gregory N

    2016-10-10

    A fundamental understanding of how polymer structure impacts internalization and delivery of biologically relevant cargoes, particularly small interfering ribonucleic acid (siRNA), is of critical importance to the successful design of improved delivery reagents. Herein we report the use of ring-opening metathesis polymerization (ROMP) methods to synthesize two series of guanidinium-rich protein transduction domain mimics (PTDMs): one based on an imide scaffold that contains one guanidinium moiety per repeat unit, and another based on a diester scaffold that contains two guanidinium moieties per repeat unit. By varying both the degree of polymerization and, in effect, the relative number of cationic charges in each PTDM, the performances of the two ROMP backbones for siRNA internalization were evaluated and compared. Internalization of fluorescently labeled siRNA into Jurkat T cells demonstrated that fluorescein isothiocyanate (FITC)-siRNA internalization had a charge content dependence, with PTDMs containing approximately 40 to 60 cationic charges facilitating the most internalization. Despite this charge content dependence, the imide scaffold yielded much lower viabilities in Jurkat T cells than the corresponding diester PTDMs with similar numbers of cationic charges, suggesting that the diester scaffold is preferred for siRNA internalization and delivery applications. These developments will not only improve our understanding of the structural factors necessary for optimal siRNA internalization, but will also guide the future development of optimized PTDMs for siRNA internalization and delivery.

  18. Optimal consistency in microRNA expression analysis using reference-gene-based normalization.

    Science.gov (United States)

    Wang, Xi; Gardiner, Erin J; Cairns, Murray J

    2015-05-01

    Normalization of high-throughput molecular expression profiles secures differential expression analysis between samples of different phenotypes or biological conditions, and facilitates comparison between experimental batches. While the same general principles apply to microRNA (miRNA) normalization, there is mounting evidence that global shifts in their expression patterns occur in specific circumstances, which pose a challenge for normalizing miRNA expression data. As an alternative to global normalization, which has the propensity to flatten large trends, normalization against constitutively expressed reference genes presents an advantage through their relative independence. Here we investigated the performance of reference-gene-based (RGB) normalization for differential miRNA expression analysis of microarray expression data, and compared the results with other normalization methods, including: quantile, variance stabilization, robust spline, simple scaling, rank invariant, and Loess regression. The comparative analyses were executed using miRNA expression in tissue samples derived from subjects with schizophrenia and non-psychiatric controls. We proposed a consistency criterion for evaluating methods by examining the overlapping of differentially expressed miRNAs detected using different partitions of the whole data. Based on this criterion, we found that RGB normalization generally outperformed global normalization methods. Thus we recommend the application of RGB normalization for miRNA expression data sets, and believe that this will yield a more consistent and useful readout of differentially expressed miRNAs, particularly in biological conditions characterized by large shifts in miRNA expression.

  19. Evaluation of automated RNA-extraction technology and a qualitative HCV assay for sensitivity and detection of HCV RNA in pool-screening systems

    NARCIS (Netherlands)

    Beld, M.; Habibuw, M. R.; Rebers, S. P.; Boom, R.; Reesink, H. W.

    2000-01-01

    BACKGROUND: The objective of this study was the evaluation of NAT technology for the detection of HCV RNA in plasma pools according to the recommendations of the Paul Ehrlich Institute (5000 IU/mL/donation) and the Committee for Proprietary Medical Products (100 IU/mL/manufacturing pool). STUDY

  20. RT-PCR amplification of RNA extracted from formalin-fixed, paraffin-embedded oral cancer sections: analysis of p53 pathway.

    Science.gov (United States)

    Tachibana, Masatsugu; Shinagawa, Yasuhiro; Kawamata, Hitoshi; Omotehara, Fumie; Horiuchi, Hideki; Ohkura, Yasuo; Kubota, Keiichi; Imai, Yutaka; Fujibayashi, Takashi; Fujimori, Takahiro

    2003-01-01

    We present a new approach towards the detection of the mRNAs in formalin-fixed, paraffin-embedded samples using a reverse transcriptase (RT)-polymerase chain reaction (PCR). The total RNAs were extracted from 10-micron-thick sections and were reverse-transcribed, then the RT-products were subjected to PCR amplification of GAPDH mRNA for screening the mRNA degradation. Next, nested PCR was performed for examining the expression of p53-related genes, p21WAF1, MDM2, p33ING1 and p14ARF. GAPDH mRNA expression was detectable in 12 out of 21 oral squamous cell carcinoma (SCC) samples. p21WAF1 mRNA expression was detectable in 5 out of 12 SCC samples, MDM2 mRNA expression was detectable in 5 our of 12 SCC samples and p33ING1 mRNA expression was detectable in 6 out of 12 SCC samples. However, the expression of p14ARF mRNA was not detectable in any of the samples. Seven out of 12 oral SCC samples showed abnormal nuclear accumulation of p53 protein by immunohistochemical staining, whereas 5 out of 12 oral SCCs showed negative staining for p53 protein. Of of p33ING1 mRNA. One of these was a verrucous carcinoma in which the p53 gene products might be inactivated by the oncoprotein E6 of human papilloma virus. Thus, the p53 tumor suppressor pathway was disrupted in most oral SCCs at the cellular levels, due to either an abnormality in p53 itself or loss of expression of p53 regulatory factors. This method would assist in making diagnosis, determining therapeutic strategy and predicting the prognosis of various cancers including oral SCCs.

  1. Drawbacks of the ancient RNA-based life-like system under primitive earth conditions.

    Science.gov (United States)

    Kawamura, Kunio

    2012-07-01

    Following the discovery of ribozymes, the "RNA world" hypothesis has become the most accepted hypothesis concerning the origin of life and genetic information. However, this hypothesis has several drawbacks. Verification of the hypothesis from different viewpoints led us to proposals from the viewpoint of the hydrothermal origin of life, solubility of RNA and related biopolymers, and the possibility of creating an evolutionary system comparable to the in vitro selection technique for functional RNA molecules based on molecular biology. Copyright © 2012 Elsevier Masson SAS. All rights reserved.

  2. Template based rodent brain extraction and atlas mapping.

    Science.gov (United States)

    Weimin Huang; Jiaqi Zhang; Zhiping Lin; Su Huang; Yuping Duan; Zhongkang Lu

    2016-08-01

    Accurate rodent brain extraction is the basic step for many translational studies using MR imaging. This paper presents a template based approach with multi-expert refinement to automatic rodent brain extraction. We first build the brain appearance model based on the learning exemplars. Together with the template matching, we encode the rodent brain position into the search space to reliably locate the rodent brain and estimate the rough segmentation. With the initial mask, a level-set segmentation and a mask-based template learning are implemented further to the brain region. The multi-expert fusion is used to generate a new mask. We finally combine the region growing based on the histogram distribution learning to delineate the final brain mask. A high-resolution rodent atlas is used to illustrate that the segmented low resolution anatomic image can be well mapped to the atlas. Tested on a public data set, all brains are located reliably and we achieve the mean Jaccard similarity score at 94.99% for brain segmentation, which is a statistically significant improvement compared to two other rodent brain extraction methods.

  3. A novel knowledge-based potential for RNA 3D structure evaluation

    Science.gov (United States)

    Yang, Yi; Gu, Qi; Zhang, Ben-Gong; Shi, Ya-Zhou; Shao, Zhi-Gang

    2018-03-01

    Ribonucleic acids (RNAs) play a vital role in biology, and knowledge of their three-dimensional (3D) structure is required to understand their biological functions. Recently structural prediction methods have been developed to address this issue, but a series of RNA 3D structures are generally predicted by most existing methods. Therefore, the evaluation of the predicted structures is generally indispensable. Although several methods have been proposed to assess RNA 3D structures, the existing methods are not precise enough. In this work, a new all-atom knowledge-based potential is developed for more accurately evaluating RNA 3D structures. The potential not only includes local and nonlocal interactions but also fully considers the specificity of each RNA by introducing a retraining mechanism. Based on extensive test sets generated from independent methods, the proposed potential correctly distinguished the native state and ranked near-native conformations to effectively select the best. Furthermore, the proposed potential precisely captured RNA structural features such as base-stacking and base-pairing. Comparisons with existing potential methods show that the proposed potential is very reliable and accurate in RNA 3D structure evaluation. Project supported by the National Science Foundation of China (Grants Nos. 11605125, 11105054, 11274124, and 11401448).

  4. Biosensor-based microRNA detection: techniques, design, performance, and challenges.

    Science.gov (United States)

    Johnson, Blake N; Mutharasan, Raj

    2014-04-07

    The current state of biosensor-based techniques for amplification-free microRNA (miRNA) detection is critically reviewed. Comparison with non-sensor and amplification-based molecular techniques (MTs), such as polymerase-based methods, is made in terms of transduction mechanism, associated protocol, and sensitivity. Challenges associated with miRNA hybridization thermodynamics which affect assay selectivity and amplification bias are briefly discussed. Electrochemical, electromechanical, and optical classes of miRNA biosensors are reviewed in terms of transduction mechanism, limit of detection (LOD), time-to-results (TTR), multiplexing potential, and measurement robustness. Current trends suggest that biosensor-based techniques (BTs) for miRNA assay will complement MTs due to the advantages of amplification-free detection, LOD being femtomolar (fM)-attomolar (aM), short TTR, multiplexing capability, and minimal sample preparation requirement. Areas of future importance in miRNA BT development are presented which include focus on achieving high measurement confidence and multiplexing capabilities.

  5. eRNA: a graphic user interface-based tool optimized for large data analysis from high-throughput RNA sequencing.

    Science.gov (United States)

    Yuan, Tiezheng; Huang, Xiaoyi; Dittmar, Rachel L; Du, Meijun; Kohli, Manish; Boardman, Lisa; Thibodeau, Stephen N; Wang, Liang

    2014-03-05

    RNA sequencing (RNA-seq) is emerging as a critical approach in biological research. However, its high-throughput advantage is significantly limited by the capacity of bioinformatics tools. The research community urgently needs user-friendly tools to efficiently analyze the complicated data generated by high throughput sequencers. We developed a standalone tool with graphic user interface (GUI)-based analytic modules, known as eRNA. The capacity of performing parallel processing and sample management facilitates large data analyses by maximizing hardware usage and freeing users from tediously handling sequencing data. The module miRNA identification" includes GUIs for raw data reading, adapter removal, sequence alignment, and read counting. The module "mRNA identification" includes GUIs for reference sequences, genome mapping, transcript assembling, and differential expression. The module "Target screening" provides expression profiling analyses and graphic visualization. The module "Self-testing" offers the directory setups, sample management, and a check for third-party package dependency. Integration of other GUIs including Bowtie, miRDeep2, and miRspring extend the program's functionality. eRNA focuses on the common tools required for the mapping and quantification analysis of miRNA-seq and mRNA-seq data. The software package provides an additional choice for scientists who require a user-friendly computing environment and high-throughput capacity for large data analysis. eRNA is available for free download at https://sourceforge.net/projects/erna/?source=directory.

  6. Extraction of high-quality epidermal RNA after ammonium thiocyanate-induced dermo-epidermal separation of 4 mm human skin biopsies

    DEFF Research Database (Denmark)

    Clemmensen, Anders; Thomassen, Mads; Clemmensen, Ole

    2009-01-01

    To obtain a separation of the epidermal and dermal compartments to examine compartment specific biological mechanisms in the skin, we incubated 4 mm human skin punch biopsies in ammonium thiocyanate. We wanted to test (i) the histological quality of the dermo-epidermal separation obtained...... by different incubation times; (ii) the amount and quality of extractable epidermal RNA and (iii) its impact on sample RNA expression profiles assessed by large-scale gene expression microarray analysis in both normal and inflamed skin. At 30-min incubation, the split between dermis and epidermis...... and almost completely separated from the dermis of 4 mm skin biopsies by 30 min incubation in 3.8% ammonium thiocyanate combined with curettage of the dermal surface, producing high-quality RNA suitable for transcriptional analysis. Our refined method of dermo-epidermal separation will undoubtedly prove...

  7. Pixel extraction based integral imaging with controllable viewing direction

    International Nuclear Information System (INIS)

    Ji, Chao-Chao; Deng, Huan; Wang, Qiong-Hua

    2012-01-01

    We propose pixel extraction based integral imaging with a controllable viewing direction. The proposed integral imaging can provide viewers three-dimensional (3D) images in a very small viewing angle. The viewing angle and the viewing direction of the reconstructed 3D images are controlled by the pixels extracted from an elemental image array. Theoretical analysis and a 3D display experiment of the viewing direction controllable integral imaging are carried out. The experimental results verify the correctness of the theory. A 3D display based on the integral imaging can protect the viewer’s privacy and has huge potential for a television to show multiple 3D programs at the same time. (paper)

  8. Csf Based Non-Ground Points Extraction from LIDAR Data

    Science.gov (United States)

    Shen, A.; Zhang, W.; Shi, H.

    2017-09-01

    Region growing is a classical method of point cloud segmentation. Based on the idea of collecting the pixels with similar properties to form regions, region growing is widely used in many fields such as medicine, forestry and remote sensing. In this algorithm, there are two core problems. One is the selection of seed points, the other is the setting of the growth constraints, in which the selection of the seed points is the foundation. In this paper, we propose a CSF (Cloth Simulation Filtering) based method to extract the non-ground seed points effectively. The experiments have shown that this method can obtain a group of seed spots compared with the traditional methods. It is a new attempt to extract seed points

  9. Visualizing RNA Secondary Structure Base Pair Binding Probabilities using Nested Concave Hulls

    OpenAIRE

    Sansen , Joris; Bourqui , Romain; Thebault , Patricia; Allali , Julien; Auber , David

    2015-01-01

    International audience; The challenge 1 of the BIOVIS 2015 design contest consists in designing an intuitive visual depiction of base pairs binding probabilities for secondary structure of ncRNA. Our representation depicts the potential nucleotide pairs binding using nested concave hulls over the computed MFE ncRNA secondary structure. Thus, it allows to identify regions with a high level of uncertainty in the MFE computation and the structures which seem to match to reality.

  10. Evaluation of RNA extraction methods and identification of putative reference genes for real-time quantitative polymerase chain reaction expression studies on olive (Olea europaea L.) fruits.

    Science.gov (United States)

    Nonis, Alberto; Vezzaro, Alice; Ruperti, Benedetto

    2012-07-11

    Genome wide transcriptomic surveys together with targeted molecular studies are uncovering an ever increasing number of differentially expressed genes in relation to agriculturally relevant processes in olive (Olea europaea L). These data need to be supported by quantitative approaches enabling the precise estimation of transcript abundance. qPCR being the most widely adopted technique for mRNA quantification, preliminary work needs to be done to set up robust methods for extraction of fully functional RNA and for the identification of the best reference genes to obtain reliable quantification of transcripts. In this work, we have assessed different methods for their suitability for RNA extraction from olive fruits and leaves and we have evaluated thirteen potential candidate reference genes on 21 RNA samples belonging to fruit developmental/ripening series and to leaves subjected to wounding. By using two different algorithms, GAPDH2 and PP2A1 were identified as the best reference genes for olive fruit development and ripening, and their effectiveness for normalization of expression of two ripening marker genes was demonstrated.

  11. Historical Feature Pattern Extraction Based Network Attack Situation Sensing Algorithm

    OpenAIRE

    Zeng, Yong; Liu, Dacheng; Lei, Zhou

    2014-01-01

    The situation sequence contains a series of complicated and multivariate random trends, which are very sudden, uncertain, and difficult to recognize and describe its principle by traditional algorithms. To solve the above questions, estimating parameters of super long situation sequence is essential, but very difficult, so this paper proposes a situation prediction method based on historical feature pattern extraction (HFPE). First, HFPE algorithm seeks similar indications from the history si...

  12. Chinese character recognition based on Gabor feature extraction and CNN

    Science.gov (United States)

    Xiong, Yudian; Lu, Tongwei; Jiang, Yongyuan

    2018-03-01

    As an important application in the field of text line recognition and office automation, Chinese character recognition has become an important subject of pattern recognition. However, due to the large number of Chinese characters and the complexity of its structure, there is a great difficulty in the Chinese character recognition. In order to solve this problem, this paper proposes a method of printed Chinese character recognition based on Gabor feature extraction and Convolution Neural Network(CNN). The main steps are preprocessing, feature extraction, training classification. First, the gray-scale Chinese character image is binarized and normalized to reduce the redundancy of the image data. Second, each image is convoluted with Gabor filter with different orientations, and the feature map of the eight orientations of Chinese characters is extracted. Third, the feature map through Gabor filters and the original image are convoluted with learning kernels, and the results of the convolution is the input of pooling layer. Finally, the feature vector is used to classify and recognition. In addition, the generalization capacity of the network is improved by Dropout technology. The experimental results show that this method can effectively extract the characteristics of Chinese characters and recognize Chinese characters.

  13. Maintaining Breast Cancer Specimen Integrity and Individual or Simultaneous Extraction of Quality DNA, RNA, and Proteins from Allprotect-Stabilized and Nonstabilized Tissue Samples

    LENUS (Irish Health Repository)

    Mee, Blanaid C.

    2011-12-29

    The Saint James\\'s Hospital Biobank was established in 2008, to develop a high-quality breast tissue BioResource, as a part of the breast cancer clinical care pathway. The aims of this work were: (1) to ascertain the quality of RNA, DNA, and protein in biobanked carcinomas and normal breast tissues, (2) to assess the efficacy of AllPrep® (Qiagen) in isolating RNA, DNA, and protein simultaneously, (3) to compare AllPrep with RNEasy® and QIAamp® (both Qiagen), and (4) to examine the effectiveness of Allprotect® (Qiagen), a new tissue stabilization medium in preserving DNA, RNA, and proteins. One hundred eleven frozen samples of carcinoma and normal breast tissue were analyzed. Tumor and normal tissue morphology were confirmed by frozen sections. Tissue type, tissue treatment (Allprotect vs. no Allprotect), extraction kit, and nucleic acid quantification were analyzed by utilizing a 4 factorial design (SPSS PASW 18 Statistics Software®). QIAamp (DNA isolation), AllPrep (DNA, RNA, and Protein isolation), and RNeasy (RNA isolation) kits were assessed and compared. Mean DNA yield and A260\\/280 values using QIAamp were 33.2 ng\\/μL and 1.86, respectively, and using AllPrep were 23.2 ng\\/μL and 1.94. Mean RNA yield and RNA Integrity Number (RIN) values with RNeasy were 73.4 ng\\/μL and 8.16, respectively, and with AllPrep were 74.8 ng\\/μL and 7.92. Allprotect-treated tissues produced higher RIN values of borderline significance (P=0.055). No discernible loss of RNA stability was detected after 6 h incubation of stabilized or nonstabilized tissues at room temperature or 4°C or in 9 freeze-thaw cycles. Allprotect requires further detailed evaluation, but we consider AllPrep to be an excellent option for the simultaneous extraction of RNA, DNA, and protein from tumor and normal breast tissues. The essential presampling procedures that maintain the diagnostic integrity of pathology specimens do not appear to compromise the quality of molecular isolates.

  14. Arduino-based automation of a DNA extraction system.

    Science.gov (United States)

    Kim, Kyung-Won; Lee, Mi-So; Ryu, Mun-Ho; Kim, Jong-Won

    2015-01-01

    There have been many studies to detect infectious diseases with the molecular genetic method. This study presents an automation process for a DNA extraction system based on microfluidics and magnetic bead, which is part of a portable molecular genetic test system. This DNA extraction system consists of a cartridge with chambers, syringes, four linear stepper actuators, and a rotary stepper actuator. The actuators provide a sequence of steps in the DNA extraction process, such as transporting, mixing, and washing for the gene specimen, magnetic bead, and reagent solutions. The proposed automation system consists of a PC-based host application and an Arduino-based controller. The host application compiles a G code sequence file and interfaces with the controller to execute the compiled sequence. The controller executes stepper motor axis motion, time delay, and input-output manipulation. It drives the stepper motor with an open library, which provides a smooth linear acceleration profile. The controller also provides a homing sequence to establish the motor's reference position, and hard limit checking to prevent any over-travelling. The proposed system was implemented and its functionality was investigated, especially regarding positioning accuracy and velocity profile.

  15. Metabarcoding monitoring analysis: the pros and cons of using co-extracted environmental DNA and RNA data to assess offshore oil production impacts on benthic communities

    Directory of Open Access Journals (Sweden)

    Olivier Laroche

    2017-05-01

    Full Text Available Sequencing environmental DNA (eDNA is increasingly being used as an alternative to traditional morphological-based identification to characterize biological assemblages and monitor anthropogenic impacts in marine environments. Most studies only assess eDNA which, compared to eRNA, can persist longer in the environment after cell death. Therefore, eRNA may provide a more immediate census of the environment due to its relatively weaker stability, leading some researchers to advocate for the use of eRNA as an additional, or perhaps superior proxy for portraying ecological changes. A variety of pre-treatment techniques for screening eDNA and eRNA derived operational taxonomic units (OTUs have been employed prior to statistical analyses, including removing singleton taxa (i.e., OTUs found only once and discarding those not present in both eDNA and eRNA datasets. In this study, we used bacterial (16S ribosomal RNA gene and eukaryotic (18S ribosomal RNA gene eDNA- and eRNA-derived data from benthic communities collected at increasing distances along a transect from an oil production platform (Taranaki, New Zealand. Macro-infauna (visual classification of benthic invertebrates and physico-chemical data were analyzed in parallel. We tested the effect of removing singleton taxa, and removing taxa not present in the eDNA and eRNA libraries from the same environmental sample (trimmed by shared OTUs, by comparing the impact of the oil production platform on alpha- and beta-diversity of the eDNA/eRNA-based biological assemblages, and by correlating these to the morphologically identified macro-faunal communities and the physico-chemical data. When trimmed by singletons, presence/absence information from eRNA data represented the best proxy to detect changes on species diversity for both bacteria and eukaryotes. However, assessment of quantitative beta-diversity from read abundance information of bacteria eRNA did not, contrary to eDNA, reveal any impact from

  16. Metabarcoding monitoring analysis: the pros and cons of using co-extracted environmental DNA and RNA data to assess offshore oil production impacts on benthic communities

    KAUST Repository

    Laroche, Olivier

    2017-05-17

    Sequencing environmental DNA (eDNA) is increasingly being used as an alternative to traditional morphological-based identification to characterize biological assemblages and monitor anthropogenic impacts in marine environments. Most studies only assess eDNA which, compared to eRNA, can persist longer in the environment after cell death. Therefore, eRNA may provide a more immediate census of the environment due to its relatively weaker stability, leading some researchers to advocate for the use of eRNA as an additional, or perhaps superior proxy for portraying ecological changes. A variety of pre-treatment techniques for screening eDNA and eRNA derived operational taxonomic units (OTUs) have been employed prior to statistical analyses, including removing singleton taxa (i.e., OTUs found only once) and discarding those not present in both eDNA and eRNA datasets. In this study, we used bacterial (16S ribosomal RNA gene) and eukaryotic (18S ribosomal RNA gene) eDNA- and eRNA-derived data from benthic communities collected at increasing distances along a transect from an oil production platform (Taranaki, New Zealand). Macro-infauna (visual classification of benthic invertebrates) and physico-chemical data were analyzed in parallel. We tested the effect of removing singleton taxa, and removing taxa not present in the eDNA and eRNA libraries from the same environmental sample (trimmed by shared OTUs), by comparing the impact of the oil production platform on alpha- and beta-diversity of the eDNA/eRNA-based biological assemblages, and by correlating these to the morphologically identified macro-faunal communities and the physico-chemical data. When trimmed by singletons, presence/absence information from eRNA data represented the best proxy to detect changes on species diversity for both bacteria and eukaryotes. However, assessment of quantitative beta-diversity from read abundance information of bacteria eRNA did not, contrary to eDNA, reveal any impact from the oil

  17. Metabarcoding monitoring analysis: the pros and cons of using co-extracted environmental DNA and RNA data to assess offshore oil production impacts on benthic communities.

    Science.gov (United States)

    Laroche, Olivier; Wood, Susanna A; Tremblay, Louis A; Lear, Gavin; Ellis, Joanne I; Pochon, Xavier

    2017-01-01

    Sequencing environmental DNA (eDNA) is increasingly being used as an alternative to traditional morphological-based identification to characterize biological assemblages and monitor anthropogenic impacts in marine environments. Most studies only assess eDNA which, compared to eRNA, can persist longer in the environment after cell death. Therefore, eRNA may provide a more immediate census of the environment due to its relatively weaker stability, leading some researchers to advocate for the use of eRNA as an additional, or perhaps superior proxy for portraying ecological changes. A variety of pre-treatment techniques for screening eDNA and eRNA derived operational taxonomic units (OTUs) have been employed prior to statistical analyses, including removing singleton taxa (i.e., OTUs found only once) and discarding those not present in both eDNA and eRNA datasets. In this study, we used bacterial (16S ribosomal RNA gene) and eukaryotic (18S ribosomal RNA gene) eDNA- and eRNA-derived data from benthic communities collected at increasing distances along a transect from an oil production platform (Taranaki, New Zealand). Macro-infauna (visual classification of benthic invertebrates) and physico-chemical data were analyzed in parallel. We tested the effect of removing singleton taxa, and removing taxa not present in the eDNA and eRNA libraries from the same environmental sample (trimmed by shared OTUs), by comparing the impact of the oil production platform on alpha- and beta-diversity of the eDNA/eRNA-based biological assemblages, and by correlating these to the morphologically identified macro-faunal communities and the physico-chemical data. When trimmed by singletons, presence/absence information from eRNA data represented the best proxy to detect changes on species diversity for both bacteria and eukaryotes. However, assessment of quantitative beta-diversity from read abundance information of bacteria eRNA did not, contrary to eDNA, reveal any impact from the oil

  18. A Meta-Path-Based Prediction Method for Human miRNA-Target Association

    Directory of Open Access Journals (Sweden)

    Jiawei Luo

    2016-01-01

    Full Text Available MicroRNAs (miRNAs are short noncoding RNAs that play important roles in regulating gene expressing, and the perturbed miRNAs are often associated with development and tumorigenesis as they have effects on their target mRNA. Predicting potential miRNA-target associations from multiple types of genomic data is a considerable problem in the bioinformatics research. However, most of the existing methods did not fully use the experimentally validated miRNA-mRNA interactions. Here, we developed RMLM and RMLMSe to predict the relationship between miRNAs and their targets. RMLM and RMLMSe are global approaches as they can reconstruct the missing associations for all the miRNA-target simultaneously and RMLMSe demonstrates that the integration of sequence information can improve the performance of RMLM. In RMLM, we use RM measure to evaluate different relatedness between miRNA and its target based on different meta-paths; logistic regression and MLE method are employed to estimate the weight of different meta-paths. In RMLMSe, sequence information is utilized to improve the performance of RMLM. Here, we carry on fivefold cross validation and pathway enrichment analysis to prove the performance of our methods. The fivefold experiments show that our methods have higher AUC scores compared with other methods and the integration of sequence information can improve the performance of miRNA-target association prediction.

  19. A Circulating microRNA Signature Predicts Age-Based Development of Lymphoma.

    Directory of Open Access Journals (Sweden)

    Afshin Beheshti

    Full Text Available Extensive epidemiological data have demonstrated an exponential rise in the incidence of non-Hodgkin lymphoma (NHL that is associated with increasing age. The molecular etiology of this remains largely unknown, which impacts the effectiveness of treatment for patients. We proposed that age-dependent circulating microRNA (miRNA signatures in the host influence diffuse large B cell lymphoma (DLBCL development. Our objective was to examine tumor development in an age-based DLBCL system using an inventive systems biology approach. We harnessed a novel murine model of spontaneous DLBCL initiation (Smurf2-deficient at two age groups: 3 and 15 months old. All Smurf2-deficient mice develop visible DLBCL tumor starting at 15 months of age. Total miRNA was isolated from serum, bone marrow and spleen and were collected for all age groups for Smurf2-deficient mice and age-matched wild-type C57BL/6 mice. Using systems biology techniques, we identified a list of 10 circulating miRNAs being regulated in both the spleen and bone marrow that were present in DLBCL forming mice starting at 3 months of age that were not present in the control mice. Furthermore, this miRNA signature was found to occur circulating in the blood and it strongly impacted JUN and MYC oncogenic signaling. In addition, quantification of the miRNA signature was performed via Droplet Digital PCR technology. It was discovered that a key miRNA signature circulates throughout a host prior to the formation of a tumor starting at 3 months old, which becomes further modulated by age and yielded calculation of a 'carcinogenic risk score'. This novel age-based circulating miRNA signature may potentially be leveraged as a DLBCL risk profile at a young age to predict future lymphoma development or disease progression as well as for potential innovative miRNA-based targeted therapeutic strategies in lymphoma.

  20. The impact of different DNA extraction kits and laboratories upon the assessment of human gut microbiota composition by 16S rRNA gene sequencing.

    Science.gov (United States)

    Kennedy, Nicholas A; Walker, Alan W; Berry, Susan H; Duncan, Sylvia H; Farquarson, Freda M; Louis, Petra; Thomson, John M; Satsangi, Jack; Flint, Harry J; Parkhill, Julian; Lees, Charlie W; Hold, Georgina L

    2014-01-01

    Determining bacterial community structure in fecal samples through DNA sequencing is an important facet of intestinal health research. The impact of different commercially available DNA extraction kits upon bacterial community structures has received relatively little attention. The aim of this study was to analyze bacterial communities in volunteer and inflammatory bowel disease (IBD) patient fecal samples extracted using widely used DNA extraction kits in established gastrointestinal research laboratories. Fecal samples from two healthy volunteers (H3 and H4) and two relapsing IBD patients (I1 and I2) were investigated. DNA extraction was undertaken using MoBio Powersoil and MP Biomedicals FastDNA SPIN Kit for Soil DNA extraction kits. PCR amplification for pyrosequencing of bacterial 16S rRNA genes was performed in both laboratories on all samples. Hierarchical clustering of sequencing data was done using the Yue and Clayton similarity coefficient. DNA extracted using the FastDNA kit and the MoBio kit gave median DNA concentrations of 475 (interquartile range 228-561) and 22 (IQR 9-36) ng/µL respectively (p<0.0001). Hierarchical clustering of sequence data by Yue and Clayton coefficient revealed four clusters. Samples from individuals H3 and I2 clustered by patient; however, samples from patient I1 extracted with the MoBio kit clustered with samples from patient H4 rather than the other I1 samples. Linear modelling on relative abundance of common bacterial families revealed significant differences between kits; samples extracted with MoBio Powersoil showed significantly increased Bacteroidaceae, Ruminococcaceae and Porphyromonadaceae, and lower Enterobacteriaceae, Lachnospiraceae, Clostridiaceae, and Erysipelotrichaceae (p<0.05). This study demonstrates significant differences in DNA yield and bacterial DNA composition when comparing DNA extracted from the same fecal sample with different extraction kits. This highlights the importance of ensuring that samples

  1. EXTRACT

    DEFF Research Database (Denmark)

    Pafilis, Evangelos; Buttigieg, Pier Luigi; Ferrell, Barbra

    2016-01-01

    The microbial and molecular ecology research communities have made substantial progress on developing standards for annotating samples with environment metadata. However, sample manual annotation is a highly labor intensive process and requires familiarity with the terminologies used. We have the...... and text-mining-assisted curation revealed that EXTRACT speeds up annotation by 15-25% and helps curators to detect terms that would otherwise have been missed.Database URL: https://extract.hcmr.gr/......., organism, tissue and disease terms. The evaluators in the BioCreative V Interactive Annotation Task found the system to be intuitive, useful, well documented and sufficiently accurate to be helpful in spotting relevant text passages and extracting organism and environment terms. Comparison of fully manual...

  2. Biomarker MicroRNAs for Diagnosis of Oral Squamous Cell Carcinoma Identified Based on Gene Expression Data and MicroRNA-mRNA Network Analysis

    Science.gov (United States)

    Zhang, Hui; Li, Tangxin; Zheng, Linqing

    2017-01-01

    Oral squamous cell carcinoma is one of the most malignant tumors with high mortality rate worldwide. Biomarker discovery is critical for early diagnosis and precision treatment of this disease. MicroRNAs are small noncoding RNA molecules which often regulate essential biological processes and are good candidates for biomarkers. By integrative analysis of both the cancer-associated gene expression data and microRNA-mRNA network, miR-148b-3p, miR-629-3p, miR-27a-3p, and miR-142-3p were screened as novel diagnostic biomarkers for oral squamous cell carcinoma based on their unique regulatory abilities in the network structure of the conditional microRNA-mRNA network and their important functions. These findings were confirmed by literature verification and functional enrichment analysis. Future experimental validation is expected for the further investigation of their molecular mechanisms. PMID:29098014

  3. Epithelial-mesenchymal transition and cancer stem cells, mediated by a long non-coding RNA, HOTAIR, are involved in cell malignant transformation induced by cigarette smoke extract

    International Nuclear Information System (INIS)

    Liu, Yi; Luo, Fei; Xu, Yuan; Wang, Bairu; Zhao, Yue; Xu, Wenchao; Shi, Le; Lu, Xiaolin; Liu, Qizhan

    2015-01-01

    The incidence of lung diseases, including cancer, caused by cigarette smoke is increasing, but the molecular mechanisms of gene regulation induced by cigarette smoke remain unclear. This report describes a long noncoding RNA (lncRNA) that is induced by cigarette smoke extract (CSE) and experiments utilizing lncRNAs to integrate inflammation with the epithelial-mesenchymal transition (EMT) in human bronchial epithelial (HBE) cells. The present study shows that, induced by CSE, IL-6, a pro-inflammatory cytokine, leads to activation of STAT3, a transcription activator. A ChIP assay determined that the interaction of STAT3 with the promoter regions of HOX transcript antisense RNA (HOTAIR) increased levels of HOTAIR. Blocking of IL-6 with anti-IL-6 antibody, decreasing STAT3, and inhibiting STAT3 activation reduced HOTAIR expression. Moreover, for HBE cells cultured in the presence of HOTAIR siRNA for 24 h, the CSE-induced EMT, formation of cancer stem cells (CSCs), and malignant transformation were reversed. Thus, IL-6, acting on STAT3 signaling, which up-regulates HOTAIR in an autocrine manner, contributes to the EMT and to CSCs induced by CSE. These data define a link between inflammation and EMT, processes involved in the malignant transformation of cells caused by CSE. This link, mediated through lncRNAs, establishes a mechanism for CSE-induced lung carcinogenesis. - Highlights: • STAT3 directly regulates the levels of LncRNA HOTAIR. • LncRNA HOTAIR mediates the link between inflammation and EMT. • LncRNA HOTAIR is involved in the malignant transformation of cells caused by CSE

  4. Automated extraction of knowledge for model-based diagnostics

    Science.gov (United States)

    Gonzalez, Avelino J.; Myler, Harley R.; Towhidnejad, Massood; Mckenzie, Frederic D.; Kladke, Robin R.

    1990-01-01

    The concept of accessing computer aided design (CAD) design databases and extracting a process model automatically is investigated as a possible source for the generation of knowledge bases for model-based reasoning systems. The resulting system, referred to as automated knowledge generation (AKG), uses an object-oriented programming structure and constraint techniques as well as internal database of component descriptions to generate a frame-based structure that describes the model. The procedure has been designed to be general enough to be easily coupled to CAD systems that feature a database capable of providing label and connectivity data from the drawn system. The AKG system is capable of defining knowledge bases in formats required by various model-based reasoning tools.

  5. DNA-, RNA-, and Protein-Based Stable-Isotope Probing for High-Throughput Biomarker Analysis of Active Microorganisms.

    Science.gov (United States)

    Jameson, Eleanor; Taubert, Martin; Coyotzi, Sara; Chen, Yin; Eyice, Özge; Schäfer, Hendrik; Murrell, J Colin; Neufeld, Josh D; Dumont, Marc G

    2017-01-01

    Stable-isotope probing (SIP) enables researchers to target active populations within complex microbial communities, which is achieved by providing growth substrates enriched in heavy isotopes, usually in the form of 13 C, 18 O, or 15 N. After growth on the substrate and subsequent extraction of microbial biomarkers, typically nucleic acids or proteins, the SIP technique is used for the recovery and analysis of isotope-labeled biomarkers from active microbial populations. In the years following the initial development of DNA- and RNA-based SIP, it was common practice to characterize labeled populations by targeted gene analysis. Such approaches usually involved fingerprint-based analyses or sequencing of clone libraries containing 16S rRNA genes or functional marker gene amplicons. Although molecular fingerprinting remains a valuable approach for rapid confirmation of isotope labeling, recent advances in sequencing technology mean that it is possible to obtain affordable and comprehensive amplicon profiles, metagenomes, or metatranscriptomes from SIP experiments. Not only can the abundance of microbial groups be inferred from metagenomes, but researchers can bin, assemble, and explore individual genomes to build hypotheses about the metabolic capabilities of labeled microorganisms. Analysis of labeled mRNA is a more recent advance that can provide independent metatranscriptome-based analysis of active microorganisms. The power of metatranscriptomics is that mRNA abundance often correlates closely with the corresponding activity of encoded enzymes, thus providing insight into microbial metabolism at the time of sampling. Together, these advances have improved the sensitivity of SIP methods and allow the use of labeled substrates at ecologically relevant concentrations. Particularly as methods improve and costs continue to drop, we expect that the integration of SIP with multiple omics-based methods will become prevalent components of microbial ecology studies

  6. Region-Based Building Rooftop Extraction and Change Detection

    Science.gov (United States)

    Tian, J.; Metzlaff, L.; d'Angelo, P.; Reinartz, P.

    2017-09-01

    Automatic extraction of building changes is important for many applications like disaster monitoring and city planning. Although a lot of research work is available based on 2D as well as 3D data, an improvement in accuracy and efficiency is still needed. The introducing of digital surface models (DSMs) to building change detection has strongly improved the resulting accuracy. In this paper, a post-classification approach is proposed for building change detection using satellite stereo imagery. Firstly, DSMs are generated from satellite stereo imagery and further refined by using a segmentation result obtained from the Sobel gradients of the panchromatic image. Besides the refined DSMs, the panchromatic image and the pansharpened multispectral image are used as input features for mean-shift segmentation. The DSM is used to calculate the nDSM, out of which the initial building candidate regions are extracted. The candidate mask is further refined by morphological filtering and by excluding shadow regions. Following this, all segments that overlap with a building candidate region are determined. A building oriented segments merging procedure is introduced to generate a final building rooftop mask. As the last step, object based change detection is performed by directly comparing the building rooftops extracted from the pre- and after-event imagery and by fusing the change indicators with the roof-top region map. A quantitative and qualitative assessment of the proposed approach is provided by using WorldView-2 satellite data from Istanbul, Turkey.

  7. REGION-BASED BUILDING ROOFTOP EXTRACTION AND CHANGE DETECTION

    Directory of Open Access Journals (Sweden)

    J. Tian

    2017-09-01

    Full Text Available Automatic extraction of building changes is important for many applications like disaster monitoring and city planning. Although a lot of research work is available based on 2D as well as 3D data, an improvement in accuracy and efficiency is still needed. The introducing of digital surface models (DSMs to building change detection has strongly improved the resulting accuracy. In this paper, a post-classification approach is proposed for building change detection using satellite stereo imagery. Firstly, DSMs are generated from satellite stereo imagery and further refined by using a segmentation result obtained from the Sobel gradients of the panchromatic image. Besides the refined DSMs, the panchromatic image and the pansharpened multispectral image are used as input features for mean-shift segmentation. The DSM is used to calculate the nDSM, out of which the initial building candidate regions are extracted. The candidate mask is further refined by morphological filtering and by excluding shadow regions. Following this, all segments that overlap with a building candidate region are determined. A building oriented segments merging procedure is introduced to generate a final building rooftop mask. As the last step, object based change detection is performed by directly comparing the building rooftops extracted from the pre- and after-event imagery and by fusing the change indicators with the roof-top region map. A quantitative and qualitative assessment of the proposed approach is provided by using WorldView-2 satellite data from Istanbul, Turkey.

  8. Thermodynamic heuristics with case-based reasoning: combined insights for RNA pseudoknot secondary structure.

    Science.gov (United States)

    Al-Khatib, Ra'ed M; Rashid, Nur'Aini Abdul; Abdullah, Rosni

    2011-08-01

    The secondary structure of RNA pseudoknots has been extensively inferred and scrutinized by computational approaches. Experimental methods for determining RNA structure are time consuming and tedious; therefore, predictive computational approaches are required. Predicting the most accurate and energy-stable pseudoknot RNA secondary structure has been proven to be an NP-hard problem. In this paper, a new RNA folding approach, termed MSeeker, is presented; it includes KnotSeeker (a heuristic method) and Mfold (a thermodynamic algorithm). The global optimization of this thermodynamic heuristic approach was further enhanced by using a case-based reasoning technique as a local optimization method. MSeeker is a proposed algorithm for predicting RNA pseudoknot structure from individual sequences, especially long ones. This research demonstrates that MSeeker improves the sensitivity and specificity of existing RNA pseudoknot structure predictions. The performance and structural results from this proposed method were evaluated against seven other state-of-the-art pseudoknot prediction methods. The MSeeker method had better sensitivity than the DotKnot, FlexStem, HotKnots, pknotsRG, ILM, NUPACK and pknotsRE methods, with 79% of the predicted pseudoknot base-pairs being correct.

  9. Sequence-based heuristics for faster annotation of non-coding RNA families.

    Science.gov (United States)

    Weinberg, Zasha; Ruzzo, Walter L

    2006-01-01

    Non-coding RNAs (ncRNAs) are functional RNA molecules that do not code for proteins. Covariance Models (CMs) are a useful statistical tool to find new members of an ncRNA gene family in a large genome database, using both sequence and, importantly, RNA secondary structure information. Unfortunately, CM searches are extremely slow. Previously, we created rigorous filters, which provably sacrifice none of a CM's accuracy, while making searches significantly faster for virtually all ncRNA families. However, these rigorous filters make searches slower than heuristics could be. In this paper we introduce profile HMM-based heuristic filters. We show that their accuracy is usually superior to heuristics based on BLAST. Moreover, we compared our heuristics with those used in tRNAscan-SE, whose heuristics incorporate a significant amount of work specific to tRNAs, where our heuristics are generic to any ncRNA. Performance was roughly comparable, so we expect that our heuristics provide a high-quality solution that--unlike family-specific solutions--can scale to hundreds of ncRNA families. The source code is available under GNU Public License at the supplementary web site.

  10. General enumeration of RNA secondary structures based on new ...

    African Journals Online (AJOL)

    Crick base pairs between AU and GC. Based on the new representation, this paper also computes the number of various types of constrained secondary structures taking the minimum stack length 1 and minimum size m for each bonding loop as ...

  11. Ontology-Based Information Extraction for Business Intelligence

    Science.gov (United States)

    Saggion, Horacio; Funk, Adam; Maynard, Diana; Bontcheva, Kalina

    Business Intelligence (BI) requires the acquisition and aggregation of key pieces of knowledge from multiple sources in order to provide valuable information to customers or feed statistical BI models and tools. The massive amount of information available to business analysts makes information extraction and other natural language processing tools key enablers for the acquisition and use of that semantic information. We describe the application of ontology-based extraction and merging in the context of a practical e-business application for the EU MUSING Project where the goal is to gather international company intelligence and country/region information. The results of our experiments so far are very promising and we are now in the process of building a complete end-to-end solution.

  12. Location Fingerprint Extraction for Magnetic Field Magnitude Based Indoor Positioning

    Directory of Open Access Journals (Sweden)

    Wenhua Shao

    2016-01-01

    Full Text Available Smartphone based indoor positioning has greatly helped people in finding their positions in complex and unfamiliar buildings. One popular positioning method is by utilizing indoor magnetic field, because this feature is stable and infrastructure-free. In this method, the magnetometer embedded on the smartphone measures indoor magnetic field and queries its position. However, the environments of the magnetometer are rather harsh. This harshness mainly consists of coarse-grained hard/soft-iron calibrations and sensor electronic noise. The two kinds of interferences decrease the position distinguishability of the magnetic field. Therefore, it is important to extract location features from magnetic fields to reduce these interferences. This paper analyzes the main interference sources of the magnetometer embedded on the smartphone. In addition, we present a feature distinguishability measurement technique to evaluate the performance of different feature extraction methods. Experiments revealed that selected fingerprints will improve position distinguishability.

  13. Growth inhibition of head and neck squamous cell carcinoma cells by sgRNA targeting the cyclin D1 mRNA based on TRUE gene silencing.

    Directory of Open Access Journals (Sweden)

    Satoshi Iizuka

    Full Text Available Head and neck squamous cell carcinoma (HNSCC exhibits increased expression of cyclin D1 (CCND1. Previous studies have shown a correlation between poor prognosis of HNSCC and cyclin D1 overexpression. tRNase ZL-utilizing efficacious gene silencing (TRUE gene silencing is one of the RNA-mediated gene expression control technologies that have therapeutic potential. This technology is based on a unique enzymatic property of mammalian tRNase ZL, which is that it can cleave any target RNA at any desired site by recognizing a pre-tRNA-like complex formed between the target RNA and an artificial small guide RNA (sgRNA. In this study, we designed several sgRNAs targeting human cyclin D1 mRNA to examine growth inhibition of HNSCC cells. Transfection of certain sgRNAs decreased levels of cyclin D1 mRNA and protein in HSC-2 and HSC-3 cells, and also inhibited their proliferation. The combination of these sgRNAs and cisplatin showed more than additive inhibition of cancer cell growth. These findings demonstrate that TRUE gene silencing of cyclin D1 leads to inhibition of the growth of HNSCC cells and suggest that these sgRNAs alone or combined with cisplatin may be a useful new therapy for HNSCCs.

  14. Spectral features based tea garden extraction from digital orthophoto maps

    Science.gov (United States)

    Jamil, Akhtar; Bayram, Bulent; Kucuk, Turgay; Zafer Seker, Dursun

    2018-05-01

    The advancements in the photogrammetry and remote sensing technologies has made it possible to extract useful tangible information from data which plays a pivotal role in various application such as management and monitoring of forests and agricultural lands etc. This study aimed to evaluate the effectiveness of spectral signatures for extraction of tea gardens from 1 : 5000 scaled digital orthophoto maps obtained from Rize city in Turkey. First, the normalized difference vegetation index (NDVI) was derived from the input images to suppress the non-vegetation areas. NDVI values less than zero were discarded and the output images was normalized in the range 0-255. Individual pixels were then mapped into meaningful objects using global region growing technique. The resulting image was filtered and smoothed to reduce the impact of noise. Furthermore, geometrical constraints were applied to remove small objects (less than 500 pixels) followed by morphological opening operator to enhance the results. These objects served as building blocks for further image analysis. Finally, for the classification stage, a range of spectral values were empirically calculated for each band and applied on candidate objects to extract tea gardens. For accuracy assessment, we employed an area based similarity metric by overlapping obtained tea garden boundaries with the manually digitized tea garden boundaries created by experts of photogrammetry. The overall accuracy of the proposed method scored 89 % for tea gardens from 10 sample orthophoto maps. We concluded that exploiting the spectral signatures using object based analysis is an effective technique for extraction of dominant tree species from digital orthophoto maps.

  15. Stability of RNA silencing-based traits after virus infection

    DEFF Research Database (Denmark)

    Jørgensen, Bodil; Albrechtsen, Merete

    2007-01-01

    with constructs based on virus coat protein (CP) genes or other viral genes has been successfully used to engineer PTGS-mediated virus resistance into a large number of crop plants and some transgenic lines have been commercially exploited. However the discovery that plant viruses encode suppressors of gene...... silencing has raised concerns that virus infection of crop plants might reverse the new silencing-based traits. Most studies of virus suppression of silencing have used model systems based on silencing of reporter genes. A few studies have analysed the effects of virus infections on plants with genetically...... engineered virus resistance based on either a simple sense or an inverted repeat construct. We decided to use genetically engineered virus resistance in potato as a model system for further studies of the effect of virus infection on genetically engineered traits. We present for the first time a comparison...

  16. SERS-based inverse molecular sentinel (iMS) nanoprobes for multiplexed detection of microRNA cancer biomarkers in biological samples

    Science.gov (United States)

    Crawford, Bridget M.; Wang, Hsin-Neng; Fales, Andrew M.; Bowie, Michelle L.; Seewaldt, Victoria L.; Vo-Dinh, Tuan

    2017-02-01

    The development of sensitive and selective biosensing techniques is of great interest for clinical diagnostics. Here, we describe the development and application of a surface enhanced Raman scattering (SERS) sensing technology, referred to as "inverse Molecular Sentinel (iMS)" nanoprobes, for the detection of nucleic acid biomarkers in biological samples. This iMS nanoprobe involves the use of plasmonic-active nanostars as the sensing platform for a homogenous assay for multiplexed detection of nucleic acid biomarkers, including DNA, RNA and microRNA (miRNA). The "OFF-to-ON" signal switch is based on a non-enzymatic strand-displacement process and the conformational change of stem-loop (hairpin) oligonucleotide probes upon target binding. Here, we demonstrate the development of iMS nanoprobes for the detection of DNA sequences as well as a modified design of the nanoprobe for the detection of short (22-nt) microRNA sequences. The application of iMS nanoprobes to detect miRNAs in real biological samples was performed with total small RNA extracted from breast cancer cell lines. The multiplex capability of the iMS technique was demonstrated using a mixture of the two differently labeled nanoprobes to detect miR-21 and miR-34a miRNA biomarkers for breast cancer. The results of this study demonstrate the feasibility of applying the iMS technique for multiplexed detection of nucleic acid biomarkers, including short miRNAs molecules.

  17. Probing electronic coupling between adenine bases in RNA strands from synchrotron radiation circular dichroism experiments

    DEFF Research Database (Denmark)

    Nielsen, Lisbeth Munksgård; Hoffmann, Søren Vrønning; Nielsen, Steen Brøndsted

    2012-01-01

    Circular dichroism spectra (176–330 nm) of RNA adenine oligomers, (rA)n (n = 1–10, 12, 15, and 20), reveal electronic coupling between two bases in short strands. The number of interacting bases in long strands is more and larger than that reported previously for the corresponding DNA strands....

  18. Network-Based Isoform Quantification with RNA-Seq Data for Cancer Transcriptome Analysis.

    Directory of Open Access Journals (Sweden)

    Wei Zhang

    2015-12-01

    Full Text Available High-throughput mRNA sequencing (RNA-Seq is widely used for transcript quantification of gene isoforms. Since RNA-Seq data alone is often not sufficient to accurately identify the read origins from the isoforms for quantification, we propose to explore protein domain-domain interactions as prior knowledge for integrative analysis with RNA-Seq data. We introduce a Network-based method for RNA-Seq-based Transcript Quantification (Net-RSTQ to integrate protein domain-domain interaction network with short read alignments for transcript abundance estimation. Based on our observation that the abundances of the neighboring isoforms by domain-domain interactions in the network are positively correlated, Net-RSTQ models the expression of the neighboring transcripts as Dirichlet priors on the likelihood of the observed read alignments against the transcripts in one gene. The transcript abundances of all the genes are then jointly estimated with alternating optimization of multiple EM problems. In simulation Net-RSTQ effectively improved isoform transcript quantifications when isoform co-expressions correlate with their interactions. qRT-PCR results on 25 multi-isoform genes in a stem cell line, an ovarian cancer cell line, and a breast cancer cell line also showed that Net-RSTQ estimated more consistent isoform proportions with RNA-Seq data. In the experiments on the RNA-Seq data in The Cancer Genome Atlas (TCGA, the transcript abundances estimated by Net-RSTQ are more informative for patient sample classification of ovarian cancer, breast cancer and lung cancer. All experimental results collectively support that Net-RSTQ is a promising approach for isoform quantification. Net-RSTQ toolbox is available at http://compbio.cs.umn.edu/Net-RSTQ/.

  19. Tree decomposition based fast search of RNA structures including pseudoknots in genomes.

    Science.gov (United States)

    Song, Yinglei; Liu, Chunmei; Malmberg, Russell; Pan, Fangfang; Cai, Liming

    2005-01-01

    Searching genomes for RNA secondary structure with computational methods has become an important approach to the annotation of non-coding RNAs. However, due to the lack of efficient algorithms for accurate RNA structure-sequence alignment, computer programs capable of fast and effectively searching genomes for RNA secondary structures have not been available. In this paper, a novel RNA structure profiling model is introduced based on the notion of a conformational graph to specify the consensus structure of an RNA family. Tree decomposition yields a small tree width t for such conformation graphs (e.g., t = 2 for stem loops and only a slight increase for pseudo-knots). Within this modelling framework, the optimal alignment of a sequence to the structure model corresponds to finding a maximum valued isomorphic subgraph and consequently can be accomplished through dynamic programming on the tree decomposition of the conformational graph in time O(k(t)N(2)), where k is a small parameter; and N is the size of the projiled RNA structure. Experiments show that the application of the alignment algorithm to search in genomes yields the same search accuracy as methods based on a Covariance model with a significant reduction in computation time. In particular; very accurate searches of tmRNAs in bacteria genomes and of telomerase RNAs in yeast genomes can be accomplished in days, as opposed to months required by other methods. The tree decomposition based searching tool is free upon request and can be downloaded at our site h t t p ://w.uga.edu/RNA-informatics/software/index.php.

  20. Kernel-based discriminant feature extraction using a representative dataset

    Science.gov (United States)

    Li, Honglin; Sancho Gomez, Jose-Luis; Ahalt, Stanley C.

    2002-07-01

    Discriminant Feature Extraction (DFE) is widely recognized as an important pre-processing step in classification applications. Most DFE algorithms are linear and thus can only explore the linear discriminant information among the different classes. Recently, there has been several promising attempts to develop nonlinear DFE algorithms, among which is Kernel-based Feature Extraction (KFE). The efficacy of KFE has been experimentally verified by both synthetic data and real problems. However, KFE has some known limitations. First, KFE does not work well for strongly overlapped data. Second, KFE employs all of the training set samples during the feature extraction phase, which can result in significant computation when applied to very large datasets. Finally, KFE can result in overfitting. In this paper, we propose a substantial improvement to KFE that overcomes the above limitations by using a representative dataset, which consists of critical points that are generated from data-editing techniques and centroid points that are determined by using the Frequency Sensitive Competitive Learning (FSCL) algorithm. Experiments show that this new KFE algorithm performs well on significantly overlapped datasets, and it also reduces computational complexity. Further, by controlling the number of centroids, the overfitting problem can be effectively alleviated.

  1. Analogy between gambling and measurement-based work extraction

    Science.gov (United States)

    Vinkler, Dror A.; Permuter, Haim H.; Merhav, Neri

    2016-04-01

    In information theory, one area of interest is gambling, where mutual information characterizes the maximal gain in wealth growth rate due to knowledge of side information; the betting strategy that achieves this maximum is named the Kelly strategy. In the field of physics, it was recently shown that mutual information can characterize the maximal amount of work that can be extracted from a single heat bath using measurement-based control protocols, i.e. using ‘information engines’. However, to the best of our knowledge, no relation between gambling and information engines has been presented before. In this paper, we briefly review the two concepts and then demonstrate an analogy between gambling, where bits are converted into wealth, and information engines, where bits representing measurements are converted into energy. From this analogy follows an extension of gambling to the continuous-valued case, which is shown to be useful for investments in currency exchange rates or in the stock market using options. Moreover, the analogy enables us to use well-known methods and results from one field to solve problems in the other. We present three such cases: maximum work extraction when the probability distributions governing the system and measurements are unknown, work extraction when some energy is lost in each cycle, e.g. due to friction, and an analysis of systems with memory. In all three cases, the analogy enables us to use known results in order to obtain new ones.

  2. Microcontroller based two axis microtron beam extraction system

    International Nuclear Information System (INIS)

    Ashoka, H.; Jathar, M.; Meshram, V.; Rao, Nageswara

    2009-01-01

    Microtron is an electron accelerator which is used to accelerate the electron beam. The Microtron consists of electro magnet with two poles separated by yoke for completion of path for magnetic flux lines. A compact Microtron capable of accelerating electrons up to 12 MeV has been developed in RRCAT. The beam from the Microtron has to be extracted from various orbits depending upon the user requirement (X-Y stage is built with an accuracy of 100 μm). This paper describes the design and development of microcontroller based two axis beam extraction system for Microtron, with a resolution of 50 μm to position the extraction tube with respect to selected orbit. Two axis motion controller is developed using current controlled micro-stepping driver mechanism, which uses Bipolar Chopper Drive for driving stepper motors. Each phase has 2A continuous driving capability. The system is provided with user selectable controls like speed, steps, direction, and mode. This system is provided with RS-232 interface, to accept commands from PC. This system also has local keyboard and LCD interface to use in Stand-alone mode (local Mode). (author)

  3. Extraction of Trivalent Actinides and Lanthanides from Californium Campaign Rework Solution Using TODGA-based Solvent Extraction System

    Energy Technology Data Exchange (ETDEWEB)

    Benker, Dennis [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Delmau, Laetitia Helene [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Dryman, Joshua Cory [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)

    2017-07-01

    This report presents the studies carried out to demonstrate the possibility of quantitatively extracting trivalent actinides and lanthanides from highly acidic solutions using a neutral ligand-based solvent extraction system. These studies stemmed from the perceived advantage of such systems over cationexchange- based solvent extraction systems that require an extensive feed adjustment to make a low-acid feed. The targeted feed solutions are highly acidic aqueous phases obtained after the dissolution of curium targets during a californium (Cf) campaign. Results obtained with actual Cf campaign solutions, but highly diluted to be manageable in a glove box, are presented, followed by results of tests run in the hot cells with Cf campaign rework solutions. It was demonstrated that a solvent extraction system based on the tetraoctyl diglycolamide molecule is capable of quantitatively extracting trivalent actinides from highly acidic solutions. This system was validated using actual feeds from a Cf campaign.

  4. RNA-based, transient modulation of gene expression in human haematopoietic stem and progenitor cells

    Science.gov (United States)

    Diener, Yvonne; Jurk, Marion; Kandil, Britta; Choi, Yeong-Hoon; Wild, Stefan; Bissels, Ute; Bosio, Andreas

    2015-01-01

    Modulation of gene expression is a useful tool to study the biology of haematopoietic stem and progenitor cells (HSPCs) and might also be instrumental to expand these cells for therapeutic approaches. Most of the studies so far have employed stable gene modification by viral vectors that are burdensome when translating protocols into clinical settings. Our study aimed at exploring new ways to transiently modify HSPC gene expression using non-integrating, RNA-based molecules. First, we tested different methods to deliver these molecules into HSPCs. The delivery of siRNAs with chemical transfection methods such as lipofection or cationic polymers did not lead to target knockdown, although we observed more than 90% fluorescent cells using a fluorochrome-coupled siRNA. Confocal microscopic analysis revealed that despite extensive washing, siRNA stuck to or in the cell surface, thereby mimicking a transfection event. In contrast, electroporation resulted in efficient, siRNA-mediated protein knockdown. For transient overexpression of proteins, we used optimised mRNA molecules with modified 5′- and 3′-UTRs. Electroporation of mRNA encoding GFP resulted in fast, efficient and persistent protein expression for at least seven days. Our data provide a broad-ranging comparison of transfection methods for hard-to-transfect cells and offer new opportunities for DNA-free, non-integrating gene modulation in HSPCs. PMID:26599627

  5. Nanosystems based on siRNA silencing HuR expression counteract diabetic retinopathy in rat.

    Science.gov (United States)

    Amadio, Marialaura; Pascale, Alessia; Cupri, Sarha; Pignatello, Rosario; Osera, Cecilia; D Agata, Velia; D Amico, Agata Grazia; Leggio, Gian Marco; Ruozi, Barbara; Govoni, Stefano; Drago, Filippo; Bucolo, Claudio

    2016-09-01

    We evaluated whether specifically and directly targeting human antigen R (HuR), a member of embryonic lethal abnormal vision (ELAV) proteins family, may represent a new potential therapeutic strategy to manage diabetic retinopathy. Nanosystems loaded with siRNA silencing HuR expression (lipoplexes), consisting of solid lipid nanoparticles (SLN) and liposomes (SUV) were prepared. Photon correlation spectroscopy analysis, Zeta potential measurement and atomic force microscopy (AFM) studies were carried out to characterize the complexation of siRNA with the lipid nanocarriers. Nanosystems were evaluated by using AFM and scanning electron microscopy. The lipoplexes were injected into the eye of streptozotocin (STZ)-induced diabetic rats. Retinal HuR and VEGF levels were detected by Western blot and ELISA, respectively. Retinal histology was also carried out. The results demonstrated that retinal HuR and VEGF are significantly increased in STZ-rats and are blunted by HuR siRNA treatment. Lipoplexes with a weak positive surface charge and with a 4:1 N/P (cationic lipid nitrogen to siRNA phosphate) ratio exert a better transfection efficiency, significantly dumping retinal HuR and VEGF levels. In conclusion, we demonstrated that siRNA can be efficiently delivered into the rat retina using lipid-based nanocarriers, and some of the lipoplexes loaded with siRNA silencing HuR expression are potential candidates to manage retinal diseases. Copyright © 2016 Elsevier Ltd. All rights reserved.

  6. Sensitive voltammetric detection of yeast RNA based on its interaction with Victoria Blue B

    Directory of Open Access Journals (Sweden)

    WEI SUN

    2009-12-01

    Full Text Available Voltammetric studies of the interaction of yeast RNA (y-RNA with Victoria Blue B (VBB are described in this paper. Furthermore, a linear sweep voltammetric method for the detection of y-RNA was established. The reaction conditions, such as acidity and amount of buffer solution, the concentration of VBB, the reaction time and temperature, etc., were carefully investigated by second order derivative linear sweep voltammetry. Under the optimal conditions, the reduction peak current of VBB at –0.75 V decreased greatly after the addition of y-RNA to the solution without any shift of the reduction peak potential. Based on the decrease of the peak current, a new quantitative method for the determination of y-RNA was developed. The effects of co-existing substances on the determination were carefully investigated and three synthetic samples were determined with satisfactory results. The stoichiometry of the VBB–y-RNA complex was calculated by linear sweep voltammetry and the interaction mechanism is discussed.

  7. tRNA acceptor-stem and anticodon bases embed separate features of amino acid chemistry

    Science.gov (United States)

    Carter, Charles W.; Wolfenden, Richard

    2016-01-01

    abstract The universal genetic code is a translation table by which nucleic acid sequences can be interpreted as polypeptides with a wide range of biological functions. That information is used by aminoacyl-tRNA synthetases to translate the code. Moreover, amino acid properties dictate protein folding. We recently reported that digital correlation techniques could identify patterns in tRNA identity elements that govern recognition by synthetases. Our analysis, and the functionality of truncated synthetases that cannot recognize the tRNA anticodon, support the conclusion that the tRNA acceptor stem houses an independent code for the same 20 amino acids that likely functioned earlier in the emergence of genetics. The acceptor-stem code, related to amino acid size, is distinct from a code in the anticodon that is related to amino acid polarity. Details of the acceptor-stem code suggest that it was useful in preserving key properties of stereochemically-encoded peptides that had developed the capacity to interact catalytically with RNA. The quantitative embedding of the chemical properties of amino acids into tRNA bases has implications for the origins of molecular biology. PMID:26595350

  8. An atlas of RNA base pairs involving modified nucleobases with optimal geometries and accurate energies

    KAUST Repository

    Chawla, Mohit

    2015-06-27

    Posttranscriptional modifications greatly enhance the chemical information of RNA molecules, contributing to explain the diversity of their structures and functions. A significant fraction of RNA experimental structures available to date present modified nucleobases, with half of them being involved in H-bonding interactions with other bases, i.e. ‘modified base pairs’. Herein we present a systematic investigation of modified base pairs, in the context of experimental RNA structures. To this end, we first compiled an atlas of experimentally observed modified base pairs, for which we recorded occurrences and structural context. Then, for each base pair, we selected a representative for subsequent quantum mechanics calculations, to find out its optimal geometry and interaction energy. Our structural analyses show that most of the modified base pairs are non Watson–Crick like and are involved in RNA tertiary structure motifs. In addition, quantum mechanics calculations quantify and provide a rationale for the impact of the different modifications on the geometry and stability of the base pairs they participate in.

  9. An atlas of RNA base pairs involving modified nucleobases with optimal geometries and accurate energies

    KAUST Repository

    Chawla, Mohit; Oliva, R.; Bujnicki, J. M.; Cavallo, Luigi

    2015-01-01

    Posttranscriptional modifications greatly enhance the chemical information of RNA molecules, contributing to explain the diversity of their structures and functions. A significant fraction of RNA experimental structures available to date present modified nucleobases, with half of them being involved in H-bonding interactions with other bases, i.e. ‘modified base pairs’. Herein we present a systematic investigation of modified base pairs, in the context of experimental RNA structures. To this end, we first compiled an atlas of experimentally observed modified base pairs, for which we recorded occurrences and structural context. Then, for each base pair, we selected a representative for subsequent quantum mechanics calculations, to find out its optimal geometry and interaction energy. Our structural analyses show that most of the modified base pairs are non Watson–Crick like and are involved in RNA tertiary structure motifs. In addition, quantum mechanics calculations quantify and provide a rationale for the impact of the different modifications on the geometry and stability of the base pairs they participate in.

  10. Capturing alternative secondary structures of RNA by decomposition of base-pairing probabilities.

    Science.gov (United States)

    Hagio, Taichi; Sakuraba, Shun; Iwakiri, Junichi; Mori, Ryota; Asai, Kiyoshi

    2018-02-19

    It is known that functional RNAs often switch their functions by forming different secondary structures. Popular tools for RNA secondary structures prediction, however, predict the single 'best' structures, and do not produce alternative structures. There are bioinformatics tools to predict suboptimal structures, but it is difficult to detect which alternative secondary structures are essential. We proposed a new computational method to detect essential alternative secondary structures from RNA sequences by decomposing the base-pairing probability matrix. The decomposition is calculated by a newly implemented software tool, RintW, which efficiently computes the base-pairing probability distributions over the Hamming distance from arbitrary reference secondary structures. The proposed approach has been demonstrated on ROSE element RNA thermometer sequence and Lysine RNA ribo-switch, showing that the proposed approach captures conformational changes in secondary structures. We have shown that alternative secondary structures are captured by decomposing base-paring probabilities over Hamming distance. Source code is available from http://www.ncRNA.org/RintW .

  11. URS DataBase: universe of RNA structures and their motifs.

    Science.gov (United States)

    Baulin, Eugene; Yacovlev, Victor; Khachko, Denis; Spirin, Sergei; Roytberg, Mikhail

    2016-01-01

    The Universe of RNA Structures DataBase (URSDB) stores information obtained from all RNA-containing PDB entries (2935 entries in October 2015). The content of the database is updated regularly. The database consists of 51 tables containing indexed data on various elements of the RNA structures. The database provides a web interface allowing user to select a subset of structures with desired features and to obtain various statistical data for a selected subset of structures or for all structures. In particular, one can easily obtain statistics on geometric parameters of base pairs, on structural motifs (stems, loops, etc.) or on different types of pseudoknots. The user can also view and get information on an individual structure or its selected parts, e.g. RNA-protein hydrogen bonds. URSDB employs a new original definition of loops in RNA structures. That definition fits both pseudoknot-free and pseudoknotted secondary structures and coincides with the classical definition in case of pseudoknot-free structures. To our knowledge, URSDB is the first database supporting searches based on topological classification of pseudoknots and on extended loop classification.Database URL: http://server3.lpm.org.ru/urs/. © The Author(s) 2016. Published by Oxford University Press.

  12. Glycerol-based deep eutectic solvents as extractants for the separation of MEK and ethanol via liquid-liquid extraction

    NARCIS (Netherlands)

    Rodriguez, N.R.; Ferré Güell, J.; Kroon, M.C.

    2016-01-01

    Four different glycerol-based deep eutectic solvents (DESs) were tested as extracting agents for the separation of the azeotropic mixture {methyl ethyl ketone + ethanol} via liquid-liquid extraction. The selected DESs for this work were: glycerol/choline chloride with molar ratios (4:1) and (2:1),

  13. Internal control for real-time polymerase chain reaction based on MS2 bacteriophage for RNA viruses diagnostics

    Directory of Open Access Journals (Sweden)

    Miriam Ribas Zambenedetti

    Full Text Available BACKGROUND Real-time reverse transcription polymerase chain reaction (RT-PCR is routinely used to detect viral infections. In Brazil, it is mandatory the use of nucleic acid tests to detect hepatitis C virus (HCV, hepatitis B virus and human immunodeficiency virus in blood banks because of the immunological window. The use of an internal control (IC is necessary to differentiate the true negative results from those consequent from a failure in some step of the nucleic acid test. OBJECTIVES The aim of this study was the construction of virus-modified particles, based on MS2 bacteriophage, to be used as IC for the diagnosis of RNA viruses. METHODS The MS2 genome was cloned into the pET47b(+ plasmid, generating pET47b(+-MS2. MS2-like particles were produced through the synthesis of MS2 RNA genome by T7 RNA polymerase. These particles were used as non-competitive IC in assays for RNA virus diagnostics. In addition, a competitive control for HCV diagnosis was developed by cloning a mutated HCV sequence into the MS2 replicase gene of pET47b(+-MS2, which produces a non-propagating MS2 particle. The utility of MS2-like particles as IC was evaluated in a one-step format multiplex real-time RT-PCR for HCV detection. FINDINGS We demonstrated that both competitive and non-competitive IC could be successfully used to monitor the HCV amplification performance, including the extraction, reverse transcription, amplification and detection steps, without compromising the detection of samples with low target concentrations. In conclusion, MS2-like particles generated by this strategy proved to be useful IC for RNA virus diagnosis, with advantage that they are produced by a low cost protocol. An attractive feature of this system is that it allows the construction of a multicontrol by the insertion of sequences from more than one pathogen, increasing its applicability for diagnosing different RNA viruses.

  14. Multispectral Image Road Extraction Based Upon Automated Map Conflation

    Science.gov (United States)

    Chen, Bin

    Road network extraction from remotely sensed imagery enables many important and diverse applications such as vehicle tracking, drone navigation, and intelligent transportation studies. There are, however, a number of challenges to road detection from an image. Road pavement material, width, direction, and topology vary across a scene. Complete or partial occlusions caused by nearby buildings, trees, and the shadows cast by them, make maintaining road connectivity difficult. The problems posed by occlusions are exacerbated with the increasing use of oblique imagery from aerial and satellite platforms. Further, common objects such as rooftops and parking lots are made of materials similar or identical to road pavements. This problem of common materials is a classic case of a single land cover material existing for different land use scenarios. This work addresses these problems in road extraction from geo-referenced imagery by leveraging the OpenStreetMap digital road map to guide image-based road extraction. The crowd-sourced cartography has the advantages of worldwide coverage that is constantly updated. The derived road vectors follow only roads and so can serve to guide image-based road extraction with minimal confusion from occlusions and changes in road material. On the other hand, the vector road map has no information on road widths and misalignments between the vector map and the geo-referenced image are small but nonsystematic. Properly correcting misalignment between two geospatial datasets, also known as map conflation, is an essential step. A generic framework requiring minimal human intervention is described for multispectral image road extraction and automatic road map conflation. The approach relies on the road feature generation of a binary mask and a corresponding curvilinear image. A method for generating the binary road mask from the image by applying a spectral measure is presented. The spectral measure, called anisotropy-tunable distance (ATD

  15. Javanese Character Feature Extraction Based on Shape Energy

    Directory of Open Access Journals (Sweden)

    Galih Hendra Wibowo

    2017-07-01

    Full Text Available Javanese character is one of Indonesia's noble culture, especially in Java. However, the number of Javanese people who are able to read the letter has decreased so that there need to be conservation efforts in the form of a system that is able to recognize the characters. One solution to these problem lies in Optical Character Recognition (OCR studies, where one of its heaviest points lies in feature extraction which is to distinguish each character. Shape Energy is one of feature extraction method with the basic idea of how the character can be distinguished simply through its skeleton. Based on the basic idea, then the development of feature extraction is done based on its components to produce an angular histogram with various variations of multiples angle. Furthermore, the performance test of this method and its basic method is performed in Javanese character dataset, which has been obtained from various images, is 240 data with 19 labels by using K-Nearest Neighbors as its classification method. Performance values were obtained based on the accuracy which is generated through the Cross-Validation process of 80.83% in the angular histogram with an angle of 20 degrees, 23% better than Shape Energy. In addition, other test results show that this method is able to recognize rotated character with the lowest performance value of 86% at 180-degree rotation and the highest performance value of 96.97% at 90-degree rotation. It can be concluded that this method is able to improve the performance of Shape Energy in the form of recognition of Javanese characters as well as robust to the rotation.

  16. Annotation-based feature extraction from sets of SBML models.

    Science.gov (United States)

    Alm, Rebekka; Waltemath, Dagmar; Wolfien, Markus; Wolkenhauer, Olaf; Henkel, Ron

    2015-01-01

    Model repositories such as BioModels Database provide computational models of biological systems for the scientific community. These models contain rich semantic annotations that link model entities to concepts in well-established bio-ontologies such as Gene Ontology. Consequently, thematically similar models are likely to share similar annotations. Based on this assumption, we argue that semantic annotations are a suitable tool to characterize sets of models. These characteristics improve model classification, allow to identify additional features for model retrieval tasks, and enable the comparison of sets of models. In this paper we discuss four methods for annotation-based feature extraction from model sets. We tested all methods on sets of models in SBML format which were composed from BioModels Database. To characterize each of these sets, we analyzed and extracted concepts from three frequently used ontologies, namely Gene Ontology, ChEBI and SBO. We find that three out of the methods are suitable to determine characteristic features for arbitrary sets of models: The selected features vary depending on the underlying model set, and they are also specific to the chosen model set. We show that the identified features map on concepts that are higher up in the hierarchy of the ontologies than the concepts used for model annotations. Our analysis also reveals that the information content of concepts in ontologies and their usage for model annotation do not correlate. Annotation-based feature extraction enables the comparison of model sets, as opposed to existing methods for model-to-keyword comparison, or model-to-model comparison.

  17. Kernel-Based Learning for Domain-Specific Relation Extraction

    Science.gov (United States)

    Basili, Roberto; Giannone, Cristina; Del Vescovo, Chiara; Moschitti, Alessandro; Naggar, Paolo

    In a specific process of business intelligence, i.e. investigation on organized crime, empirical language processing technologies can play a crucial role. The analysis of transcriptions on investigative activities, such as police interrogatories, for the recognition and storage of complex relations among people and locations is a very difficult and time consuming task, ultimately based on pools of experts. We discuss here an inductive relation extraction platform that opens the way to much cheaper and consistent workflows. The presented empirical investigation shows that accurate results, comparable to the expert teams, can be achieved, and parametrization allows to fine tune the system behavior for fitting domain-specific requirements.

  18. Contour extraction of echocardiographic images based on pre-processing

    Energy Technology Data Exchange (ETDEWEB)

    Hussein, Zinah Rajab; Rahmat, Rahmita Wirza; Abdullah, Lili Nurliyana [Department of Multimedia, Faculty of Computer Science and Information Technology, Department of Computer and Communication Systems Engineering, Faculty of Engineering University Putra Malaysia 43400 Serdang, Selangor (Malaysia); Zamrin, D M [Department of Surgery, Faculty of Medicine, National University of Malaysia, 56000 Cheras, Kuala Lumpur (Malaysia); Saripan, M Iqbal

    2011-02-15

    In this work we present a technique to extract the heart contours from noisy echocardiograph images. Our technique is based on improving the image before applying contours detection to reduce heavy noise and get better image quality. To perform that, we combine many pre-processing techniques (filtering, morphological operations, and contrast adjustment) to avoid unclear edges and enhance low contrast of echocardiograph images, after implementing these techniques we can get legible detection for heart boundaries and valves movement by traditional edge detection methods.

  19. Contour extraction of echocardiographic images based on pre-processing

    International Nuclear Information System (INIS)

    Hussein, Zinah Rajab; Rahmat, Rahmita Wirza; Abdullah, Lili Nurliyana; Zamrin, D M; Saripan, M Iqbal

    2011-01-01

    In this work we present a technique to extract the heart contours from noisy echocardiograph images. Our technique is based on improving the image before applying contours detection to reduce heavy noise and get better image quality. To perform that, we combine many pre-processing techniques (filtering, morphological operations, and contrast adjustment) to avoid unclear edges and enhance low contrast of echocardiograph images, after implementing these techniques we can get legible detection for heart boundaries and valves movement by traditional edge detection methods.

  20. Morphological operation based dense houses extraction from DSM

    OpenAIRE

    Li, Y.; Zhu, L.; Tachibana, K.; Shimamura, H.

    2014-01-01

    This paper presents a method of reshaping and extraction of markers and masks of the dense houses from the DSM based on mathematical morphology (MM). Houses in a digital surface model (DSM) are almost joined together in high-density housing areas, and most segmentation methods cannot completely separate them. We propose to label the markers of the buildings firstly and segment them into masks by watershed then. To avoid detecting more than one marker for a house or no marker at all d...

  1. A window-based time series feature extraction method.

    Science.gov (United States)

    Katircioglu-Öztürk, Deniz; Güvenir, H Altay; Ravens, Ursula; Baykal, Nazife

    2017-10-01

    This study proposes a robust similarity score-based time series feature extraction method that is termed as Window-based Time series Feature ExtraCtion (WTC). Specifically, WTC generates domain-interpretable results and involves significantly low computational complexity thereby rendering itself useful for densely sampled and populated time series datasets. In this study, WTC is applied to a proprietary action potential (AP) time series dataset on human cardiomyocytes and three precordial leads from a publicly available electrocardiogram (ECG) dataset. This is followed by comparing WTC in terms of predictive accuracy and computational complexity with shapelet transform and fast shapelet transform (which constitutes an accelerated variant of the shapelet transform). The results indicate that WTC achieves a slightly higher classification performance with significantly lower execution time when compared to its shapelet-based alternatives. With respect to its interpretable features, WTC has a potential to enable medical experts to explore definitive common trends in novel datasets. Copyright © 2017 Elsevier Ltd. All rights reserved.

  2. Answer Extraction Based on Merging Score Strategy of Hot Terms

    Institute of Scientific and Technical Information of China (English)

    LE Juan; ZHANG Chunxia; NIU Zhendong

    2016-01-01

    Answer extraction (AE) is one of the key technologies in developing the open domain Question&an-swer (Q&A) system . Its task is to yield the highest score to the expected answer based on an effective answer score strategy. We introduce an answer extraction method by Merging score strategy (MSS) based on hot terms. The hot terms are defined according to their lexical and syn-tactic features to highlight the role of the question terms. To cope with the syntactic diversities of the corpus, we propose four improved candidate answer score algorithms. Each of them is based on the lexical function of hot terms and their syntactic relationships with the candidate an-swers. Two independent corpus score algorithms are pro-posed to tap the role of the corpus in ranking the candi-date answers. Six algorithms are adopted in MSS to tap the complementary action among the corpus, the candi-date answers and the questions. Experiments demonstrate the effectiveness of the proposed strategy.

  3. Improving information extraction using a probability-based approach

    DEFF Research Database (Denmark)

    Kim, S.; Ahmed, Saeema; Wallace, K.

    2007-01-01

    Information plays a crucial role during the entire life-cycle of a product. It has been shown that engineers frequently consult colleagues to obtain the information they require to solve problems. However, the industrial world is now more transient and key personnel move to other companies...... or retire. It is becoming essential to retrieve vital information from archived product documents, if it is available. There is, therefore, great interest in ways of extracting relevant and sharable information from documents. A keyword-based search is commonly used, but studies have shown...... the recall, while maintaining the high precision, a learning approach that makes identification decisions based on a probability model, rather than simply looking up the presence of the pre-defined variations, looks promising. This paper presents the results of developing such a probability-based entity...

  4. Model-Based Extracted Water Desalination System for Carbon Sequestration

    Energy Technology Data Exchange (ETDEWEB)

    Dees, Elizabeth M. [General Electric Global Research Center, Niskayuna, NY (United States); Moore, David Roger [General Electric Global Research Center, Niskayuna, NY (United States); Li, Li [Pennsylvania State Univ., University Park, PA (United States); Kumar, Manish [Pennsylvania State Univ., University Park, PA (United States)

    2017-05-28

    Over the last 1.5 years, GE Global Research and Pennsylvania State University defined a model-based, scalable, and multi-stage extracted water desalination system that yields clean water, concentrated brine, and, optionally, salt. The team explored saline brines that ranged across the expected range for extracted water for carbon sequestration reservoirs (40,000 up to 220,000 ppm total dissolved solids, TDS). In addition, the validated the system performance at pilot scale with field-sourced water using GE’s pre-pilot and lab facilities. This project encompassed four principal tasks, in addition to Project Management and Planning: 1) identify a deep saline formation carbon sequestration site and a partner that are suitable for supplying extracted water; 2) conduct a techno-economic assessment and down-selection of pre-treatment and desalination technologies to identify a cost-effective system for extracted water recovery; 3) validate the downselected processes at the lab/pre-pilot scale; and 4) define the scope of the pilot desalination project. Highlights from each task are described below: Deep saline formation characterization The deep saline formations associated with the five DOE NETL 1260 Phase 1 projects were characterized with respect to their mineralogy and formation water composition. Sources of high TDS feed water other than extracted water were explored for high TDS desalination applications, including unconventional oil and gas and seawater reverse osmosis concentrate. Technoeconomic analysis of desalination technologies Techno-economic evaluations of alternate brine concentration technologies, including humidification-dehumidification (HDH), membrane distillation (MD), forward osmosis (FO), turboexpander-freeze, solvent extraction and high pressure reverse osmosis (HPRO), were conducted. These technologies were evaluated against conventional falling film-mechanical vapor recompression (FF-MVR) as a baseline desalination process. Furthermore, a

  5. RNA-Seq-based toxicogenomic assessment of fresh frozen and formalin-fixed tissues yields similar mechanistic insights.

    Science.gov (United States)

    Auerbach, Scott S; Phadke, Dhiral P; Mav, Deepak; Holmgren, Stephanie; Gao, Yuan; Xie, Bin; Shin, Joo Heon; Shah, Ruchir R; Merrick, B Alex; Tice, Raymond R

    2015-07-01

    Formalin-fixed, paraffin-embedded (FFPE) pathology specimens represent a potentially vast resource for transcriptomic-based biomarker discovery. We present here a comparison of results from a whole transcriptome RNA-Seq analysis of RNA extracted from fresh frozen and FFPE livers. The samples were derived from rats exposed to aflatoxin B1 (AFB1 ) and a corresponding set of control animals. Principal components analysis indicated that samples were separated in the two groups representing presence or absence of chemical exposure, both in fresh frozen and FFPE sample types. Sixty-five percent of the differentially expressed transcripts (AFB1 vs. controls) in fresh frozen samples were also differentially expressed in FFPE samples (overlap significance: P < 0.0001). Genomic signature and gene set analysis of AFB1 differentially expressed transcript lists indicated highly similar results between fresh frozen and FFPE at the level of chemogenomic signatures (i.e., single chemical/dose/duration elicited transcriptomic signatures), mechanistic and pathology signatures, biological processes, canonical pathways and transcription factor networks. Overall, our results suggest that similar hypotheses about the biological mechanism of toxicity would be formulated from fresh frozen and FFPE samples. These results indicate that phenotypically anchored archival specimens represent a potentially informative resource for signature-based biomarker discovery and mechanistic characterization of toxicity. Copyright © 2014 John Wiley & Sons, Ltd.

  6. Nanotechnology based approaches for detection and delivery of microRNA in healthcare and crop protection.

    Science.gov (United States)

    Chaudhary, Vrantika; Jangra, Sumit; Yadav, Neelam R

    2018-04-13

    Nanobiotechnology has the potential to revolutionize diverse sectors including medicine, agriculture, food, textile and pharmaceuticals. Disease diagnostics, therapeutics and crop protection strategies are fast emerging using nanomaterials preferably nanobiomaterials. It has potential for development of novel nanobiomolecules which offer several advantages over conventional treatment methods. RNA nanoparticles with many unique features are promising candidates in disease treatment. The miRNAs are involved in many biochemical and developmental pathways and their regulation in plants and animals. These appear to be a powerful tool for controlling various pathological diseases in human, plants and animals, however there are challenges associated with miRNA based nanotechnology. Several advancements made in the field of miRNA therapeutics make it an attractive approach, but a lot more has to be explored in nanotechnology assisted miRNA therapy. The miRNA based technologies can be employed for detection and combating crop diseases as well. Despite these potential advantages, nanobiotechnology applications in the agricultural sector are still in its infancy and have not yet made its mark in comparison with healthcare sector. The review provides a platform to discuss nature, role and use of miRNAs in nanobiotechnology applications.

  7. incaRNAfbinv: a web server for the fragment-based design of RNA sequences

    Science.gov (United States)

    Drory Retwitzer, Matan; Reinharz, Vladimir; Ponty, Yann; Waldispühl, Jérôme; Barash, Danny

    2016-01-01

    Abstract In recent years, new methods for computational RNA design have been developed and applied to various problems in synthetic biology and nanotechnology. Lately, there is considerable interest in incorporating essential biological information when solving the inverse RNA folding problem. Correspondingly, RNAfbinv aims at including biologically meaningful constraints and is the only program to-date that performs a fragment-based design of RNA sequences. In doing so it allows the design of sequences that do not necessarily exactly fold into the target, as long as the overall coarse-grained tree graph shape is preserved. Augmented by the weighted sampling algorithm of incaRNAtion, our web server called incaRNAfbinv implements the method devised in RNAfbinv and offers an interactive environment for the inverse folding of RNA using a fragment-based design approach. It takes as input: a target RNA secondary structure; optional sequence and motif constraints; optional target minimum free energy, neutrality and GC content. In addition to the design of synthetic regulatory sequences, it can be used as a pre-processing step for the detection of novel natural occurring RNAs. The two complementary methodologies RNAfbinv and incaRNAtion are merged together and fully implemented in our web server incaRNAfbinv, available at http://www.cs.bgu.ac.il/incaRNAfbinv. PMID:27185893

  8. RNA-Based TWIST1 Inhibition via Dendrimer Complex to Reduce Breast Cancer Cell Metastasis

    Directory of Open Access Journals (Sweden)

    James Finlay

    2015-01-01

    Full Text Available Breast cancer is the leading cause of cancer-related deaths among women in the United States, and survival rates are lower for patients with metastases and/or triple-negative breast cancer (TNBC; ER, PR, and Her2 negative. Understanding the mechanisms of cancer metastasis is therefore crucial to identify new therapeutic targets and develop novel treatments to improve patient outcomes. A potential target is the TWIST1 transcription factor, which is often overexpressed in aggressive breast cancers and is a master regulator of cellular migration through epithelial-mesenchymal transition (EMT. Here, we demonstrate an siRNA-based TWIST1 silencing approach with delivery using a modified poly(amidoamine (PAMAM dendrimer. Our results demonstrate that SUM1315 TNBC cells efficiently take up PAMAM-siRNA complexes, leading to significant knockdown of TWIST1 and EMT-related target genes. Knockdown lasts up to one week after transfection and leads to a reduction in migration and invasion, as determined by wound healing and transwell assays. Furthermore, we demonstrate that PAMAM dendrimers can deliver siRNA to xenograft orthotopic tumors and siRNA remains in the tumor for at least four hours after treatment. These results suggest that further development of dendrimer-based delivery of siRNA for TWIST1 silencing may lead to a valuable adjunctive therapy for patients with TNBC.

  9. Monitoring of the antiviral potential of bee venom and wax extracts against Adeno-7 (DNA) and Rift Valley fever virus (RNA) viruses models.

    Science.gov (United States)

    Hassan, Mostafa I; Mohamed, Aly F; Amer, Moner A; Hammad, Kotb M; Riad, Saber A

    2015-04-01

    This study monitored the antiviral potential of bee venom and four wax extracts, ethanol white and black beeswax (EWW/EBW) and acetone white and black beeswax (AWW/ABW) extracts. Two different virus models namely Adeno-7 as DNA model and RVFV as RNA virus models. End point calculation assay was used to calculate virus depletion titer. The depletion of viral infectivity titer of ABW to Adeno-7 virus showed strong antiviral activity recorded a depletion of viral infectivity titer (1.66 log (10)/ ml) that gave equal action with bee venom and more than interferon IFN (1 log (10)/ ml). On the other hand, antiviral activity of EBW showed a moderate potential, while AWW showed no antiviral activity. Finally EWW showed synergetic activity against Adeno-7 virus activity. Thus, activity of wax extracts to RVFV was arranged in order of IFN bee venom > AWW & EBW > EWW and ABW recorded 3.34, 0.65, 0.5, 0.34 respectively. It is the first time to study the beeswax effect against DNA and RNA virus' models; acetone black beeswax recorded a depletion titer 1.66 log (10)/ml.

  10. Generating unstable resonances for extraction schemes based on transverse splitting

    Directory of Open Access Journals (Sweden)

    M. Giovannozzi

    2009-02-01

    Full Text Available A few years ago, a novel multiturn extraction scheme was proposed, based on particle trapping inside stable resonances. Numerical simulations and experimental tests have confirmed the feasibility of such a scheme for low order resonances. While the third-order resonance is generically unstable and those higher than fourth order are generically stable, the fourth-order resonance can be either stable or unstable depending on the specifics of the system under consideration. By means of the normal form, a general approach to control the stability of the fourth-order resonance has been derived. This approach is based on the control of the amplitude detuning and the general form for a lattice with an arbitrary number of sextupole and octupole families is derived in this paper. Numerical simulations have confirmed the analytical results and have shown that, when crossing the unstable fourth-order resonance, the region around the center of the phase space is depleted and particles are trapped in only the four stable islands. A four-turn extraction could be designed using this technique.

  11. Quantum Jarzynski equality of measurement-based work extraction.

    Science.gov (United States)

    Morikuni, Yohei; Tajima, Hiroyasu; Hatano, Naomichi

    2017-03-01

    Many studies of quantum-size heat engines assume that the dynamics of an internal system is unitary and that the extracted work is equal to the energy loss of the internal system. Both assumptions, however, should be under scrutiny. In the present paper, we analyze quantum-scale heat engines, employing the measurement-based formulation of the work extraction recently introduced by Hayashi and Tajima [M. Hayashi and H. Tajima, arXiv:1504.06150]. We first demonstrate the inappropriateness of the unitary time evolution of the internal system (namely, the first assumption above) using a simple two-level system; we show that the variance of the energy transferred to an external system diverges when the dynamics of the internal system is approximated to a unitary time evolution. Second, we derive the quantum Jarzynski equality based on the formulation of Hayashi and Tajima as a relation for the work measured by an external macroscopic apparatus. The right-hand side of the equality reduces to unity for "natural" cyclic processes but fluctuates wildly for noncyclic ones, exceeding unity often. This fluctuation should be detectable in experiments and provide evidence for the present formulation.

  12. Liquid-phase extraction coupled with metal-organic frameworks-based dispersive solid phase extraction of herbicides in peanuts.

    Science.gov (United States)

    Li, Na; Wang, Zhibing; Zhang, Liyuan; Nian, Li; Lei, Lei; Yang, Xiao; Zhang, Hanqi; Yu, Aimin

    2014-10-01

    Liquid-phase extraction coupled with metal-organic frameworks-based dispersive solid phase extraction was developed and applied to the extraction of pesticides in high fatty matrices. The herbicides were ultrasonically extracted from peanut using ethyl acetate as extraction solvent. The separation of the analytes from a large amount of co-extractive fat was achieved by dispersive solid-phase extraction using MIL-101(Cr) as sorbent. In this step, the analytes were adsorbed on MIL-101(Cr) and the fat remained in bulk. The herbicides were separated and determined by high-performance liquid chromatography. The experimental parameters, including type and volume of extraction solvent, ultrasonication time, volume of hexane and eluting solvent, amount of MIL-101(Cr) and dispersive solid phase extraction time, were optimized. The limits of detection for herbicides range from 0.98 to 1.9 μg/kg. The recoveries of the herbicides are in the range of 89.5-102.7% and relative standard deviations are equal or lower than 7.0%. The proposed method is simple, effective and suitable for treatment of the samples containing high content of fat. Copyright © 2014 Elsevier B.V. All rights reserved.

  13. A mRNA-Responsive G-Quadruplex-Based Drug Release System

    Directory of Open Access Journals (Sweden)

    Hidenobu Yaku

    2015-04-01

    Full Text Available G-quadruplex-based drug delivery carriers (GDDCs were designed to capture and release a telomerase inhibitor in response to a target mRNA. Hybridization between a loop on the GDDC structure and the mRNA should cause the G-quadruplex structure of the GDDC to unfold and release the bound inhibitor, anionic copper(II phthalocyanine (CuAPC. As a proof of concept, GDDCs were designed with a 10-30-mer loop, which can hybridize with a target sequence in epidermal growth factor receptor (EGFR mRNA. Structural analysis using circular dichroism (CD spectroscopy showed that the GDDCs form a (3 + 1 type G-quadruplex structure in 100 mM KCl and 10 mM MgCl2 in the absence of the target RNA. Visible absorbance titration experiments showed that the GDDCs bind to CuAPC with Ka values of 1.5 × 105 to 5.9 × 105 M−1 (Kd values of 6.7 to 1.7 μM at 25 °C, depending on the loop length. Fluorescence titration further showed that the G-quadruplex structure unfolds upon binding to the target RNA with Ka values above 1.0 × 108 M−1 (Kd values below 0.01 μM at 25 °C. These results suggest the carrier can sense and bind to the target RNA, which should result in release of the bound drug. Finally, visible absorbance titration experiments demonstrated that the GDDC release CuAPC in response to the target RNA.

  14. Construction of permanently inducible miRNA-based expression vectors using site-specific recombinases

    Directory of Open Access Journals (Sweden)

    Garwick-Coppens Sara E

    2011-11-01

    Full Text Available Abstract Background RNA interference (RNAi is a conserved gene silencing mechanism mediated by small inhibitory microRNAs (miRNAs. Promoter-driven miRNA expression vectors have emerged as important tools for delivering natural or artificially designed miRNAs to eukaryotic cells and organisms. Such systems can be used to query the normal or pathogenic functions of natural miRNAs or messenger RNAs, or to therapeutically silence disease genes. Results As with any molecular cloning procedure, building miRNA-based expression constructs requires a time investment and some molecular biology skills. To improve efficiency and accelerate the construction process, we developed a method to rapidly generate miRNA expression vectors using recombinases instead of more traditional cut-and-paste molecular cloning techniques. In addition to streamlining the construction process, our cloning strategy provides vectors with added versatility. In our system, miRNAs can be constitutively expressed from the U6 promoter, or inducibly expressed by Cre recombinase. We also engineered a built-in mechanism to destroy the vector with Flp recombinase, if desired. Finally, to further simplify the construction process, we developed a software package that automates the prediction and design of optimal miRNA sequences using our system. Conclusions We designed and tested a modular system to rapidly clone miRNA expression cassettes. Our strategy reduces the hands-on time required to successfully generate effective constructs, and can be implemented in labs with minimal molecular cloning expertise. This versatile system provides options that permit constitutive or inducible miRNA expression, depending upon the needs of the end user. As such, it has utility for basic or translational applications.

  15. Modifying a standard method allows simultaneous extraction of RNA and protein, enabling detection of enzymes in the rat retina with low expressions and protein levels.

    Science.gov (United States)

    Agardh, Elisabet; Gustavsson, Carin; Hagert, Per; Nilsson, Marie; Agardh, Carl-David

    2006-02-01

    The aim of the study was to evaluate messenger RNA and protein expression in limited amounts of tissue with low protein content. The Chomczynski method was used for simultaneous extraction of RNA, and protein was modified in the protein isolation step. Template mass and cycling time for the complementary DNA synthesis step of real-time reverse transcription-polymerase chain reaction (RT-PCR) for analysis of catalase, copper/zinc superoxide dismutase, manganese superoxide dismutase, the catalytic subunit of glutamylcysteine ligase, glutathione peroxidase 1, and the endogenous control cyclophilin B (CypB) were optimized before PCR. Polymerase chain reaction accuracy and efficacy were demonstrated by calculating the regression (R2) values of the separate amplification curves. Appropriate antibodies, blocking buffers, and running conditions were established for Western blot, and protein detection and multiplex assays with CypB were performed for each target. During the extraction procedure, the protein phase was dissolved in a modified washing buffer containing 0.1% sodium dodecyl sulfate, followed by ultrafiltration. Enzyme expression on real-time RT-PCR was accomplished with high reliability and reproducibility (R2, 0.990-0.999), and all enzymes except for glutathione peroxidase 1 were detectable in individual retinas on Western blot. Western blot multiplexing with CypB was possible for all targets. In conclusion, connecting gene expression directly to protein levels in the individual rat retina was possible by simultaneous extraction of RNA and protein. Real-time RT-PCR and Western blot allowed accurate detection of retinal protein expressions and levels.

  16. Nanoparticle-based delivery of small interfering RNA: challenges for cancer therapy

    Directory of Open Access Journals (Sweden)

    Miele E

    2012-07-01

    Full Text Available Evelina Miele,1,* Gian Paolo Spinelli,2,* Ermanno Miele,3 Enzo Di Fabrizio,3,6 Elisabetta Ferretti,4 Silverio Tomao,2 Alberto Gulino,1,5 1Department of Molecular Medicine, 2Department of Medico-Surgical Sciences and Biotechnologies, Sapienza University of Rome, Rome, 3Nanostructures, Istituto Italiano di Tecnologia, via Morego, 30, 16163 Genova, 4Department of Experimental Medicine, Sapienza University of Rome, Rome, 5Center for Life Nanoscience, Istituto Italiano di Tecnologia, Rome, Italy, 6BIONEM lab, University of Magna Graecia, Campus S. Venuta, Viale Europa 88100 Catanzaro, Italy *These authors contributed equally to this workAbstract: During recent decades there have been remarkable advances and profound changes in cancer therapy. Many therapeutic strategies learned at the bench, including monoclonal antibodies and small molecule inhibitors, have been used at the bedside, leading to important successes. One of the most important advances in biology has been the discovery that small interfering RNA (siRNA is able to regulate the expression of genes, by a phenomenon known as RNA interference (RNAi. RNAi is one of the most rapidly growing fields of research in biology and therapeutics. Much research effort has gone into the application of this new discovery in the treatment of various diseases, including cancer. However, even though these molecules may have potential and strong utility, some limitations make their clinical application difficult, including delivery problems, side effects due to off-target actions, disturbance of physiological functions of the cellular machinery involved in gene silencing, and induction of the innate immune response. Many researchers have attempted to overcome these limitations and to improve the safety of potential RNAi-based therapeutics. Nanoparticles, which are nanostructured entities with tunable size, shape, and surface, as well as biological behavior, provide an ideal opportunity to modify current

  17. Model-based Extracted Water Desalination System for Carbon Sequestration

    Energy Technology Data Exchange (ETDEWEB)

    Gettings, Rachel; Dees, Elizabeth

    2017-03-23

    The focus of this research effort centered around water recovery from high Total Dissolved Solids (TDS) extracted waters (180,000 mg/L) using a combination of water recovery (partial desalination) technologies. The research goals of this project were as follows: 1. Define the scope and test location for pilot-scale implementation of the desalination system, 2.Define a scalable, multi-stage extracted water desalination system that yields clean water, concentrated brine, and, salt from saline brines, and 3. Validate overall system performance with field-sourced water using GE pre-pilot lab facilities. Conventional falling film-mechanical vapor recompression (FF-MVR) technology was established as a baseline desalination process. A quality function deployment (QFD) method was used to compare alternate high TDS desalination technologies to the base case FF-MVR technology, including but not limited to: membrane distillation (MD), forward osmosis (FO), and high pressure reverse osmosis (HPRO). Technoeconomic analysis of high pressure reverse osmosis (HPRO) was performed comparing the following two cases: 1. a hybrid seawater RO (SWRO) plus HPRO system and 2. 2x standard seawater RO system, to achieve the same total pure water recovery rate. Pre-pilot-scale tests were conducted using field production water to validate key process steps for extracted water pretreatment. Approximately 5,000 gallons of field produced water was processed through, microfiltration, ultrafiltration, and steam regenerable sorbent operations. Improvements in membrane materials of construction were considered as necessary next steps to achieving further improvement in element performance at high pressure. Several modifications showed promising results in their ability to withstand close to 5,000 PSI without gross failure.

  18. Emerging RNA-based drugs: siRNAs, microRNAs and derivates.

    Science.gov (United States)

    Pereira, Tiago Campos; Lopes-Cendes, Iscia

    2012-09-01

    An emerging new category of therapeutic agents based on ribonucleic acid has emerged and shown very promising in vitro, animal and pre-clinical results, known as small interfering RNAs (siRNAs), microRNAs mimics (miRNA mimics) and their derivates. siRNAs are small RNA molecules that promote potent and specific silencing of mutant, exogenous or aberrant genes through a mechanism known as RNA interference. These agents have called special attention to medicine since they have been used to experimentally treat a series of neurological conditions with distinct etiologies such as prion, viral, bacterial, fungal, genetic disorders and others. siRNAs have also been tested in other scenarios such as: control of anxiety, alcohol consumption, drug-receptor blockage and inhibition of pain signaling. Although in a much earlier stage, miRNAs mimics, anti-miRs and small activating RNAs (saRNAs) also promise novel therapeutic approaches to control gene expression. In this review we intend to introduce clinicians and medical researchers to the most recent advances in the world of siRNA- and miRNA-mediated gene control, its history, applications in cells, animals and humans, delivery methods (an yet unsolved hurdle), current status and possible applications in future clinical practice.

  19. Sequence-specific inhibition of Dicer measured with a force-based microarray for RNA ligands.

    Science.gov (United States)

    Limmer, Katja; Aschenbrenner, Daniela; Gaub, Hermann E

    2013-04-01

    Malfunction of protein translation causes many severe diseases, and suitable correction strategies may become the basis of effective therapies. One major regulatory element of protein translation is the nuclease Dicer that cuts double-stranded RNA independently of the sequence into pieces of 19-22 base pairs starting the RNA interference pathway and activating miRNAs. Inhibiting Dicer is not desirable owing to its multifunctional influence on the cell's gene regulation. Blocking specific RNA sequences by small-molecule binding, however, is a promising approach to affect the cell's condition in a controlled manner. A label-free assay for the screening of site-specific interference of small molecules with Dicer activity is thus needed. We used the Molecular Force Assay (MFA), recently developed in our lab, to measure the activity of Dicer. As a model system, we used an RNA sequence that forms an aptamer-binding site for paromomycin, a 615-dalton aminoglycoside. We show that Dicer activity is modulated as a function of concentration and incubation time: the addition of paromomycin leads to a decrease of Dicer activity according to the amount of ligand. The measured dissociation constant of paromomycin to its aptamer was found to agree well with literature values. The parallel format of the MFA allows a large-scale search and analysis for ligands for any RNA sequence.

  20. A plasmonic colorimetric strategy for visual miRNA detection based on hybridization chain reaction

    Science.gov (United States)

    Miao, Jie; Wang, Jingsheng; Guo, Jinyang; Gao, Huiguang; Han, Kun; Jiang, Chengmin; Miao, Peng

    2016-08-01

    In this work, a novel colorimetric strategy for miRNA analysis is proposed based on hybridization chain reaction (HCR)-mediated localized surface plasmon resonance (LSPR) variation of silver nanoparticles (AgNPs). miRNA in the sample to be tested is able to release HCR initiator from a solid interface to AgNPs colloid system by toehold exchange-mediated strand displacement, which then triggers the consumption of fuel strands with single-stranded tails for HCR. The final produced long nicked double-stranded DNA loses the ability to protect AgNPs from salt-induced aggregation. The stability variation of the colloid system can then be monitored by recording corresponding UV-vis spectrum and initial miRNA level is thus determined. This sensing system involves only four DNA strands which is quite simple. The practical utility is confirmed to be excellent by employing different biological samples.

  1. Contactless physiological signals extraction based on skin color magnification

    Science.gov (United States)

    Suh, Kun Ha; Lee, Eui Chul

    2017-11-01

    Although the human visual system is not sufficiently sensitive to perceive blood circulation, blood flow caused by cardiac activity makes slight changes on human skin surfaces. With advances in imaging technology, it has become possible to capture these changes through digital cameras. However, it is difficult to obtain clear physiological signals from such changes due to its fineness and noise factors, such as motion artifacts and camera sensing disturbances. We propose a method for extracting physiological signals with improved quality from skin colored-videos recorded with a remote RGB camera. The results showed that our skin color magnification method reveals the hidden physiological components remarkably in the time-series signal. A Korea Food and Drug Administration-approved heart rate monitor was used for verifying the resulting signal synchronized with the actual cardiac pulse, and comparisons of signal peaks showed correlation coefficients of almost 1.0. In particular, our method can be an effective preprocessing before applying additional postfiltering techniques to improve accuracy in image-based physiological signal extractions.

  2. Tie Points Extraction for SAR Images Based on Differential Constraints

    Science.gov (United States)

    Xiong, X.; Jin, G.; Xu, Q.; Zhang, H.

    2018-04-01

    Automatically extracting tie points (TPs) on large-size synthetic aperture radar (SAR) images is still challenging because the efficiency and correct ratio of the image matching need to be improved. This paper proposes an automatic TPs extraction method based on differential constraints for large-size SAR images obtained from approximately parallel tracks, between which the relative geometric distortions are small in azimuth direction and large in range direction. Image pyramids are built firstly, and then corresponding layers of pyramids are matched from the top to the bottom. In the process, the similarity is measured by the normalized cross correlation (NCC) algorithm, which is calculated from a rectangular window with the long side parallel to the azimuth direction. False matches are removed by the differential constrained random sample consensus (DC-RANSAC) algorithm, which appends strong constraints in azimuth direction and weak constraints in range direction. Matching points in the lower pyramid images are predicted with the local bilinear transformation model in range direction. Experiments performed on ENVISAT ASAR and Chinese airborne SAR images validated the efficiency, correct ratio and accuracy of the proposed method.

  3. TIE POINTS EXTRACTION FOR SAR IMAGES BASED ON DIFFERENTIAL CONSTRAINTS

    Directory of Open Access Journals (Sweden)

    X. Xiong

    2018-04-01

    Full Text Available Automatically extracting tie points (TPs on large-size synthetic aperture radar (SAR images is still challenging because the efficiency and correct ratio of the image matching need to be improved. This paper proposes an automatic TPs extraction method based on differential constraints for large-size SAR images obtained from approximately parallel tracks, between which the relative geometric distortions are small in azimuth direction and large in range direction. Image pyramids are built firstly, and then corresponding layers of pyramids are matched from the top to the bottom. In the process, the similarity is measured by the normalized cross correlation (NCC algorithm, which is calculated from a rectangular window with the long side parallel to the azimuth direction. False matches are removed by the differential constrained random sample consensus (DC-RANSAC algorithm, which appends strong constraints in azimuth direction and weak constraints in range direction. Matching points in the lower pyramid images are predicted with the local bilinear transformation model in range direction. Experiments performed on ENVISAT ASAR and Chinese airborne SAR images validated the efficiency, correct ratio and accuracy of the proposed method.

  4. Fractal Complexity-Based Feature Extraction Algorithm of Communication Signals

    Science.gov (United States)

    Wang, Hui; Li, Jingchao; Guo, Lili; Dou, Zheng; Lin, Yun; Zhou, Ruolin

    How to analyze and identify the characteristics of radiation sources and estimate the threat level by means of detecting, intercepting and locating has been the central issue of electronic support in the electronic warfare, and communication signal recognition is one of the key points to solve this issue. Aiming at accurately extracting the individual characteristics of the radiation source for the increasingly complex communication electromagnetic environment, a novel feature extraction algorithm for individual characteristics of the communication radiation source based on the fractal complexity of the signal is proposed. According to the complexity of the received signal and the situation of environmental noise, use the fractal dimension characteristics of different complexity to depict the subtle characteristics of the signal to establish the characteristic database, and then identify different broadcasting station by gray relation theory system. The simulation results demonstrate that the algorithm can achieve recognition rate of 94% even in the environment with SNR of -10dB, and this provides an important theoretical basis for the accurate identification of the subtle features of the signal at low SNR in the field of information confrontation.

  5. Mobility of radionuclides based on sequential extraction of soils

    International Nuclear Information System (INIS)

    Salbu, B.; Oughton, D.H.; Lien, H.N.; Oestby, G.; Strand, P.

    1992-01-01

    Since 1989, core samples of soil and vegetation from semi-natural pastures have been collected at selected sites in Norway during the growing season. The activity concentrations in soil and vegetation as well as transfer coefficients vary significantly between regions, within regions and even within sampling plot areas. In order to differentiate between mobil and inert fractions of radioactive and stable isotopes of Cs and Sr in soils, samples were extracted sequentially using agents with increasing dissolution power. The reproducibility of the sequential extraction technique is good and the data obtained seems most informative. As the distribution pattern for radioactive and stable isotopes of Cs and Sr are similar, a high degree of isotopic exchange is indicated. Based on easily leachable fractions, mobility factors are calculated. In general the mobility of 90 Sr is higher than for 137 Cs. Mobility factors are not significantly influenced by seasonal variations, but a decrease in the mobile fraction in soil with time is indicated. Mobility factors should be considered useful for modelling purposes. (au)

  6. New miRNA labeling method for bead-based quantification

    Directory of Open Access Journals (Sweden)

    Lanfranchi Gerolamo

    2010-06-01

    Full Text Available Abstract Background microRNAs (miRNAs are small single-stranded non-coding RNAs that act as crucial regulators of gene expression. Different methods have been developed for miRNA expression profiling in order to better understand gene regulation in normal and pathological conditions. miRNAs expression values obtained from large scale methodologies such as microarrays still need a validation step with alternative technologies. Results Here we have applied with an innovative approach, the Luminex® xMAP™ technology validate expression data of differentially expressed miRNAs obtained from high throughput arrays. We have developed a novel labeling system of small RNA molecules (below 200 nt, optimizing the sensitive cloning method for miRNAs, termed miRNA amplification profiling (mRAP. The Luminex expression patterns of three miRNAs (miR-23a, miR-27a and miR-199a in seven different cell lines have been validated by TaqMan miRNA assay. In all cases, bead-based meas were confirmed by the data obtained by TaqMan and microarray technologies. Conclusions We demonstrate that the measure of individual miRNA by the bead-based method is feasible, high speed, sensitive and low cost. The Luminex® xMAP™ technology also provides flexibility, since the central reaction can be scaled up with additional miRNA capturing beads, allowing validation of many differentially expressed miRNAs obtained from microarrays in a single experiment. We propose this technology as an alternative method to qRT-PCR for validating miRNAs expression data obtained with high-throughput technologies.

  7. Base substitutions at scissile bond sites are sufficient to alter RNA-binding and cleavage activity of RNase III.

    Science.gov (United States)

    Kim, Kyungsub; Sim, Se-Hoon; Jeon, Che Ok; Lee, Younghoon; Lee, Kangseok

    2011-02-01

    RNase III, a double-stranded RNA-specific endoribonuclease, degrades bdm mRNA via cleavage at specific sites. To better understand the mechanism of cleavage site selection by RNase III, we performed a genetic screen for sequences containing mutations at the bdm RNA cleavage sites that resulted in altered mRNA stability using a transcriptional bdm'-'cat fusion construct. While most of the isolated mutants showed the increased bdm'-'cat mRNA stability that resulted from the inability of RNase III to cleave the mutated sequences, one mutant sequence (wt-L) displayed in vivo RNA stability similar to that of the wild-type sequence. In vivo and in vitro analyses of the wt-L RNA substrate showed that it was cut only once on the RNA strand to the 5'-terminus by RNase III, while the binding constant of RNase III to this mutant substrate was moderately increased. A base substitution at the uncleaved RNase III cleavage site in wt-L mutant RNA found in another mutant lowered the RNA-binding affinity by 11-fold and abolished the hydrolysis of scissile bonds by RNase III. Our results show that base substitutions at sites forming the scissile bonds are sufficient to alter RNA cleavage as well as the binding activity of RNase III. © 2010 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.

  8. Towards a more appropriate water based extraction for the assessment of organic contaminant availability

    International Nuclear Information System (INIS)

    Hickman, Zachary A.; Reid, Brian J.

    2005-01-01

    This study correlated extractabilities of 37 d aged phenanthrene residues in four dissimilar soils with the fraction that was available for earthworm (Lumbricus rubellus) accumulation and microorganism (Pseudomonas sp.) mineralisation. Extractability was determined using two established techniques, namely (1) a water based extraction using CO 2 equilibrated water and (2) an aqueous based hydroxypropyl-β-cyclodextrin (HPCD) extraction. Results showed no relationship between earthworm accumulation and phenanthrene extractability using either HPCD (r 2 =0.07; slope=-4.76; n=5) or the water based extraction (r 2 =0.31; slope=-5.34; n=5). Earthworm accumulation was overestimated by both techniques. In contrast, the fraction of phenanthrene extractable using both the HPCD technique and the water based extraction correlated strongly with microbial mineralisation. However, the slopes of these linear relationships were 0.48 (r 2 =0.96; n=10), and 0.99 (r 2 =0.88; n=10) for the water based extraction and HPCD, respectively. Thus, the HPCD extraction provided values that were numerically close to the mineralisation values, whilst the water based extraction values were approximately half the mineralisation values. It is submitted that HPCD extraction provided an appropriate method of assessing the fraction of contaminant available for microbial mineralisation in these dissimilar soils. - No significant difference was found between microbially mineralised phenanthrene and extractability using hydroxypropyl-β-cyclodextrin in four dissimilar soils; the water-only extraction removed half of this fraction

  9. COSMO-RS-based extractant screening for phenol extraction as model system

    NARCIS (Netherlands)

    Burghoff, B.; Goetheer, E.L.V.; Haan, A.B. de

    2008-01-01

    The focus of this investigation is the development of a fast and reliable extractant screening approach. Phenol extraction is selected as the model process. A quantum chemical conductor-like screening model for real solvents (COSMO-RS) is combined with molecular design considerations. For this

  10. Towards a “Sample-In, Answer-Out” Point-of-Care Platform for Nucleic Acid Extraction and Amplification: Using an HPV E6/E7 mRNA Model System

    Directory of Open Access Journals (Sweden)

    Anja Gulliksen

    2012-01-01

    Full Text Available The paper presents the development of a “proof-of-principle” hands-free and self-contained diagnostic platform for detection of human papillomavirus (HPV E6/E7 mRNA in clinical specimens. The automated platform performs chip-based sample preconcentration, nucleic acid extraction, amplification, and real-time fluorescent detection with minimal user interfacing. It consists of two modular prototypes, one for sample preparation and one for amplification and detection; however, a common interface is available to facilitate later integration into one single module. Nucleic acid extracts (n=28 from cervical cytology specimens extracted on the sample preparation chip were tested using the PreTect HPV-Proofer and achieved an overall detection rate for HPV across all dilutions of 50%–85.7%. A subset of 6 clinical samples extracted on the sample preparation chip module was chosen for complete validation on the NASBA chip module. For 4 of the samples, a 100% amplification for HPV 16 or 33 was obtained at the 1 : 10 dilution for microfluidic channels that filled correctly. The modules of a “sample-in, answer-out” diagnostic platform have been demonstrated from clinical sample input through sample preparation, amplification and final detection.

  11. Gravitational wave extraction based on Cauchy-characteristic extraction and characteristic evolution

    Energy Technology Data Exchange (ETDEWEB)

    Babiuc, Maria [Department of Physics and Astronomy, University of Pittsburgh, Pittsburgh, PA 15260 (United States); Szilagyi, Bela [Department of Physics and Astronomy, University of Pittsburgh, Pittsburgh, PA 15260 (United States); Max-Planck-Institut fuer Gravitationsphysik, Albert-Einstein-Institut, Am Muehlenberg 1, D-14476 Golm (Germany); Hawke, Ian [Max-Planck-Institut fuer Gravitationsphysik, Albert-Einstein-Institut, Am Muehlenberg 1, D-14476 Golm (Germany); School of Mathematics, University of Southampton, Southampton SO17 1BJ (United Kingdom); Zlochower, Yosef [Department of Physics and Astronomy, and Center for Gravitational Wave Astronomy, University of Texas at Brownsville, Brownsville, TX 78520 (United States)

    2005-12-07

    We implement a code to find the gravitational news at future null infinity by using data from a Cauchy code as boundary data for a characteristic code. This technique of Cauchy-characteristic extraction (CCE) allows for the unambiguous extraction of gravitational waves from numerical simulations. We first test the technique on non-radiative spacetimes: Minkowski spacetime, perturbations of Minkowski spacetime and static black hole spacetimes in various gauges. We show the convergence and limitations of the algorithm and illustrate its success in cases where other wave extraction methods fail. We further apply our techniques to a standard radiative test case for wave extraction, a linearized Teukolsky wave, presenting our results in comparison to the Zerilli technique, and we argue for the advantages of our method of extraction.

  12. Development of a flat membrane based device for electromembrane extraction

    DEFF Research Database (Denmark)

    Huang, Chuixiu; Eibak, Lars Erik Eng; Gjelstad, Astrid

    2014-01-01

    this EME device, exhaustive extraction of the basic drugs quetiapine, citalopram, amitriptyline, methadone and sertraline was investigated from both acidified water samples and human plasma. The volume of acceptor solution, extraction time, and extraction voltage were found to be important factors...

  13. A quantitative analysis of secondary RNA structure using domination based parameters on trees

    Directory of Open Access Journals (Sweden)

    Zou Yue

    2006-03-01

    Full Text Available Abstract Background It has become increasingly apparent that a comprehensive database of RNA motifs is essential in order to achieve new goals in genomic and proteomic research. Secondary RNA structures have frequently been represented by various modeling methods as graph-theoretic trees. Using graph theory as a modeling tool allows the vast resources of graphical invariants to be utilized to numerically identify secondary RNA motifs. The domination number of a graph is a graphical invariant that is sensitive to even a slight change in the structure of a tree. The invariants selected in this study are variations of the domination number of a graph. These graphical invariants are partitioned into two classes, and we define two parameters based on each of these classes. These parameters are calculated for all small order trees and a statistical analysis of the resulting data is conducted to determine if the values of these parameters can be utilized to identify which trees of orders seven and eight are RNA-like in structure. Results The statistical analysis shows that the domination based parameters correctly distinguish between the trees that represent native structures and those that are not likely candidates to represent RNA. Some of the trees previously identified as candidate structures are found to be "very" RNA like, while others are not, thereby refining the space of structures likely to be found as representing secondary RNA structure. Conclusion Search algorithms are available that mine nucleotide sequence databases. However, the number of motifs identified can be quite large, making a further search for similar motif computationally difficult. Much of the work in the bioinformatics arena is toward the development of better algorithms to address the computational problem. This work, on the other hand, uses mathematical descriptors to more clearly characterize the RNA motifs and thereby reduce the corresponding search space. These

  14. Matrix factorization-based data fusion for the prediction of lncRNA-disease associations.

    Science.gov (United States)

    Fu, Guangyuan; Wang, Jun; Domeniconi, Carlotta; Yu, Guoxian

    2018-05-01

    Long non-coding RNAs (lncRNAs) play crucial roles in complex disease diagnosis, prognosis, prevention and treatment, but only a small portion of lncRNA-disease associations have been experimentally verified. Various computational models have been proposed to identify lncRNA-disease associations by integrating heterogeneous data sources. However, existing models generally ignore the intrinsic structure of data sources or treat them as equally relevant, while they may not be. To accurately identify lncRNA-disease associations, we propose a Matrix Factorization based LncRNA-Disease Association prediction model (MFLDA in short). MFLDA decomposes data matrices of heterogeneous data sources into low-rank matrices via matrix tri-factorization to explore and exploit their intrinsic and shared structure. MFLDA can select and integrate the data sources by assigning different weights to them. An iterative solution is further introduced to simultaneously optimize the weights and low-rank matrices. Next, MFLDA uses the optimized low-rank matrices to reconstruct the lncRNA-disease association matrix and thus to identify potential associations. In 5-fold cross validation experiments to identify verified lncRNA-disease associations, MFLDA achieves an area under the receiver operating characteristic curve (AUC) of 0.7408, at least 3% higher than those given by state-of-the-art data fusion based computational models. An empirical study on identifying masked lncRNA-disease associations again shows that MFLDA can identify potential associations more accurately than competing models. A case study on identifying lncRNAs associated with breast, lung and stomach cancers show that 38 out of 45 (84%) associations predicted by MFLDA are supported by recent biomedical literature and further proves the capability of MFLDA in identifying novel lncRNA-disease associations. MFLDA is a general data fusion framework, and as such it can be adopted to predict associations between other biological

  15. Site-Specific Incorporation of Functional Components into RNA by an Unnatural Base Pair Transcription System

    Directory of Open Access Journals (Sweden)

    Rie Kawai

    2012-03-01

    Full Text Available Toward the expansion of the genetic alphabet, an unnatural base pair between 7-(2-thienylimidazo[4,5-b]pyridine (Ds and pyrrole-2-carbaldehyde (Pa functions as a third base pair in replication and transcription, and provides a useful tool for the site-specific, enzymatic incorporation of functional components into nucleic acids. We have synthesized several modified-Pa substrates, such as alkylamino-, biotin-, TAMRA-, FAM-, and digoxigenin-linked PaTPs, and examined their transcription by T7 RNA polymerase using Ds-containing DNA templates with various sequences. The Pa substrates modified with relatively small functional groups, such as alkylamino and biotin, were efficiently incorporated into RNA transcripts at the internal positions, except for those less than 10 bases from the 3′-terminus. We found that the efficient incorporation into a position close to the 3′-terminus of a transcript depended on the natural base contexts neighboring the unnatural base, and that pyrimidine-Ds-pyrimidine sequences in templates were generally favorable, relative to purine-Ds-purine sequences. The unnatural base pair transcription system provides a method for the site-specific functionalization of large RNA molecules.

  16. Study design requirements for RNA sequencing-based breast cancer diagnostics.

    Science.gov (United States)

    Mer, Arvind Singh; Klevebring, Daniel; Grönberg, Henrik; Rantalainen, Mattias

    2016-02-01

    Sequencing-based molecular characterization of tumors provides information required for individualized cancer treatment. There are well-defined molecular subtypes of breast cancer that provide improved prognostication compared to routine biomarkers. However, molecular subtyping is not yet implemented in routine breast cancer care. Clinical translation is dependent on subtype prediction models providing high sensitivity and specificity. In this study we evaluate sample size and RNA-sequencing read requirements for breast cancer subtyping to facilitate rational design of translational studies. We applied subsampling to ascertain the effect of training sample size and the number of RNA sequencing reads on classification accuracy of molecular subtype and routine biomarker prediction models (unsupervised and supervised). Subtype classification accuracy improved with increasing sample size up to N = 750 (accuracy = 0.93), although with a modest improvement beyond N = 350 (accuracy = 0.92). Prediction of routine biomarkers achieved accuracy of 0.94 (ER) and 0.92 (Her2) at N = 200. Subtype classification improved with RNA-sequencing library size up to 5 million reads. Development of molecular subtyping models for cancer diagnostics requires well-designed studies. Sample size and the number of RNA sequencing reads directly influence accuracy of molecular subtyping. Results in this study provide key information for rational design of translational studies aiming to bring sequencing-based diagnostics to the clinic.

  17. BRAKER1: Unsupervised RNA-Seq-Based Genome Annotation with GeneMark-ET and AUGUSTUS.

    Science.gov (United States)

    Hoff, Katharina J; Lange, Simone; Lomsadze, Alexandre; Borodovsky, Mark; Stanke, Mario

    2016-03-01

    Gene finding in eukaryotic genomes is notoriously difficult to automate. The task is to design a work flow with a minimal set of tools that would reach state-of-the-art performance across a wide range of species. GeneMark-ET is a gene prediction tool that incorporates RNA-Seq data into unsupervised training and subsequently generates ab initio gene predictions. AUGUSTUS is a gene finder that usually requires supervised training and uses information from RNA-Seq reads in the prediction step. Complementary strengths of GeneMark-ET and AUGUSTUS provided motivation for designing a new combined tool for automatic gene prediction. We present BRAKER1, a pipeline for unsupervised RNA-Seq-based genome annotation that combines the advantages of GeneMark-ET and AUGUSTUS. As input, BRAKER1 requires a genome assembly file and a file in bam-format with spliced alignments of RNA-Seq reads to the genome. First, GeneMark-ET performs iterative training and generates initial gene structures. Second, AUGUSTUS uses predicted genes for training and then integrates RNA-Seq read information into final gene predictions. In our experiments, we observed that BRAKER1 was more accurate than MAKER2 when it is using RNA-Seq as sole source for training and prediction. BRAKER1 does not require pre-trained parameters or a separate expert-prepared training step. BRAKER1 is available for download at http://bioinf.uni-greifswald.de/bioinf/braker/ and http://exon.gatech.edu/GeneMark/ katharina.hoff@uni-greifswald.de or borodovsky@gatech.edu Supplementary data are available at Bioinformatics online. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  18. Taxonomic resolutions based on 18S rRNA genes: a case study of subclass copepoda.

    Directory of Open Access Journals (Sweden)

    Shu Wu

    Full Text Available Biodiversity studies are commonly conducted using 18S rRNA genes. In this study, we compared the inter-species divergence of variable regions (V1-9 within the copepod 18S rRNA gene, and tested their taxonomic resolutions at different taxonomic levels. Our results indicate that the 18S rRNA gene is a good molecular marker for the study of copepod biodiversity, and our conclusions are as follows: 1 18S rRNA genes are highly conserved intra-species (intra-species similarities are close to 100%; and could aid in species-level analyses, but with some limitations; 2 nearly-whole-length sequences and some partial regions (around V2, V4, and V9 of the 18S rRNA gene can be used to discriminate between samples at both the family and order levels (with a success rate of about 80%; 3 compared with other regions, V9 has a higher resolution at the genus level (with an identification success rate of about 80%; and 4 V7 is most divergent in length, and would be a good candidate marker for the phylogenetic study of Acartia species. This study also evaluated the correlation between similarity thresholds and the accuracy of using nuclear 18S rRNA genes for the classification of organisms in the subclass Copepoda. We suggest that sample identification accuracy should be considered when a molecular sequence divergence threshold is used for taxonomic identification, and that the lowest similarity threshold should be determined based on a pre-designated level of acceptable accuracy.

  19. Spectroelectrochemical detection of microRNA-155 based on functional RNA immobilization onto ITO/GNP nanopattern.

    Science.gov (United States)

    Mohammadniaei, Mohsen; Yoon, Jinho; Lee, Taek; Choi, Jeong-Woo

    2018-05-20

    We fabricated a microRNA biosensor using the combination of surface enhanced Raman spectroscopy (SERS) and electrochemical (EC) techniques. For the first time, the weaknesses of each techniques for microRNA detection was compensated by the other ones to give rise to the specific and wide-range detection of miR-155. A single stranded 3' methylene blue (MB) and 5' thiol-modified RNA (MB-ssRNA-SH) was designed to detect the target miR-155 and immobilized onto the gold nanoparticle-modified ITO (ITO/GNP). Upon the invasion of target strand, the double-stranded RNA transformed rapidly to an upright structure resulting in a notable decrease in SERS and redox signals of the MB. For the first time, by combination of SERS and EC techniques in a single platform we extended the dynamic range of both techniques from 10 pM to 450 nM (SERS: 10 pM-5 nM and EC: 5 nM-450 nM). As well, the SERS technique improved the detection limit of the EC method from 100 pM to 100 fM, while the EC method covered single-mismatch detection which was the SERS deficiency. The fabricated single-step biosensor possessing a good capability of miRNA detection in human serum, could be employed throughout the broad ranges of biomedical and bioelectronics applications. Copyright © 2018 Elsevier B.V. All rights reserved.

  20. MITIE: Simultaneous RNA-Seq-based transcript identification and quantification in multiple samples.

    Science.gov (United States)

    Behr, Jonas; Kahles, André; Zhong, Yi; Sreedharan, Vipin T; Drewe, Philipp; Rätsch, Gunnar

    2013-10-15

    High-throughput sequencing of mRNA (RNA-Seq) has led to tremendous improvements in the detection of expressed genes and reconstruction of RNA transcripts. However, the extensive dynamic range of gene expression, technical limitations and biases, as well as the observed complexity of the transcriptional landscape, pose profound computational challenges for transcriptome reconstruction. We present the novel framework MITIE (Mixed Integer Transcript IdEntification) for simultaneous transcript reconstruction and quantification. We define a likelihood function based on the negative binomial distribution, use a regularization approach to select a few transcripts collectively explaining the observed read data and show how to find the optimal solution using Mixed Integer Programming. MITIE can (i) take advantage of known transcripts, (ii) reconstruct and quantify transcripts simultaneously in multiple samples, and (iii) resolve the location of multi-mapping reads. It is designed for genome- and assembly-based transcriptome reconstruction. We present an extensive study based on realistic simulated RNA-Seq data. When compared with state-of-the-art approaches, MITIE proves to be significantly more sensitive and overall more accurate. Moreover, MITIE yields substantial performance gains when used with multiple samples. We applied our system to 38 Drosophila melanogaster modENCODE RNA-Seq libraries and estimated the sensitivity of reconstructing omitted transcript annotations and the specificity with respect to annotated transcripts. Our results corroborate that a well-motivated objective paired with appropriate optimization techniques lead to significant improvements over the state-of-the-art in transcriptome reconstruction. MITIE is implemented in C++ and is available from http://bioweb.me/mitie under the GPL license.

  1. Distant Supervision for Relation Extraction with Ranking-Based Methods

    Directory of Open Access Journals (Sweden)

    Yang Xiang

    2016-05-01

    Full Text Available Relation extraction has benefited from distant supervision in recent years with the development of natural language processing techniques and data explosion. However, distant supervision is still greatly limited by the quality of training data, due to its natural motivation for greatly reducing the heavy cost of data annotation. In this paper, we construct an architecture called MIML-sort (Multi-instance Multi-label Learning with Sorting Strategies, which is built on the famous MIML framework. Based on MIML-sort, we propose three ranking-based methods for sample selection with which we identify relation extractors from a subset of the training data. Experiments are set up on the KBP (Knowledge Base Propagation corpus, one of the benchmark datasets for distant supervision, which is large and noisy. Compared with previous work, the proposed methods produce considerably better results. Furthermore, the three methods together achieve the best F1 on the official testing set, with an optimal enhancement of F1 from 27.3% to 29.98%.

  2. Polyphenolic extracts from cowpea (Vigna unguiculata) protect colonic myofibroblasts (CCD18Co cells) from lipopolysaccharide (LPS)-induced inflammation--modulation of microRNA 126.

    Science.gov (United States)

    Ojwang, Leonnard O; Banerjee, Nivedita; Noratto, Giuliana D; Angel-Morales, Gabriela; Hachibamba, Twambo; Awika, Joseph M; Mertens-Talcott, Susanne U

    2015-01-01

    Cowpea (Vigna unguiculata) is a drought tolerant crop with several agronomic advantages over other legumes. This study evaluated varieties from four major cowpea phenotypes (black, red, light brown and white) containing different phenolic profiles for their anti-inflammatory property on non-malignant colonic myofibroblasts (CCD18Co) cells challenged with an endotoxin (lipopolysaccharide, LPS). Intracellular reactive oxygen species (ROS) assay on the LPS-stimulated cells revealed antioxidative potential of black and red cowpea varieties. Real-time qRT-PCR analysis in LPS-stimulated cells revealed down-regulation of proinflammatory cytokines (IL-8, TNF-α, VCAM-1), transcription factor NF-κB and modulation of microRNA-126 (specific post-transcriptional regulator of VCAM-1) by cowpea polyphenolics. The ability of cowpea polyphenols to modulate miR-126 signaling and its target gene VCAM-1 were studied in LPS-stimulated endothelial cells transfected with a specific inhibitor of miR-126, and treated with 10 mg GAE/L black cowpea extract where the extract in part reversed the effect of the miR-126 inhibitor. This suggests that cowpea may exert their anti-inflammatory activities at least in part through induction of miR-126 that then down-regulate VCAM-1 mRNA and protein expressions. Overall, Cowpea therefore is promising as an anti-inflammatory dietary component.

  3. Covariant Evolutionary Event Analysis for Base Interaction Prediction Using a Relational Database Management System for RNA.

    Science.gov (United States)

    Xu, Weijia; Ozer, Stuart; Gutell, Robin R

    2009-01-01

    With an increasingly large amount of sequences properly aligned, comparative sequence analysis can accurately identify not only common structures formed by standard base pairing but also new types of structural elements and constraints. However, traditional methods are too computationally expensive to perform well on large scale alignment and less effective with the sequences from diversified phylogenetic classifications. We propose a new approach that utilizes coevolutional rates among pairs of nucleotide positions using phylogenetic and evolutionary relationships of the organisms of aligned sequences. With a novel data schema to manage relevant information within a relational database, our method, implemented with a Microsoft SQL Server 2005, showed 90% sensitivity in identifying base pair interactions among 16S ribosomal RNA sequences from Bacteria, at a scale 40 times bigger and 50% better sensitivity than a previous study. The results also indicated covariation signals for a few sets of cross-strand base stacking pairs in secondary structure helices, and other subtle constraints in the RNA structure.

  4. Ionization and fragmentation of DNA-RNA bases: a density functional theory study

    International Nuclear Information System (INIS)

    Sadr-Arani, Leila

    2014-01-01

    Ionizing radiation (IR) cross human tissue, deposit energy and dissipate fragmenting molecules. The resulting fragments may be highlighted by mass spectrometry. Despite the amount of information obtained experimentally by the interpretation of the mass spectrum, experience alone cannot answer all the questions of the mechanism of fragmentation of DNA/RNA bases and a theoretical study is a complement to this information. A theoretical study allows us to know the weakest bonds in the molecule during ionization and thus may help to provide mechanisms of dissociation and produced fragments. The purpose of this work, using the DFT with the PBE functional, is to study the ionization and fragmentation mechanisms of DNA/RNA bases (Uracil, Cytosine, Adenine and Guanine) and to identify the cations corresponding to each peak in mass spectra. For all RNA bases, the retro Diels-Alder reaction (elimination of HNCO or NCO*) is a major route for dissociating, with the exception of adenine for which there is no atom oxygen in its structure. Loss of NH 3 (NH 2 *) molecule is another common way to all bases that contain amine group. The possibility of the loss of hydrogen from the cations is also investigated, as well as the dissociation of dehydrogenated cations and protonated uracil. This work shows the interest of providing DFT calculation in the interpretation of mass spectra of DNA bases. (author)

  5. lncRNATargets: A platform for lncRNA target prediction based on nucleic acid thermodynamics.

    Science.gov (United States)

    Hu, Ruifeng; Sun, Xiaobo

    2016-08-01

    Many studies have supported that long noncoding RNAs (lncRNAs) perform various functions in various critical biological processes. Advanced experimental and computational technologies allow access to more information on lncRNAs. Determining the functions and action mechanisms of these RNAs on a large scale is urgently needed. We provided lncRNATargets, which is a web-based platform for lncRNA target prediction based on nucleic acid thermodynamics. The nearest-neighbor (NN) model was used to calculate binging-free energy. The main principle of NN model for nucleic acid assumes that identity and orientation of neighbor base pairs determine stability of a given base pair. lncRNATargets features the following options: setting of a specific temperature that allow use not only for human but also for other animals or plants; processing all lncRNAs in high throughput without RNA size limitation that is superior to any other existing tool; and web-based, user-friendly interface, and colored result displays that allow easy access for nonskilled computer operators and provide better understanding of results. This technique could provide accurate calculation on the binding-free energy of lncRNA-target dimers to predict if these structures are well targeted together. lncRNATargets provides high accuracy calculations, and this user-friendly program is available for free at http://www.herbbol.org:8001/lrt/ .

  6. The use of 125iodine-labeled RNA for detection of the RNA binding to ribosomes

    International Nuclear Information System (INIS)

    Mori, Tomohiko; Fukuda, Mitsuru

    1975-01-01

    The in vitro labeling of RNA with radioactive iodine is the efficient method to obtain the RNA with high specific activity. The present paper reports on the application of this technique to the production of iodine-labeled RNA for use in the experiment of binding RNA to ribosomes. Tobacco mosaic virus (TMV) RNA was used as natural mRNA, and E. coli S-30 preparation was used as a source of ribosomes. The TMV-RNA was prepared by bentonite-phenol extraction from TMV, and the method used for the iodation of RNA was based on the procedure described by Getz et al. The iodine-labeled RNA was incubated in a cell-free protein synthesizing system (S-30) prepared from E. coli K-12. After the incubation, the reaction mixture was layered onto sucrose gradient, centrifuged, and fractionated into 18 fractions. Optical density at 260 nm was measured, and radioactivity was counted, for each fraction. The binding of mRNA to ribosomes occurred even at 0 deg C, and the occurrence of the nonspecific binding was also shown. Consequently, the specific binding, i.e. the formation of the initiation complex being involved in amino acid incorporation, may be estimated by subtracting the radioactivity associated with monosomes in the presence of both rRNA and ATA from that in the presence of rRNA only. It was shown that the iodine-labeled RNA can be used for the studies of binding RNA to ribosomes. (Kako, I.)

  7. Characters Feature Extraction Based on Neat Oracle Bone Rubbings

    OpenAIRE

    Lei Guo

    2013-01-01

    In order to recognize characters on the neat oracle bone rubbings, a new mesh point feature extraction algorithm was put forward in this paper by researching and improving of the existing coarse mesh feature extraction algorithm and the point feature extraction algorithm. Some improvements of this algorithm were as followings: point feature was introduced into the coarse mesh feature, the absolute address was converted to relative address, and point features have been changed grid and positio...

  8. The Research of Disease Spots Extraction Based on Evolutionary Algorithm

    Directory of Open Access Journals (Sweden)

    Kangshun Li

    2017-01-01

    Full Text Available According to the characteristics of maize disease spot performance in the image, this paper designs two-histogram segmentation method based on evolutionary algorithm, which combined with the analysis of image of maize diseases and insect pests, with full consideration of color and texture characteristic of the lesion of pests and diseases, the chroma and gray image, composed of two tuples to build a two-dimensional histogram, solves the problem of one-dimensional histograms that cannot be clearly divided into target and background bimodal distribution and improved the traditional two-dimensional histogram application in pest damage lesion extraction. The chromosome coding suitable for the characteristics of lesion image is designed based on second segmentation of the genetic algorithm Otsu. Determining initial population with analysis results of lesion image, parallel selection, optimal preservation strategy, and adaptive mutation operator are used to improve the search efficiency. Finally, by setting the fluctuation threshold, we continue to search for the best threshold in the range of fluctuations for implementation of global search and local search.

  9. Generating Unstable Resonances for Extraction Schemes Based on Transverse Splitting

    CERN Document Server

    Giovannozzi, M; Turchetti, G

    2009-01-01

    A few years ago, a novel multi-turn extraction scheme was proposed, based on particle trapping inside stable resonances. Numerical simulations and experimental tests have confirmed the feasibility of such a scheme for low order resonances. While the third-order resonance is generically unstable and those higher than fourth-order are generically stable, the fourth-order resonance can be either stable or unstable depending on the specifics of the system under consideration. By means of the Normal Form a general approach to control the stability of the fourth-order resonance has been derived. This approach is based on the control of the amplitude detuning and the general form for a lattice with an arbitrary number of sextupole and octupole families is derived in this paper. Numerical simulations have confirmed the analytical results and have shown that, when crossing the unstable fourth-order resonance, the region around the centre of the phase space is depleted and particles are trapped in only the four stable ...

  10. Iris recognition based on key image feature extraction.

    Science.gov (United States)

    Ren, X; Tian, Q; Zhang, J; Wu, S; Zeng, Y

    2008-01-01

    In iris recognition, feature extraction can be influenced by factors such as illumination and contrast, and thus the features extracted may be unreliable, which can cause a high rate of false results in iris pattern recognition. In order to obtain stable features, an algorithm was proposed in this paper to extract key features of a pattern from multiple images. The proposed algorithm built an iris feature template by extracting key features and performed iris identity enrolment. Simulation results showed that the selected key features have high recognition accuracy on the CASIA Iris Set, where both contrast and illumination variance exist.

  11. Fine-grained parallelism accelerating for RNA secondary structure prediction with pseudoknots based on FPGA.

    Science.gov (United States)

    Xia, Fei; Jin, Guoqing

    2014-06-01

    PKNOTS is a most famous benchmark program and has been widely used to predict RNA secondary structure including pseudoknots. It adopts the standard four-dimensional (4D) dynamic programming (DP) method and is the basis of many variants and improved algorithms. Unfortunately, the O(N(6)) computing requirements and complicated data dependency greatly limits the usefulness of PKNOTS package with the explosion in gene database size. In this paper, we present a fine-grained parallel PKNOTS package and prototype system for accelerating RNA folding application based on FPGA chip. We adopted a series of storage optimization strategies to resolve the "Memory Wall" problem. We aggressively exploit parallel computing strategies to improve computational efficiency. We also propose several methods that collectively reduce the storage requirements for FPGA on-chip memory. To the best of our knowledge, our design is the first FPGA implementation for accelerating 4D DP problem for RNA folding application including pseudoknots. The experimental results show a factor of more than 50x average speedup over the PKNOTS-1.08 software running on a PC platform with Intel Core2 Q9400 Quad CPU for input RNA sequences. However, the power consumption of our FPGA accelerator is only about 50% of the general-purpose micro-processors.

  12. Rank-Based miRNA Signatures for Early Cancer Detection

    Directory of Open Access Journals (Sweden)

    Mario Lauria

    2014-01-01

    Full Text Available We describe a new signature definition and analysis method to be used as biomarker for early cancer detection. Our new approach is based on the construction of a reference map of transcriptional signatures of both healthy and cancer affected individuals using circulating miRNA from a large number of subjects. Once such a map is available, the diagnosis for a new patient can be performed by observing the relative position on the map of his/her transcriptional signature. To demonstrate its efficacy for this specific application we report the results of the application of our method to published datasets of circulating miRNA, and we quantify its performance compared to current state-of-the-art methods. A number of additional features make this method an ideal candidate for large-scale use, for example, as a mass screening tool for early cancer detection or for at-home diagnostics. Specifically, our method is minimally invasive (because it works well with circulating miRNA, it is robust with respect to lab-to-lab protocol variability and batch effects (it requires that only the relative ranking of expression value of miRNA in a profile be accurate not their absolute values, and it is scalable to a large number of subjects. Finally we discuss the need for HPC capability in a widespread application of our or similar methods.

  13. A DNA sequence obtained by replacement of the dopamine RNA aptamer bases is not an aptamer.

    Science.gov (United States)

    Álvarez-Martos, Isabel; Ferapontova, Elena E

    2017-08-05

    A unique specificity of the aptamer-ligand biorecognition and binding facilitates bioanalysis and biosensor development, contributing to discrimination of structurally related molecules, such as dopamine and other catecholamine neurotransmitters. The aptamer sequence capable of specific binding of dopamine is a 57 nucleotides long RNA sequence reported in 1997 (Biochemistry, 1997, 36, 9726). Later, it was suggested that the DNA homologue of the RNA aptamer retains the specificity of dopamine binding (Biochem. Biophys. Res. Commun., 2009, 388, 732). Here, we show that the DNA sequence obtained by the replacement of the RNA aptamer bases for their DNA analogues is not able of specific biorecognition of dopamine, in contrast to the original RNA aptamer sequence. This DNA sequence binds dopamine and structurally related catecholamine neurotransmitters non-specifically, as any DNA sequence, and, thus, is not an aptamer and cannot be used neither for in vivo nor in situ analysis of dopamine in the presence of structurally related neurotransmitters. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. A Sensitive and Specific PCR Based Method for Identification of Cryptosporidium Sp. Using New Primers from 18S Ribosomal RNA

    Directory of Open Access Journals (Sweden)

    M Heydarnezhadi

    2011-09-01

    Full Text Available Background: The main goal of the present study was to develop a new sensitive and specific PCR based method for Identification of Cryptosporidium sp. using novel primers from 18S ribosomal RNA. Cryptosporidi­osis in high-risk host groups particularly in neonates and immuno-compromised individuals may result in death. To the best of our knowledge this is the first study regarding develop a new PCR based method to diagnose the cryptosporidiosis in Iran.Methods: A total of 850 human fecal samples from patients clinically suspected to cryptosporidiosis and 100 healthy and diarrheic cattle stool specimens were collected. The simplified formol-ether concentration method was carried out for all samples. They were then examined microscopically by modified Ziehl-Neel­sen staining method. Total DNA was extracted by QIA amp DNA stool mini kit was carried out by using designed prim­ers.Results: Twenty nine cases of cryptosporidiosis infection in human and 30 samples from cattle microscopi­cally were posi­tive. The described primary and nested PCR method could detect all Cryptospori­dium positive samples from human and cattle. Regards to suspected negative samples in pri­mary PCR examination, the Nested PCR could ap­prove two more positive results. Furthermore, Nested PCR analysis was able to detect one more case which was nega­tive in both microscopically examination and primary PCR. Specificity of the test was 100%. Sensitivity of Nested PCR in comparison to our gold standard; microscopy after Ridley concentration modified ziehl-Neelsen, was 100 %.Conclusion: Our developed PCR based method by using new primers devised from 18S ribosomal RNA revealed the ability for identification of the Cryptosporidium species such as C. parvum and C. huminis with high specificity and sensitivity.

  15. Associations between HIV-RNA-based indicators and virological and clinical outcomes

    DEFF Research Database (Denmark)

    Laut, Kamilla G; Shepherd, Leah C; Pedersen, Court

    2016-01-01

    OBJECTIVES: To evaluate and compare the performance of six HIV-RNA-based quality of care indicators for predicting short-term and long-term outcomes. DESIGN: Multinational cohort study. METHODS: We included EuroSIDA patients on antiretroviral therapy (ART) with at least three viral load measureme......OBJECTIVES: To evaluate and compare the performance of six HIV-RNA-based quality of care indicators for predicting short-term and long-term outcomes. DESIGN: Multinational cohort study. METHODS: We included EuroSIDA patients on antiretroviral therapy (ART) with at least three viral load...... measurements after baseline (the latest of 01/01/2001 or entry into EuroSIDA). Using multivariate Poisson regression, we modelled the association between short-term (resistance, triple-class failure) and long-term (all-cause mortality, any AIDS/non-AIDS clinical event) outcomes and the indicators: viraemia...

  16. Averrhoa carambola free phenolic extract ameliorates nonalcoholic hepatic steatosis by modulating mircoRNA-34a, mircoRNA-33 and AMPK pathways in leptin receptor-deficient db/db mice.

    Science.gov (United States)

    Pang, Daorui; You, Lijun; Zhou, Lin; Li, Tong; Zheng, Bisheng; Liu, Rui Hai

    2017-12-13

    The objective of the present study is to investigate the hepatic steatosis relieving effect of Averrhoa carambola free phenolic extract (ACF) on leptin receptor-deficient (db/db) mice and elucidate the modulation hepatic lipogenesis mechanisms. The serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) assays, accompanying hematoxylin and eosin (H&E) staining, were applied to identify the alleviation of liver histopathological changes. Serum and hepatic lipid assays, combined with oil red O staining, were used to investigate the amelioration of lipid accumulation. Further assessments by quantitative real-time PCR and western blot assays were used to elucidate the suppression of the fatty acid and triglyceride (TG) synthesis mechanisms underlying ACF protection. These results indicated that ACF treatment significantly reduced the liver TG of db/db mice (p < 0.05). The mechanisms are partly through phosphorylation of AMPK α and down-regulation of SREBP-1c expression, and further down-regulation of FAS and SCD1 (p < 0.05). In addition, the expression levels of mircoRNA-34a and mircoRNA-33, which modulate this signaling pathway, were significantly down-regulated by ACF treatment (p < 0.05). Collectively, these results revealed that ACF exhibited a potent hepatic steatosis relieving effect partly by inhibiting the signal transduction of hepatic lipogenesis.

  17. Polyethylene glycol and polyvinylpirrolidone effect on bacterial rRNA extraction and hybridization from cells exposed to tannins Efeito de polietilenoglicol e polivinilpirrolidona na extração e hibridização de rRNA bacteriano de células expostas a taninos

    OpenAIRE

    Pedro Braga Arcuri; Michael Larry Thonney; Peter Schofield; Alice Nelson Pell

    2003-01-01

    In order to detect fluctuations in ruminal microbial populations due to forage tannins using 16S ribosomal RNA (rRNA) probes, recovery of intact rRNA is required. The objective of this work was to evaluate the effect of polyethylene glycol (PEG) and polyvinylpirrolidone (PVP) on extraction of bacterial rRNA, in the presence of tannins from tropical legume forages and other sources, that hybridize with oligonucleotide probes. Ruminococcus albus 8 cells were exposed to 8 g/L tannic acid or 1 g/...

  18. Dissection of functional lncRNAs in Alzheimer's disease by construction and analysis of lncRNA-mRNA networks based on competitive endogenous RNAs.

    Science.gov (United States)

    Wang, Lian-Kun; Chen, Xiao-Feng; He, Dan-Dan; Li, You; Fu, Jin

    2017-04-08

    Alzheimer's disease (AD) is a neurodegenerative disorder that is the most common cause of dementia in the elderly, and intracellular neurofibrillary tangles (NFTs) are one of the pathological features of AD. Recent studies have suggested long noncoding RNAs (lncRNAs) play important roles in AD. Competing endogenous RNAs (ceRNAs) is a mechanism that has recently been proposed, in which lncRNAs compete for common miRNA-binding sites with mRNAs. However, the roles of lncRNAs and ceRNA in AD NFTs is limited. In this study, we constructed a global triple network based on ceRNA theory, then an AD NFT lncRNA-mRNA network (NFTLMN) was generated. By analyzing the NFTLMN, three lncRNAs (AP000265.1, KB-1460A1.5 and RP11-145M9.4), which are highly related with AD NFTs were identified. To further explore the cross-talk between mRNAs and lncRNAs, a clustering module analysis was performed on the NFTLMN and two AD NFT related modules were identified. Our study provides a better understanding of the molecular basis of AD NFTs and may offer novel treatment strategies for AD. Copyright © 2016. Published by Elsevier Inc.

  19. Extraction of alkaloids for NMR-based profiling

    DEFF Research Database (Denmark)

    Yilmaz, Ali; Nyberg, Nils; Jaroszewski, Jerzy W.

    2012-01-01

    A museum collection of Cinchona cortex samples (n = 117), from the period 1850–1950, were extracted with a mixture of chloroform-d1, methanol-d4, water-d2, and perchloric acid in the ratios 5:5:1:1. The extracts were directly analyzed using 1H NMR spectroscopy (600 MHz) and the spectra evaluated...

  20. Ionic Liquid-Based Ultrasonic/Microwave-Assisted Extraction of ...

    African Journals Online (AJOL)

    Conclusion: Compared with traditional methods, IL-UMAE method uses Ionic liquid-solvent which greatly shortens the extraction time. IL-UMAE as a simple, effective and environmentally friendly approach shows a broad prospect for active ingredient extraction. Keywords: Dioscorea zingiberensis Steroidal saponins, Ionic ...

  1. The development of simple field based procedures for extraction of ...

    African Journals Online (AJOL)

    Field and laboratory experiments were conducted to develop procedures for extracting volatiles from the vine of Adenia cissampeloides which could effect the highest yield at the lowest extraction costs and also could be produced at the cottage industry level. The participatory rural appraisal technique was used to ensure ...

  2. Sample-based XPath Ranking for Web Information Extraction

    NARCIS (Netherlands)

    Jundt, Oliver; van Keulen, Maurice

    Web information extraction typically relies on a wrapper, i.e., program code or a configuration that specifies how to extract some information from web pages at a specific website. Manually creating and maintaining wrappers is a cumbersome and error-prone task. It may even be prohibitive as some

  3. BEaST: brain extraction based on nonlocal segmentation technique

    NARCIS (Netherlands)

    Eskildsen, Simon F.; Coupé, Pierrick; Fonov, Vladimir; Manjón, José V.; Leung, Kelvin K.; Guizard, Nicolas; Wassef, Shafik N.; Østergaard, Lasse Riis; Collins, D. Louis; Saradha, A.; Abdi, Hervé; Abdulkadir, Ahmed; Acharya, Deepa; Achuthan, Anusha; Adluru, Nagesh; Aghajanian, Jania; Agrusti, Antonella; Agyemang, Alex; Ahdidan, Jamila; Ahmad, Duaa; Ahmed, Shiek; Aisen, Paul; Akhondi-Asl, Alireza; Aksu, Yaman; Alberca, Roman; Alcauter, Sarael; Alexander, Daniel; Alin, Aylin; Almeida, Fabio; Alvarez-Linera, Juan; Amlien, Inge; Anand, Shyam; Anderson, Dallas; Ang, Amma; Angersbach, Steve; Ansarian, Reza; Aoyama, Eiji; Appannah, Arti; Arfanakis, Konstantinos; Armor, Tom; Arrighi, Michael; Arumughababu, S. Vethanayaki; Arunagiri, Vidhya; Ashe-McNalley, Cody; Ashford, Wes; Le Page, Aurelie; Avants, Brian; Chen, Wei; Richard, Edo; Schmand, Ben

    2012-01-01

    Brain extraction is an important step in the analysis of brain images. The variability in brain morphology and the difference in intensity characteristics due to imaging sequences make the development of a general purpose brain extraction algorithm challenging. To address this issue, we propose a

  4. Evolutionary patterns of RNA-based duplication in non-mammalian chordates.

    Directory of Open Access Journals (Sweden)

    Ming Chen

    Full Text Available The role of RNA-based duplication, or retroposition, in the evolution of new gene functions in mammals, plants, and Drosophila has been widely reported. However, little is known about RNA-based duplication in non-mammalian chordates. In this study, we screened ten non-mammalian chordate genomes for retrocopies and investigated their evolutionary patterns. We identified numerous retrocopies in these species. Examination of the age distribution of these retrocopies revealed no burst of young retrocopies in ancient chordate species. Upon comparing these non-mammalian chordate species to the mammalian species, we observed that a larger fraction of the non-mammalian retrocopies was under strong evolutionary constraints than mammalian retrocopies are, as evidenced by signals of purifying selection and expression profiles. For the Western clawed frog, Medaka, and Sea squirt, many retrogenes have evolved gonad and brain expression patterns, similar to what was observed in human. Testing of retrogene movement in the Medaka genome, where the nascent sex chrosomes have been well assembled, did not reveal any significant gene movement. Taken together, our analyses demonstrate that RNA-based duplication generates many functional genes and can make a significant contribution to the evolution of non-mammalian genomes.

  5. Classification of Textures Using Filter Based Local Feature Extraction

    Directory of Open Access Journals (Sweden)

    Bocekci Veysel Gokhan

    2016-01-01

    Full Text Available In this work local features are used in feature extraction process in image processing for textures. The local binary pattern feature extraction method from textures are introduced. Filtering is also used during the feature extraction process for getting discriminative features. To show the effectiveness of the algorithm before the extraction process, three different noise are added to both train and test images. Wiener filter and median filter are used to remove the noise from images. We evaluate the performance of the method with Naïve Bayesian classifier. We conduct the comparative analysis on benchmark dataset with different filtering and size. Our experiments demonstrate that feature extraction process combine with filtering give promising results on noisy images.

  6. Fully Convolutional Network Based Shadow Extraction from GF-2 Imagery

    Science.gov (United States)

    Li, Z.; Cai, G.; Ren, H.

    2018-04-01

    There are many shadows on the high spatial resolution satellite images, especially in the urban areas. Although shadows on imagery severely affect the information extraction of land cover or land use, they provide auxiliary information for building extraction which is hard to achieve a satisfactory accuracy through image classification itself. This paper focused on the method of building shadow extraction by designing a fully convolutional network and training samples collected from GF-2 satellite imagery in the urban region of Changchun city. By means of spatial filtering and calculation of adjacent relationship along the sunlight direction, the small patches from vegetation or bridges have been eliminated from the preliminary extracted shadows. Finally, the building shadows were separated. The extracted building shadow information from the proposed method in this paper was compared with the results from the traditional object-oriented supervised classification algorihtms. It showed that the deep learning network approach can improve the accuracy to a large extent.

  7. Text feature extraction based on deep learning: a review.

    Science.gov (United States)

    Liang, Hong; Sun, Xiao; Sun, Yunlei; Gao, Yuan

    2017-01-01

    Selection of text feature item is a basic and important matter for text mining and information retrieval. Traditional methods of feature extraction require handcrafted features. To hand-design, an effective feature is a lengthy process, but aiming at new applications, deep learning enables to acquire new effective feature representation from training data. As a new feature extraction method, deep learning has made achievements in text mining. The major difference between deep learning and conventional methods is that deep learning automatically learns features from big data, instead of adopting handcrafted features, which mainly depends on priori knowledge of designers and is highly impossible to take the advantage of big data. Deep learning can automatically learn feature representation from big data, including millions of parameters. This thesis outlines the common methods used in text feature extraction first, and then expands frequently used deep learning methods in text feature extraction and its applications, and forecasts the application of deep learning in feature extraction.

  8. GIMDA: Graphlet interaction-based MiRNA-disease association prediction.

    Science.gov (United States)

    Chen, Xing; Guan, Na-Na; Li, Jian-Qiang; Yan, Gui-Ying

    2018-03-01

    MicroRNAs (miRNAs) have been confirmed to be closely related to various human complex diseases by many experimental studies. It is necessary and valuable to develop powerful and effective computational models to predict potential associations between miRNAs and diseases. In this work, we presented a prediction model of Graphlet Interaction for MiRNA-Disease Association prediction (GIMDA) by integrating the disease semantic similarity, miRNA functional similarity, Gaussian interaction profile kernel similarity and the experimentally confirmed miRNA-disease associations. The related score of a miRNA to a disease was calculated by measuring the graphlet interactions between two miRNAs or two diseases. The novelty of GIMDA lies in that we used graphlet interaction to analyse the complex relationships between two nodes in a graph. The AUCs of GIMDA in global and local leave-one-out cross-validation (LOOCV) turned out to be 0.9006 and 0.8455, respectively. The average result of five-fold cross-validation reached to 0.8927 ± 0.0012. In case study for colon neoplasms, kidney neoplasms and prostate neoplasms based on the database of HMDD V2.0, 45, 45, 41 of the top 50 potential miRNAs predicted by GIMDA were validated by dbDEMC and miR2Disease. Additionally, in the case study of new diseases without any known associated miRNAs and the case study of predicting potential miRNA-disease associations using HMDD V1.0, there were also high percentages of top 50 miRNAs verified by the experimental literatures. © 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

  9. Artificial terraced field extraction based on high resolution DEMs

    Science.gov (United States)

    Na, Jiaming; Yang, Xin; Xiong, Liyang; Tang, Guoan

    2017-04-01

    With the increase of human activities, artificial landforms become one of the main terrain features with special geographical and hydrological value. Terraced field, as the most important artificial landscapes of the loess plateau, plays an important role in conserving soil and water. With the development of digital terrain analysis (DTA), there is a current and future need in developing a robust, repeatable and cost-effective research methodology for terraced fields. In this paper, a novel method using bidirectional DEM shaded relief is proposed for terraced field identification based on high resolution DEM, taking Zhifanggou watershed, Shannxi province as the study area. Firstly, 1m DEM is obtained by low altitude aerial photogrammetry using Unmanned Aerial Vehicle (UAV), and 0.1m DOM is also obtained as the test data. Then, the positive and negative terrain segmentation is done to acquire the area of terraced field. Finally, a bidirectional DEM shaded relief is simulated to extract the ridges of each terraced field stages. The method in this paper can get not only polygon feature of the terraced field areas but also line feature of terraced field ridges. The accuracy is 89.7% compared with the artificial interpretation result from DOM. And additional experiment shows that this method has a strong robustness as well as high accuracy.

  10. Morphological operation based dense houses extraction from DSM

    Science.gov (United States)

    Li, Y.; Zhu, L.; Tachibana, K.; Shimamura, H.

    2014-08-01

    This paper presents a method of reshaping and extraction of markers and masks of the dense houses from the DSM based on mathematical morphology (MM). Houses in a digital surface model (DSM) are almost joined together in high-density housing areas, and most segmentation methods cannot completely separate them. We propose to label the markers of the buildings firstly and segment them into masks by watershed then. To avoid detecting more than one marker for a house or no marker at all due to its higher neighbour, the DSM is morphologically reshaped. It is carried out by a MM operation using the certain disk shape SE of the similar size to the houses. The sizes of the houses need to be estimated before reshaping. A granulometry generated by opening-by-reconstruction to the NDSM is proposed to detect the scales of the off-terrain objects. It is a histogram of the global volume of the top hats of the convex objects in the continuous scales. The obvious step change in the profile means that there are many objects of similar sizes occur at this scale. In reshaping procedure, the slices of the object are derived by morphological filtering at the detected continuous scales and reconstructed in pile as the dome. The markers are detected on the basis of the domes.

  11. Historical feature pattern extraction based network attack situation sensing algorithm.

    Science.gov (United States)

    Zeng, Yong; Liu, Dacheng; Lei, Zhou

    2014-01-01

    The situation sequence contains a series of complicated and multivariate random trends, which are very sudden, uncertain, and difficult to recognize and describe its principle by traditional algorithms. To solve the above questions, estimating parameters of super long situation sequence is essential, but very difficult, so this paper proposes a situation prediction method based on historical feature pattern extraction (HFPE). First, HFPE algorithm seeks similar indications from the history situation sequence recorded and weighs the link intensity between occurred indication and subsequent effect. Then it calculates the probability that a certain effect reappears according to the current indication and makes a prediction after weighting. Meanwhile, HFPE method gives an evolution algorithm to derive the prediction deviation from the views of pattern and accuracy. This algorithm can continuously promote the adaptability of HFPE through gradual fine-tuning. The method preserves the rules in sequence at its best, does not need data preprocessing, and can track and adapt to the variation of situation sequence continuously.

  12. Historical Feature Pattern Extraction Based Network Attack Situation Sensing Algorithm

    Directory of Open Access Journals (Sweden)

    Yong Zeng

    2014-01-01

    Full Text Available The situation sequence contains a series of complicated and multivariate random trends, which are very sudden, uncertain, and difficult to recognize and describe its principle by traditional algorithms. To solve the above questions, estimating parameters of super long situation sequence is essential, but very difficult, so this paper proposes a situation prediction method based on historical feature pattern extraction (HFPE. First, HFPE algorithm seeks similar indications from the history situation sequence recorded and weighs the link intensity between occurred indication and subsequent effect. Then it calculates the probability that a certain effect reappears according to the current indication and makes a prediction after weighting. Meanwhile, HFPE method gives an evolution algorithm to derive the prediction deviation from the views of pattern and accuracy. This algorithm can continuously promote the adaptability of HFPE through gradual fine-tuning. The method preserves the rules in sequence at its best, does not need data preprocessing, and can track and adapt to the variation of situation sequence continuously.

  13. mRNA-based vaccines synergize with radiation therapy to eradicate established tumors

    International Nuclear Information System (INIS)

    Fotin-Mleczek, Mariola; Zanzinger, Kai; Heidenreich, Regina; Lorenz, Christina; Kowalczyk, Aleksandra; Kallen, Karl-Josef; Huber, Stephan M

    2014-01-01

    The eradication of large, established tumors by active immunotherapy is a major challenge because of the numerous cancer evasion mechanisms that exist. This study aimed to establish a novel combination therapy consisting of messenger RNA (mRNA)-based cancer vaccines and radiation, which would facilitate the effective treatment of established tumors with aggressive growth kinetics. The combination of a tumor-specific mRNA-based vaccination with radiation was tested in two syngeneic tumor models, a highly immunogenic E.G7-OVA and a low immunogenic Lewis lung cancer (LLC). The molecular mechanism induced by the combination therapy was evaluated via gene expression arrays as well as flow cytometry analyses of tumor infiltrating cells. In both tumor models we demonstrated that a combination of mRNA-based immunotherapy with radiation results in a strong synergistic anti-tumor effect. This was manifested as either complete tumor eradication or delay in tumor growth. Gene expression analysis of mouse tumors revealed a variety of substantial changes at the tumor site following radiation. Genes associated with antigen presentation, infiltration of immune cells, adhesion, and activation of the innate immune system were upregulated. A combination of radiation and immunotherapy induced significant downregulation of tumor associated factors and upregulation of tumor suppressors. Moreover, combination therapy significantly increased CD4 + , CD8 + and NKT cell infiltration of mouse tumors. Our data provide a scientific rationale for combining immunotherapy with radiation and provide a basis for the development of more potent anti-cancer therapies. The online version of this article (doi:10.1186/1748-717X-9-180) contains supplementary material, which is available to authorized users

  14. Fluorescence-based codetection with protein markers reveals distinct cellular compartments for altered MicroRNA expression in solid tumors

    DEFF Research Database (Denmark)

    Sempere, Lorenzo F; Preis, Meir; Yezefski, Todd

    2010-01-01

    of altered miRNA expression in solid tumors, we developed a sensitive fluorescence-based in situ hybridization (ISH) method to visualize miRNA accumulation within individual cells in formalin-fixed, paraffin-embedded tissue specimens. This ISH method was implemented to be compatible with routine clinical...

  15. An improved approach for flow-based cloud point extraction.

    Science.gov (United States)

    Frizzarin, Rejane M; Rocha, Fábio R P

    2014-04-11

    Novel strategies are proposed to circumvent the main drawbacks of flow-based cloud point extraction (CPE). The surfactant-rich phase (SRP) was directly retained into the optical path of the spectrophotometric cell, thus avoiding its dilution previously to the measurement and yielding higher sensitivity. Solenoid micro-pumps were exploited to improve mixing by the pulsed flow and also to modulate the flow-rate for retention and removal of the SRP, thus avoiding the elution step, often carried out with organic solvents. The heat released and the increase of the salt concentration provided by an on-line neutralization reaction were exploited to induce the cloud point without an external heating device. These innovations were demonstrated by the spectrophotometric determination of iron, yielding a linear response from 10 to 200 μg L(-1) with a coefficient of variation of 2.3% (n=7). Detection limit and sampling rate were estimated at 5 μg L(-1) (95% confidence level) and 26 samples per hour, respectively. The enrichment factor was 8.9 and the procedure consumed only 6 μg of TAN and 390 μg of Triton X-114 per determination. At the 95% confidence level, the results obtained for freshwater samples agreed with the reference procedure and those obtained for digests of bovine muscle, rice flour, brown bread and tort lobster agreed with the certified reference values. The proposed procedure thus shows advantages in relation to previously proposed approaches for flow-based CPE, being a fast and environmental friendly alternative for on-line separation and pre-concentration. Copyright © 2014 Elsevier B.V. All rights reserved.

  16. Linking phylogenetic identities of bacteria to starch fermentation in an in vitro model of the large intestine by RNA-based stable isotope probing.

    Science.gov (United States)

    Kovatcheva-Datchary, Petia; Egert, Markus; Maathuis, Annet; Rajilić-Stojanović, Mirjana; de Graaf, Albert A; Smidt, Hauke; de Vos, Willem M; Venema, Koen

    2009-04-01

    Carbohydrates, including starches, are an important energy source for humans, and are known for their interactions with the microbiota in the digestive tract. Largely, those interactions are thought to promote human health. Using 16S ribosomal RNA (rRNA)-based stable isotope probing (SIP), we identified starch-fermenting bacteria under human colon-like conditions. To the microbiota of the TIM-2 in vitro model of the human colon 7.4 g l(-1) of [U-(13)C]-starch was added. RNA extracted from lumen samples after 0 (control), 2, 4 and 8 h was subjected to density-gradient ultracentrifugation. Terminal-restriction fragment length polymorphism (T-RFLP) fingerprinting and phylogenetic analyses of the labelled and unlabelled 16S rRNA suggested populations related to Ruminococcus bromii, Prevotella spp. and Eubacterium rectale to be involved in starch metabolism. Additionally, 16S rRNA related to that of Bifidobacterium adolescentis was abundant in all analysed fractions. While this might be due to the enrichment of high-GC RNA in high-density fractions, it could also indicate an active role in starch fermentation. Comparison of the T-RFLP fingerprints of experiments performed with labelled and unlabelled starch revealed Ruminococcus bromii as the primary degrader in starch fermentation in the studied model, as it was found to solely predominate in the labelled fractions. LC-MS analyses of the lumen and dialysate samples showed that, for both experiments, starch fermentation primarily yielded acetate, butyrate and propionate. Integration of molecular and metabolite data suggests metabolic cross-feeding in the system, where populations related to Ruminococcus bromii are the primary starch degrader, while those related to Prevotella spp., Bifidobacterium adolescentis and Eubacterium rectale might be further involved in the trophic chain.

  17. 34A, miRNA-944, miRNA-101 and miRNA-218 in cervical cancer

    African Journals Online (AJOL)

    RNAs (21 - 24 nucleotides in length) that are critical for many important processes such as development, ... RNA extraction and reverse transcription. Total RNA was extracted from each of the experimental groups using ... used as an endogenous control to normalize the expression of miRNA-143, miRNA-34A, miRNA-.

  18. Social network extraction based on Web: 1. Related superficial methods

    Science.gov (United States)

    Khairuddin Matyuso Nasution, Mahyuddin

    2018-01-01

    Often the nature of something affects methods to resolve the related issues about it. Likewise, methods to extract social networks from the Web, but involve the structured data types differently. This paper reveals several methods of social network extraction from the same sources that is Web: the basic superficial method, the underlying superficial method, the description superficial method, and the related superficial methods. In complexity we derive the inequalities between methods and so are their computations. In this case, we find that different results from the same tools make the difference from the more complex to the simpler: Extraction of social network by involving co-occurrence is more complex than using occurrences.

  19. Whole Blood mRNA Expression-Based Prognosis of Metastatic Renal Cell Carcinoma.

    Science.gov (United States)

    Giridhar, Karthik V; Sosa, Carlos P; Hillman, David W; Sanhueza, Cristobal; Dalpiaz, Candace L; Costello, Brian A; Quevedo, Fernando J; Pitot, Henry C; Dronca, Roxana S; Ertz, Donna; Cheville, John C; Donkena, Krishna Vanaja; Kohli, Manish

    2017-11-03

    The Memorial Sloan Kettering Cancer Center (MSKCC) prognostic score is based on clinical parameters. We analyzed whole blood mRNA expression in metastatic clear cell renal cell carcinoma (mCCRCC) patients and compared it to the MSKCC score for predicting overall survival. In a discovery set of 19 patients with mRCC, we performed whole transcriptome RNA sequencing and selected eighteen candidate genes for further evaluation based on associations with overall survival and statistical significance. In an independent validation of set of 47 patients with mCCRCC, transcript expression of the 18 candidate genes were quantified using a customized NanoString probeset. Cox regression multivariate analysis confirmed that two of the candidate genes were significantly associated with overall survival. Higher expression of BAG1 [hazard ratio (HR) of 0.14, p < 0.0001, 95% confidence interval (CI) 0.04-0.36] and NOP56 (HR 0.13, p < 0.0001, 95% CI 0.05-0.34) were associated with better prognosis. A prognostic model incorporating expression of BAG1 and NOP56 into the MSKCC score improved prognostication significantly over a model using the MSKCC prognostic score only ( p < 0.0001). Prognostic value of using whole blood mRNA gene profiling in mCCRCC is feasible and should be prospectively confirmed in larger studies.

  20. Whole Blood mRNA Expression-Based Prognosis of Metastatic Renal Cell Carcinoma

    Directory of Open Access Journals (Sweden)

    Karthik V. Giridhar

    2017-11-01

    Full Text Available The Memorial Sloan Kettering Cancer Center (MSKCC prognostic score is based on clinical parameters. We analyzed whole blood mRNA expression in metastatic clear cell renal cell carcinoma (mCCRCC patients and compared it to the MSKCC score for predicting overall survival. In a discovery set of 19 patients with mRCC, we performed whole transcriptome RNA sequencing and selected eighteen candidate genes for further evaluation based on associations with overall survival and statistical significance. In an independent validation of set of 47 patients with mCCRCC, transcript expression of the 18 candidate genes were quantified using a customized NanoString probeset. Cox regression multivariate analysis confirmed that two of the candidate genes were significantly associated with overall survival. Higher expression of BAG1 [hazard ratio (HR of 0.14, p < 0.0001, 95% confidence interval (CI 0.04–0.36] and NOP56 (HR 0.13, p < 0.0001, 95% CI 0.05–0.34 were associated with better prognosis. A prognostic model incorporating expression of BAG1 and NOP56 into the MSKCC score improved prognostication significantly over a model using the MSKCC prognostic score only (p < 0.0001. Prognostic value of using whole blood mRNA gene profiling in mCCRCC is feasible and should be prospectively confirmed in larger studies.

  1. MicroRNA-based Therapy in Animal Models of Selected Gastrointestinal Cancers

    Directory of Open Access Journals (Sweden)

    Jana Merhautova

    2016-09-01

    Full Text Available Gastrointestinal cancer accounts for the 20 most frequent cancer diseases worldwide and there is a constant urge to bring new therapeutics with new mechanism of action into the clinical practice. Quantity of in vitro and in vivo evidences indicate, that exogenous change in pathologically imbalanced microRNAs (miRNAs is capable of transforming the cancer cell phenotype. This review analyzed preclinical miRNA-based therapy attempts in animal models of gastric, pancreatic, gallbladder, and colorectal cancer. From more than 400 original articles, 26 was found to assess the effect of miRNA mimics, precursors, expression vectors, or inhibitors administered locally or systemically being an approach with relatively high translational potential. We have focused on mapping available information on animal model used (animal strain, cell line, xenograft method, pharmacological aspects (oligonucleotide chemistry, delivery system, posology, route of administration and toxicology assessments. We also summarize findings in the field pharmacokinetics and toxicity of miRNA-based therapy.□

  2. Double-stranded RNA transcribed from vector-based oligodeoxynucleotide acts as transcription factor decoy

    International Nuclear Information System (INIS)

    Xiao, Xiao; Gang, Yi; Wang, Honghong; Wang, Jiayin; Zhao, Lina; Xu, Li; Liu, Zhiguo

    2015-01-01

    Highlights: • A shRNA vector based transcription factor decoy, VB-ODN, was designed. • VB-ODN for NF-κB inhibited cell viability in HEK293 cells. • VB-ODN inhibited expression of downstream genes of target transcription factors. • VB-ODN may enhance nuclear entry ratio for its feasibility of virus production. - Abstract: In this study, we designed a short hairpin RNA vector-based oligodeoxynucleotide (VB-ODN) carrying transcription factor (TF) consensus sequence which could function as a decoy to block TF activity. Specifically, VB-ODN for Nuclear factor-κB (NF-κB) could inhibit cell viability and decrease downstream gene expression in HEK293 cells without affecting expression of NF-κB itself. The specific binding between VB-ODN produced double-stranded RNA and NF-κB was evidenced by electrophoretic mobility shift assay. Moreover, similar VB-ODNs designed for three other TFs also inhibit their downstream gene expression but not that of themselves. Our study provides a new design of decoy for blocking TF activity

  3. Predicting microRNA-disease associations using label propagation based on linear neighborhood similarity.

    Science.gov (United States)

    Li, Guanghui; Luo, Jiawei; Xiao, Qiu; Liang, Cheng; Ding, Pingjian

    2018-05-12

    Interactions between microRNAs (miRNAs) and diseases can yield important information for uncovering novel prognostic markers. Since experimental determination of disease-miRNA associations is time-consuming and costly, attention has been given to designing efficient and robust computational techniques for identifying undiscovered interactions. In this study, we present a label propagation model with linear neighborhood similarity, called LPLNS, to predict unobserved miRNA-disease associations. Additionally, a preprocessing step is performed to derive new interaction likelihood profiles that will contribute to the prediction since new miRNAs and diseases lack known associations. Our results demonstrate that the LPLNS model based on the known disease-miRNA associations could achieve impressive performance with an AUC of 0.9034. Furthermore, we observed that the LPLNS model based on new interaction likelihood profiles could improve the performance to an AUC of 0.9127. This was better than other comparable methods. In addition, case studies also demonstrated our method's outstanding performance for inferring undiscovered interactions between miRNAs and diseases, especially for novel diseases. Copyright © 2018. Published by Elsevier Inc.

  4. Double-stranded RNA transcribed from vector-based oligodeoxynucleotide acts as transcription factor decoy

    Energy Technology Data Exchange (ETDEWEB)

    Xiao, Xiao [State Key Laboratory of Cancer Biology and Xijing Hospital of Digestive Diseases, Xijing Hospital, Fourth Military Medical University, Xi’an 710032, Shaanxi Province (China); Gang, Yi [State Key Laboratory of Cancer Biology and Xijing Hospital of Digestive Diseases, Xijing Hospital, Fourth Military Medical University, Xi’an 710032, Shaanxi Province (China); Department of Infectious Diseases, Tangdu Hospital, Fourth Military Medical University, Xi’an 710038, Shaanxi Province (China); Wang, Honghong [No. 518 Hospital of Chinese People’s Liberation Army, Xi’an 710043, Shaanxi Province (China); Wang, Jiayin [The Genome Institute, Washington University in St. Louis, St. Louis, MO 63108 (United States); Zhao, Lina [Department of Radiation Oncology, Xijing Hospital, Fourth Military Medical University, Xi’an 710032, Shaanxi Province (China); Xu, Li, E-mail: lxuhelen@163.com [State Key Laboratory of Cancer Biology and Xijing Hospital of Digestive Diseases, Xijing Hospital, Fourth Military Medical University, Xi’an 710032, Shaanxi Province (China); Liu, Zhiguo, E-mail: liuzhiguo@fmmu.edu.cn [State Key Laboratory of Cancer Biology and Xijing Hospital of Digestive Diseases, Xijing Hospital, Fourth Military Medical University, Xi’an 710032, Shaanxi Province (China)

    2015-02-06

    Highlights: • A shRNA vector based transcription factor decoy, VB-ODN, was designed. • VB-ODN for NF-κB inhibited cell viability in HEK293 cells. • VB-ODN inhibited expression of downstream genes of target transcription factors. • VB-ODN may enhance nuclear entry ratio for its feasibility of virus production. - Abstract: In this study, we designed a short hairpin RNA vector-based oligodeoxynucleotide (VB-ODN) carrying transcription factor (TF) consensus sequence which could function as a decoy to block TF activity. Specifically, VB-ODN for Nuclear factor-κB (NF-κB) could inhibit cell viability and decrease downstream gene expression in HEK293 cells without affecting expression of NF-κB itself. The specific binding between VB-ODN produced double-stranded RNA and NF-κB was evidenced by electrophoretic mobility shift assay. Moreover, similar VB-ODNs designed for three other TFs also inhibit their downstream gene expression but not that of themselves. Our study provides a new design of decoy for blocking TF activity.

  5. Distinct summer and winter bacterial communities in the active layer of Svalbard permafrost revealed by DNA- and RNA-based analyses

    Science.gov (United States)

    Schostag, Morten; Stibal, Marek; Jacobsen, Carsten S.; Bælum, Jacob; Taş, Neslihan; Elberling, Bo; Jansson, Janet K.; Semenchuk, Philipp; Priemé, Anders

    2015-01-01

    The active layer of soil overlaying permafrost in the Arctic is subjected to dramatic annual changes in temperature and soil chemistry, which likely affect bacterial activity and community structure. We studied seasonal variations in the bacterial community of active layer soil from Svalbard (78°N) by co-extracting DNA and RNA from 12 soil cores collected monthly over a year. PCR amplicons of 16S rRNA genes (DNA) and reverse transcribed transcripts (cDNA) were quantified and sequenced to test for the effect of low winter temperature and seasonal variation in concentration of easily degradable organic matter on the bacterial communities. The copy number of 16S rRNA genes and transcripts revealed no distinct seasonal changes indicating potential bacterial activity during winter despite soil temperatures well below −10°C. Multivariate statistical analysis of the bacterial diversity data (DNA and cDNA libraries) revealed a season-based clustering of the samples, and, e.g., the relative abundance of potentially active Cyanobacteria peaked in June and Alphaproteobacteria increased over the summer and then declined from October to November. The structure of the bulk (DNA-based) community was significantly correlated with pH and dissolved organic carbon, while the potentially active (RNA-based) community structure was not significantly correlated with any of the measured soil parameters. A large fraction of the 16S rRNA transcripts was assigned to nitrogen-fixing bacteria (up to 24% in June) and phototrophic organisms (up to 48% in June) illustrating the potential importance of nitrogen fixation in otherwise nitrogen poor Arctic ecosystems and of phototrophic bacterial activity on the soil surface. PMID:25983731

  6. [TYPING OF LEPTOSPIRA SPP. STRAINS BASED ON 16S rRNA].

    Science.gov (United States)

    Ostankova, Yu V; Semenov, A V; Stoyanova, N A; Tokarevich, N K; Lyubimova, N E; Petrova, O A; Ananina, Yu V; Petrov, E M

    2016-01-01

    Comparative typing of Leptospira spp. strain collection based on analysis of 16S RNA fragment. 2 pairs of primers were used for PCR, that jointly flank 1423b.p. sized fragment. Sequences of Leptospira spp. strain 16S rRNA, presented in the international database, were used for phylogenetic analysis. A high similarity, including interspecies, of the 16S fragment in Leptospira spp. strains was shown independently of the source, serovar and serogroup. Heterogeneity of the primary matrix, spontaneous mutations of hotspots and erroneous nucleotide couplings, characteristic for 16S sequence of pathogenic Leptospira spp. strains, are discussed. Molecular-genetic characteristic of certain reference Leptospira spp. strains by 16S sequence is obtained. Results of the studies give evidence on expedience of introduction into clinical practice of identification of Leptospira spp. by 16S sequence directly from the clinical material, that would allow to significantly reduce identification time, dismiss complex type-specific sera and other labor-intensive methods.

  7. A compilation of Web-based research tools for miRNA analysis.

    Science.gov (United States)

    Shukla, Vaibhav; Varghese, Vinay Koshy; Kabekkodu, Shama Prasada; Mallya, Sandeep; Satyamoorthy, Kapaettu

    2017-09-01

    Since the discovery of microRNAs (miRNAs), a class of noncoding RNAs that regulate the gene expression posttranscriptionally in sequence-specific manner, there has been a release of number of tools useful for both basic and advanced applications. This is because of the significance of miRNAs in many pathophysiological conditions including cancer. Numerous bioinformatics tools that have been developed for miRNA analysis have their utility for detection, expression, function, target prediction and many other related features. This review provides a comprehensive assessment of web-based tools for the miRNA analysis that does not require prior knowledge of any computing languages. © The Author 2017. Published by Oxford University Press. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  8. Ratiometric FRET-based detection of DNA and micro-RNA in solution

    International Nuclear Information System (INIS)

    Matveeva, Evgenia G.; Gryczynski, Zygmunt; Stewart, Donald R.; Gryczynski, Ignacy

    2009-01-01

    A ratiometric method for detecting DNA oligomers in bulk solution based on Foerster resonance energy transfer (FRET) is described. The two fluorescence signals (green and red), originating from Cy3 (donor, green) and Cy5 (acceptor, red) labels, are simultaneously detected from the pre-hybridized Cy3oligomerY:Cy5oligomerX system. The ratio of red to green intensities is sensitive to the presence of the single-stranded complimentary oligomer, which replaces single-stranded Cy3oligomerY in the donor:acceptor complex and perturbs the FRET. The detection scheme is generally applicable to the detection of DNA and RNA, and particularly micro-RNA. The proposed method is applicable to various double-stranded various lengths targets (manipulation of the sample preparation conditions, such as temperature, incubation time, denaturizing agent, may be needed).

  9. RNA aptamer-based electrochemical biosensor for selective and label-free analysis of dopamine

    DEFF Research Database (Denmark)

    Farjami, Elahe; Campos, Rui; Nielsen, Jesper Sejrup

    2013-01-01

    , including dopamine precursors and metabolites and other neurotransmitters (NT). Here we report an electrochemical RNA aptamer-based biosensor for analysis of dopamine in the presence of other NT. The biosensor exploits a specific binding of dopamine by the RNA aptamer, immobilized at a cysteamine......, norepinephrine, 3,4-dihydroxy-phenylalanine (l-DOPA), 3,4-dihydroxyphenylacetic acid (DOPAC), methyldopamine, and tyramine, which gave negligible signals under conditions of experiments (electroanalysis at 0.185 V vs Ag/AgCl). The interference from ascorbic and uric acids was eliminated by application...... as a general strategy not to restrict the conformational freedom and binding properties of surface-bound aptamers and, thus, be applicable for the development of other aptasensors...

  10. Identification of threats using linguistics-based knowledge extraction.

    Energy Technology Data Exchange (ETDEWEB)

    Chew, Peter A.

    2008-09-01

    One of the challenges increasingly facing intelligence analysts, along with professionals in many other fields, is the vast amount of data which needs to be reviewed and converted into meaningful information, and ultimately into rational, wise decisions by policy makers. The advent of the world wide web (WWW) has magnified this challenge. A key hypothesis which has guided us is that threats come from ideas (or ideology), and ideas are almost always put into writing before the threats materialize. While in the past the 'writing' might have taken the form of pamphlets or books, today's medium of choice is the WWW, precisely because it is a decentralized, flexible, and low-cost method of reaching a wide audience. However, a factor which complicates matters for the analyst is that material published on the WWW may be in any of a large number of languages. In 'Identification of Threats Using Linguistics-Based Knowledge Extraction', we have sought to use Latent Semantic Analysis (LSA) and other similar text analysis techniques to map documents from the WWW, in whatever language they were originally written, to a common language-independent vector-based representation. This then opens up a number of possibilities. First, similar documents can be found across language boundaries. Secondly, a set of documents in multiple languages can be visualized in a graphical representation. These alone offer potentially useful tools and capabilities to the intelligence analyst whose knowledge of foreign languages may be limited. Finally, we can test the over-arching hypothesis--that ideology, and more specifically ideology which represents a threat, can be detected solely from the words which express the ideology--by using the vector-based representation of documents to predict additional features (such as the ideology) within a framework based on supervised learning. In this report, we present the results of a three-year project of the same name. We believe

  11. A double base change in alternate base pairs induced by ultraviolet irradiation in a glycine transfer RNA gene

    International Nuclear Information System (INIS)

    Coleman, R.D.; Dunst, R.W.; Hill, C.W.; Pennsylvania State Univ., Hershey

    1980-01-01

    The glyUsusub(AGA) mutation affects Escherichia coli tRNAsup(Gly)sub(GGG), changing it to an AGA missense suppressor tRNA. Sequence studies have shown that the mutation involves a double base substitution at the first and third positions of the tRNA anticodon, the result being a change in the anticodon from CCC to UCU. A system has been developed to facilitate the detection of this novel mutation, and we have shown that ultraviolet irradiation and N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) are effective in causing the double base change. A single observation of the mutation occuring spontaneously has been made also. The frequency of MNNG-induced glyUsusub(AGA) mutations is compatible with their being caused by two separate mutagenic events. The frequency of UV-induced glyUsusub(AGA) mutations, however, strongly suggests that the occurence of one base substitution strongly enhances the chance of finding the second substitution at the alternate position. In addition to the double change in the anticodon, the glyUsusub(AGA) tRNA differs from tRNAsup(gly)sub(GGG) in that it bears a modification of the A adjacent to the 3' position of the anticodon. Most likely, this modified base is N-[9-(β-D-ribofuranosyl)-purin-6-ylcarbamoyl] threonine. (orig.) 891 AJ/orig. 892 BRE [de

  12. Analysis of energy-based algorithms for RNA secondary structure prediction

    Directory of Open Access Journals (Sweden)

    Hajiaghayi Monir

    2012-02-01

    Full Text Available Abstract Background RNA molecules play critical roles in the cells of organisms, including roles in gene regulation, catalysis, and synthesis of proteins. Since RNA function depends in large part on its folded structures, much effort has been invested in developing accurate methods for prediction of RNA secondary structure from the base sequence. Minimum free energy (MFE predictions are widely used, based on nearest neighbor thermodynamic parameters of Mathews, Turner et al. or those of Andronescu et al. Some recently proposed alternatives that leverage partition function calculations find the structure with maximum expected accuracy (MEA or pseudo-expected accuracy (pseudo-MEA methods. Advances in prediction methods are typically benchmarked using sensitivity, positive predictive value and their harmonic mean, namely F-measure, on datasets of known reference structures. Since such benchmarks document progress in improving accuracy of computational prediction methods, it is important to understand how measures of accuracy vary as a function of the reference datasets and whether advances in algorithms or thermodynamic parameters yield statistically significant improvements. Our work advances such understanding for the MFE and (pseudo-MEA-based methods, with respect to the latest datasets and energy parameters. Results We present three main findings. First, using the bootstrap percentile method, we show that the average F-measure accuracy of the MFE and (pseudo-MEA-based algorithms, as measured on our largest datasets with over 2000 RNAs from diverse families, is a reliable estimate (within a 2% range with high confidence of the accuracy of a population of RNA molecules represented by this set. However, average accuracy on smaller classes of RNAs such as a class of 89 Group I introns used previously in benchmarking algorithm accuracy is not reliable enough to draw meaningful conclusions about the relative merits of the MFE and MEA-based algorithms

  13. Lentiviral transgenic microRNA-based shRNA suppressed mouse cytochromosome P450 3A (CYP3A expression in a dose-dependent and inheritable manner.

    Directory of Open Access Journals (Sweden)

    Yong Wang

    Full Text Available Cytochomosome P450 enzymes (CYP are heme-containing monooxygenases responsible for oxidative metabolism of many exogenous and endogenous compounds including drugs. The species difference of CYP limits the extent to which data obtained from animals can be translated to humans in pharmacodynamics or pharmacokinetics studies. Transgenic expression of human CYP in animals lacking or with largely reduced endogenous CYP counterparts is recognized as an ideal strategy to correct CYP species difference. CYP3A is the most abundant CYP subfamily both in human and mammals. In this study, we designed a microRNA-based shRNA (miR-shRNA simultaneously targeting four members of mouse CYP3A subfamily (CYP3A11, CYP3A16, CYP3A41 and CYP3A44, and transgenic mice expressing the designed miR-shRNA were generated by lentiviral transgenesis. Results showed that the CYP3A expression level in transgenic mice was markedly reduced compared to that in wild type or unrelated miR-shRNA transgenic mice, and was inversely correlated to the miR-shRNA expression level. The CYP3A expression levels in transgenic offspring of different generations were also remarkably lower compared to those of controls, and moreover the inhibition rate of CYP3A expression remained comparable over generations. The ratio of the targeted CYP3A transcriptional levels was comparable between knockdown and control mice of the same gender as detected by RT-PCR DGGE analysis. These data suggested that transgenic miR-shRNA suppressed CYP3A expression in a dose-dependent and inheritable manner, and transcriptional levels of the targeted CYP3As were suppressed to a similar extent. The observed knockdown efficacy was further confirmed by enzymatic activity analysis, and data showed that CYP3A activities in transgenic mice were markedly reduced compared to those in wild-type or unrelated miR-shRNA transgenic controls (1.11±0.71 vs 5.85±1.74, 5.9±2.4; P<0.01. This work laid down a foundation to further knock

  14. A New Augmentation Based Algorithm for Extracting Maximal Chordal Subgraphs.

    Science.gov (United States)

    Bhowmick, Sanjukta; Chen, Tzu-Yi; Halappanavar, Mahantesh

    2015-02-01

    A graph is chordal if every cycle of length greater than three contains an edge between non-adjacent vertices. Chordal graphs are of interest both theoretically, since they admit polynomial time solutions to a range of NP-hard graph problems, and practically, since they arise in many applications including sparse linear algebra, computer vision, and computational biology. A maximal chordal subgraph is a chordal subgraph that is not a proper subgraph of any other chordal subgraph. Existing algorithms for computing maximal chordal subgraphs depend on dynamically ordering the vertices, which is an inherently sequential process and therefore limits the algorithms' parallelizability. In this paper we explore techniques to develop a scalable parallel algorithm for extracting a maximal chordal subgraph. We demonstrate that an earlier attempt at developing a parallel algorithm may induce a non-optimal vertex ordering and is therefore not guaranteed to terminate with a maximal chordal subgraph. We then give a new algorithm that first computes and then repeatedly augments a spanning chordal subgraph. After proving that the algorithm terminates with a maximal chordal subgraph, we then demonstrate that this algorithm is more amenable to parallelization and that the parallel version also terminates with a maximal chordal subgraph. That said, the complexity of the new algorithm is higher than that of the previous parallel algorithm, although the earlier algorithm computes a chordal subgraph which is not guaranteed to be maximal. We experimented with our augmentation-based algorithm on both synthetic and real-world graphs. We provide scalability results and also explore the effect of different choices for the initial spanning chordal subgraph on both the running time and on the number of edges in the maximal chordal subgraph.

  15. Sparsity-based shrinkage approach for practicability improvement of H-LBP-based edge extraction

    Energy Technology Data Exchange (ETDEWEB)

    Zhao, Chenyi [School of Physics, Northeast Normal University, Changchun 130024 (China); Qiao, Shuang, E-mail: qiaos810@nenu.edu.cn [School of Physics, Northeast Normal University, Changchun 130024 (China); Sun, Jianing, E-mail: sunjn118@nenu.edu.cn [School of Mathematics and Statistics, Northeast Normal University, Changchun 130024 (China); Zhao, Ruikun; Wu, Wei [Jilin Cancer Hospital, Changchun 130021 (China)

    2016-07-21

    The local binary pattern with H function (H-LBP) technique enables fast and efficient edge extraction in digital radiography. In this paper, we reformulate the model of H-LBP and propose a novel sparsity-based shrinkage approach, in which the threshold can be adapted to the data sparsity. Using this model, we upgrade fast H-LBP framework and apply it to real digital radiography. The experiments show that the method improved using the new shrinkage approach can avoid elaborately artificial modulation of parameters and possess greater robustness in edge extraction compared with the other current methods without increasing processing time. - Highlights: • An novel sparsity-based shrinkage approach for edge extraction on digital radiography is proposed. • The threshold of SS-LBP can be adaptive to the data sparsity. • SS-LBP is the development of AH-LBP and H-LBP. • Without boosting processing time and losing processing efficiency, SS-LBP can avoid elaborately artificial modulation of parameters provides. • SS-LBP has more robust performance in edge extraction compared with the existing methods.

  16. Towards Antiviral shRNAs Based on the AgoshRNA Design.

    Directory of Open Access Journals (Sweden)

    Ying Poi Liu

    Full Text Available RNA interference (RNAi can be induced by intracellular expression of a short hairpin RNA (shRNA. Processing of the shRNA requires the RNaseIII-like Dicer enzyme to remove the loop and to release the biologically active small interfering RNA (siRNA. Dicer is also involved in microRNA (miRNA processing to liberate the mature miRNA duplex, but recent studies indicate that miR-451 is not processed by Dicer. Instead, this miRNA is processed by the Argonaute 2 (Ago2 protein, which also executes the subsequent cleavage of a complementary mRNA target. Interestingly, shRNAs that structurally resemble miR-451 can also be processed by Ago2 instead of Dicer. The key determinant of these "AgoshRNA" molecules is a relatively short basepaired stem, which avoids Dicer recognition and consequently allows alternative processing by Ago2. AgoshRNA processing yields a single active RNA strand, whereas standard shRNAs produce a duplex with guide and passenger strands and the latter may cause adverse off-target effects. In this study, we converted previously tested active anti-HIV-1 shRNA molecules into AgoshRNA. We tested several designs that could potentially improve AgoshRNA activity, including extension of the complementarity between the guide strand and the mRNA target and reduction of the thermodynamic stability of the hairpins. We demonstrate that active AgoshRNAs can be generated. However, the RNAi activity is reduced compared to the matching shRNAs. Despite reduced RNAi activity, comparison of an active AgoshRNA and the matching shRNA in a sensitive cell toxicity assay revealed that the AgoshRNA is much less toxic.

  17. Predicting microRNA precursors with a generalized Gaussian components based density estimation algorithm

    Directory of Open Access Journals (Sweden)

    Wu Chi-Yeh

    2010-01-01

    Full Text Available Abstract Background MicroRNAs (miRNAs are short non-coding RNA molecules, which play an important role in post-transcriptional regulation of gene expression. There have been many efforts to discover miRNA precursors (pre-miRNAs over the years. Recently, ab initio approaches have attracted more attention because they do not depend on homology information and provide broader applications than comparative approaches. Kernel based classifiers such as support vector machine (SVM are extensively adopted in these ab initio approaches due to the prediction performance they achieved. On the other hand, logic based classifiers such as decision tree, of which the constructed model is interpretable, have attracted less attention. Results This article reports the design of a predictor of pre-miRNAs with a novel kernel based classifier named the generalized Gaussian density estimator (G2DE based classifier. The G2DE is a kernel based algorithm designed to provide interpretability by utilizing a few but representative kernels for constructing the classification model. The performance of the proposed predictor has been evaluated with 692 human pre-miRNAs and has been compared with two kernel based and two logic based classifiers. The experimental results show that the proposed predictor is capable of achieving prediction performance comparable to those delivered by the prevailing kernel based classification algorithms, while providing the user with an overall picture of the distribution of the data set. Conclusion Software predictors that identify pre-miRNAs in genomic sequences have been exploited by biologists to facilitate molecular biology research in recent years. The G2DE employed in this study can deliver prediction accuracy comparable with the state-of-the-art kernel based machine learning algorithms. Furthermore, biologists can obtain valuable insights about the different characteristics of the sequences of pre-miRNAs with the models generated by the G

  18. Stormbow: A Cloud-Based Tool for Reads Mapping and Expression Quantification in Large-Scale RNA-Seq Studies.

    Science.gov (United States)

    Zhao, Shanrong; Prenger, Kurt; Smith, Lance

    2013-01-01

    RNA-Seq is becoming a promising replacement to microarrays in transcriptome profiling and differential gene expression study. Technical improvements have decreased sequencing costs and, as a result, the size and number of RNA-Seq datasets have increased rapidly. However, the increasing volume of data from large-scale RNA-Seq studies poses a practical challenge for data analysis in a local environment. To meet this challenge, we developed Stormbow, a cloud-based software package, to process large volumes of RNA-Seq data in parallel. The performance of Stormbow has been tested by practically applying it to analyse 178 RNA-Seq samples in the cloud. In our test, it took 6 to 8 hours to process an RNA-Seq sample with 100 million reads, and the average cost was $3.50 per sample. Utilizing Amazon Web Services as the infrastructure for Stormbow allows us to easily scale up to handle large datasets with on-demand computational resources. Stormbow is a scalable, cost effective, and open-source based tool for large-scale RNA-Seq data analysis. Stormbow can be freely downloaded and can be used out of box to process Illumina RNA-Seq datasets.

  19. Semantic Information Extraction of Lanes Based on Onboard Camera Videos

    Science.gov (United States)

    Tang, L.; Deng, T.; Ren, C.

    2018-04-01

    In the field of autonomous driving, semantic information of lanes is very important. This paper proposes a method of automatic detection of lanes and extraction of semantic information from onboard camera videos. The proposed method firstly detects the edges of lanes by the grayscale gradient direction, and improves the Probabilistic Hough transform to fit them; then, it uses the vanishing point principle to calculate the lane geometrical position, and uses lane characteristics to extract lane semantic information by the classification of decision trees. In the experiment, 216 road video images captured by a camera mounted onboard a moving vehicle were used to detect lanes and extract lane semantic information. The results show that the proposed method can accurately identify lane semantics from video images.

  20. A Karnaugh-Map based fingerprint minutiae extraction method

    Directory of Open Access Journals (Sweden)

    Sunil Kumar Singla

    2010-07-01

    Full Text Available Fingerprint is one of the most promising method among all the biometric techniques and has been used for thepersonal authentication for a long time because of its wide acceptance and reliability. Features (Minutiae are extracted fromthe fingerprint in question and are compared with the features already stored in the database for authentication. Crossingnumber (CN is the most commonly used minutiae extraction method for fingerprints. In this paper, a new Karnaugh-Mapbased fingerprint minutiae extraction method has been proposed and discussed. In the proposed algorithm the 8 neighborsof a pixel in a 33 window are arranged as 8 bits of a byte and corresponding hexadecimal (hex value is calculated. Thesehex values are simplified using standard Karnaugh-Map (K-map technique to obtain the minimized logical expression.Experiments conducted on the FVC2002/Db1_a database reveals that the developed method is better than the crossingnumber (CN method.

  1. An albumin-mediated cholesterol design-based strategy for tuning siRNA pharmacokinetics and gene silencing.

    Science.gov (United States)

    Bienk, Konrad; Hvam, Michael Lykke; Pakula, Malgorzata Maria; Dagnæs-Hansen, Frederik; Wengel, Jesper; Malle, Birgitte Mølholm; Kragh-Hansen, Ulrich; Cameron, Jason; Bukrinski, Jens Thostrup; Howard, Kenneth A

    2016-06-28

    Major challenges for the clinical translation of small interfering RNA (siRNA) include overcoming the poor plasma half-life, site-specific delivery and modulation of gene silencing. In this work, we exploit the intrinsic transport properties of human serum albumin to tune the blood circulatory half-life, hepatic accumulation and gene silencing; based on the number of siRNA cholesteryl modifications. We demonstrate by a gel shift assay a strong and specific affinity of recombinant human serum albumin (rHSA) towards cholesteryl-modified siRNA (Kd>1×10(-7)M) dependent on number of modifications. The rHSA/siRNA complex exhibited reduced nuclease degradation and reduced induction of TNF-α production by human peripheral blood mononuclear cells. The increased solubility of heavily cholesteryl modified siRNA in the presence of rHSA facilitated duplex annealing and consequent interaction that allowed in vivo studies using multiple cholesteryl modifications. A structural-activity-based screen of in vitro EGFP-silencing was used to select optimal siRNA designs containing cholesteryl modifications within the sense strand that were used for in vivo studies. We demonstrate plasma half-life extension in NMRI mice from t1/2 12min (naked) to t1/2 45min (single cholesteryl) and t1/2 71min (double cholesteryl) using fluorescent live bioimaging. The biodistribution showed increased accumulation in the liver for the double cholesteryl modified siRNA that correlated with an increase in hepatic Factor VII gene silencing of 28% (rHSA/siRNA) compared to 4% (naked siRNA) 6days post-injection. This work presents a novel albumin-mediated cholesteryl design-based strategy for tuning pharmacokinetics and systemic gene silencing. Copyright © 2016 Elsevier B.V. All rights reserved.

  2. Comparison of direct boiling method with commercial kits for extracting fecal microbiome DNA by Illumina sequencing of 16S rRNA tags.

    Science.gov (United States)

    Peng, Xin; Yu, Ke-Qiang; Deng, Guan-Hua; Jiang, Yun-Xia; Wang, Yu; Zhang, Guo-Xia; Zhou, Hong-Wei

    2013-12-01

    Low cost and high throughput capacity are major advantages of using next generation sequencing (NGS) techniques to determine metagenomic 16S rRNA tag sequences. These methods have significantly changed our view of microorganisms in the fields of human health and environmental science. However, DNA extraction using commercial kits has shortcomings of high cost and time constraint. In the present study, we evaluated the determination of fecal microbiomes using a direct boiling method compared with 5 different commercial extraction methods, e.g., Qiagen and MO BIO kits. Principal coordinate analysis (PCoA) using UniFrac distances and clustering showed that direct boiling of a wide range of feces concentrations gave a similar pattern of bacterial communities as those obtained from most of the commercial kits, with the exception of the MO BIO method. Fecal concentration by boiling method affected the estimation of α-diversity indices, otherwise results were generally comparable between boiling and commercial methods. The operational taxonomic units (OTUs) determined through direct boiling showed highly consistent frequencies with those determined through most of the commercial methods. Even those for the MO BIO kit were also obtained by the direct boiling method with high confidence. The present study suggested that direct boiling could be used to determine the fecal microbiome and using this method would significantly reduce the cost and improve the efficiency of the sample preparation for studying gut microbiome diversity. © 2013 Elsevier B.V. All rights reserved.

  3. Spliceosomal small nuclear RNAs of Tetrahymena thermophila and some possible snRNA-snRNA base-pairing interactions

    DEFF Research Database (Denmark)

    Orum, H; Nielsen, Henrik; Engberg, J

    1991-01-01

    We have identified and characterized the full set of spliceosomal small nuclear RNAs (snRNAs; U1, U2, U4, U5 and U6) from the ciliated protozoan Tetrahymena thermophila. With the exception of U4 snRNA, the sizes of the T. thermophila snRNAs are closely similar to their metazoan homologues. The T....... thermophila snRNAs all have unique 5' ends, which start with an adenine residue. In contrast, with the exception of U6, their 3' ends show some size heterogeneity. The primary sequences of the T. thermophila snRNAs contain the sequence motifs shown, or proposed, to be of functional importance in other...

  4. Optimization of β-cyclodextrin-based flavonol extraction from apple pomace using response surface methodology.

    Science.gov (United States)

    Parmar, Indu; Sharma, Sowmya; Rupasinghe, H P Vasantha

    2015-04-01

    The present study investigated five cyclodextrins (CDs) for the extraction of flavonols from apple pomace powder and optimized β-CD based extraction of total flavonols using response surface methodology. A 2(3) central composite design with β-CD concentration (0-5 g 100 mL(-1)), extraction temperature (20-72 °C), extraction time (6-48 h) and second-order quadratic model for the total flavonol yield (mg 100 g(-1) DM) was selected to generate the response surface curves. The optimal conditions obtained were: β-CD concentration, 2.8 g 100 mL(-1); extraction temperature, 45 °C and extraction time, 25.6 h that predicted the extraction of 166.6 mg total flavonols 100 g(-1) DM. The predicted amount was comparable to the experimental amount of 151.5 mg total flavonols 100 g(-1) DM obtained from optimal β-CD based parameters, thereby giving a low absolute error and adequacy of fitted model. In addition, the results from optimized extraction conditions showed values similar to those obtained through previously established solvent based sonication assisted flavonol extraction procedure. To the best of our knowledge, this is the first study to optimize aqueous β-CD based flavonol extraction which presents an environmentally safe method for value-addition to under-utilized bio resources.

  5. Classification and energetics of the base-phosphate interactions in RNA

    Czech Academy of Sciences Publication Activity Database

    Zirbel, C.L.; Šponer, Judit E.; Šponer, Jiří; Stombaugh, J.; Leontis, N.B.

    2009-01-01

    Roč. 37, č. 15 (2009), s. 4898-4918 ISSN 0305-1048 R&D Projects: GA AV ČR(CZ) IAA400550701; GA AV ČR(CZ) IAA400040802; GA AV ČR(CZ) 1QS500040581; GA MŠk(CZ) LC06030 Grant - others:GA ČR(CZ) GA203/09/1476 Program:GA Institutional research plan: CEZ:AV0Z50040507; CEZ:AV0Z50040702 Keywords : RNA * base * phosphate Subject RIV: BO - Biophysics Impact factor: 7.479, year: 2009

  6. Semantics-based information extraction for detecting economic events

    NARCIS (Netherlands)

    A.C. Hogenboom (Alexander); F. Frasincar (Flavius); K. Schouten (Kim); O. van der Meer

    2013-01-01

    textabstractAs today's financial markets are sensitive to breaking news on economic events, accurate and timely automatic identification of events in news items is crucial. Unstructured news items originating from many heterogeneous sources have to be mined in order to extract knowledge useful for

  7. Rule set transferability for object-based feature extraction

    NARCIS (Netherlands)

    Anders, N.S.; Seijmonsbergen, Arie C.; Bouten, Willem

    2015-01-01

    Cirques are complex landforms resulting from glacial erosion and can be used to estimate Equilibrium Line Altitudes and infer climate history. Automated extraction of cirques may help research on glacial geomorphology and climate change. Our objective was to test the transferability of an

  8. Ecosystem-based design rules for marine sand extraction sites

    NARCIS (Netherlands)

    Jong, de Maarten F.; Borsje, Bas W.; Baptist, Martin J.; Wal, van der Jan Tjalling; Lindeboom, Han J.; Hoekstra, Piet

    2016-01-01

    The demand for marine sand in the Netherlands as well as globally is increasing. Over the last decades, only shallow sand extraction of 2m below the seabed was allowed on the Dutch Continental Shelf (DCS). To guarantee sufficient supply and to decrease the surface area of direct impact, the Dutch

  9. Ecosystem-based design rules for marine sand extraction sites

    NARCIS (Netherlands)

    de Jong, Maarten F.; Borsje, Bas W.; Baptist, Martin J.; van der Wal, Jan Tjalling; Lindeboom, Han J.; Hoekstra, Piet

    2016-01-01

    The demand for marine sand in the Netherlands as well as globally is increasing. Over the last decades, only shallow sand extraction of 2m below the seabed was allowed on the Dutch Continental Shelf (DCS). To guarantee sufficient supply and to decrease the surface area of direct impact, the Dutch

  10. Human disease MiRNA inference by combining target information based on heterogeneous manifolds.

    Science.gov (United States)

    Ding, Pingjian; Luo, Jiawei; Liang, Cheng; Xiao, Qiu; Cao, Buwen

    2018-04-01

    The emergence of network medicine has provided great insight into the identification of disease-related molecules, which could help with the development of personalized medicine. However, the state-of-the-art methods could neither simultaneously consider target information and the known miRNA-disease associations nor effectively explore novel gene-disease associations as a by-product during the process of inferring disease-related miRNAs. Computational methods incorporating multiple sources of information offer more opportunities to infer disease-related molecules, including miRNAs and genes in heterogeneous networks at a system level. In this study, we developed a novel algorithm, named inference of Disease-related MiRNAs based on Heterogeneous Manifold (DMHM), to accurately and efficiently identify miRNA-disease associations by integrating multi-omics data. Graph-based regularization was utilized to obtain a smooth function on the data manifold, which constitutes the main principle of DMHM. The novelty of this framework lies in the relatedness between diseases and miRNAs, which are measured via heterogeneous manifolds on heterogeneous networks integrating target information. To demonstrate the effectiveness of DMHM, we conducted comprehensive experiments based on HMDD datasets and compared DMHM with six state-of-the-art methods. Experimental results indicated that DMHM significantly outperformed the other six methods under fivefold cross validation and de novo prediction tests. Case studies have further confirmed the practical usefulness of DMHM. Copyright © 2018 Elsevier Inc. All rights reserved.

  11. Tuning iteration space slicing based tiled multi-core code implementing Nussinov's RNA folding.

    Science.gov (United States)

    Palkowski, Marek; Bielecki, Wlodzimierz

    2018-01-15

    RNA folding is an ongoing compute-intensive task of bioinformatics. Parallelization and improving code locality for this kind of algorithms is one of the most relevant areas in computational biology. Fortunately, RNA secondary structure approaches, such as Nussinov's recurrence, involve mathematical operations over affine control loops whose iteration space can be represented by the polyhedral model. This allows us to apply powerful polyhedral compilation techniques based on the transitive closure of dependence graphs to generate parallel tiled code implementing Nussinov's RNA folding. Such techniques are within the iteration space slicing framework - the transitive dependences are applied to the statement instances of interest to produce valid tiles. The main problem at generating parallel tiled code is defining a proper tile size and tile dimension which impact parallelism degree and code locality. To choose the best tile size and tile dimension, we first construct parallel parametric tiled code (parameters are variables defining tile size). With this purpose, we first generate two nonparametric tiled codes with different fixed tile sizes but with the same code structure and then derive a general affine model, which describes all integer factors available in expressions of those codes. Using this model and known integer factors present in the mentioned expressions (they define the left-hand side of the model), we find unknown integers in this model for each integer factor available in the same fixed tiled code position and replace in this code expressions, including integer factors, with those including parameters. Then we use this parallel parametric tiled code to implement the well-known tile size selection (TSS) technique, which allows us to discover in a given search space the best tile size and tile dimension maximizing target code performance. For a given search space, the presented approach allows us to choose the best tile size and tile dimension in

  12. Investigation of antineoplastic activity of chewing tablets based on dry oat extract and quercetin

    Directory of Open Access Journals (Sweden)

    Ярослав Ростиславович Андрійчук

    2015-07-01

    Full Text Available One of the main goals of domestic pharmaceutical science is development of new medicines. Thus, new tablet drug was created based on dry oat extract and quercetin. Investigation of antineoplastic activity was performed. Antineoplastic activity of investigational drug based on dry oat extract and quercetin was experimentally proved.

  13. Application of ionic liquids based enzyme-assisted extraction of chlorogenic acid from Eucommia ulmoides leaves

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Tingting; Sui, Xiaoyu, E-mail: suixiaoyu@outlook.com; Li, Li; Zhang, Jie; Liang, Xin; Li, Wenjing; Zhang, Honglian; Fu, Shuang

    2016-01-15

    A new approach for ionic liquid based enzyme-assisted extraction (ILEAE) of chlorogenic acid (CGA) from Eucommia ulmoides is presented in which enzyme pretreatment was used in ionic liquids aqueous media to enhance extraction yield. For this purpose, the solubility of CGA and the activity of cellulase were investigated in eight 1-alkyl-3-methylimidazolium ionic liquids. Cellulase in 0.5 M [C6mim]Br aqueous solution was found to provide better performance in extraction. The factors of ILEAE procedures including extraction time, extraction phase pH, extraction temperatures and enzyme concentrations were investigated. Moreover, the novel developed approach offered advantages in term of yield and efficiency compared with other conventional extraction techniques. Scanning electronic microscopy of plant samples indicated that cellulase treated cell wall in ionic liquid solution was subjected to extract, which led to more efficient extraction by reducing mass transfer barrier. The proposed ILEAE method would develope a continuous process for enzyme-assisted extraction including enzyme incubation and solvent extraction process. In this research, we propose a novel view for enzyme-assisted extraction of plant active component, besides concentrating on enzyme facilitated cell wall degradation, focusing on improvement of bad permeability of ionic liquids solutions. - Highlights: • An ionic liquid based enzyme-assisted extraction method of natural product was explored. • ILEAE utilizes enzymatic treatment to improve permeability of ionic liquids solution. • Enzyme incubation and solvent extraction process were ongoing simultaneously. • ILEAE process simplified operating process and suitable for more complete extraction.

  14. Application of ionic liquids based enzyme-assisted extraction of chlorogenic acid from Eucommia ulmoides leaves

    International Nuclear Information System (INIS)

    Liu, Tingting; Sui, Xiaoyu; Li, Li; Zhang, Jie; Liang, Xin; Li, Wenjing; Zhang, Honglian; Fu, Shuang

    2016-01-01

    A new approach for ionic liquid based enzyme-assisted extraction (ILEAE) of chlorogenic acid (CGA) from Eucommia ulmoides is presented in which enzyme pretreatment was used in ionic liquids aqueous media to enhance extraction yield. For this purpose, the solubility of CGA and the activity of cellulase were investigated in eight 1-alkyl-3-methylimidazolium ionic liquids. Cellulase in 0.5 M [C6mim]Br aqueous solution was found to provide better performance in extraction. The factors of ILEAE procedures including extraction time, extraction phase pH, extraction temperatures and enzyme concentrations were investigated. Moreover, the novel developed approach offered advantages in term of yield and efficiency compared with other conventional extraction techniques. Scanning electronic microscopy of plant samples indicated that cellulase treated cell wall in ionic liquid solution was subjected to extract, which led to more efficient extraction by reducing mass transfer barrier. The proposed ILEAE method would develope a continuous process for enzyme-assisted extraction including enzyme incubation and solvent extraction process. In this research, we propose a novel view for enzyme-assisted extraction of plant active component, besides concentrating on enzyme facilitated cell wall degradation, focusing on improvement of bad permeability of ionic liquids solutions. - Highlights: • An ionic liquid based enzyme-assisted extraction method of natural product was explored. • ILEAE utilizes enzymatic treatment to improve permeability of ionic liquids solution. • Enzyme incubation and solvent extraction process were ongoing simultaneously. • ILEAE process simplified operating process and suitable for more complete extraction.

  15. A novel technique for extracting clouds base height using ground based imaging

    Directory of Open Access Journals (Sweden)

    E. Hirsch

    2011-01-01

    Full Text Available The height of a cloud in the atmospheric column is a key parameter in its characterization. Several remote sensing techniques (passive and active, either ground-based or on space-borne platforms and in-situ measurements are routinely used in order to estimate top and base heights of clouds. In this article we present a novel method that combines thermal imaging from the ground and sounded wind profile in order to derive the cloud base height. This method is independent of cloud types, making it efficient for both low boundary layer and high clouds. In addition, using thermal imaging ensures extraction of clouds' features during daytime as well as at nighttime. The proposed technique was validated by comparison to active sounding by ceilometers (which is a standard ground based method, to lifted condensation level (LCL calculations, and to MODIS products obtained from space. As all passive remote sensing techniques, the proposed method extracts only the height of the lowest cloud layer, thus upper cloud layers are not detected. Nevertheless, the information derived from this method can be complementary to space-borne cloud top measurements when deep-convective clouds are present. Unlike techniques such as LCL, this method is not limited to boundary layer clouds, and can extract the cloud base height at any level, as long as sufficient thermal contrast exists between the radiative temperatures of the cloud and its surrounding air parcel. Another advantage of the proposed method is its simplicity and modest power needs, making it particularly suitable for field measurements and deployment at remote locations. Our method can be further simplified for use with visible CCD or CMOS camera (although nighttime clouds will not be observed.

  16. Deciphering chicken gut microbial dynamics based on high-throughput 16S rRNA metagenomics analyses.

    Science.gov (United States)

    Mohd Shaufi, Mohd Asrore; Sieo, Chin Chin; Chong, Chun Wie; Gan, Han Ming; Ho, Yin Wan

    2015-01-01

    Chicken gut microbiota has paramount roles in host performance, health and immunity. Understanding the topological difference in gut microbial community composition is crucial to provide knowledge on the functions of each members of microbiota to the physiological maintenance of the host. The gut microbiota profiling of the chicken was commonly performed previously using culture-dependent and early culture-independent methods which had limited coverage and accuracy. Advances in technology based on next-generation sequencing (NGS), offers unparalleled coverage and depth in determining microbial gut dynamics. Thus, the aim of this study was to investigate the ileal and caecal microbiota development as chicken aged, which is important for future effective gut modulation. Ileal and caecal contents of broiler chicken were extracted from 7, 14, 21 and 42-day old chicken. Genomic DNA was then extracted and amplified based on V3 hyper-variable region of 16S rRNA. Bioinformatics, ecological and statistical analyses such as Principal Coordinate Analysis (PCoA) was performed in mothur software and plotted using PRIMER 6. Additional analyses for predicted metagenomes were performed through PICRUSt and STAMP software package based on Greengenes databases. A distinctive difference in bacterial communities was observed between ilea and caeca as the chicken aged (P microbial communities in the caeca were more diverse in comparison to the ilea communities. The potentially pathogenic bacteria such as Clostridium were elevated as the chicken aged and the population of beneficial microbe such as Lactobacillus was low at all intervals. On the other hand, based on predicted metagenomes analysed, clear distinction in functions and roles of gut microbiota such as gene pathways related to nutrient absorption (e.g. sugar and amino acid metabolism), and bacterial proliferation and colonization (e.g. bacterial motility proteins, two-component system and bacterial secretion system) were

  17. Sorting live stem cells based on Sox2 mRNA expression.

    Directory of Open Access Journals (Sweden)

    Hans M Larsson

    Full Text Available While cell sorting usually relies on cell-surface protein markers, molecular beacons (MBs offer the potential to sort cells based on the presence of any expressed mRNA and in principle could be extremely useful to sort rare cell populations from primary isolates. We show here how stem cells can be purified from mixed cell populations by sorting based on MBs. Specifically, we designed molecular beacons targeting Sox2, a well-known stem cell marker for murine embryonic (mES and neural stem cells (NSC. One of our designed molecular beacons displayed an increase in fluorescence compared to a nonspecific molecular beacon both in vitro and in vivo when tested in mES and NSCs. We sorted Sox2-MB(+SSEA1(+ cells from a mixed population of 4-day retinoic acid-treated mES cells and effectively isolated live undifferentiated stem cells. Additionally, Sox2-MB(+ cells isolated from primary mouse brains were sorted and generated neurospheres with higher efficiency than Sox2-MB(- cells. These results demonstrate the utility of MBs for stem cell sorting in an mRNA-specific manner.

  18. Model-based design of RNA hybridization networks implemented in living cells.

    Science.gov (United States)

    Rodrigo, Guillermo; Prakash, Satya; Shen, Shensi; Majer, Eszter; Daròs, José-Antonio; Jaramillo, Alfonso

    2017-09-19

    Synthetic gene circuits allow the behavior of living cells to be reprogrammed, and non-coding small RNAs (sRNAs) are increasingly being used as programmable regulators of gene expression. However, sRNAs (natural or synthetic) are generally used to regulate single target genes, while complex dynamic behaviors would require networks of sRNAs regulating each other. Here, we report a strategy for implementing such networks that exploits hybridization reactions carried out exclusively by multifaceted sRNAs that are both targets of and triggers for other sRNAs. These networks are ultimately coupled to the control of gene expression. We relied on a thermodynamic model of the different stable conformational states underlying this system at the nucleotide level. To test our model, we designed five different RNA hybridization networks with a linear architecture, and we implemented them in Escherichia coli. We validated the network architecture at the molecular level by native polyacrylamide gel electrophoresis, as well as the network function at the bacterial population and single-cell levels with a fluorescent reporter. Our results suggest that it is possible to engineer complex cellular programs based on RNA from first principles. Because these networks are mainly based on physical interactions, our designs could be expanded to other organisms as portable regulatory resources or to implement biological computations. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  19. Direct RNA-based detection of CTX-M β-lactamases in human blood samples.

    Science.gov (United States)

    Stein, Claudia; Makarewicz, Oliwia; Pfeifer, Yvonne; Brandt, Christian; Pletz, Mathias W

    2015-05-01

    Bloodstream infections with ESBL-producers are associated with increased mortality, which is due to delayed appropriate treatment resulting in clinical failure. Current routine diagnostics for detection of bloodstream infections consists of blood culture followed by species identification and susceptibility testing. In attempts to improve and accelerate diagnostic procedures, PCR-based methods have been developed. These methods focus on species identification covering only a limited number of ESBL coding genes. Therefore, they fail to cover the steadily further evolving genetic diversity of clinically relevant β-lactamases. We have recently designed a fast and novel RNA targeting method to detect and specify CTX-M alleles from bacterial cultures, based on an amplification-pyrosequencing approach. We further developed this assay towards a diagnostic tool for clinical use and evaluated its sensitivity and specificity when applied directly to human blood samples. An optimized protocol for mRNA isolation allows detection of specific CTX-M groups from as little as 100 CFU/mL blood via reverse transcription, amplification, and pyrosequencing directly from human EDTA blood samples as well as from pre-incubated human blood cultures with a turnaround time for test results of <7 h. Copyright © 2015 Elsevier GmbH. All rights reserved.

  20. GA Based Optimal Feature Extraction Method for Functional Data Classification

    OpenAIRE

    Jun Wan; Zehua Chen; Yingwu Chen; Zhidong Bai

    2010-01-01

    Classification is an interesting problem in functional data analysis (FDA), because many science and application problems end up with classification problems, such as recognition, prediction, control, decision making, management, etc. As the high dimension and high correlation in functional data (FD), it is a key problem to extract features from FD whereas keeping its global characters, which relates to the classification efficiency and precision to heavens. In this paper...

  1. In Situ Dark Adaptation Enhances the Efficiency of DNA Extraction from Mature Pin Oak (Quercus palustris Leaves, Facilitating the Identification of Partial Sequences of the 18S rRNA and Isoprene Synthase (IspS Genes

    Directory of Open Access Journals (Sweden)

    Csengele E. Barta

    2017-10-01

    Full Text Available Mature oak (Quercus spp. leaves, although abundantly available during the plants’ developmental cycle, are rarely exploited as viable sources of genomic DNA. These leaves are rich in metabolites difficult to remove during standard DNA purification, interfering with downstream molecular genetics applications. The current work assessed whether in situ dark adaptation, to deplete sugar reserves and inhibit secondary metabolite synthesis could compensate for the difficulties encountered when isolating DNA from mature leaves rich in secondary metabolites. We optimized a rapid, commercial kit based method to extract genomic DNA from dark- and light-adapted leaves. We demonstrated that in situ dark adaptation increases the yield and quality of genomic DNA obtained from mature oak leaves, yielding templates of sufficiently high quality for direct downstream applications, such as PCR amplification and gene identification. The quality of templates isolated from dark-adapted pin oak leaves particularly improved the amplification of larger fragments in our experiments. From DNA extracts prepared with our optimized method, we identified for the first time partial segments of the genes encoding 18S rRNA and isoprene synthase (IspS from pin oak (Quercus palustris, whose full genome has not yet been sequenced.

  2. RCK: accurate and efficient inference of sequence- and structure-based protein-RNA binding models from RNAcompete data.

    Science.gov (United States)

    Orenstein, Yaron; Wang, Yuhao; Berger, Bonnie

    2016-06-15

    Protein-RNA interactions, which play vital roles in many processes, are mediated through both RNA sequence and structure. CLIP-based methods, which measure protein-RNA binding in vivo, suffer from experimental noise and systematic biases, whereas in vitro experiments capture a clearer signal of protein RNA-binding. Among them, RNAcompete provides binding affinities of a specific protein to more than 240 000 unstructured RNA probes in one experiment. The computational challenge is to infer RNA structure- and sequence-based binding models from these data. The state-of-the-art in sequence models, Deepbind, does not model structural preferences. RNAcontext models both sequence and structure preferences, but is outperformed by GraphProt. Unfortunately, GraphProt cannot detect structural preferences from RNAcompete data due to the unstructured nature of the data, as noted by its developers, nor can it be tractably run on the full RNACompete dataset. We develop RCK, an efficient, scalable algorithm that infers both sequence and structure preferences based on a new k-mer based model. Remarkably, even though RNAcompete data is designed to be unstructured, RCK can still learn structural preferences from it. RCK significantly outperforms both RNAcontext and Deepbind in in vitro binding prediction for 244 RNAcompete experiments. Moreover, RCK is also faster and uses less memory, which enables scalability. While currently on par with existing methods in in vivo binding prediction on a small scale test, we demonstrate that RCK will increasingly benefit from experimentally measured RNA structure profiles as compared to computationally predicted ones. By running RCK on the entire RNAcompete dataset, we generate and provide as a resource a set of protein-RNA structure-based models on an unprecedented scale. Software and models are freely available at http://rck.csail.mit.edu/ bab@mit.edu Supplementary data are available at Bioinformatics online. © The Author 2016. Published by

  3. 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.

  4. An analytical platform for mass spectrometry-based identification and chemical analysis of RNA in ribonucleoprotein complexes.

    Science.gov (United States)

    Taoka, Masato; Yamauchi, Yoshio; Nobe, Yuko; Masaki, Shunpei; Nakayama, Hiroshi; Ishikawa, Hideaki; Takahashi, Nobuhiro; Isobe, Toshiaki

    2009-11-01

    We describe here a mass spectrometry (MS)-based analytical platform of RNA, which combines direct nano-flow reversed-phase liquid chromatography (RPLC) on a spray tip column and a high-resolution LTQ-Orbitrap mass spectrometer. Operating RPLC under a very low flow rate with volatile solvents and MS in the negative mode, we could estimate highly accurate mass values sufficient to predict the nucleotide composition of a approximately 21-nucleotide small interfering RNA, detect post-transcriptional modifications in yeast tRNA, and perform collision-induced dissociation/tandem MS-based structural analysis of nucleolytic fragments of RNA at a sub-femtomole level. Importantly, the method allowed the identification and chemical analysis of small RNAs in ribonucleoprotein (RNP) complex, such as the pre-spliceosomal RNP complex, which was pulled down from cultured cells with a tagged protein cofactor as bait. We have recently developed a unique genome-oriented database search engine, Ariadne, which allows tandem MS-based identification of RNAs in biological samples. Thus, the method presented here has broad potential for automated analysis of RNA; it complements conventional molecular biology-based techniques and is particularly suited for simultaneous analysis of the composition, structure, interaction, and dynamics of RNA and protein components in various cellular RNP complexes.

  5. 16S rRNA-based detection of oral pathogens in coronary atherosclerotic plaque

    Directory of Open Access Journals (Sweden)

    Mahendra Jaideep

    2010-01-01

    Full Text Available Background: Atherosclerosis develops as a response of the vessel wall to injury. Chronic bacterial infections have been associated with an increased risk for atherosclerosis and coronary artery disease. The ability of oral pathogens to colonize in coronary atheromatous plaque is well known. Aim: The aim of this study was to detect the presence of Treponema denticola, Porphyromonas gingivalis and Campylobacter rectus in the subgingival and atherosclerotic plaques of patients with coronary artery disease. Materials and Methods: Fifty-one patients in the age group of 40-80 years with coronary artery disease were selected for the study. DNA was extracted from the plaque samples. The specific primers for T. denticola, C. rectus and P. gingivalis were used to amplify a part of the 16S rRNA gene by polymerase chain reaction. Statistical Analysis Used: Chi-square analysis, correlation coefficient and prevalence percentage of the microorganisms were carried out for the analysis. Results: Of the 51 patients, T. denticola, C. rectus and P. gingivalis were detected in 49.01%, 21.51% and 45.10% of the atherosclerotic plaque samples. Conclusions: Our study revealed the presence of bacterial DNA of the oral pathogenic microorganisms in coronary atherosclerotic plaques. The presence of the bacterial DNA in the coronary atherosclerotic plaques in significant proportion may suggest the possible relationship between periodontal bacterial infection and genesis of coronary atherosclerosis.

  6. Ionic liquid-based microwave-assisted extraction of flavonoids from Bauhinia championii (Benth.) Benth.

    Science.gov (United States)

    Xu, Wei; Chu, Kedan; Li, Huang; Zhang, Yuqin; Zheng, Haiyin; Chen, Ruilan; Chen, Lidian

    2012-12-03

    An ionic liquids (IL)-based microwave-assisted approach for extraction and determination of flavonoids from Bauhinia championii (Benth.) Benth. was proposed for the first time. Several ILs with different cations and anions and the microwave-assisted extraction (MAE) conditions, including sample particle size, extraction time and liquid-solid ratio, were investigated. Two M 1-butyl-3-methylimidazolium bromide ([bmim] Br) solution with 0.80 M HCl was selected as the optimal solvent. Meanwhile the optimized conditions a ratio of liquid to material of 30:1, and the extraction for 10 min at 70 °C. Compared with conventional heat-reflux extraction (CHRE) and the regular MAE, IL-MAE exhibited a higher extraction yield and shorter extraction time (from 1.5 h to 10 min). The optimized extraction samples were analysed by LC-MS/MS. IL extracts of Bauhinia championii (Benth.)Benth consisted mainly of flavonoids, among which myricetin, quercetin and kaempferol, β-sitosterol, triacontane and hexacontane were identified. The study indicated that IL-MAE was an efficient and rapid method with simple sample preparation. LC-MS/MS was also used to determine the chemical composition of the ethyl acetate/MAE extract of Bauhinia championii (Benth.) Benth, and it maybe become a rapid method to determine the composition of new plant extracts.

  7. Theragnosis-based combined cancer therapy using doxorubicin-conjugated microRNA-221 molecular beacon.

    Science.gov (United States)

    Lee, Jonghwan; Choi, Kyung-Ju; Moon, Sung Ung; Kim, Soonhag

    2016-01-01

    Recently, microRNA (miRNA or miR) has emerged as a new cancer biomarker because of its high expression level in various cancer types and its role in the control of tumor suppressor genes. In cancer studies, molecular imaging and treatment based on target cancer markers have been combined to facilitate simultaneous cancer diagnosis and therapy. In this study, for combined therapy with diagnosis of cancer, we developed a doxorubicin-conjugated miR-221 molecular beacon (miR-221 DOXO MB) in a single platform composed of three different nucleotides: miR-221 binding sequence, black hole quencher 1 (BHQ1), and doxorubicin binding site. Imaging of endogenous miR-221 was achieved by specific hybridization between miR-221 and the miR-221 binding site in miR-221 DOXO MB. The presence of miR-221 triggered detachment of the quencher oligo and subsequent activation of a fluorescent signal of miR-221 DOXO MB. Simultaneous cancer therapy in C6 astrocytoma cells and nude mice was achieved by inhibition of miRNA-221 function that downregulates tumor suppressor genes. The detection of miR-221 expression and inhibition of miR-221 function by miR-221 DOXO MB provide the feasibility as a cancer theragnostic probe. Furthermore, a cytotoxic effect was induced by unloading of doxorubicin intercalated into miR-221 DOXO MB inside cells. Loss of miR-221 function and cytotoxicity induced by the miR-221 DOXO MB provides combined therapeutic efficacy against cancers. This method could be used as a new theragnostic probe with enhanced therapy to detect and inhibit many cancer-related miRNAs. Copyright © 2015 Elsevier Ltd. All rights reserved.

  8. A hybrid model based on neural networks for biomedical relation extraction.

    Science.gov (United States)

    Zhang, Yijia; Lin, Hongfei; Yang, Zhihao; Wang, Jian; Zhang, Shaowu; Sun, Yuanyuan; Yang, Liang

    2018-05-01

    Biomedical relation extraction can automatically extract high-quality biomedical relations from biomedical texts, which is a vital step for the mining of biomedical knowledge hidden in the literature. Recurrent neural networks (RNNs) and convolutional neural networks (CNNs) are two major neural network models for biomedical relation extraction. Neural network-based methods for biomedical relation extraction typically focus on the sentence sequence and employ RNNs or CNNs to learn the latent features from sentence sequences separately. However, RNNs and CNNs have their own advantages for biomedical relation extraction. Combining RNNs and CNNs may improve biomedical relation extraction. In this paper, we present a hybrid model for the extraction of biomedical relations that combines RNNs and CNNs. First, the shortest dependency path (SDP) is generated based on the dependency graph of the candidate sentence. To make full use of the SDP, we divide the SDP into a dependency word sequence and a relation sequence. Then, RNNs and CNNs are employed to automatically learn the features from the sentence sequence and the dependency sequences, respectively. Finally, the output features of the RNNs and CNNs are combined to detect and extract biomedical relations. We evaluate our hybrid model using five public (protein-protein interaction) PPI corpora and a (drug-drug interaction) DDI corpus. The experimental results suggest that the advantages of RNNs and CNNs in biomedical relation extraction are complementary. Combining RNNs and CNNs can effectively boost biomedical relation extraction performance. Copyright © 2018 Elsevier Inc. All rights reserved.

  9. Extraction of CT dose information from DICOM metadata: automated Matlab-based approach.

    Science.gov (United States)

    Dave, Jaydev K; Gingold, Eric L

    2013-01-01

    The purpose of this study was to extract exposure parameters and dose-relevant indexes of CT examinations from information embedded in DICOM metadata. DICOM dose report files were identified and retrieved from a PACS. An automated software program was used to extract from these files information from the structured elements in the DICOM metadata relevant to exposure. Extracting information from DICOM metadata eliminated potential errors inherent in techniques based on optical character recognition, yielding 100% accuracy.

  10. An investigation of paper based microfluidic devices for size based separation and extraction applications.

    Science.gov (United States)

    Zhong, Z W; Wu, R G; Wang, Z P; Tan, H L

    2015-09-01

    Conventional microfluidic devices are typically complex and expensive. The devices require the use of pneumatic control systems or highly precise pumps to control the flow in the devices. This work investigates an alternative method using paper based microfluidic devices to replace conventional microfluidic devices. Size based separation and extraction experiments conducted were able to separate free dye from a mixed protein and dye solution. Experimental results showed that pure fluorescein isothiocyanate could be separated from a solution of mixed fluorescein isothiocyanate and fluorescein isothiocyanate labeled bovine serum albumin. The analysis readings obtained from a spectrophotometer clearly show that the extracted tartrazine sample did not contain any amount of Blue-BSA, because its absorbance value was 0.000 measured at a wavelength of 590nm, which correlated to Blue-BSA. These demonstrate that paper based microfluidic devices, which are inexpensive and easy to implement, can potentially replace their conventional counterparts by the use of simple geometry designs and the capillary action. These findings will potentially help in future developments of paper based microfluidic devices. Copyright © 2015 Elsevier B.V. All rights reserved.

  11. Elucidating the Small Regulatory RNA Repertoire of the Sea Anemone Anemonia viridis Based on Whole Genome and Small RNA Sequencing.

    Science.gov (United States)

    Urbarova, Ilona; Patel, Hardip; Forêt, Sylvain; Karlsen, Bård Ove; Jørgensen, Tor Erik; Hall-Spencer, Jason M; Johansen, Steinar D

    2018-02-01

    Cnidarians harbor a variety of small regulatory RNAs that include microRNAs (miRNAs) and PIWI-interacting RNAs (piRNAs), but detailed information is limited. Here, we report the identification and expression of novel miRNAs and putative piRNAs, as well as their genomic loci, in the symbiotic sea anemone Anemonia viridis. We generated a draft assembly of the A. viridis genome with putative size of 313 Mb that appeared to be composed of about 36% repeats, including known transposable elements. We detected approximately equal fractions of DNA transposons and retrotransposons. Deep sequencing of small RNA libraries constructed from A. viridis adults sampled at a natural CO2 gradient off Vulcano Island, Italy, identified 70 distinct miRNAs. Eight were homologous to previously reported miRNAs in cnidarians, whereas 62 appeared novel. Nine miRNAs were recognized as differentially expressed along the natural seawater pH gradient. We found a highly abundant and diverse population of piRNAs, with a substantial fraction showing ping-pong signatures. We identified nearly 22% putative piRNAs potentially targeting transposable elements within the A. viridis genome. The A. viridis genome appeared similar in size to that of other hexacorals with a very high divergence of transposable elements resembling that of the sea anemone genus Exaiptasia. The genome encodes and expresses a high number of small regulatory RNAs, which include novel miRNAs and piRNAs. Differentially expressed small RNAs along the seawater pH gradient indicated regulatory gene responses to environmental stressors. © The Author(s) 2018. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  12. RNA-based ovarian cancer research from 'a gene to systems biomedicine' perspective.

    Science.gov (United States)

    Gov, Esra; Kori, Medi; Arga, Kazim Yalcin

    2017-08-01

    Ovarian cancer remains the leading cause of death from a gynecologic malignancy, and treatment of this disease is harder than any other type of female reproductive cancer. Improvements in the diagnosis and development of novel and effective treatment strategies for complex pathophysiologies, such as ovarian cancer, require a better understanding of disease emergence and mechanisms of progression through systems medicine approaches. RNA-level analyses generate new information that can help in understanding the mechanisms behind disease pathogenesis, to identify new biomarkers and therapeutic targets and in new drug discovery. Whole RNA sequencing and coding and non-coding RNA expression array datasets have shed light on the mechanisms underlying disease progression and have identified mRNAs, miRNAs, and lncRNAs involved in ovarian cancer progression. In addition, the results from these analyses indicate that various signalling pathways and biological processes are associated with ovarian cancer. Here, we present a comprehensive literature review on RNA-based ovarian cancer research and highlight the benefits of integrative approaches within the systems biomedicine concept for future ovarian cancer research. We invite the ovarian cancer and systems biomedicine research fields to join forces to achieve the interdisciplinary caliber and rigor required to find real-life solutions to common, devastating, and complex diseases such as ovarian cancer. CAF: cancer-associated fibroblasts; COG: Cluster of Orthologous Groups; DEA: disease enrichment analysis; EOC: epithelial ovarian carcinoma; ESCC: oesophageal squamous cell carcinoma; GSI: gamma secretase inhibitor; GO: Gene Ontology; GSEA: gene set enrichment analyzes; HAS: Hungarian Academy of Sciences; lncRNAs: long non-coding RNAs; MAPK/ERK: mitogen-activated protein kinase/extracellular signal-regulated kinases; NGS: next-generation sequencing; ncRNAs: non-coding RNAs; OvC: ovarian cancer; PI3K

  13. Hardware Prototyping of Neural Network based Fetal Electrocardiogram Extraction

    Science.gov (United States)

    Hasan, M. A.; Reaz, M. B. I.

    2012-01-01

    The aim of this paper is to model the algorithm for Fetal ECG (FECG) extraction from composite abdominal ECG (AECG) using VHDL (Very High Speed Integrated Circuit Hardware Description Language) for FPGA (Field Programmable Gate Array) implementation. Artificial Neural Network that provides efficient and effective ways of separating FECG signal from composite AECG signal has been designed. The proposed method gives an accuracy of 93.7% for R-peak detection in FHR monitoring. The designed VHDL model is synthesized and fitted into Altera's Stratix II EP2S15F484C3 using the Quartus II version 8.0 Web Edition for FPGA implementation.

  14. RNAPattMatch: a web server for RNA sequence/structure motif detection based on pattern matching with flexible gaps

    Science.gov (United States)

    Drory Retwitzer, Matan; Polishchuk, Maya; Churkin, Elena; Kifer, Ilona; Yakhini, Zohar; Barash, Danny

    2015-01-01

    Searching for RNA sequence-structure patterns is becoming an essential tool for RNA practitioners. Novel discoveries of regulatory non-coding RNAs in targeted organisms and the motivation to find them across a wide range of organisms have prompted the use of computational RNA pattern matching as an enhancement to sequence similarity. State-of-the-art programs differ by the flexibility of patterns allowed as queries and by their simplicity of use. In particular—no existing method is available as a user-friendly web server. A general program that searches for RNA sequence-structure patterns is RNA Structator. However, it is not available as a web server and does not provide the option to allow flexible gap pattern representation with an upper bound of the gap length being specified at any position in the sequence. Here, we introduce RNAPattMatch, a web-based application that is user friendly and makes sequence/structure RNA queries accessible to practitioners of various background and proficiency. It also extends RNA Structator and allows a more flexible variable gaps representation, in addition to analysis of results using energy minimization methods. RNAPattMatch service is available at http://www.cs.bgu.ac.il/rnapattmatch. A standalone version of the search tool is also available to download at the site. PMID:25940619

  15. Nuclear factor 90 uses an ADAR2-like binding mode to recognize specific bases in dsRNA.

    Science.gov (United States)

    Jayachandran, Uma; Grey, Heather; Cook, Atlanta G

    2016-02-29

    Nuclear factors 90 and 45 (NF90 and NF45) form a protein complex involved in the post-transcriptional control of many genes in vertebrates. NF90 is a member of the dsRNA binding domain (dsRBD) family of proteins. RNA binding partners identified so far include elements in 3' untranslated regions of specific mRNAs and several non-coding RNAs. In NF90, a tandem pair of dsRBDs separated by a natively unstructured segment confers dsRNA binding activity. We determined a crystal structure of the tandem dsRBDs of NF90 in complex with a synthetic dsRNA. This complex shows surprising similarity to the tandem dsRBDs from an adenosine-to-inosine editing enzyme, ADAR2 in complex with a substrate RNA. Residues involved in unusual base-specific recognition in the minor groove of dsRNA are conserved between NF90 and ADAR2. These data suggest that, like ADAR2, underlying sequences in dsRNA may influence how NF90 recognizes its target RNAs. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  16. Confirming the RNAi-mediated mechanism of action of siRNA-based cancer therapeutics in mice.

    Science.gov (United States)

    Judge, Adam D; Robbins, Marjorie; Tavakoli, Iran; Levi, Jasna; Hu, Lina; Fronda, Anna; Ambegia, Ellen; McClintock, Kevin; MacLachlan, Ian

    2009-03-01

    siRNAs that specifically silence the expression of cancer-related genes offer a therapeutic approach in oncology. However, it remains critical to determine the true mechanism of their therapeutic effects. Here, we describe the preclinical development of chemically modified siRNA targeting the essential cell-cycle proteins polo-like kinase 1 (PLK1) and kinesin spindle protein (KSP) in mice. siRNA formulated in stable nucleic acid lipid particles (SNALP) displayed potent antitumor efficacy in both hepatic and subcutaneous tumor models. This was correlated with target gene silencing following a single intravenous administration that was sufficient to cause extensive mitotic disruption and tumor cell apoptosis. Our siRNA formulations induced no measurable immune response, minimizing the potential for nonspecific effects. Additionally, RNAi-specific mRNA cleavage products were found in tumor cells, and their presence correlated with the duration of target mRNA silencing. Histological biomarkers confirmed that RNAi-mediated gene silencing effectively inhibited the target's biological activity. This report supports an RNAi-mediated mechanism of action for siRNA antitumor effects, suggesting a new methodology for targeting other key genes in cancer development with siRNA-based therapeutics.

  17. In-vivo quantification of primary microRNA processing by Drosha with a luciferase based system

    International Nuclear Information System (INIS)

    Allegra, Danilo; Mertens, Daniel

    2011-01-01

    Research highlights: → Posttranscriptional regulation of miRNA processing is difficult to quantify. → Our in-vivo processing assay can quantify Drosha cleavage in live cells. → It is based on luciferase reporters fused with pri-miRNAs. → The assay validates the processing defect caused by a mutation in pri-16-1. → It is a sensitive method to quantify pri-miRNA cleavage by Drosha in live cells. -- Abstract: The RNAse III Drosha is responsible for the first step of microRNA maturation, the cleavage of primary miRNA to produce the precursor miRNA. Processing by Drosha is finely regulated and influences the amount of mature microRNA in a cell. We describe in the present work a method to quantify Drosha processing activity in-vivo, which is applicable to any microRNA. With respect to other methods for measuring Drosha activity, our system is faster and scalable, can be used with any cellular system and does not require cell sorting or use of radioactive isotopes. This system is useful to study regulation of Drosha activity in physiological and pathological conditions.

  18. REMap: Operon map of M. tuberculosis based on RNA sequence data.

    Science.gov (United States)

    Pelly, Shaaretha; Winglee, Kathryn; Xia, Fang Fang; Stevens, Rick L; Bishai, William R; Lamichhane, Gyanu

    2016-07-01

    A map of the transcriptional organization of genes of an organism is a basic tool that is necessary to understand and facilitate a more accurate genetic manipulation of the organism. Operon maps are largely generated by computational prediction programs that rely on gene conservation and genome architecture and may not be physiologically relevant. With the widespread use of RNA sequencing (RNAseq), the prediction of operons based on actual transcriptome sequencing rather than computational genomics alone is much needed. Here, we report a validated operon map of Mycobacterium tuberculosis, developed using RNAseq data from both the exponential and stationary phases of growth. At least 58.4% of M. tuberculosis genes are organized into 749 operons. Our prediction algorithm, REMap (RNA Expression Mapping of operons), considers the many cases of transcription coverage of intergenic regions, and avoids dependencies on functional annotation and arbitrary assumptions about gene structure. As a result, we demonstrate that REMap is able to more accurately predict operons, especially those that contain long intergenic regions or functionally unrelated genes, than previous operon prediction programs. The REMap algorithm is publicly available as a user-friendly tool that can be readily modified to predict operons in other bacteria. Copyright © 2016 Elsevier Ltd. All rights reserved.

  19. Comparação de três protocolos distintos para extração de RNA de amostras fixadas em formalina e emblocadas em parafina Comparison of three different protocols for extracting RNA from formalin-fixed paraffin-embedded tissues

    Directory of Open Access Journals (Sweden)

    Gisele Rodrigues Gouveia

    2011-12-01

    Full Text Available INTRODUÇÃO: Tecidos fixados em formalina e emblocados em parafina (FFEP são importantes fontes de amostras para estudos retrospectivos. Apesar de sua capacidade de preservação de proteínas e morfologia celular, a formalina interfere negativamente em testes de biologia molecular por fragmentar e modificar quimicamente os ácidos nucleicos, particularmente o ácido ribonucleico (RNA. OBJETIVO: Comparar a eficiência de três diferentes protocolos de extração de RNA para análise de expressão gênica de tecidos FFEP. MATERIAL E MÉTODOS: Amostras de linfonodo humano FEEP foram submetidas à extração de RNA utilizando-se os kits Ambion e Arcturus Bioscience e o método clássico de Trizol. Após a extração, o RNA foi quantificado e testado quanto à sua capacidade de amplificaç��o pela reação em cadeia da polimerase em tempo real (RT-PCR utilizando primers do gene endógeno gliceraldeído-3 fosfato desidrogenase (GAPDH. DISCUSSÃO/CONCLUSÃO: Todos os protocolos testados produziram quantidades adequadas e suficientes de RNA total, entretanto, somente os protocolos com uso dos kits Ambion e Arcturus produziram RNA capaz de ser amplificado pela PCR.INTRODUCTION: Formalin fixed paraffin embedded (FFPE tissues are an important sample source for retrospective studies. Despite its ability to preserve proteins and cell morphology, formalin hinders Molecular Biology tests once it fragments and chemically modifies nucleic acids, particularly RNA. OBJECTIVE: To compare the efficiency of three different RNA extraction protocols for gene expression analysis of FFEP tissues. MATERIAL AND METHODS: RNA was extracted from FFPE samples of human lymph by means of Ambion and Arcturus Bioscience kits and the classical Trizol method. After extraction, RNA was quantified and tested for amplification through real time polymerase chain reaction (RT-PCR using glyceraldehydes-3 phosphate dehydrogenase (GAPDH endogenous gene primers. DISCUSSION

  20. A green deep eutectic solvent-based aqueous two-phase system for protein extracting

    International Nuclear Information System (INIS)

    Xu, Kaijia; Wang, Yuzhi; Huang, Yanhua; Li, Na; Wen, Qian

    2015-01-01

    Highlights: • A strategy for the protein purification with a deep eutectic solvent(DES)-based aqueous two-phase system. • Choline chloride-glycerin DES was selected as the extraction solvent. • Bovine serum albumin and trypsin were used as the analytes. • Aggregation phenomenon was detected in the mechanism research. - Abstract: As a new type of green solvent, deep eutectic solvent (DES) has been applied for the extraction of proteins with an aqueous two-phase system (ATPS) in this work. Four kinds of choline chloride (ChCl)-based DESs were synthesized to extract bovine serum albumin (BSA), and ChCl-glycerol was selected as the suitable extraction solvent. Single factor experiments have been done to investigate the effects of the extraction process, including the amount of DES, the concentration of salt, the mass of protein, the shaking time, the temperature and PH value. Experimental results show 98.16% of the BSA could be extracted into the DES-rich phase in a single-step extraction under the optimized conditions. A high extraction efficiency of 94.36% was achieved, while the conditions were applied to the extraction of trypsin (Try). Precision, repeatability and stability experiments were studied and the relative standard deviations (RSD) of the extraction efficiency were 0.4246% (n = 3), 1.6057% (n = 3) and 1.6132% (n = 3), respectively. Conformation of BSA was not changed during the extraction process according to the investigation of UV–vis spectra, FT-IR spectra and CD spectra of BSA. The conductivity, dynamic light scattering (DLS) and transmission electron microscopy (TEM) were used to explore the mechanism of the extraction. It turned out that the formation of DES–protein aggregates play a significant role in the separation process. All the results suggest that ChCl-based DES-ATPS are supposed to have the potential to provide new possibilities in the separation of proteins

  1. Image segmentation-based robust feature extraction for color image watermarking

    Science.gov (United States)

    Li, Mianjie; Deng, Zeyu; Yuan, Xiaochen

    2018-04-01

    This paper proposes a local digital image watermarking method based on Robust Feature Extraction. The segmentation is achieved by Simple Linear Iterative Clustering (SLIC) based on which an Image Segmentation-based Robust Feature Extraction (ISRFE) method is proposed for feature extraction. Our method can adaptively extract feature regions from the blocks segmented by SLIC. This novel method can extract the most robust feature region in every segmented image. Each feature region is decomposed into low-frequency domain and high-frequency domain by Discrete Cosine Transform (DCT). Watermark images are then embedded into the coefficients in the low-frequency domain. The Distortion-Compensated Dither Modulation (DC-DM) algorithm is chosen as the quantization method for embedding. The experimental results indicate that the method has good performance under various attacks. Furthermore, the proposed method can obtain a trade-off between high robustness and good image quality.

  2. Isogeometric finite element data structures based on Bézier extraction of T-splines

    NARCIS (Netherlands)

    Scott, M.A.; Borden, M.J.; Verhoosel, C.V.; Sederberg, T.W.; Hughes, T.J.R.

    2011-01-01

    We develop finite element data structures for T-splines based on Bézier extraction generalizing our previous work for NURBS. As in traditional finite element analysis, the extracted Bézier elements are defined in terms of a fixed set of polynomial basis functions, the so-called Bernstein basis. The

  3. Human Milk MicroRNA and Total RNA Differ Depending on Milk Fractionation.

    Science.gov (United States)

    Alsaweed, Mohammed; Hepworth, Anna R; Lefèvre, Christophe; Hartmann, Peter E; Geddes, Donna T; Hassiotou, Foteini

    2015-10-01

    MicroRNA have been recently discovered in human milk signifying potentially important functions for both the lactating breast and the infant. Whilst human milk microRNA have started to be explored, little data exist on the evaluation of sample processing, and analysis to ensure that a full spectrum of microRNA can be obtained. Human milk comprises three main fractions: cells, skim milk, and lipids. Typically, the skim milk fraction has been measured in isolation despite evidence that the lipid fraction may contain more microRNA. This study aimed to standardize isolation of microRNA and total RNA from all three fractions of human milk to determine the most appropriate sampling and analysis procedure for future studies. Three different methods from eight commercially available kits were tested for their efficacy in extracting total RNA and microRNA from the lipid, skim, and cell fractions of human milk. Each fraction yielded different concentrations of RNA and microRNA, with the highest quantities found in the cell and lipid fractions, and the lowest in skim milk. The column-based phenol-free method was the most efficient extraction method for all three milk fractions. Two microRNAs were expressed and validated in the three milk fractions by qPCR using the three recommended extraction kits for each fraction. High expression levels were identified in the skim and lipid milk factions for these microRNAs. These results suggest that careful consideration of both the human milk sample preparation and extraction protocols should be made prior to embarking upon research in this area. © 2015 The Authors. Journal of Cellular Biochemistry Published by Wiley Periodicals, Inc.

  4. Text extraction method for historical Tibetan document images based on block projections

    Science.gov (United States)

    Duan, Li-juan; Zhang, Xi-qun; Ma, Long-long; Wu, Jian

    2017-11-01

    Text extraction is an important initial step in digitizing the historical documents. In this paper, we present a text extraction method for historical Tibetan document images based on block projections. The task of text extraction is considered as text area detection and location problem. The images are divided equally into blocks and the blocks are filtered by the information of the categories of connected components and corner point density. By analyzing the filtered blocks' projections, the approximate text areas can be located, and the text regions are extracted. Experiments on the dataset of historical Tibetan documents demonstrate the effectiveness of the proposed method.

  5. Optical Sensing Material for pH Detection based on the Use of Roselle Extract

    International Nuclear Information System (INIS)

    Nurul Huda Abd Karim; Musa Ahmad; Mohammad Osman Herman; Ahmad Mahir Mokhtar

    2008-01-01

    This research assessed the potential of natural colour extract of Hibiscus Sabdariffa L. (roselle) as sensing material.The pH sensor was developed based on the use of natural reddish colour in roselle calyx, delphinidin-3-sambubioside immobilised in a glass fibre filter paper. In free solution, roselle extract was characterised by using UV-visible spectrophotometer to study the effect of pH, extract concentration, response time, repeatability and photo stability. The study showed that natural colour extract can be used as sensing material for the development of an optical pH sensor. (author)

  6. Evaluation of DNA Extraction Methods Suitable for PCR-based Detection and Genotyping of Clostridium botulinum

    DEFF Research Database (Denmark)

    Auricchio, Bruna; Anniballi, Fabrizio; Fiore, Alfonsina

    2013-01-01

    in terms of cost, time, labor, and supplies. Eleven botulinum toxin–producing clostridia strains and 25 samples (10 food, 13 clinical, and 2 environmental samples) naturally contaminated with botulinum toxin–producing clostridia were used to compare 4 DNA extraction procedures: Chelex® 100 matrix, Phenol......Sufficient quality and quantity of extracted DNA is critical to detecting and performing genotyping of Clostridium botulinum by means of PCR-based methods. An ideal extraction method has to optimize DNA yield, minimize DNA degradation, allow multiple samples to be extracted, and be efficient...

  7. Efficient delivery of Notch1 siRNA to SKOV3 cells by cationic cholesterol derivative-based liposome

    Directory of Open Access Journals (Sweden)

    Zhao Y

    2016-10-01

    Full Text Available Yun-Chun Zhao,1 Li Zhang,2 Shi-Sen Feng,3 Lu Hong,3 Hai-Li Zheng,3 Li-Li Chen,4 Xiao-Ling Zheng,1 Yi-Qing Ye,1 Meng-Dan Zhao,1 Wen-Xi Wang,3 Cai-Hong Zheng1 1Pharmacy Department, Women’s Hospital, 2Pharmacy Department, The Second Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, People’s Republic of China; 3Department of Pharmaceutic Preparation, College of Pharmaceutical Science, Zhejiang University of Technology, Hangzhou, 4Department of Gynecologic Oncology, Women’s Hospital, School of Medicine, Zhejiang University, Hangzhou, People’s Republic of China Abstract: A novel cationic cholesterol derivative-based small interfering RNA (siRNA interference strategy was suggested to inhibit Notch1 activation in SKOV3 cells for the gene therapy of ovarian cancer. The cationic cholesterol derivative, N-(cholesterylhemisuccinoyl-amino-3-propyl-N, N-dimethylamine (DMAPA-chems liposome, was incubated with siRNA at different nitrogen-to-phosphate ratios to form stabilized, near-spherical siRNA/DMAPA-chems nanoparticles with sizes of 100–200 nm and zeta potentials of 40–50 mV. The siRNA/DMAPA-chems nanoparticles protected siRNA from nuclease degradation in 25% fetal bovine serum. The nanoparticles exhibited high cell uptake and Notch1 gene knockdown efficiency in SKOV3 cells at an nitrogen-to-phosphate ratio of 100 and an siRNA concentration of 50 nM. They also inhibited the growth and promoted the apoptosis of SKOV3 cells. These results may provide the potential for using cationic cholesterol derivatives as efficient nonviral siRNA carriers for the suppression of Notch1 activation in ovarian cancer cells. Keywords: siRNA, cationic cholesterol derivative, Notch1, ovarian cancer cells

  8. Polyethylenimine-based polyplex delivery of self-replicating RNA vaccines.

    Science.gov (United States)

    Démoulins, Thomas; Milona, Panagiota; Englezou, Pavlos C; Ebensen, Thomas; Schulze, Kai; Suter, Rolf; Pichon, Chantal; Midoux, Patrick; Guzmán, Carlos A; Ruggli, Nicolas; McCullough, Kenneth C

    2016-04-01

    Self-amplifying replicon RNA (RepRNA) are large molecules (12-14 kb); their self-replication amplifies mRNA template numbers, affording several rounds of antigen production, effectively increasing vaccine antigen payloads. Their sensitivity to RNase-sensitivity and inefficient uptake by dendritic cells (DCs) - absolute requirements for vaccine design - were tackled by condensing RepRNA into synthetic, nanoparticulate, polyethylenimine (PEI)-polyplex delivery vehicles. Polyplex-delivery formulations for small RNA molecules cannot be transferred to RepRNA due to its greater size and complexity; the N:P charge ratio and impact of RepRNA folding would influence polyplex condensation, post-delivery decompaction and the cytosolic release essential for RepRNA translation. Polyplex-formulations proved successful for delivery of RepRNA encoding influenza virus hemagglutinin and nucleocapsid to DCs. Cytosolic translocation was facilitated, leading to RepRNA translation. This efficacy was confirmed in vivo, inducing both humoral and cellular immune responses. Accordingly, this paper describes the first PEI-polyplexes providing efficient delivery of the complex and large, self-amplifying RepRNA vaccines. The use of self-amplifying replicon RNA (RepRNA) to increase vaccine antigen payloads can potentially be useful in effective vaccine design. Nonetheless, its use is limited by the degradation during the uptake process. Here, the authors attempted to solve this problem by packaging RepRNA using polyethylenimine (PEI)-polyplex delivery vehicles. The efficacy was confirmed in vivo by the appropriate humoral and cellular immune responses. This novel delivery method may prove to be very useful for future vaccine design. Copyright © 2015 Elsevier Inc. All rights reserved.

  9. Stability of non-Watson-Crick G-A/A-G base pair in synthetic DNA and RNA oligonucleotides.

    Science.gov (United States)

    Ito, Yuko; Sone, Yumiko; Mizutani, Takaharu

    2004-03-01

    A non-Watson-Crick G-A/A-G base pair is found in SECIS (selenocysteine-insertion sequence) element in the 3'-untranslated region of Se-protein mRNAs and in the functional site of the hammerhead ribozyme. We studied the stability of G-A/A-G base pair (bold) in 17mer GT(U)GACGGAAACCGGAAC synthetic DNA and RNA oligonucleotides by thermal melting experiments and gel electrophoresis. The measured Tm value of DNA oligonucleotide having G-A/A-G pair showed an intermediate value (58 degrees C) between that of Watson-Crick G-C/C-G base pair (75 degrees C) and that of G-G/A-A of non-base-pair (40 degrees C). Similar thermal melting patterns were obtained with RNA oligonucleotides. This result indicates that the secondary structure of oligonucleotide having G-A/A-G base pair is looser than that of the G-C type Watson-Crick base pair. In the comparison between RNA and DNA having G-A/A-G base pair, the Tm value of the RNA oligonucleotide was 11 degrees C lower than that of DNA, indicating that DNA has a more rigid structure than RNA. The stained pattern of oligonucleotide on polyacrylamide gel clarified that the mobility of the DNA oligonucleotide G-A/A-G base pair changed according to the urea concentration from the rigid state (near the mobility of G-C/C-G oligonucleotide) in the absence of urea to the random state (near the mobility of G-G/A-A oligonucleotide) in 7 M urea. However, the RNA oligonucleotide with G-A/A-G pair moved at an intermediate mobility between that of oligonucleotide with G-C/C-G and of the oligonucleotide with G-G/A-A, and the mobility pattern did not depend on urea concentration. Thus, DNA and RNA oligonucleotides with the G-A/A-G base pair showed a pattern indicating an intermediate structure between the rigid Watson-Crick base pair and the random structure of non-base pair. RNA with G-A/A-G base pair has the intermediate structure not influenced by urea concentration. Finally, this study indicated that the intermediate rigidity imparted by Non

  10. A rapid extraction of landslide disaster information research based on GF-1 image

    Science.gov (United States)

    Wang, Sai; Xu, Suning; Peng, Ling; Wang, Zhiyi; Wang, Na

    2015-08-01

    In recent years, the landslide disasters occurred frequently because of the seismic activity. It brings great harm to people's life. It has caused high attention of the state and the extensive concern of society. In the field of geological disaster, landslide information extraction based on remote sensing has been controversial, but high resolution remote sensing image can improve the accuracy of information extraction effectively with its rich texture and geometry information. Therefore, it is feasible to extract the information of earthquake- triggered landslides with serious surface damage and large scale. Taking the Wenchuan county as the study area, this paper uses multi-scale segmentation method to extract the landslide image object through domestic GF-1 images and DEM data, which uses the estimation of scale parameter tool to determine the optimal segmentation scale; After analyzing the characteristics of landslide high-resolution image comprehensively and selecting spectrum feature, texture feature, geometric features and landform characteristics of the image, we can establish the extracting rules to extract landslide disaster information. The extraction results show that there are 20 landslide whose total area is 521279.31 .Compared with visual interpretation results, the extraction accuracy is 72.22%. This study indicates its efficient and feasible to extract earthquake landslide disaster information based on high resolution remote sensing and it provides important technical support for post-disaster emergency investigation and disaster assessment.

  11. Polymerase-free measurement of microRNA-122 with single base specificity using single molecule arrays: Detection of drug-induced liver injury.

    Directory of Open Access Journals (Sweden)

    David M Rissin

    Full Text Available We have developed a single probe method for detecting microRNA from human serum using single molecule arrays, with sequence specificity down to a single base, and without the use of amplification by polymerases. An abasic peptide nucleic acid (PNA probe-containing a reactive amine instead of a nucleotide at a specific position in the sequence-for detecting a microRNA was conjugated to superparamagnetic beads. These beads were incubated with a sample containing microRNA, a biotinylated reactive nucleobase-containing an aldehyde group-that was complementary to the missing base in the probe sequence, and a reducing agent. When a target molecule with an exact match in sequence hybridized to the capture probe, the reactive nucleobase was covalently attached to the backbone of the probe by a dynamic covalent chemical reaction. Single molecules of the biotin-labeled probe were then labeled with streptavidin-β-galactosidase (SβG, the beads were resuspended in a fluorogenic enzyme substrate, loaded into an array of femtoliter wells, and sealed with oil. The array was imaged fluorescently to determine which beads were associated with single enzymes, and the average number of enzymes per bead was determined. The assay had a limit of detection of 500 fM, approximately 500 times more sensitive than a corresponding analog bead-based assay, with target specificity down to a single base mis-match. This assay was used to measure microRNA-122 (miR-122-an established biomarker of liver toxicity-extracted from the serum of patients who had acute liver injury due to acetaminophen, and control healthy patients. All patients with liver injury had higher levels of miR-122 in their serum compared to controls, and the concentrations measured correlated well with those determined using RT-qPCR. This approach allows rapid quantification of circulating microRNA with single-based specificity and a limit of quantification suitable for clinical use.

  12. High Quality RNA Isolation from Leaf, Shell, Root Tissues and Callus of Hazelnut (Corylus avellana L.

    Directory of Open Access Journals (Sweden)

    Hossein Khosravi

    2017-12-01

    Full Text Available Extraction of high quality RNA is a critical step in molecular genetics studies. Hazelnut is one of the most important nuts plants in the world. The presence of the taxol and other taxanes in hazelnut plant necessitates explaining their biosynthesis pathway and identifying the candidate genes. Therefore, an easy and practical method is necessary for RNA extraction from hazelnuts. Hazelnut has high levels of phenolic compounds. High amounts of polyphenolic and polysaccharide compounds in plants could be causing problems in RNA extraction procedures.  To avoid these problems, a simple and efficient method can be used based on cetyltrimethylammonium bromide (CTAB extraction buffer and lithium chloride for extraction of high quality RNA from different parts of hazelnut plant. Using this method, a high-quality RNA sample (light absorbed in the A260/A280 was 2.04

  13. Fusion of Pixel-based and Object-based Features for Road Centerline Extraction from High-resolution Satellite Imagery

    Directory of Open Access Journals (Sweden)

    CAO Yungang

    2016-10-01

    Full Text Available A novel approach for road centerline extraction from high spatial resolution satellite imagery is proposed by fusing both pixel-based and object-based features. Firstly, texture and shape features are extracted at the pixel level, and spectral features are extracted at the object level based on multi-scale image segmentation maps. Then, extracted multiple features are utilized in the fusion framework of Dempster-Shafer evidence theory to roughly identify the road network regions. Finally, an automatic noise removing algorithm combined with the tensor voting strategy is presented to accurately extract the road centerline. Experimental results using high-resolution satellite imageries with different scenes and spatial resolutions showed that the proposed approach compared favorably with the traditional methods, particularly in the aspect of eliminating the salt noise and conglutination phenomenon.

  14. The optimal extraction of feature algorithm based on KAZE

    Science.gov (United States)

    Yao, Zheyi; Gu, Guohua; Qian, Weixian; Wang, Pengcheng

    2015-10-01

    As a novel method of 2D features extraction algorithm over the nonlinear scale space, KAZE provide a special method. However, the computation of nonlinear scale space and the construction of KAZE feature vectors are more expensive than the SIFT and SURF significantly. In this paper, the given image is used to build the nonlinear space up to a maximum evolution time through the efficient Additive Operator Splitting (AOS) techniques and the variable conductance diffusion. Changing the parameter can improve the construction of nonlinear scale space and simplify the image conductivities for each dimension space, with the predigest computation. Then, the detection for points of interest can exhibit a maxima of the scale-normalized determinant with the Hessian response in the nonlinear scale space. At the same time, the detection of feature vectors is optimized by the Wavelet Transform method, which can avoid the second Gaussian smoothing in the KAZE Features and cut down the complexity of the algorithm distinctly in the building and describing vectors steps. In this way, the dominant orientation is obtained, similar to SURF, by summing the responses within a sliding circle segment covering an angle of π/3 in the circular area of radius 6σ with a sampling step of size σ one by one. Finally, the extraction in the multidimensional patch at the given scale, centered over the points of interest and rotated to align its dominant orientation to a canonical direction, is able to simplify the description of feature by reducing the description dimensions, just as the PCA-SIFT method. Even though the features are somewhat more expensive to compute than SIFT due to the construction of nonlinear scale space, but compared to SURF, the result revels a step forward in performance in detection, description and application against the previous ways by the following contrast experiments.

  15. Biofluid-based microRNA Biomarkers for Parkinsons Disease: an Overview and Update

    Directory of Open Access Journals (Sweden)

    Sapana Shinde

    2015-02-01

    Full Text Available Parkinson's disease (PD is a highly debilitating motor disorder and is the second most common neurodegenerative disease after Alzheimer's disease. Its current method of diagnosis mainly relies on subjective clinical rating scales in the presence of clinical motor features. Early detection of PD is a known challenge as neuronal cell death may range from 50% to 80% when a patient is first diagnosed with PD. Therefore, there is an urgent need to identify and develop biomarkers for early detection of this progressive disease. This mini review focuses on the recent developments of biofluid-based microRNAs (miRNAs as molecular biomarkers for PD. A comprehensive list of miRNA biomarkers found in blood, plasma, serum, and cerebral spinal fluid is presented. Challenges and future perspectives of using these PD-related molecular biomarkers in a “real-world” clinical setting are also discussed.

  16. MKRMDA: multiple kernel learning-based Kronecker regularized least squares for MiRNA-disease association prediction.

    Science.gov (United States)

    Chen, Xing; Niu, Ya-Wei; Wang, Guang-Hui; Yan, Gui-Ying

    2017-12-12

    Recently, as the research of microRNA (miRNA) continues, there are plenty of experimental evidences indicating that miRNA could be associated with various human complex diseases development and progression. Hence, it is necessary and urgent to pay more attentions to the relevant study of predicting diseases associated miRNAs, which may be helpful for effective prevention, diagnosis and treatment of human diseases. Especially, constructing computational methods to predict potential miRNA-disease associations is worthy of more studies because of the feasibility and effectivity. In this work, we developed a novel computational model of multiple kernels learning-based Kronecker regularized least squares for MiRNA-disease association prediction (MKRMDA), which could reveal potential miRNA-disease associations by automatically optimizing the combination of multiple kernels for disease and miRNA. MKRMDA obtained AUCs of 0.9040 and 0.8446 in global and local leave-one-out cross validation, respectively. Meanwhile, MKRMDA achieved average AUCs of 0.8894 ± 0.0015 in fivefold cross validation. Furthermore, we conducted three different kinds of case studies on some important human cancers for further performance evaluation. In the case studies of colonic cancer, esophageal cancer and lymphoma based on known miRNA-disease associations in HMDDv2.0 database, 76, 94 and 88% of the corresponding top 50 predicted miRNAs were confirmed by experimental reports, respectively. In another two kinds of case studies for new diseases without any known associated miRNAs and diseases only with known associations in HMDDv1.0 database, the verified ratios of two different cancers were 88 and 94%, respectively. All the results mentioned above adequately showed the reliable prediction ability of MKRMDA. We anticipated that MKRMDA could serve to facilitate further developments in the field and the follow-up investigations by biomedical researchers.

  17. MicroRNA prediction using a fixed-order Markov model based on the secondary structure pattern.

    Directory of Open Access Journals (Sweden)

    Wei Shen

    Full Text Available Predicting miRNAs is an arduous task, due to the diversity of the precursors and complexity of enzyme processes. Although several prediction approaches have reached impressive performances, few of them could achieve a full-function recognition of mature miRNA directly from the candidate hairpins across species. Therefore, researchers continue to seek a more powerful model close to biological recognition to miRNA structure. In this report, we describe a novel miRNA prediction algorithm, known as FOMmiR, using a fixed-order Markov model based on the secondary structural pattern. For a training dataset containing 809 human pre-miRNAs and 6441 human pseudo-miRNA hairpins, the model's parameters were defined and evaluated. The results showed that FOMmiR reached 91% accuracy on the human dataset through 5-fold cross-validation. Moreover, for the independent test datasets, the FOMmiR presented an outstanding prediction in human and other species including vertebrates, Drosophila, worms and viruses, even plants, in contrast to the well-known algorithms and models. Especially, the FOMmiR was not only able to distinguish the miRNA precursors from the hairpins, but also locate the position and strand of the mature miRNA. Therefore, this study provides a new generation of miRNA prediction algorithm, which successfully realizes a full-function recognition of the mature miRNAs directly from the hairpin sequences. And it presents a new understanding of the biological recognition based on the strongest signal's location detected by FOMmiR, which might be closely associated with the enzyme cleavage mechanism during the miRNA maturation.

  18. A simple experimental arrangement for measuring the vapour pressures and sublimation enthalpies by the Knudsen effusion method: Application to DNA and RNA bases

    International Nuclear Information System (INIS)

    Barros, A.L.F. de; Medina, A.; Zappa, F.; Pereira, J.M.; Bessa, E.; Martins, M.H.P.; Coelho, L.F.S.; Wolff, W.; Castro Faria, N.V. de

    2006-01-01

    We measured the vapour pressure of several DNA and RNA bases-uracil, adenine, guanine, thymine and cytosine-in the 300-450 K range. In each case the sample mass loss rate was measured as function of temperature with a simple setup consisting of a commercial film deposition system and a homemade oven. Afterwards vapour pressure values were extracted from these data using the Knudsen effusion method. Sublimation enthalpy values, obtained from vapour pressure data by applying the Clausius-Clapeyron equation, are in very good agreement with literature values. The results suggest that crystal-based film thickness monitors may be useful in on-line cross-section measurements, monitoring the gas target thickness. They also show the viability of using this oven for producing a biomolecular gas target

  19. An automated procedure for covariation-based detection of RNA structure

    International Nuclear Information System (INIS)

    Winker, S.; Overbeek, R.; Woese, C.R.; Olsen, G.J.; Pfluger, N.

    1989-12-01

    This paper summarizes our investigations into the computational detection of secondary and tertiary structure of ribosomal RNA. We have developed a new automated procedure that not only identifies potential bondings of secondary and tertiary structure, but also provides the covariation evidence that supports the proposed bondings, and any counter-evidence that can be detected in the known sequences. A small number of previously unknown bondings have been detected in individual RNA molecules (16S rRNA and 7S RNA) through the use of our automated procedure. Currently, we are systematically studying mitochondrial rRNA. Our goal is to detect tertiary structure within 16S rRNA and quaternary structure between 16S and 23S rRNA. Our ultimate hope is that automated covariation analysis will contribute significantly to a refined picture of ribosome structure. Our colleagues in biology have begun experiments to test certain hypotheses suggested by an examination of our program's output. These experiments involve sequencing key portions of the 23S ribosomal RNA for species in which the known 16S ribosomal RNA exhibits variation (from the dominant pattern) at the site of a proposed bonding. The hope is that the 23S ribosomal RNA of these species will exhibit corresponding complementary variation or generalized covariation. 24 refs

  20. An automated procedure for covariation-based detection of RNA structure

    Energy Technology Data Exchange (ETDEWEB)

    Winker, S.; Overbeek, R.; Woese, C.R.; Olsen, G.J.; Pfluger, N.

    1989-12-01

    This paper summarizes our investigations into the computational detection of secondary and tertiary structure of ribosomal RNA. We have developed a new automated procedure that not only identifies potential bondings of secondary and tertiary structure, but also provides the covariation evidence that supports the proposed bondings, and any counter-evidence that can be detected in the known sequences. A small number of previously unknown bondings have been detected in individual RNA molecules (16S rRNA and 7S RNA) through the use of our automated procedure. Currently, we are systematically studying mitochondrial rRNA. Our goal is to detect tertiary structure within 16S rRNA and quaternary structure between 16S and 23S rRNA. Our ultimate hope is that automated covariation analysis will contribute significantly to a refined picture of ribosome structure. Our colleagues in biology have begun experiments to test certain hypotheses suggested by an examination of our program's output. These experiments involve sequencing key portions of the 23S ribosomal RNA for species in which the known 16S ribosomal RNA exhibits variation (from the dominant pattern) at the site of a proposed bonding. The hope is that the 23S ribosomal RNA of these species will exhibit corresponding complementary variation or generalized covariation. 24 refs.

  1. Viral RNA testing and automation on the bead-based CBNE detection microsystem.

    Energy Technology Data Exchange (ETDEWEB)

    Galambos, Paul C.; Bourdon, Christopher Jay; Farrell, Cara M.; Rossito, Paul (University of California at Davis); McClain, Jaime L.; Derzon, Mark Steven; Cullor, James Sterling (University of California at Davis); Rahimian, Kamayar

    2008-09-01

    We developed prototype chemistry for nucleic acid hybridization on our bead-based diagnostics platform and we established an automatable bead handling protocol capable of 50 part-per-billion (ppb) sensitivity. We are working towards a platform capable of parallel, rapid (10 minute), raw sample testing for orthogonal (in this case nucleic acid and immunoassays) identification of biological (and other) threats in a single sensor microsystem. In this LDRD we developed the nucleic acid chemistry required for nucleic acid hybridization. Our goal is to place a non-cell associated RNA virus (Bovine Viral Diarrhea, BVD) on the beads for raw sample testing. This key pre-requisite to showing orthogonality (nucleic acid measurements can be performed in parallel with immunoassay measurements). Orthogonal detection dramatically reduces false positives. We chose BVD because our collaborators (UC-Davis) can supply samples from persistently infected animals; and because proof-of-concept field testing can be performed with modification of the current technology platform at the UC Davis research station. Since BVD is a cattle-prone disease this research dovetails with earlier immunoassay work on Botulinum toxin simulant testing in raw milk samples. Demonstration of BVD RNA detection expands the repertoire of biological macromolecules that can be adapted to our bead-based detection. The resources of this late start LDRD were adequate to partially demonstrate the conjugation of the beads to the nucleic acids. It was never expected to be adequate for a full live virus test but to motivate that additional investment. In addition, we were able to reduce the LOD (Limit of Detection) for the botulinum toxin stimulant to 50 ppb from the earlier LOD of 1 ppm. A low LOD combined with orthogonal detection provides both low false negatives and low false positives. The logical follow-on steps to this LDRD research are to perform live virus identification as well as concurrent nucleic acid and

  2. A novel murmur-based heart sound feature extraction technique using envelope-morphological analysis

    Science.gov (United States)

    Yao, Hao-Dong; Ma, Jia-Li; Fu, Bin-Bin; Wang, Hai-Yang; Dong, Ming-Chui

    2015-07-01

    Auscultation of heart sound (HS) signals serves as an important primary approach to diagnose cardiovascular diseases (CVDs) for centuries. Confronting the intrinsic drawbacks of traditional HS auscultation, computer-aided automatic HS auscultation based on feature extraction technique has witnessed explosive development. Yet, most existing HS feature extraction methods adopt acoustic or time-frequency features which exhibit poor relationship with diagnostic information, thus restricting the performance of further interpretation and analysis. Tackling such a bottleneck problem, this paper innovatively proposes a novel murmur-based HS feature extraction method since murmurs contain massive pathological information and are regarded as the first indications of pathological occurrences of heart valves. Adapting discrete wavelet transform (DWT) and Shannon envelope, the envelope-morphological characteristics of murmurs are obtained and three features are extracted accordingly. Validated by discriminating normal HS and 5 various abnormal HS signals with extracted features, the proposed method provides an attractive candidate in automatic HS auscultation.

  3. SRMDAP: SimRank and Density-Based Clustering Recommender Model for miRNA-Disease Association Prediction

    Directory of Open Access Journals (Sweden)

    Xiaoying Li

    2018-01-01

    Full Text Available Aberrant expression of microRNAs (miRNAs can be applied for the diagnosis, prognosis, and treatment of human diseases. Identifying the relationship between miRNA and human disease is important to further investigate the pathogenesis of human diseases. However, experimental identification of the associations between diseases and miRNAs is time-consuming and expensive. Computational methods are efficient approaches to determine the potential associations between diseases and miRNAs. This paper presents a new computational method based on the SimRank and density-based clustering recommender model for miRNA-disease associations prediction (SRMDAP. The AUC of 0.8838 based on leave-one-out cross-validation and case studies suggested the excellent performance of the SRMDAP in predicting miRNA-disease associations. SRMDAP could also predict diseases without any related miRNAs and miRNAs without any related diseases.

  4. A novel method of predicting microRNA-disease associations based on microRNA, disease, gene and environment factor networks.

    Science.gov (United States)

    Peng, Wei; Lan, Wei; Zhong, Jiancheng; Wang, Jianxin; Pan, Yi

    2017-07-15

    MicroRNAs have been reported to have close relationship with diseases due to their deregulation of the expression of target mRNAs. Detecting disease-related microRNAs is helpful for disease therapies. With the development of high throughput experimental techniques, a large number of microRNAs have been sequenced. However, it is still a big challenge to identify which microRNAs are related to diseases. Recently, researchers are interesting in combining multiple-biological information to identify the associations between microRNAs and diseases. In this work, we have proposed a novel method to predict the microRNA-disease associations based on four biological properties. They are microRNA, disease, gene and environment factor. Compared with previous methods, our method makes predictions not only by using the prior knowledge of associations among microRNAs, disease, environment factors and genes, but also by using the internal relationship among these biological properties. We constructed four biological networks based on the similarity of microRNAs, diseases, environment factors and genes, respectively. Then random walking was implemented on the four networks unequally. In the walking course, the associations can be inferred from the neighbors in the same networks. Meanwhile the association information can be transferred from one network to another. The results of experiment showed that our method achieved better prediction performance than other existing state-of-the-art methods. Copyright © 2017 Elsevier Inc. All rights reserved.

  5. Parameter extraction and estimation based on the PV panel outdoor ...

    African Journals Online (AJOL)

    The experimental data obtained are validated and compared with the estimated results obtained through simulation based on the manufacture's data sheet. The simulation is based on the Newton-Raphson iterative method in MATLAB environment. This approach aids the computation of the PV module's parameters at any ...

  6. A green deep eutectic solvent-based aqueous two-phase system for protein extracting.

    Science.gov (United States)

    Xu, Kaijia; Wang, Yuzhi; Huang, Yanhua; Li, Na; Wen, Qian

    2015-03-15

    As a new type of green solvent, deep eutectic solvent (DES) has been applied for the extraction of proteins with an aqueous two-phase system (ATPS) in this work. Four kinds of choline chloride (ChCl)-based DESs were synthesized to extract bovine serum albumin (BSA), and ChCl-glycerol was selected as the suitable extraction solvent. Single factor experiments have been done to investigate the effects of the extraction process, including the amount of DES, the concentration of salt, the mass of protein, the shaking time, the temperature and PH value. Experimental results show 98.16% of the BSA could be extracted into the DES-rich phase in a single-step extraction under the optimized conditions. A high extraction efficiency of 94.36% was achieved, while the conditions were applied to the extraction of trypsin (Try). Precision, repeatability and stability experiments were studied and the relative standard deviations (RSD) of the extraction efficiency were 0.4246% (n=3), 1.6057% (n=3) and 1.6132% (n=3), respectively. Conformation of BSA was not changed during the extraction process according to the investigation of UV-vis spectra, FT-IR spectra and CD spectra of BSA. The conductivity, dynamic light scattering (DLS) and transmission electron microscopy (TEM) were used to explore the mechanism of the extraction. It turned out that the formation of DES-protein aggregates play a significant role in the separation process. All the results suggest that ChCl-based DES-ATPS are supposed to have the potential to provide new possibilities in the separation of proteins. Copyright © 2015 Elsevier B.V. All rights reserved.

  7. Diversity of ribosomal 16S DNA- and RNA-based bacterial community in an office building drinking water system.

    Science.gov (United States)

    Inkinen, J; Jayaprakash, B; Santo Domingo, J W; Keinänen-Toivola, M M; Ryu, H; Pitkänen, T

    2016-06-01

    Next-generation sequencing of 16S ribosomal RNA genes (rDNA) and ribosomal RNA (rRNA) was used to characterize water and biofilm microbiome collected from a drinking water distribution system of an office building after its first year of operation. The total bacterial community (rDNA) and active bacterial members (rRNA) sequencing databases were generated by Illumina MiSeq PE250 platform. As estimated by Chao1 index, species richness in cold water system was lower (180-260) in biofilms (Sphingomonas spp., Methylobacterium spp., Limnohabitans spp., Rhizobiales order) than in waters (250-580), (also Methylotenera spp.) (P = 0·005, n = 20). Similarly species richness (Chao1) was slightly higher (210-580) in rDNA libraries compared to rRNA libraries (150-400; P = 0·054, n = 24). Active Mycobacterium spp. was found in cross-linked polyethylene (PEX), but not in corresponding copper pipeline biofilm. Nonpathogenic Legionella spp. was found in rDNA libraries but not in rRNA libraries. Microbial communities differed between water and biofilms, between cold and hot water systems, locations in the building and between water rRNA and rDNA libraries, as shown by clear clusters in principal component analysis (PcoA). By using the rRNA method, we found that not all bacterial community members were active (e.g. Legionella spp.), whereas other members showed increased activity in some locations; for example, Pseudomonas spp. in hot water circulations' biofilm and order Rhizobiales and Limnohabitans spp. in stagnated locations' water and biofilm. rRNA-based methods may be better than rDNA-based methods for evaluating human health implications as rRNA methods can be used to describe the active bacterial fraction. This study indicates that copper as a pipeline material might have an adverse impact on the occurrence of Mycobacterium spp. The activity of Legionella spp. maybe questionable when detected solely by using DNA-based methods. © 2016 The Society for Applied

  8. Hybridization-based reconstruction of small non-coding RNA transcripts from deep sequencing data.

    Science.gov (United States)

    Ragan, Chikako; Mowry, Bryan J; Bauer, Denis C

    2012-09-01

    Recent advances in RNA sequencing technology (RNA-Seq) enables comprehensive profiling of RNAs by producing millions of short sequence reads from size-fractionated RNA libraries. Although conventional tools for detecting and distinguishing non-coding RNAs (ncRNAs) from reference-genome data can be applied to sequence data, ncRNA detection can be improved by harnessing the full information content provided by this new technology. Here we present NorahDesk, the first unbiased and universally applicable method for small ncRNAs detection from RNA-Seq data. NorahDesk utilizes the coverage-distribution of small RNA sequence data as well as thermodynamic assessments of secondary structure to reliably predict and annotate ncRNA classes. Using publicly available mouse sequence data from brain, skeletal muscle, testis and ovary, we evaluated our method with an emphasis on the performance for microRNAs (miRNAs) and piwi-interacting small RNA (piRNA). We compared our method with Dario and mirDeep2 and found that NorahDesk produces longer transcripts with higher read coverage. This feature makes it the first method particularly suitable for the prediction of both known and novel piRNAs.

  9. Evaluating the effect of disturbed ensemble distributions on SCFG based statistical sampling of RNA secondary structures

    Directory of Open Access Journals (Sweden)

    Scheid Anika

    2012-07-01

    Full Text Available Abstract Background Over the past years, statistical and Bayesian approaches have become increasingly appreciated to address the long-standing problem of computational RNA structure prediction. Recently, a novel probabilistic method for the prediction of RNA secondary structures from a single sequence has been studied which is based on generating statistically representative and reproducible samples of the entire ensemble of feasible structures for a particular input sequence. This method samples the possible foldings from a distribution implied by a sophisticated (traditional or length-dependent stochastic context-free grammar (SCFG that mirrors the standard thermodynamic model applied in modern physics-based prediction algorithms. Specifically, that grammar represents an exact probabilistic counterpart to the energy model underlying the Sfold software, which employs a sampling extension of the partition function (PF approach to produce statistically representative subsets of the Boltzmann-weighted ensemble. Although both sampling approaches have the same worst-case time and space complexities, it has been indicated that they differ in performance (both with respect to prediction accuracy and quality of generated samples, where neither of these two competing approaches generally outperforms the other. Results In this work, we will consider the SCFG based approach in order to perform an analysis on how the quality of generated sample sets and the corresponding prediction accuracy changes when different degrees of disturbances are incorporated into the needed sampling probabilities. This is motivated by the fact that if the results prove to be resistant to large errors on the distinct sampling probabilities (compared to the exact ones, then it will be an indication that these probabilities do not need to be computed exactly, but it may be sufficient and more efficient to approximate them. Thus, it might then be possible to decrease the worst

  10. Detection of Sialic Acid-Utilising Bacteria in a Caecal Community Batch Culture Using RNA-Based Stable Isotope Probing

    Directory of Open Access Journals (Sweden)

    Wayne Young

    2015-03-01

    Full Text Available Sialic acids are monosaccharides typically found on cell surfaces and attached to soluble proteins, or as essential components of ganglioside structures that play a critical role in brain development and neural transmission. Human milk also contains sialic acid conjugated to oligosaccharides, glycolipids, and glycoproteins. These nutrients can reach the large bowel where they may be metabolised by the microbiota. However, little is known about the members of the microbiota involved in this function. To identify intestinal bacteria that utilise sialic acid within a complex intestinal community, we cultured the caecal microbiota from piglets in the presence of 13C-labelled sialic acid. Using RNA-based stable isotope probing, we identified bacteria that consumed 13C-sialic acid by fractionating total RNA in isopycnic buoyant density gradients followed by 16S rRNA gene analysis. Addition of sialic acid caused significant microbial community changes. A relative rise in Prevotella and Lactobacillus species was accompanied by a corresponding reduction in the genera Escherichia/Shigella, Ruminococcus and Eubacterium. Inspection of isotopically labelled RNA sequences suggests that the labelled sialic acid was consumed by a wide range of bacteria. However, species affiliated with the genus Prevotella were clearly identified as the most prolific users, as solely their RNA showed significantly higher relative shares among the most labelled RNA species. Given the relevance of sialic acid in nutrition, this study contributes to a better understanding of their microbial transformation in the intestinal tract with potential implications for human health.

  11. A Method of Road Extraction from High-resolution Remote Sensing Images Based on Shape Features

    Directory of Open Access Journals (Sweden)

    LEI Xiaoqi

    2016-02-01

    Full Text Available Road extraction from high-resolution remote sensing image is an important and difficult task.Since remote sensing images include complicated information,the methods that extract roads by spectral,texture and linear features have certain limitations.Also,many methods need human-intervention to get the road seeds(semi-automatic extraction,which have the great human-dependence and low efficiency.The road-extraction method,which uses the image segmentation based on principle of local gray consistency and integration shape features,is proposed in this paper.Firstly,the image is segmented,and then the linear and curve roads are obtained by using several object shape features,so the method that just only extract linear roads are rectified.Secondly,the step of road extraction is carried out based on the region growth,the road seeds are automatic selected and the road network is extracted.Finally,the extracted roads are regulated by combining the edge information.In experiments,the images that including the better gray uniform of road and the worse illuminated of road surface were chosen,and the results prove that the method of this study is promising.

  12. ASAP: a web-based platform for the analysis and interactive visualization of single-cell RNA-seq data.

    Science.gov (United States)

    Gardeux, Vincent; David, Fabrice P A; Shajkofci, Adrian; Schwalie, Petra C; Deplancke, Bart

    2017-10-01

    Single-cell RNA-sequencing (scRNA-seq) allows whole transcriptome profiling of thousands of individual cells, enabling the molecular exploration of tissues at the cellular level. Such analytical capacity is of great interest to many research groups in the world, yet these groups often lack the expertise to handle complex scRNA-seq datasets. We developed a fully integrated, web-based platform aimed at the complete analysis of scRNA-seq data post genome alignment: from the parsing, filtering and normalization of the input count data files, to the visual representation of the data, identification of cell clusters, differentially expressed genes (including cluster-specific marker genes), and functional gene set enrichment. This Automated Single-cell Analysis Pipeline (ASAP) combines a wide range of commonly used algorithms with sophisticated visualization tools. Compared with existing scRNA-seq analysis platforms, researchers (including those lacking computational expertise) are able to interact with the data in a straightforward fashion and in real time. Furthermore, given the overlap between scRNA-seq and bulk RNA-seq analysis workflows, ASAP should conceptually be broadly applicable to any RNA-seq dataset. As a validation, we demonstrate how we can use ASAP to simply reproduce the results from a single-cell study of 91 mouse cells involving five distinct cell types. The tool is freely available at asap.epfl.ch and R/Python scripts are available at github.com/DeplanckeLab/ASAP. bart.deplancke@epfl.ch. Supplementary data are available at Bioinformatics online. © The Author(s) 2017. Published by Oxford University Press.

  13. High Class-Imbalance in pre-miRNA Prediction: A Novel Approach Based on deepSOM.

    Science.gov (United States)

    Stegmayer, Georgina; Yones, Cristian; Kamenetzky, Laura; Milone, Diego H

    2017-01-01

    The computational prediction of novel microRNA within a full genome involves identifying sequences having the highest chance of being a miRNA precursor (pre-miRNA). These sequences are usually named candidates to miRNA. The well-known pre-miRNAs are usually only a few in comparison to the hundreds of thousands of potential candidates to miRNA that have to be analyzed, which makes this task a high class-imbalance classification problem. The classical way of approaching it has been training a binary classifier in a supervised manner, using well-known pre-miRNAs as positive class and artificially defining the negative class. However, although the selection of positive labeled examples is straightforward, it is very difficult to build a set of negative examples in order to obtain a good set of training samples for a supervised method. In this work, we propose a novel and effective way of approaching this problem using machine learning, without the definition of negative examples. The proposal is based on clustering unlabeled sequences of a genome together with well-known miRNA precursors for the organism under study, which allows for the quick identification of the best candidates to miRNA as those sequences clustered with known precursors. Furthermore, we propose a deep model to overcome the problem of having very few positive class labels. They are always maintained in the deep levels as positive class while less likely pre-miRNA sequences are filtered level after level. Our approach has been compared with other methods for pre-miRNAs prediction in several species, showing effective predictivity of novel miRNAs. Additionally, we will show that our approach has a lower training time and allows for a better graphical navegability and interpretation of the results. A web-demo interface to try deepSOM is available at http://fich.unl.edu.ar/sinc/web-demo/deepsom/.

  14. Vision-Based Interest Point Extraction Evaluation in Multiple Environments

    National Research Council Canada - National Science Library

    McKeehan, Zachary D

    2008-01-01

    Computer-based vision is becoming a primary sensor mechanism in many facets of real world 2-D and 3-D applications, including autonomous robotics, augmented reality, object recognition, motion tracking, and biometrics...

  15. Effect of single base changes and the absence of modified bases in 16S RNA on the reconstitution and function of Escherichia coli 30S ribosomes

    International Nuclear Information System (INIS)

    Denman, R.; Krzyzosiak, W.; Nurse, K.; Ofengand, J.

    1987-01-01

    The gene coding for E. coli 16S rRNA was placed in pUC19 under the control of the strong class III T7 promoter, phi 10, by ligation of the 1490 bp BclI/BstEII fragment of the rrnB operon with appropriate synthetic oligodeoxynucleotides. Such constructs allowed efficient in vitro synthesis of full-length transcripts (up to 900 mol RNA/mol template) free of modified bases. The synthetic RNA could be assembled into 30S subunits upon addition of E. coli 30S ribosomal proteins. The particles co-sedimented with authentic 30S particles and were electron microscopically indistinguishable from them. Upon addition of 50S subunits, codon-dependent P-site binding of tRNA and codon-dependent polypeptide synthesis were >80% of 30S reconstituted from natural 16S RNA and >50% of isolated 30S. UV-induced crosslinking of P-site bound AcVal-tRNA to residue C 1400 was preserved. Changing C 1400 to A had little effect on reconstitution, P-site binding, or polypeptide synthesis. However, the substitution of C 1499 by G markedly inhibited assembly. The effect on P-site binding and polypeptide synthesis is under study. These results show (1) none of the modified bases of 16S RNA are essential for protein synthesis, (2) substitution of A for C 1400 has little functional effect, and (3) position 1400 may be important for ribosome assembly

  16. Sieve-based relation extraction of gene regulatory networks from biological literature.

    Science.gov (United States)

    Žitnik, Slavko; Žitnik, Marinka; Zupan, Blaž; Bajec, Marko

    2015-01-01

    Relation extraction is an essential procedure in literature mining. It focuses on extracting semantic relations between parts of text, called mentions. Biomedical literature includes an enormous amount of textual descriptions of biological entities, their interactions and results of related experiments. To extract them in an explicit, computer readable format, these relations were at first extracted manually from databases. Manual curation was later replaced with automatic or semi-automatic tools with natural language processing capabilities. The current challenge is the development of information extraction procedures that can directly infer more complex relational structures, such as gene regulatory networks. We develop a computational approach for extraction of gene regulatory networks from textual data. Our method is designed as a sieve-based system and uses linear-chain conditional random fields and rules for relation extraction. With this method we successfully extracted the sporulation gene regulation network in the bacterium Bacillus subtilis for the information extraction challenge at the BioNLP 2013 conference. To enable extraction of distant relations using first-order models, we transform the data into skip-mention sequences. We infer multiple models, each of which is able to extract different relationship types. Following the shared task, we conducted additional analysis using different system settings that resulted in reducing the reconstruction error of bacterial sporulation network from 0.73 to 0.68, measured as the slot error rate between the predicted and the reference network. We observe that all relation extraction sieves contribute to the predictive performance of the proposed approach. Also, features constructed by considering mention words and their prefixes and suffixes are the most important features for higher accuracy of extraction. Analysis of distances between different mention types in the text shows that our choice of transforming

  17. Improved External Base Resistance Extraction for Submicrometer InP/InGaAs DHBT Models

    DEFF Research Database (Denmark)

    Johansen, Tom Keinicke; Krozer, Viktor; Nodjiadjim, Virginie

    2011-01-01

    An improved direct parameter extraction method is proposed for III–V heterojunction bipolar transistor (HBT) external base resistance $R_{\\rm bx}$ extraction from forward active $S$-parameters. The method is formulated taking into account the current dependence of the intrinsic base–collector cap......An improved direct parameter extraction method is proposed for III–V heterojunction bipolar transistor (HBT) external base resistance $R_{\\rm bx}$ extraction from forward active $S$-parameters. The method is formulated taking into account the current dependence of the intrinsic base...... factor given as the ratio of the emitter to the collector area. The determination of the parameters $I_{p}$ and $X_{0}$ from experimental $S$-parameters is described. The method is applied to high-speed submicrometer InP/InGaAs DHBT devices and leads to small-signal equivalent circuit models, which...

  18. Post-processing of Deep Web Information Extraction Based on Domain Ontology

    Directory of Open Access Journals (Sweden)

    PENG, T.

    2013-11-01

    Full Text Available Many methods are utilized to extract and process query results in deep Web, which rely on the different structures of Web pages and various designing modes of databases. However, some semantic meanings and relations are ignored. So, in this paper, we present an approach for post-processing deep Web query results based on domain ontology which can utilize the semantic meanings and relations. A block identification model (BIM based on node similarity is defined to extract data blocks that are relevant to specific domain after reducing noisy nodes. Feature vector of domain books is obtained by result set extraction model (RSEM based on vector space model (VSM. RSEM, in combination with BIM, builds the domain ontology on books which can not only remove the limit of Web page structures when extracting data information, but also make use of semantic meanings of domain ontology. After extracting basic information of Web pages, a ranking algorithm is adopted to offer an ordered list of data records to users. Experimental results show that BIM and RSEM extract data blocks and build domain ontology accurately. In addition, relevant data records and basic information are extracted and ranked. The performances precision and recall show that our proposed method is feasible and efficient.

  19. RNA isolation from bloodstains collected on FTA cards - application in clinical and forensic genetics.

    Science.gov (United States)

    Skonieczna, Katarzyna; Styczyński, Jan; Krenska, Anna; Wysocki, Mariusz; Jakubowska, Aneta; Grzybowski, Tomasz

    2016-01-01

    Aim of the study: In recent years, RNA analysis has been increasingly used in clinical and forensic genetics. Nevertheless, a major limitation of RNA-based applications is very low RNA stability in biological material, due to the RNAse activity. This highlights the need for improving the methods of RNA collection and storage. Technological approaches such as FTA Classic Cards (Whatman) could provide a solution for the problem of RNA degradation. However, different methods of RNA isolation from FTA cards could have diverse effects on RNA quantity and quality. The purpose of this research was to analyze the utility of three different methods of RNA isolation from peripheral blood collected on FTA Classic Cards (Whatman). The study also aimed at assessing RNA stability in bloodstains deposited on FTA cards. Material and methods: The study was performed on peripheral bloodstains collected from 59 individuals on FTA Classic Cards (Whatman). RNA was isolated with High Pure RNA Isolation Kit (Roche Diagnostics), Universal RNA/miRNA Purification (EURx) and TRIzol Reagent (Life Technologies). RNA was subjected to quantitative analysis followed by reverse transcription and Real - Time PCR reaction. Results: The study has shown that FTA Classic Cards (Whatman) are useful tools for storing bloodstains at room temperature for RNA analysis. Moreover, the method of RNA extraction employing TRIzol Reagent (Life Technologies) provides the highest efficiency and reproducibility for samples stored for no more than 2 years. Conclusions: The FTA cards are suitable for collecting and storing bloodstains for RNA analysis in clinical and forensic genetics.

  20. 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.

  1. Fault Features Extraction and Identification based Rolling Bearing Fault Diagnosis

    International Nuclear Information System (INIS)

    Qin, B; Sun, G D; Zhang L Y; Wang J G; HU, J

    2017-01-01

    For the fault classification model based on extreme learning machine (ELM), the diagnosis accuracy and stability of rolling bearing is greatly influenced by a critical parameter, which is the number of nodes in hidden layer of ELM. An adaptive adjustment strategy is proposed based on vibrational mode decomposition, permutation entropy, and nuclear kernel extreme learning machine to determine the tunable parameter. First, the vibration signals are measured and then decomposed into different fault feature models based on variation mode decomposition. Then, fault feature of each model is formed to a high dimensional feature vector set based on permutation entropy. Second, the ELM output function is expressed by the inner product of Gauss kernel function to adaptively determine the number of hidden layer nodes. Finally, the high dimension feature vector set is used as the input to establish the kernel ELM rolling bearing fault classification model, and the classification and identification of different fault states of rolling bearings are carried out. In comparison with the fault classification methods based on support vector machine and ELM, the experimental results show that the proposed method has higher classification accuracy and better generalization ability. (paper)

  2. Optimization of Critical Hairpin Features Allows miRNA-based Gene Knockdown Upon Single-copy Transduction

    Directory of Open Access Journals (Sweden)

    Renier Myburgh

    2014-01-01

    Full Text Available Gene knockdown using micro RNA (miRNA-based vector constructs is likely to become a prominent gene therapy approach. It was the aim of this study to improve the efficiency of gene knockdown through optimizing the structure of miRNA mimics. Knockdown of two target genes was analyzed: CCR5 and green fluorescent protein. We describe here a novel and optimized miRNA mimic design called mirGE comprising a lower stem length of 13 base pairs (bp, positioning of the targeting strand on the 5′ side of the miRNA, together with nucleotide mismatches in upper stem positions 1 and 12 placed on the passenger strand. Our mirGE proved superior to miR-30 in four aspects: yield of targeting strand incorporation into RNA-induced silencing complex (RISC; incorporation into RISC of correct targeting strand; precision of cleavage by Drosha; and ratio of targeting strand over passenger strand. A triple mirGE hairpin cassette targeting CCR5 was constructed. It allowed CCR5 knockdown with an efficiency of over 90% upon single-copy transduction. Importantly, single-copy expression of this construct rendered transduced target cells, including primary human macrophages, resistant to infection with a CCR5-tropic strain of HIV. Our results provide new insights for a better knockdown efficiency of constructs containing miRNA. Our results also provide the proof-of-principle that cells can be rendered HIV resistant through single-copy vector transduction, rendering this approach more compatible with clinical applications.

  3. Quaternary ammonium based task specific ionic liquid for the efficient and selective extraction of neptunium

    Energy Technology Data Exchange (ETDEWEB)

    Gupta, Nishesh Kumar [National Institute of Technology, Odisha (India). Dept. of Chemistry; Sengupta, Arijit [Bhabha Atomic Research Centre, Mumbai (India). Radiochemistry Div.; Biswas, Sujoy [Bhabha Atomic Research Centre, Mumbai (India). Uranium Extraction Div.

    2017-07-01

    Liquid-liquid extraction of neptunium from aqueous acidic solution using quaternary ammonium based task specific ionic liquid (TSIL) was investigated. The extraction of Np was predominated by the 'cation exchange' mechanism via [NpO{sub 2}.Hpth]{sup +} species for NpO{sub 2}{sup 2+}, while NpO{sub 2}{sup +} was extracted in ionic liquid as [NpO{sub 2}.H.Hpth]{sup +}. The extraction process was thermodynamically spontaneous while kinetically slower. Na{sub 2}CO{sub 3} as strippant showed quantitative back extraction of neptunium ions from TSIL. TSIL showed excellent radiolytic stability upto 500 kGy gamma exposure. Finally, the TSIL was employed for the processing of simulated high level waste solutions revealing high selectivity of TSIL towards neptunium.

  4. Optimization of PEG-based extraction of polysaccharides from Dendrobium nobile Lindl. and bioactivity study.

    Science.gov (United States)

    Zhang, Yi; Wang, Hongxin; Wang, Peng; Ma, ChaoYang; He, GuoHua; Rahman, Md Ramim Tanver

    2016-11-01

    Polyethylene glycol (PEG) as a green solvent was employed to extract polysaccharide. The optimal conditions for PEG-based ultrasonic extraction of Dendrobium nobile Lindl. polysaccharide (JCP) were determined by response surface methodology. Under the optimal conditions: extraction temperature of 58.5°C; ultrasound power of 193W, and the concentration of polyethylene glycol-200 (PEG-200) solution of 45%, the highest JCP yield was obtained as 15.23±0.57%, which was close to the predicted yield, 15.57%. UV and FT-IR analysis revealed the general characteristic absorption peaks of both JCP with water extraction (JCP w ) and PEG-200 solvent extraction (JCP p ). Thermal analysis of both JCPs was performed with Thermal Gravimetric Analyzer (TGA) and Differential Scanning Calorimeter (DSC). Antioxidant activities of two polysaccharides were also compared and no significant difference in vitro was obtained. Copyright © 2016 Elsevier B.V. All rights reserved.

  5. siMacro: A Fast and Easy Data Processing Tool for Cell-Based Genomewide siRNA Screens

    Directory of Open Access Journals (Sweden)

    Nitin Kumar Singh

    2013-03-01

    Full Text Available Growing numbers of studies employ cell line-based systematic short interfering RNA (siRNA screens to study gene functions and to identify drug targets. As multiple sources of variations that are unique to siRNA screens exist, there is a growing demand for a computational tool that generates normalized values and standardized scores. However, only a few tools have been available so far with limited usability. Here, we present siMacro, a fast and easy-to-use Microsoft Office Excel-based tool with a graphic user interface, designed to process single-condition or two-condition synthetic screen datasets. siMacro normalizes position and batch effects, censors outlier samples, and calculates Z-scores and robust Z-scores, with a spreadsheet output of >120,000 samples in under 1 minute.

  6. Andrographolide antagonizes cigarette smoke extract-induced inflammatory response and oxidative stress in human alveolar epithelial A549 cells through induction of microRNA-218.

    Science.gov (United States)

    Li, Ying-jie; Yu, Chang-hai; Li, Jing-bo; Wu, Xi-ya

    2013-12-01

    Andrographolide is a major bioactive labdane diterpenoid isolated from Andrographis paniculata and has protective effects against cigarette smoke (CS)-induced lung injury. This study was done to determine whether such protective effects were mediated through modulation of microRNA (miR)-218 expression. Therefore, we exposed human alveolar epithelial A549 cells to cigarette smoke extract (CSE) with or without andrographolide pretreatment and measured the level of glutathione, nuclear factor-kappaB (NF-κB) activation, proinflammatory cytokine production, and miR-218 expression. We found that andrographolide pretreatment significantly restored the glutathione level in CSE-exposed A549 cells, coupled with reduced inhibitor κB (IκB)-α phosphorylation and p65 nuclear translocation and interleukin (IL)-8 and IL-6 secretion. The miR-218 expression was significantly upregulated by andrographolide pretreatment. To determine the biological role of miR-218, we overexpressed and downregulated its expression using miR-218 mimic and anti-miR-218 inhibitor, respectively. We observed that miR-218 overexpression led to a marked reduction in IκB-α phosphorylation, p65 nuclear accumulation, and NF-κB-dependent transcriptional activity in CSE-treated A549 cells. In contrast, miR-218 silencing enhanced IκB-α phosphorylation and p65 nuclear accumulation in cells with andrographolide pretreatment and reversed andrographolide-mediated reduction of IL-6 and IL-8 production. In addition, depletion of miR-218 significantly reversed the upregulation of glutathione levels in A549 cells by andrographolide. Taken together, our results demonstrate that andrographolide mitigates CSE-induced inflammatory response in A549 cells, largely through inhibition of NF-κB activation via upregulation of miR-218, and thus has preventive benefits in CS-induced inflammatory lung diseases.

  7. A Robust Gradient Based Method for Building Extraction from LiDAR and Photogrammetric Imagery

    Directory of Open Access Journals (Sweden)

    Fasahat Ullah Siddiqui

    2016-07-01

    Full Text Available Existing automatic building extraction methods are not effective in extracting buildings which are small in size and have transparent roofs. The application of large area threshold prohibits detection of small buildings and the use of ground points in generating the building mask prevents detection of transparent buildings. In addition, the existing methods use numerous parameters to extract buildings in complex environments, e.g., hilly area and high vegetation. However, the empirical tuning of large number of parameters reduces the robustness of building extraction methods. This paper proposes a novel Gradient-based Building Extraction (GBE method to address these limitations. The proposed method transforms the Light Detection And Ranging (LiDAR height information into intensity image without interpolation of point heights and then analyses the gradient information in the image. Generally, building roof planes have a constant height change along the slope of a roof plane whereas trees have a random height change. With such an analysis, buildings of a greater range of sizes with a transparent or opaque roof can be extracted. In addition, a local colour matching approach is introduced as a post-processing stage to eliminate trees. This stage of our proposed method does not require any manual setting and all parameters are set automatically from the data. The other post processing stages including variance, point density and shadow elimination are also applied to verify the extracted buildings, where comparatively fewer empirically set parameters are used. The performance of the proposed GBE method is evaluated on two benchmark data sets by using the object and pixel based metrics (completeness, correctness and quality. Our experimental results show the effectiveness of the proposed method in eliminating trees, extracting buildings of all sizes, and extracting buildings with and without transparent roof. When compared with current state

  8. Ionic liquid-based microwave-assisted extraction of rutin from Chinese medicinal plants.

    Science.gov (United States)

    Zeng, Huan; Wang, Yuzhi; Kong, Jinhuan; Nie, Chan; Yuan, Ya

    2010-12-15

    An ionic liquid-based microwave-assisted extraction (ILMAE) method has been developed for the effective extraction of rutin from Chinese medicinal plants including Saururus chinensis (Lour.) Bail. (S. chinensis) and Flos Sophorae. A series of 1-butyl-3-methylimidazolium ionic liquids with different anions were investigated. The results indicated that the characteristics of anions have remarkable effects on the extraction efficiency of rutin and among the investigated ionic liquids, 1-butyl-3-methylimidazolium bromide ([bmim]Br) aqueous solution was the best. In addition, the ILMAE procedures for the two kinds of medicinal herbs were also optimized by means of a series of single factor experiments and an L(9) (3(4)) orthogonal design. Compared with the optimal ionic liquid-based heating extraction (ILHE), marinated extraction (ILME), ultrasonic-assisted extraction (ILUAE), the optimized approach of ILMAE gained higher extraction efficiency which is 4.879 mg/g in S. chinensis with RSD 1.33% and 171.82 mg/g in Flos Sophorae with RSD 1.47% within the shortest extraction time. Reversed phase high performance liquid chromatography (RP-HPLC) with ultraviolet detection was employed for the analysis of rutin in Chinese medicinal plants. Under the optimum conditions, the average recoveries of rutin from S. chinensis and Flos Sophorae were 101.23% and 99.62% with RSD lower than 3%, respectively. The developed approach is linear at concentrations from 42 to 252 mg L(-1) of rutin solution, with the regression coefficient (r) at 0.99917. Moreover, the extraction mechanism of ILMAE and the microstructures and chemical structures of the two researched samples before and after extraction were also investigated. With the help of LC-MS, it was future demonstrated that the two researched herbs do contain active ingredient of rutin and ionic liquids would not influence the structure of rutin. Copyright © 2010 Elsevier B.V. All rights reserved.

  9. RNA-Seq-Based Transcript Structure Analysis with TrBorderExt.

    Science.gov (United States)

    Wang, Yejun; Sun, Ming-An; White, Aaron P

    2018-01-01

    RNA-Seq has become a routine strategy for genome-wide gene expression comparisons in bacteria. Despite lower resolution in transcript border parsing compared with dRNA-Seq, TSS-EMOTE, Cappable-seq, Term-seq, and others, directional RNA-Seq still illustrates its advantages: low cost, quantification and transcript border analysis with a medium resolution (±10-20 nt). To facilitate mining of directional RNA-Seq datasets especially with respect to transcript structure analysis, we developed a tool, TrBorderExt, which can parse transcript start sites and termination sites accurately in bacteria. A detailed protocol is described in this chapter for how to use the software package step by step to identify bacterial transcript borders from raw RNA-Seq data. The package was developed with Perl and R programming languages, and is accessible freely through the website: http://www.szu-bioinf.org/TrBorderExt .

  10. Characterization of RNA from Exosomes and Other Extracellular Vesicles Isolated by a Novel Spin Column-Based Method

    Science.gov (United States)

    Enderle, Daniel; Spiel, Alexandra; Coticchia, Christine M.; Berghoff, Emily; Mueller, Romy; Schlumpberger, Martin; Sprenger-Haussels, Markus; Shaffer, Jonathan M.; Lader, Eric; Skog, Johan; Noerholm, Mikkel

    2015-01-01

    Exosomes and other extracellular vesicles (commonly referred to as EVs) have generated a lot of attention for their potential applications in both diagnostics and therapeutics. The contents of these vesicles are the subject of intense research, and the relatively recent discovery of RNA inside EVs has raised interest in the biological function of these RNAs as well as their potential as biomarkers for cancer and other diseases. Traditional ultracentrifugation-based protocols to isolate EVs are labor-intensive and subject to significant variability. Various attempts to develop methods with robust, reproducible performance have not yet been completely successful. Here, we report the development and characterization of a spin column-based method for the isolation of total RNA from EVs in serum and plasma. This method isolates highly pure RNA of equal or higher quantity compared to ultracentrifugation, with high specificity for vesicular over non-vesicular RNA. The spin columns have a capacity to handle up to 4 mL sample volume, enabling detection of low-abundance transcripts in serum and plasma. We conclude that the method is an improvement over traditional methods in providing a faster, more standardized way to achieve reliable high quality RNA preparations from EVs in biofluids such as serum and plasma. The first kit utilizing this new method has recently been made available by Qiagen as “exoRNeasy Serum/Plasma Maxi Kit”. PMID:26317354

  11. EMSAR: estimation of transcript abundance from RNA-seq data by mappability-based segmentation and reclustering.

    Science.gov (United States)

    Lee, Soohyun; Seo, Chae Hwa; Alver, Burak Han; Lee, Sanghyuk; Park, Peter J

    2015-09-03

    RNA-seq has been widely used for genome-wide expression profiling. RNA-seq data typically consists of tens of millions of short sequenced reads from different transcripts. However, due to sequence similarity among genes and among isoforms, the source of a given read is often ambiguous. Existing approaches for estimating expression levels from RNA-seq reads tend to compromise between accuracy and computational cost. We introduce a new approach for quantifying transcript abundance from RNA-seq data. EMSAR (Estimation by Mappability-based Segmentation And Reclustering) groups reads according to the set of transcripts to which they are mapped and finds maximum likelihood estimates using a joint Poisson model for each optimal set of segments of transcripts. The method uses nearly all mapped reads, including those mapped to multiple genes. With an efficient transcriptome indexing based on modified suffix arrays, EMSAR minimizes the use of CPU time and memory while achieving accuracy comparable to the best existing methods. EMSAR is a method for quantifying transcripts from RNA-seq data with high accuracy and low computational cost. EMSAR is available at https://github.com/parklab/emsar.

  12. A PCR-Based Method to Construct Lentiviral Vector Expressing Double Tough Decoy for miRNA Inhibition.

    Directory of Open Access Journals (Sweden)

    Huiling Qiu

    Full Text Available DNA vector-encoded Tough Decoy (TuD miRNA inhibitor is attracting increased attention due to its high efficiency in miRNA suppression. The current methods used to construct TuD vectors are based on synthesizing long oligonucleotides (~90 mer, which have been costly and problematic because of mutations during synthesis. In this study, we report a PCR-based method for the generation of double Tough Decoy (dTuD vector in which only two sets of shorter oligonucleotides (< 60 mer were used. Different approaches were employed to test the inhibitory potency of dTuDs. We demonstrated that dTuD is the most efficient method in miRNA inhibition in vitro and in vivo. Using this method, a mini dTuD library against 88 human miRNAs was constructed and used for a high-throughput screening (HTS of AP-1 pathway-related miRNAs. Seven miRNAs (miR-18b-5p, -101-3p, -148b-3p, -130b-3p, -186-3p, -187-3p and -1324 were identified as candidates involved in AP-1 pathway regulation. This novel method allows for an accurate and cost-effective generation of dTuD miRNA inhibitor, providing a powerful tool for efficient miRNA suppression in vitro and in vivo.

  13. Multigenic lentiviral vectors for combined and tissue-specific expression of miRNA- and protein-based antiangiogenic factors

    Directory of Open Access Journals (Sweden)

    Anne Louise Askou

    Full Text Available Lentivirus-based gene delivery vectors carrying multiple gene cassettes are powerful tools in gene transfer studies and gene therapy, allowing coexpression of multiple therapeutic factors and, if desired, fluorescent reporters. Current strategies to express transgenes and microRNA (miRNA clusters from a single vector have certain limitations that affect transgene expression levels and/or vector titers. In this study, we describe a novel vector design that facilitates combined expression of therapeutic RNA- and protein-based antiangiogenic factors as well as a fluorescent reporter from back-to-back RNApolII-driven expression cassettes. This configuration allows effective production of intron-embedded miRNAs that are released upon transduction of target cells. Exploiting such multigenic lentiviral vectors, we demonstrate robust miRNA-directed downregulation of vascular endothelial growth factor (VEGF expression, leading to reduced angiogenesis, and parallel impairment of angiogenic pathways by codelivering the gene encoding pigment epithelium-derived factor (PEDF. Notably, subretinal injections of lentiviral vectors reveal efficient retinal pigment epithelium-specific gene expression driven by the VMD2 promoter, verifying that multigenic lentiviral vectors can be produced with high titers sufficient for in vivo applications. Altogether, our results suggest the potential applicability of combined miRNA- and protein-encoding lentiviral vectors in antiangiogenic gene therapy, including new combination therapies for amelioration of age-related macular degeneration.

  14. Transgenic Sugarcane Resistant to Sorghum mosaic virus Based on Coat Protein Gene Silencing by RNA Interference

    Directory of Open Access Journals (Sweden)

    Jinlong Guo

    2015-01-01

    Full Text Available As one of the critical diseases of sugarcane, sugarcane mosaic disease can lead to serious decline in stalk yield and sucrose content. It is mainly caused by Potyvirus sugarcane mosaic virus (SCMV and/or Sorghum mosaic virus (SrMV, with additional differences in viral strains. RNA interference (RNAi is a novel strategy for producing viral resistant plants. In this study, based on multiple sequence alignment conducted on genomic sequences of different strains and isolates of SrMV, the conserved region of coat protein (CP genes was selected as the target gene and the interference sequence with size of 423 bp in length was obtained through PCR amplification. The RNAi vector pGII00-HACP with an expression cassette containing both hairpin interference sequence and cp4-epsps herbicide-tolerant gene was transferred to sugarcane cultivar ROC22 via Agrobacterium-mediated transformation. After herbicide screening, PCR molecular identification, and artificial inoculation challenge, anti-SrMV positive transgenic lines were successfully obtained. SrMV resistance rate of the transgenic lines with the interference sequence was 87.5% based on SrMV challenge by artificial inoculation. The genetically modified SrMV-resistant lines of cultivar ROC22 provide resistant germplasm for breeding lines and can also serve as resistant lines having the same genetic background for study of resistance mechanisms.

  15. RNA interference-based therapeutics: new strategies to fight infectious disease.

    Science.gov (United States)

    López-Fraga, M; Wright, N; Jiménez, A

    2008-12-01

    For many years, there has been an ongoing search for new compounds that can selectively alter gene expression as a new way to treat human disease by addressing targets that are otherwise "undruggable" with traditional pharmaceutical approaches involving small molecules or proteins. RNA interference (RNAi) strategies have raised a lot of attention and several compounds are currently being tested in clinical trials. Viruses are the obvious target for RNAi-therapy, as most are difficult to treat with conventional drugs, they become rapidly resistant to drug treatment and their genes differ substantially from human genes, minimizing side effects. Antisense strategy offers very high target specificity, i.e., any viral sequence could potentially be targeted using the complementary oligonucleotide sequence. Consequently, new antisense-based therapeutics have the potential to lead a revolution in the anti-infective drug development field. Additionally, the relatively short turnaround for efficacy testing of potential RNAi molecules and that any pathogen is theoretically amenable to rapid targeting, make them invaluable tools for treating a wide range of diseases. This review will focus on some of the current efforts to treat infectious disease with RNAi-based therapies and some of the obstacles that have appeared on the road to successful clinical intervention.

  16. Toxicity Thresholds Based on EDTA Extractable Nickel and Barley Root Elongation in Chinese Soils

    Directory of Open Access Journals (Sweden)

    Guangyun Zhu

    2018-04-01

    Full Text Available The uncertainty in the risk assessment of trace metal elements in soils when total metal contents are used can be decreased by assessing their availability and/or extractability when the soils have a high background value or different sources of trace metal elements. In this study, the added water-soluble nickel (Ni toxicity to barley root elongation was studied in 17 representative Chinese soil samples with and without artificial rainwater leaching. The extractability of added Ni in soils was estimated by three sequential extractions with ethylenediaminetetraacetic acid (EDTA. The results showed that the effective concentration of EDTA extractable Ni (EC50, which caused 50% inhibition of barley root elongation, ranged from 46 to 1019 mg/kg in unleached soils and 24 to 1563 mg/kg in leached soils. Regression models for EDTA extractable Ni and total Ni added to soils against soil properties indicated that EDTA extractable Ni was significantly correlated with the total Ni added to soils and that pH was the most important control factor. Regression models for toxicity thresholds based on EDTA extractable Ni against soil properties showed that soil citrate dithionate extractable Fe was more important than soil pH in predicting Ni toxicity. These results can be used to accurately assess the risk of contaminated soils with high background values and/or different Ni sources.

  17. Extraction optimization and pixel-based chemometric analysis of semi-volatile organic compounds in groundwater

    DEFF Research Database (Denmark)

    Christensen, Peter; Tomasi, Giorgio; Kristensen, Mette

    2017-01-01

    . In this study, we tested the combination of solid phase extraction (SPE) with dispersive liquid-liquid micro extraction (DLLME), or with stir bar sorptive extraction (SBSE), as an extraction method for semi-VOCs in groundwater. Combining SPE with DLLME or SBSE resulted in better separation of peaks...... in an unresolved complex mixture. SPE-DLLME was chosen as the preferred extraction method. SPE-DLLME covered a larger polarity range (logKo/w 2.0-11.2), had higher extraction efficiency at logKo/w 2.0-3.8 and 5.8-11.2, and was faster compared to SPE-SBSE. SPE-DLLME extraction combined with chemical analysis by gas...... chromatography-mass spectrometry (GC-MS) and pixel-based data analysis of summed extraction ion chromatograms (sEICs) was tested as a new method for chemical fingerprinting of semi-VOCs in 15 groundwater samples. The results demonstrate that SPE-DLLME-GC-MS provides an excellent compromise between compound...

  18. Extraction or adsorption? Voltammetric assessment of protamine transfer at ionophore-based polymeric membranes.

    Science.gov (United States)

    Garada, Mohammed B; Kabagambe, Benjamin; Amemiya, Shigeru

    2015-01-01

    Cation-exchange extraction of polypeptide protamine from water into an ionophore-based polymeric membrane has been hypothesized as the origin of a potentiometric sensor response to this important heparin antidote. Here, we apply ion-transfer voltammetry not only to confirm protamine extraction into ionophore-doped polymeric membranes but also to reveal protamine adsorption at the membrane/water interface. Protamine adsorption is thermodynamically more favorable than protamine extraction as shown by cyclic voltammetry at plasticized poly(vinyl chloride) membranes containing dinonylnaphthalenesulfonate as a protamine-selective ionophore. Reversible adsorption of protamine at low concentrations down to 0.038 μg/mL is demonstrated by stripping voltammetry. Adsorptive preconcentration of protamine at the membrane/water interface is quantitatively modeled by using the Frumkin adsorption isotherm. We apply this model to ensure that stripping voltammograms are based on desorption of all protamine molecules that are transferred across the interface during a preconcentration step. In comparison to adsorption, voltammetric extraction of protamine requires ∼0.2 V more negative potentials, where a potentiometric super-Nernstian response to protamine is also observed. This agreement confirms that the potentiometric protamine response is based on protamine extraction. The voltammetrically reversible protamine extraction results in an apparently irreversible potentiometric response to protamine because back-extraction of protamine from the membrane extremely slows down at the mixed potential based on cation-exchange extraction of protamine. Significantly, this study demonstrates the advantages of ion-transfer voltammetry over potentiometry to quantitatively and mechanistically assess protamine transfer at ionophore-based polymeric membranes as foundation for reversible, selective, and sensitive detection of protamine.

  19. Automated identification of RNA 3D modules with discriminative power in RNA structural alignments

    DEFF Research Database (Denmark)

    Theis, Corinna; Höner zu Siederdissen, Christian; Hofacker, Ivo L.

    2013-01-01

    Recent progress in predicting RNA structure is moving towards filling the 'gap' in 2D RNA structure prediction where, for example, predicted internal loops often form non-canonical base pairs. This is increasingly recognized with the steady increase of known RNA 3D modules. There is a general...... comparative evidence. Subsequently, the modules, initially represented by a graph, are turned into models for the RMDetect program, which allows to test their discriminative power using real and randomized Rfam alignments. An initial extraction of 22495 3D modules in all PDB files results in 977 internal loop...

  20. Biosensor method and system based on feature vector extraction

    Science.gov (United States)

    Greenbaum, Elias [Knoxville, TN; Rodriguez, Jr., Miguel; Qi, Hairong [Knoxville, TN; Wang, Xiaoling [San Jose, CA

    2012-04-17

    A method of biosensor-based detection of toxins comprises the steps of providing at least one time-dependent control signal generated by a biosensor in a gas or liquid medium, and obtaining a time-dependent biosensor signal from the biosensor in the gas or liquid medium to be monitored or analyzed for the presence of one or more toxins selected from chemical, biological or radiological agents. The time-dependent biosensor signal is processed to obtain a plurality of feature vectors using at least one of amplitude statistics and a time-frequency analysis. At least one parameter relating to toxicity of the gas or liquid medium is then determined from the feature vectors based on reference to the control signal.

  1. Patent Keyword Extraction Algorithm Based on Distributed Representation for Patent Classification

    Directory of Open Access Journals (Sweden)

    Jie Hu

    2018-02-01

    Full Text Available Many text mining tasks such as text retrieval, text summarization, and text comparisons depend on the extraction of representative keywords from the main text. Most existing keyword extraction algorithms are based on discrete bag-of-words type of word representation of the text. In this paper, we propose a patent keyword extraction algorithm (PKEA based on the distributed Skip-gram model for patent classification. We also develop a set of quantitative performance measures for keyword extraction evaluation based on information gain and cross-validation, based on Support Vector Machine (SVM classification, which are valuable when human-annotated keywords are not available. We used a standard benchmark dataset and a homemade patent dataset to evaluate the performance of PKEA. Our patent dataset includes 2500 patents from five distinct technological fields related to autonomous cars (GPS systems, lidar systems, object recognition systems, radar systems, and vehicle control systems. We compared our method with Frequency, Term Frequency-Inverse Document Frequency (TF-IDF, TextRank and Rapid Automatic Keyword Extraction (RAKE. The experimental results show that our proposed algorithm provides a promising way to extract keywords from patent texts for patent classification.

  2. Pyridinium ionic liquid-based liquid-solid extraction of inorganic and organic iodine from Laminaria.

    Science.gov (United States)

    Peng, Li-Qing; Yu, Wen-Yan; Xu, Jing-Jing; Cao, Jun

    2018-01-15

    A simple, green and effective extraction method, namely, pyridinium ionic liquid- (IL) based liquid-solid extraction (LSE), was first designed to extract the main inorganic and organic iodine compounds (I - , monoiodo-tyrosine (MIT) and diiodo-tyrosine (DIT)). The optimal extraction conditions were as follows: ultrasonic intensity 100W, IL ([EPy]Br) concentration 200mM, extraction time 30min, liquid/solid ratio 10mL/g, and pH value 6.5. The morphologies of Laminaria were studied by scanning electron microscopy and transmission electron microscopy. The recovery values of I - , MIT and DIT from Laminaria were in the range of 88% to 94%, and limits of detection were in the range of 59.40 to 283.6ng/g. The proposed method was applied to the extraction and determination of iodine compounds in three Laminaria. The results showed that IL-based LSE could be a promising method for rapid extraction of bioactive iodine from complex food matrices. Copyright © 2017 Elsevier Ltd. All rights reserved.

  3. Feature Extraction from 3D Point Cloud Data Based on Discrete Curves

    Directory of Open Access Journals (Sweden)

    Yi An

    2013-01-01

    Full Text Available Reliable feature extraction from 3D point cloud data is an important problem in many application domains, such as reverse engineering, object recognition, industrial inspection, and autonomous navigation. In this paper, a novel method is proposed for extracting the geometric features from 3D point cloud data based on discrete curves. We extract the discrete curves from 3D point cloud data and research the behaviors of chord lengths, angle variations, and principal curvatures at the geometric features in the discrete curves. Then, the corresponding similarity indicators are defined. Based on the similarity indicators, the geometric features can be extracted from the discrete curves, which are also the geometric features of 3D point cloud data. The threshold values of the similarity indicators are taken from [0,1], which characterize the relative relationship and make the threshold setting easier and more reasonable. The experimental results demonstrate that the proposed method is efficient and reliable.

  4. Extraction of Plant-based Capsules for Microencapsulation Applications.

    Science.gov (United States)

    Potroz, Michael G; Mundargi, Raghavendra C; Park, Jae Hyeon; Tan, Ee-Lin; Cho, Nam-Joon

    2016-11-09

    Microcapsules derived from plant-based spores or pollen provide a robust platform for a diverse range of microencapsulation applications. Sporopollenin exine capsules (SECs) are obtained when spores or pollen are processed so as to remove the internal sporoplasmic contents. The resulting hollow microcapsules exhibit a high degree of micromeritic uniformity and retain intricate microstructural features related to the particular plant species. Herein, we demonstrate a streamlined process for the production of SECs from Lycopodium clavatum spores and for the loading of hydrophilic compounds into these SECs. The current SEC isolation procedure has been recently optimized to significantly reduce the processing requirements which are conventionally used in SEC isolation, and to ensure the production of intact microcapsules. Natural L. clavatum spores are defatted with acetone, treated with phosphoric acid, and extensively washed to remove sporoplasmic contents. After acetone defatting, a single processing step using 85% phosphoric acid has been shown to remove all sporoplasmic contents. By limiting the acid processing time to 30 hr, it is possible to isolate clean SECs and avoid SEC fracturing, which has been shown to occur with prolonged processing time. Extensive washing with water, dilute acids, dilute bases, and solvents ensures that all sporoplasmic material and chemical residues are adequately removed. The vacuum loading technique is utilized to load a model protein (Bovine Serum Albumin) as a representative hydrophilic compound. Vacuum loading provides a simple technique to load various compounds without the need for harsh solvents or undesirable chemicals which are often required in other microencapsulation protocols. Based on these isolation and loading protocols, SECs provide a promising material for use in a diverse range of microencapsulation applications, such as, therapeutics, foods, cosmetics, and personal care products.

  5. Single channel blind source separation based on ICA feature extraction

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    A new technique is proposed to solve the blind source separation (BSS) given only a single channel observation. The basis functions and the density of the coefficients of source signals learned by ICA are used as the prior knowledge. Based on the learned prior information the learning rules of single channel BSS are presented by maximizing the joint log likelihood of the mixed sources to obtain source signals from single observation,in which the posterior density of the given measurements is maximized. The experimental results exhibit a successful separation performance for mixtures of speech and music signals.

  6. A Small RNA-Based Immune System Defends Germ Cells against Mobile Genetic Elements

    Directory of Open Access Journals (Sweden)

    Astrid D. Haase

    2016-01-01

    Full Text Available Transposons are mobile genetic elements that threaten the survival of species by destabilizing the germline genomes. Limiting the spread of these selfish elements is imperative. Germ cells employ specialized small regulatory RNA pathways to restrain transposon activity. PIWI proteins and Piwi-interacting RNAs (piRNAs silence transposons at the transcriptional and posttranscriptional level with loss-of-function mutant animals universally exhibiting sterility often associated with germ cell defects. This short review aims to illustrate basic strategies of piRNA-guided defense against transposons. Mechanisms of piRNA silencing are most readily studied in Drosophila melanogaster, which serves as a model to delineate molecular concepts and as a reference for mammalian piRNA systems. PiRNA pathways utilize two major strategies to handle the challenges of transposon control: (1 the hard-wired molecular memory of prior transpositions enables recognition of mobile genetic elements and discriminates transposons from host genes; (2 a feed-forward adaptation mechanism shapes piRNA populations to selectively combat the immediate threat of transposon transcripts. In flies, maternally contributed PIWI-piRNA complexes bolster both of these lines of defense and ensure transgenerational immunity. While recent studies have provided a conceptual framework of what could be viewed as an ancient immune system, we are just beginning to appreciate its many molecular innovations.

  7. Wavelet-Based Feature Extraction in Fault Diagnosis for Biquad High-Pass Filter Circuit

    OpenAIRE

    Yuehai Wang; Yongzheng Yan; Qinyong Wang

    2016-01-01

    Fault diagnosis for analog circuit has become a prominent factor in improving the reliability of integrated circuit due to its irreplaceability in modern integrated circuits. In fact fault diagnosis based on intelligent algorithms has become a popular research topic as efficient feature extraction and selection are a critical and intricate task in analog fault diagnosis. Further, it is extremely important to propose some general guidelines for the optimal feature extraction and selection. In ...

  8. All-in-one detector of circulating mRNA based on a smartphone

    Science.gov (United States)

    Cmiel, Vratislav; Gumulec, Jaromir; Svoboda, Ondrej; Raudenska, Martina; Hudcova, Kristyna; Sekora, Jiri; Balogh, Jaroslav; Masarik, Michal; Provaznik, Ivo

    2016-03-01

    Metallothionein is significantly elevated in various tumors, notably in prostate cancer on both mRNA and protein level. We demonstrated a strong predictive potential of free circulating metallothionein 2A isoform mRNA for patients with this cancer. Circulating mRNA detection relies on expensive equipment and requires high level of expertise. In this work we developed compact "all-in-one" laboratory system which replace microvolume spectrophotometer, thermocycler and realtime PCR machines. We managed to design and construct a microprocessor controlled heating/cooling chamber that ensures required temperature gradient. The chamber includes implemented optical system to enable fluorescence excitation and fluorescence analysis using a smart-phone.

  9. On the conformational stability of the smallest RNA kissing complexes maintained through two G·C base pairs

    International Nuclear Information System (INIS)

    Chu, Wally; Weerasekera, Akila; Kim, Chul-Hyun

    2017-01-01

    Two identical 5′GACG3′ tetra-loop motifs with different stem sequences (called H2 and H3) are found in the 5′ end region of Moloney Murine Leukemia Virus (MMLV) genomic RNA. They play important roles in RNA dimerization and encapsidation through two identical tetra-loops (5′GACG3′) forming a loop-to-loop kissing complex, the smallest RNA kissing complex ever found in nature. We examined the effects of a loop-closing base pair as well as a stem sequence on the conformational stability of the kissing complex. UV melting analysis and gel electrophoresis were performed on eight RNA sequences mimicking the H2 and H3 hairpin tetra-loops with variation in loop-closing base pairs. Our results show that changing the loop-closing base pair from the wildtype (5′A·U3′ for H3, 5′U·A3′ for H2) to 5′G·C3’/5′C·G3′ has significant effect on the stability of the kissing complexes: the substitution to 5′C·G3′ significantly decreases both thermal and mechanical stability, while switching to the 5′G·C3′ significantly increases the mechanical stability only. The kissing complexes with the wildtype loop-closing base pairs (5′A·U3′ for H3 and 5′U·A3′ for H2) show different stability when attached to a different stem sequence (H2 stem vs. H3 stem). This suggests that not only the loop-closing base pair itself, but also the stem sequence, affects the conformational stability of the RNA kissing complex. - Highlights: • Thermodynamic parameters of the smallest RNA kissing interactions were measured. • The effects of loop-closing base pairs on the RNA kissing complex was investigated. • Changing the base pair to 5′CG3′ decreases the stability of the kissing complex. • Changing it to 5′GC3′ increases the mechanical resilience of the kissing complex. • Difference in its stem sequence also affects the stability of the kissing complex.

  10. Feature Fusion Based Road Extraction for HJ-1-C SAR Image

    Directory of Open Access Journals (Sweden)

    Lu Ping-ping

    2014-06-01

    Full Text Available Road network extraction in SAR images is one of the key tasks of military and civilian technologies. To solve the issues of road extraction of HJ-1-C SAR images, a road extraction algorithm is proposed based on the integration of ratio and directional information. Due to the characteristic narrow dynamic range and low signal to noise ratio of HJ-1-C SAR images, a nonlinear quantization and an image filtering method based on a multi-scale autoregressive model are proposed here. A road extraction algorithm based on information fusion, which considers ratio and direction information, is also proposed. By processing Radon transformation, main road directions can be extracted. Cross interferences can be suppressed, and the road continuity can then be improved by the main direction alignment and secondary road extraction. The HJ-1-C SAR image acquired in Wuhan, China was used to evaluate the proposed method. The experimental results show good performance with correctness (80.5% and quality (70.1% when applied to a SAR image with complex content.

  11. Support patient search on pathology reports with interactive online learning based data extraction.

    Science.gov (United States)

    Zheng, Shuai; Lu, James J; Appin, Christina; Brat, Daniel; Wang, Fusheng

    2015-01-01

    Structural reporting enables semantic understanding and prompt retrieval of clinical findings about patients. While synoptic pathology reporting provides templates for data entries, information in pathology reports remains primarily in narrative free text form. Extracting data of interest from narrative pathology reports could significantly improve the representation of the information and enable complex structured queries. However, manual extraction is tedious and error-prone, and automated tools are often constructed with a fixed training dataset and not easily adaptable. Our goal is to extract data from pathology reports to support advanced patient search with a highly adaptable semi-automated data extraction system, which can adjust and self-improve by learning from a user's interaction with minimal human effort. We have developed an online machine learning based information extraction system called IDEAL-X. With its graphical user interface, the system's data extraction engine automatically annotates values for users to review upon loading each report text. The system analyzes users' corrections regarding these annotations with online machine learning, and incrementally enhances and refines the learning model as reports are processed. The system also takes advantage of customized controlled vocabularies, which can be adaptively refined during the online learning process to further assist the data extraction. As the accuracy of automatic annotation improves overtime, the effort of human annotation is gradually reduced. After all reports are processed, a built-in query engine can be applied to conveniently define queries based on extracted structured data. We have evaluated the system with a dataset of anatomic pathology reports from 50 patients. Extracted data elements include demographical data, diagnosis, genetic marker, and procedure. The system achieves F-1 scores of around 95% for the majority of tests. Extracting data from pathology reports could enable